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Lecture Notes

Veterinary Histology

VMED 7123

Dr. Charlotte L. Ownby

Fall Semester 2004

For Lecture Notes with Color Images use go to: http://www.cvm.okstate.edu/instruction/mm_curr/histology/Hi

stologyReference/index.htm Under Course Outline, double click on Organ System of Interest

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Digestive System I - lips, tongue, salivary glands, esophagus, stomach, small and large intestines

The digestive system includes the gastrointestinal tract as well as associated organs like the pancreas and liver. Digestive System I will cover the oral cavity (lips, tongue, major salivary glands) and the gastrointestinal tract, i.e. esophagus, stomach, small and large intestines. The digestive system consists throughout most of its length of a series of tubular organs lined with specific types of epithelium to fulfill specific functions related to the digestion and absorption of nutrients from a food source and the elimination of waste products.

Organ FuFunctionnction

Lips Ingestion and fragmentation of food

Teeth Fragmentation of food

Tongue Fragmentation and swallowing

Salivary Glands Fragmentation and moistening of food; swallowing

Esophagus Passage of food from oral cavity to the stomach

Stomach Completion of fragmentation and beginning of digestion

Small Intestine - duodenum Digestion; emulsificaton of fats by enzymes from the pancreas and bile from the liver

Small Intestine - jejunum & ileum

Completion of digestion and absorption

Large Intestine- cecum Absorption of water from liquid residue

Large Intestine - colon Absorption of water from liquid residue

Large Intestine - rectum Storage of feces prior to defecation

Anus Route for defecation of feces outside the body

Oral Cavity Organs that make up the oral cavity include the lips, teeth, tongue and major salivary glands. These organs function to obtain and ingest food, fragment it into smaller particles, moisten and swallow it. Teeth will not be covered in this course.

Lips

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The lips aid in obtaining food and placing it in the mouth so that the teeth and tongue can manipulate it and begin fragmenting it. Lips are covered by a stratified squamous epithelium that is usually keratinized on the outer surface and contains many hairs whereas the epithelium on the inner surface is more moist and non-keratinized.

Tongue The tongue is a highly muscular organ used to manipulate food in the mouth and for the sense of taste. It is covered with stratified squamous epithelium that in the anterior part forms specialized structures known as papillae that are involved in the manipulation of food as well as in the sense of taste. The skeletal muscle of the tongue is unique in that it runs in three different directions allowing for a wide range of movements needed to properly manipulate foodstuffs. The types, numbers and distribution of papillae in the tongue vary greatly among species. In domestic animals there are usually five different types of papillae.

1. Filiform papillae are highly keratinized, sharply pointed and aid in mechanically breaking up food material. They are numerous in ruminants and cats where they are used in lapping milk.

2. Fungiform papillae are smooth with a rounded surface. They help manipulate the food but also have taste buds on their lateral surfaces.

3. Conical papillae are somewhat larger than fungiform papillae, are used in manipulating and breaking down ingested food. They can be distinguished from fungiform papillae by their larger size, tendency to project above other papillae and they do not have taste buds.

4. Foliate papillae, covered with non-keratinized stratified squamous epithelium, are leaf-shaped structures defined by an invagination of the mucous membrane on their sides. Many taste buds on their lateral surfaces indicate their role in gustation. They are absent in ruminants but well developed in the horse and dog.

5. Circumvallate papillae are the largest (up to 1/8 inch diameter) papillae. They are surrounded by a deep indentation of the mucous membrane and are not numerous. They do not rise above the surface of the tongue. Many taste buds are located on their sides. Serous von Ebner's glands empty into the "moat" around these papillae and help keep it free of food particles.

Salivary Glands

The salivary glands all empty their secretions into the buccal cavity. They vary as to their distance from the buccal cavity, their size and the nature of their secretory products. They can also be divided into major and minor glands. We will consider only the major salivary glands of which there are three: parotid, sublingual and submandibular. These glands all have the tubuloalveolar glandular structure and all are compound, i.e., composed of numerous secretory endpieces connected by an elaborate system of branching ducts. In general saliva is a dilute,

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hypotonic solution containing various enzymes (esp. amylase and lysozyme) and other proteins such as antibodies, glycoproteins as well as electrolytes. Saliva in the buccal cavity is the combined secretion of the numerous salivary glands, both major and minor. The secretions of salivary cells can be either of a serous type, i.e., watery and rich in enzymes and antibodies or mucous, i.e., viscid containing more glycoproteins. Individual salivary glands may contain mostly cells of the serous type, of the mucous type or a mixture of both types. The final composition of saliva at any given time depends on the proportion contributed by specific salivary glands and is determined in the major glands by the parasympathetic nervous system resulting from physical, chemical and psychological stimuli.

Salivary Gland Type of Secretory Cells Parotid Serous Sublingual Mucous Submandibular Mixed

Basic Plan of the Digestive Tube

From the esophagus to the anus, the digestive system is basically a tube very similar to other tubular organs in the body. All such tubular organs are composed of several tissue layers arranged around a lumen. In a "generic" tubular organ, these layers are as follows (from the lumen to the ablumenal layer).

• Tunica mucosa: This layer is composed of epithelium, connective tissue and muscle. These tissues can usually be found in distinct layers as follows:

lamina epithelialis mucosae: consists only of epithelium lamina propria mucosae: consists of either loose areolar or reticular

connective tissue lamina muscularis mucosae: consists of smooth muscle

• Tunica submucosa: consists of loose connective tissue, nerves, blood vessels, and glands in some organs

• Tunica muscularis: consists of at least two layers, an inner circular and an outer longitudinal with parasympathetic ganglia located between the layers

• Tunica adventitia or tunica serosa: consists of loose connective tissue

If the organ is surrounded by other tissues, this layer is called a tunica adventitia and its connective tissue blends with that of the surrounding tissues.

If the organ is suspended in the body cavity, this layer is called a tunica serosa and it is covered by a simple squamous epithelium that is called mesothelium.

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Esophagus

The esophagus connects the oral cavity with the stomach allowing and aiding in the movement of food particles to the stomach. It is a muscular tube having the layers described above for the typical tubular organ. In the esophagus the layers are specialized for the function of further fragmenting food particles.

Layers of the Esophagus

• Tunica mucosa:

o lamina epithelialis: consists of stratified squamous epithelium that can be highly folded in an empty organ; may be highly keratinized in animals that ingest hard, dry materials such as herbivores

o lamina propria: consists of loose connective tissue which often has scattered lymph nodules esp. in pigs and humans

o lamina muscularis mucosae: consists of smooth muscle; distribution and continuity is highly species variable as follows: (1) continuous in human (2) separate muscle bundles that fuse in horses, ruminants and cats, (3) absent in cervical part in dogs, (4) absent in pigs in cervical region but complete near the stomach

• Tunica submucosa: consists of loose connective tissue that is very elastic allowing for expansion when food is present; ties the overlying epithelium to the underlying muscle layers; seromucous glands present in most species and numerous in the dog but absent in horses and cats. Lymphoid nodules may be present in the pig esophagus.

• Tunica muscularis: consists of smooth and/or skeletal muscle; inner circular and outer longitudinal layers usually begin as skeletal muscle at the cervical end (voluntary control of swallowing) changing to smooth near the distal end close to the stomach; skeletal muscle throughout in ruminants and the dog.

• Tunica serosa/adventitia: consists of typical loose connective tissue that blends into the connective tissue of surrounding tissues.

Stomach

The stomach connects the esophagus to the intestines and in most species serves not only to continue the breakdown of foodstuffs via the use of digestive enzymes and acid but it also as a storage depot for food. Usually food remains in the stomach a few hours during which it is converted into a liquid material called chyme.

• Stomachs are either simple or compound, i.e., consisting of one chamber or many chambers. Simple stomachs are composed primarily of glands, that is the tunica mucosa is filled with glands.

• Ruminant stomachs are compound stomachs containing both non-glandular and glandular regions. The non-glandular regions include the reticulum, rumen and

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the omasum. The glandular region is the abomasum which has its own regions similar to those found in a simple stomach.

Regional variation in the glands of the tunica mucosa of the stomach

Not all regions of the stomach mucosa have the same histological structure. They vary as follows:

• cardia: contains many mucus-secreting glands • fundus: mostly glands secreting acid-peptic gastric juices; some mucus-secreting glands • pylorus: contains two different types of mucus-secreting glands; also contains endocrine

cells secreting gastrin

Wall of the Glandular Stomach

• Tunica mucosa: in the empty stomach, this layer is thrown into deep longitudinal folds called rugae that extend from the lamina muscularis mucosae to the lumen; in the full stomach the rugae are much reduced in size as a result of distension of the tunica mucosa to accomodate the presence of a large amount of food material

• Tunica submucosa: typical loose connective tissue contains parasympathetic ganglia located in submucosal plexuses also known as Meissner's plexuses

• Tunica muscularis: typical smooth muscle consisting of at least two layers, an inner circular layer and an outer longitudinal layer; parasympathetic ganglia located between the two muscle layers in the myenteric or Auerbach's plexus

• Tunica serosa: typical small amount of loose connective tissue with overlying simple squamous epithelium or mesothelium

Layers of the Tunica Mucosa of the Stomach - Fundic Region

• Lamina epithelialis: consists of simple columnar epithelium that forms branched, tubular glands; organized into gastric pits that open onto the lumen and gastric glands that empty into the base of the gastric pits

• Lamina propria: consists of loose areolar connective tissue that in the glandular stomach is minimal between gastric glands and difficult to see in sections; highly vascular containing many blood and lymphatic capillaries

• Stratum compactum: consists of dense connective tissue containing thick collagen fibers; located between the lamina propria and the lamina muscularis mucosae; prominent in carnivores where it probably helps prevent the perforation of the wall of the stomach by sharp objects such as bones that might be present in the lumen

• Lamina muscularis mucosae: consists of several layers of smooth muscle oriented both longitudinally and circularly; usually not very thick

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Four cell types in the gastric gland

• Surface mucous cells: line the gastric pit and secrete mucous and bicarbonate ions to protect the epithelium from digestion by gastric juice (contains HCl and pepsin) present in the stomach lumen

• Neck mucous cells: found dispersed between the parietal cells; secret a mucus that is thinner than that secreted by the surface mucous cells; mucus protects other glandular cells from action of proteases and HCl

• Parietal (oxyntic) cells: found throughout the gastric gland; round cells that contain distinct eosinophilic (pink) cytoplasm and round, prominent nucleus; secrete HCl and intrinsic factor, needed for absorption of vitamin B12.

• Chief (peptic, zymogenic) cells: found mostly near the base of the gastric glands; very basophilic (purple) containing basally positioned nucleus and prominent basophilic apical cytoplasm filled with many ribosomes; secrete pepsinogen, which is converted to pepsin in the acidic milieu of the stomach.

• Neuroendocrine cells: difficult to distinguish by conventional light microscopy; several types are present; some secrete serotonin, gastrin, glucagon, and somatostatin, among other hormones

• Stem cells: located primarily in the neck region; difficult to identify in routine H&E sections; undergo mitosis to form more cells then differentiate into the other cell types present in the gland

Parasympathetic Ganglia

Aggregations of parasympathetic ganglion cells are found throughout the digestive tube in two locations. Some are located in the submucosa and are usually called Meissner's plexus; others are located between the inner circular and outer longitundinal layers of smooth muscle in the tunica muscularis. The latter ones are usually called myenteric or Auerbach's plexus. Postganglionic fibers from Meissner's plexus innervate the lamina muscularis mucosae whereas postganglionic fibers from the myenteric plexus innervate the smooth muscle of the tunica muscularis. The two layers of smooth muscle in the tunica muscularis inherently contract in a wave of peristalsis that helps move stomach contents toward the small intestine. However, contractions of the smooth muscle are regulated by the autonomic nervous system as well as other factors such as hormones released into the stomach. An increase in peristalsis results from an increase in parasympathetic stimulation; a decrease in peristalsis results from an increase in sympathetic stimulation.

Meissner's plexus and the myenteric plexus both consist of the cell bodies of parasympathetic ganglion cells that are easily identified by their large size in comparison with other cells in the area and also by the large, round nucleus that contains a prominent nucleolus. These cell bodies are found in the midst of unmyelinated nerve fibers and near areas of myelinated axons.

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Compound stomach - found in ruminants and has four parts Chamber Histology Function

Rumen

(part of forestomach)

non-glandular; keratinized stratified squamous epithelium

mechanical and chemical breakdown of food; breakdown of food by microbes; production of volatile fatty acids; absorption of volatile fatty acids, lactic acid, ammonia, inorganic ions and water

Reticulum

(part of forestomach)

non-glandular; keratinized stratified squamous epithelium "

Omasum

(part of forestomach)

non-glandular; keratinized stratified squamous epithelium "

Abomasum glandular; simple columnar glandular epithelium enzymatic digestion

Rumen

• Tunica mucosa: characterized by the presence of long (1.0-1.5 cm long) conical projections called papillae that extend in to the lumen

o Lamina epithelialis mucosae - keratinized stratified squamous. o Lamina propria - typical; no glands o Lamina muscularis mucosae- absent; NOTE: It is easy to confuse a thickened

layer of connective tissue that extends into the papilla with a lamina muscularis mucosae but this tissue is connective tissue, not smooth muscle.

• Tunica submucosa: merges with lamina propria; no glands or lymphoid aggregates.

• Tunica muscularis: typical • Tunica serosa: typical

Reticulum

Similar to rumen, except as noted below:

• Tunica mucosa: when viewed from the lumen of the reticulum, the mucosa looks like a "honeycomb" or reticulum. The basis of this honeycomb is a series of connected vertical

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primary folds that give rise to secondary and tertiary papillae which project into the lumen.

• Lamina muscularis mucosa: a layer of smooth muscle extends from the tips of the papillae down to the position of the lamina muscularis mucosa although in the reticulum this layer is not quite typical. However, the smooth muscle in the reticulum is continuous with the smooth muscle of the lamina muscularis mucosa in the esophagus.

• Other tunics are typical

Omasum

• This part of the non-glandular region of the compound stomach is notable for the complexity of the foldings of its tunica mucosa

• These folds or laminae are covered with a highly keratinized stratified squamous epithelium

• Underlying this epithelium is the sparse loose connective tissue of the lamina propria. • The laminae muscularis mucosa extends into the primary laminae usually in two layers.

In between these two layers of the laminae muscularis mucosae there is a layer of smooth muscle belonging to the tunica muscularis. These three layers of smooth muscle intertwine as they extend toward the tip of the laminae and eventually fuse to form one large mass of muscle at the tip.

Abomasum

The abomasum is the glandular part of the compound stomach and histologically it is essentially the same as a simple stomach.

Small Intestine

The small intestine is a typical tubular organ in that it has all of the typical tunics and layers. However, the tunica mucosa is especially modified to fulfill the function of absorption. Also, the three regions of the small intestine, the duodenum, the jejunum, and the ileum, each have special modifications to the wall to enable each region to better perform its particular function. In the small intestine digestion occurs in the lumen as well as at the surface of the lining epithelial cells. Pancreatic enzymes such as trypsin, chymotrypsin, elastase, carboxypeptidases, peptide hydrolases, amylase and lipases are adsorbed onto the membrane surface of the epithelial cells where they mix with the chyme present in the lumen catalyzing the breakdown of proteins, carbohydrates and lipids. The smaller breakdown products are then absorbed by the lining epithelial cells that are called enterocytes.

Layers of the Small Intestine

• Tunica mucosa: This layer protrudes out into the lumen as projections called villi and it dips down to the underlying lamina muscularis mucosae forming pockets called crypts.

o Lamina epithelialis mucosae - simple columnar epithelium - villus - a villus contains enterocytes (absorption), goblet cells (protective mucus) in its upper region and neuroendocrine cells (local hormones)

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- crypt - a crypt (crypt of Lieberkühn) contains goblet cells, paneth cells (defensive), neuroendocrine cells, stem cells, intraepithelial lymphocytes (defensive)

o Lamina propria - loose connective tissue rich in blood and lymphatic vessels present in the core of the villi and between crypts

o Lamina muscularis mucosae- thin layer of smooth muscle located at the base of the crypts

• Tunica submucosa: This layer blends with the lamina propria and is typical. In the duodenum it has coiled branched glands known as Brunner's glands, the ducts of which open into the base of the crypts.

• Tunica muscularis: typical consisting of an inner circular layer and an outer longitudinal layer

• Tunica serosa: typical

Enteroendocrine cells: These cells secrete hormones such as secretin, somatostatin, enteroglucagon and serotonin; one hormone per type of cell. Paneth cells: These remarkable cells contain large granules that contain defensins (antimicrobial peptides) as well as lysozymes and phospholipase A. These chemicals represent the "first-line" of defense against microbes that enter through the digestive tract. Compared to the other cells present in the epithelial lining, Paneth cells are long-lived, i.e., weeks versus a few days for the other cells.

Specializations to enhance absorption ability

The small intestine has all of the "layers" of a typical tubular organ but the tunica mucosa is highly specialized to perform the function of absorption. To fulfill this function it uses several strategies to increase the surface area of the plasma membrane of the absorptive epithelial cells.

• individual cells have numerous projections of their apical plasma membranes called microvilli

• the lamina epithelialis and lamina propria together form folds that project out into the lumen called villi

• the tunica mucosa and tunica submucosa together form large transverse folds into the lumen called plicae circulares

• the small intestine is extremely long (usually several meters)

Regional variations in the small intestine

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Duodenum

• presence of Brunner's glands in the submucosa o serous in pig and horse o mucous in ruminant and dog o mixed in cat

• presence of chyme in the small intestine induces cells of Brunner's glands to secrete alkaline mucus that neutralizes gastric acid and pepsin and further promotes digestion

• no plicae circulares • longest villi of all three • regions • highest number of goblet cells

Jejunum

• no glands in the submucosa • no lymphoid nodules

Ileum

• permanent aggregated lymphoid nodules in the submucosa • shortest villi; least number of goblet cells

Large Intestine

Unlike the small intestine, there are no plicae circulares or villi in the large intestine so the surface of the tunica mucosa is more uniform and flatter than that of the small intestine.

• Tunica mucosa:

o lamina epithelialis -simple columnar epithelium that forms straight tubular glands lined with absorptive columnar cells (recovering water and salt) and numerous goblet cells (producing mucus to facilitate passage of dry waste material); stem cells and lymphocytes are also present

o lamina propria- loose connective tissue that contains numerous blood and lymphatic vessels, collagen, lymphocytes and plasma cells

o lamina muscularis mucosae- present beneath the base of the crypts and prominent; undergoes rhythmic contractions mixed in cat

• Tunica submucosa: typical • Tunica muscularis: inner circular and outer longitudinal layers; outer longitudinal layer

is organized into three separate bands known as taenia coli; movement of more solid waste to the rectum

• Tunica serosa is typical.

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Commensal bacteria reside in the large intestine and play a role in the continued digestion of food. Digestive System II – Liver, gall bladder, pancreas

Other organs that are part of the digestive system include the liver, gall bladder and pancreas.

Liver

The liver is the largest gland in the body; it is multifunctional. To understand the function of the liver it is necessary to understand the blood supply to the liver. It is supplied by the hepatic artery in the typical manner but it is the only digestive organ drained by the inferior vena cava. Other digestive organs such as the small intestine, parts of the large intestine, stomach and pancreas are drained by the hepatic portal system which takes the blood directly to the liver. Thus, the liver receives oxygen poor, nutrient rich blood from the hepatic portal system and oxygen rich blood from the hepatic artery.

Functions of the liver.

Digestive and Metabolic Functions

• synthesis and secretion of bile • storage of glycogen and lipid reserves • maintaining normal blood glucose, amino acid and fatty acid concentrations • synthesis and release of cholesterol bound to transport proteins • inactivation of toxins • storage of iron reserves • storage of fat-soluble vitamins

Non-Digestive Functions

• synthesis of plasma proteins • synthesis of clotting factors • synthesis of the inactive angiotensinogen • phagocytosis of damaged red blood cells • storage of blood • breakdown of circulating hormones (insulin and epinephrine) and immunoglobulins • inactivation of lipid-soluble drugs

General organization of the liver

Structurally the liver is divided into lobules by loose connective tissue septae. These septae are more prominent in some domestic animals than in others; the pig has the most prominent septae and they are readily apparent grossly. For a long time the lobule as defined by these septae was thought to be the basic functional unit of the liver but now it seems that another unit, i.e., the

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hepatic acinus, might better represent the functional unit of the liver. Both the hepatic lobule and the hepatic acinus will be described but first the basic histology of the liver will be described.

At low magnification the liver looks relatively homogeneous and on first examination little organization can be discerned. A closer look reveals the presence of "lobules" or groups of hepatocytes arranged around a blood vessel, the central vein, and defined by loose connective tissue in which the portal canals are found. This type of organization is most easily seen in the pig liver.

Hepatocytes are one of the primary functional cells of the liver. They are located in flat irregular plates that are arranged radially like the spokes of a wheel around a branch of the hepatic vein, called the central vein or central venule since it really has the structure of a venule.

Portal canal: Three structures are found gouped together in the loose connective tissue surrounding the plates of hepatocytes. These include branches of the hepatic artery, the hepatic portal vein (venule) and the intralobular bile ductule. This group of three structures has been called a portal triad but now is called a portal canal.

Portal canal:

• branch of the hepatic artery • branch of the hepatic portal vein (venule) • section of an intralobular bile ductule

Hepatocytes are arranged in rows that radiate out from the central vein. These rows are one cell wide and are surrounded by sinusoidal capillaries or sinusoids. This arrangement ensures that each hepatocyte is in very close contact with blood flowing through the sinusoids, i.e. bathed in blood.

The endothelial cells lining sinusoids are fenestrated and in most species lack a basal lamina. Gaps are also present between the endothelial cells. Taken together these two properties make the sinusoids extremely leaky and allow for the extremely close contact between the blood and the surface of hepatocytes. Many materials in the blood, except for whole blood cells, can pass between the spaces in the sinusoidal lining.

Although sinusoidal endothelial cells lie very close to hepatocytes, they do not actually make contact. A narrow space is present between the surface of the hepatocyte and the surface of the endothelial cell. This is called the space of Disse; it is filled with numerous microvilli from the hepatocytes. As in other areas of the body, these structures serve to increase the surface area of the cell membrane that comes in contact with the blood facilitating exchange of molecules between hepatocytes and the blood.

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What is the basic functional unit of the liver?

• Hepatic lobule: The hepatic lobule is defined as having a central vein (CV) at its center with its edges defined by portal canals (PC). This model only takes into consideration the flow of blood in one direction, i.e., from the branch of the hepatic artery located in the portal canal toward the central vein. Yet, in the liver, blood actually flows from branches of the hepatic artery in several directions. Thus, the hepatic lobule does not define a "functional unit" of the liver very well.

• Hepatic acinus: More recent terminology identifies the hepatic acinus as the "functional unit" of the liver. This definition recognizes both the real pattern of blood flow in the liver and the role of hepatocytes as secretory cells, secreting bile. The hepatic acinus has three zones.

o Hepatocytes in Zone 1 are the first to receive blood and it is high in oxygen.

o Hepatocytes in Zone 2 are the second cells to receive blood and it is lower in oxygen.

o Hepatocytes in Zone 3 are the last to receive blood from a branch of the hepatic artery and it is lowest in oxygen.

o Thus, the cells with the highest metabolic potential are found in Zone 1 and those with the least are found in Zone 3. Importantly, the cells in Zone 3 are the most susceptible to ischemic conditions due to the already low level of oxygen that reaches them through the blood.

Secretion of bile in the liver

Bile is produced and secreted by hepatocytes into a special "duct" called a bile canaliculus. This "duct" is actually just a space formed between two hepatocytes that is separated from the connective tissue space around the hepatocytes by the presence of tight junctions. The bile canaliculi empty into branches of the bile ductules which eventually empty into the hepatic duct that carries the bile out of the liver to the gall bladder for concentration and storage. In the duct system, bile flows in the direction opposite to the flow of blood in the sinusoids.

Gall Bladder

The gall bladder receives bile from the liver. Bile is composed of bile salts that emulsify fats forming water-soluble complexes with lipids (micelles) to facilitate the absorption of fat. Bile salts in the small intestine also activate lipases in the intestine.

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Functions of the gall bladder

• storage of bile • concentration of bile • acidification of bile • send bile to the duodenum in response to cholecystokinin

secreted by enteroendocrine cells in small intestine; horse does not have a gall bladder and bile is continuously received from the liver

Gall bladder structure The gall bladder is a sac that is lined with a simple columnar epithelium and has a tunica muscularis containing smooth muscle that is innervated by both the parasympathetic and sympathetic branches of the autonomic nervous system. Tunics (layers) of the Gall Bladder

• Tunica mucosa: When the gall bladder is empty, this layer is extremely folded. When full, this layer is smoother but still has some short folds.

o lamina epithelialis: composed of simple columnar epithelial cells with numerous microvilli on their luminal surfaces and connected by tight junctions near luminal surfaces.

o lamina propria: composed of loose connective tissue rich in reticular and elastic fibers to support the large shape changes that occur in the lamina epithelialis; lamina propria may contain simple tubuloalveolar glands especially in ruminants. May be mucous or serous depending on species.

o lamina muscularis mucosae: not present

Tunica submucosa: present and typical

Tunica muscularis: contains much smooth muscle, poorly organized

Tunica serosa: present and typical

Pancreas

The pancreas contains both exocrine and endocrine components that secrete digestive enzymes and peptide hormones respectively. These two components are very different structurally and functionally but are intermingled within the gland. However, the organization of the exocrine part into acini make it fairly easy to recognize in histological sections as does the organization of the endocrine part around areas of high vascularity.

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Organization of the pancreas

The bulk of the pancreas by volume consists of exocrine cells that secrete an alkaline solution of digestive enzymes. This secretion moves through a duct system that eventually leads to the pancreatic duct. Only about 5% of the volume of the pancreas consists of endocrine cells. These cells secrete peptide hormones that play a role in controlling carbohydrate metabolism. The endocrine cells are closely associated with large numbers of blood capillaries into which they secrete the peptide hormones.

The exocrine pancreas . . .

The exocrine portion of the pancreas is a compound acinar gland. It has many small lobules, each of which is surrounded by connective tissue septa through which run blood vessels, nerves, lymphatics, and interlobular ducts.

Exocrine secretion by the pancreas is controlled by hormones and nerves.

• When the hormone, secretin, is released from neuroendocrine cells in the duodenum of the small intestine, the pancreas secretion is watery and rich in bicarbonate. This "basic" secretion helps to neutralize the acidic chyme as it comes into the small intestine.

• When cholecystokinin-pancreozymin (CCK) is released by neuroendocrine cells in the duodenum the pancreas secretes a product rich in enzymes that breakdown proteins, carbohydrates, lipids and nucleic acids in the lumen of the small intestine.

• When gastrin is secreted by pyloric neuroendocrine cells the pancreatic secretion is rich in digestive enzymes. Two of the digestive enzymes secreted by the pancreas are trypsin and chymotrypsin; they are secreted as non-active, pro- or zymogen forms and are subsequently activated by enterokinase in the lumen of the duodenum to avoid digestion of the pancreatic acinar cells.

A compound acinar gland

• Acini: The secretory cells of the pancreas are arranged around a small lumen. The pancreatic acinar cells produce the digestive enzymes in the typical pattern of protein synthesis. However these cells are highly active in protein synthesis for export and this high activity is reflected in their bizonal staining properties with the typical dyes used for histology, i.e., hematoxylin and eosin. The basal region of these secretory cells usually stains intensely with hematoxylin reflecting the presence of large amounts of endoplasmic reticulum where the protein is being synthesized on ribosomes. These proteins move from the rough endoplasmic reticulum to the Golgi apparatus where they are glycosylated, then from the Golgi as secretory granules. In these granules the enzymes are found in an inactive or zymogen form. They are activated after release in to the duct system. The presence of numerous zymogen granules containing high concentrations of protein is reflected in the intense eosin staining in the apical region of

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the secretory cells. These granules are most abundant during fasting or between meals and least abundant after a meal has been ingested.

• Ducts: The secretory product of the acinar cells is carried out of the pancreas by a duct system as in other exocrine glands.

o Intercalated duct. The first part of the duct system is called the intercalated duct or intralobular duct. It is lined with low cuboidal epithelial cells that secrete bicarbonate ion into the secretory product. This duct actually extends into the acinar lumen, where its walls consist of the pale staining centroacinar cells. Intercalated ducts empty into the larger interlobular ducts.

o Interlobular ducts. These ducts are lined with a low columnar epithelium that may contain goblet cells. Interlobular ducts empty into the main pancreatic ducts that exit the pancreas.

The endocrine pancreas . . . The cells of the endocrine portion of the pancreas are arranged either in round-to-oval shaped areas rich in blood vessels known as the islets of Langerhans or they may be scattered throughout the exocrine portions of the pancreas near the acini or ducts. There are several different types of cells in the islet or other regions, each secreting a different peptide hormone. It is not possible to distinguish among these cells with routine hematoxylin and eosin stain used for histological preparations. Immunocytochemistry is necessary to identify which cells are secreting a particular peptide. This is done by staining with an antibody made to the specific peptide that is combined with a label that can be visualized at the light microscopic level such as immunoperoxidase. Examples of peptide hormones secreted by the endocrine pancreas:

• insulin - increases uptake of glucose by most cells; reduces blood level • glucagon - decreases uptake of glucose; increases blood level • somatostatin - many effects of gastrointestinal function; may inhibit insulin and glucagon

secretion • vasoactive intestinal peptide • pancreatic polypeptide • motilin, serotonin, substance P

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Hematopoiesis

Hematopoiesis is the process by which immature precursor cells develop into mature blood cells. The currently accepted theory on how this process works is called the monophyletic theory which simply means that a single type of stem cell gives rise to all the mature blood cells in the body. This stem cell is called the pluripotential (pluripotent) stem cell.

SITES OF HEMATOPOIESIS

Age of animal Site of hematopoiesis Embryo yolk sac then liver

3rd to 7th month spleen

4th and 5th months marrow cavity - esp. granulocytes and platelets

7th month marrow cavity - erythrocytes

Birth mostly bone marrow; spleen and liver when needed

Birth to maturity number of active sites in bone marrow decreases but retain ability for hematopoiesis

Adult bone marrow of skull, ribs, sternum, vertebral column, pelvis, proximal ends of femurs

STRUCTURE AND FUNCTION OF BONE MARROW

Bone marrow has a vascular compartment and an extravascular compartment. The vascular compartment is supplied by a nutrient artery which branches into central longitudinal arteries which send out radial branches that eventually open into sinuses. These sinuses converge into a central vein that carries the blood out of the bone marrow into the general circulation. Hematopoiesis takes place in the extravascular compartment. The extra vascular compartment consists of a stroma of reticular connective tissue and a parenchyma of developing blood cells, plasma cells, macrophages and fat cells. The high activity of the bone marrow is demonstrated by its daily output of mature blood cells: 2.5 billion erythrocytes, 2.5 billion platelets, 50-100 billion granulocytes. The numbers of lymphocytes and monocytes is also very large.

Bone marrow is the site for other important activities in addition to hematopoiesis. These include the removal of aged and defective erythrocytes and the differentiation of B lymphocytes. It is also the site of numerous plasma cells.

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THE PROCESS OF HEMATOPOIESIS

The monophyletic theory of hematopoiesis states that pluripotent stem cells multiply to produce more pluripotent stem cells, thus ensuring the steady and lasting supply of stem cells. Some of the pluripotent stem cells differentiate into precursor cells that are at least partially committed to become one type of mature blood cell.

Pluripotent stem cells multiply slowly into one of five possible unipotential stem cells, which then multiply rapidly into the precursor of the specific mature blood cell for which they are destined.

Although the pluripotent stem cells and the unipotential stem cells cannot be distinguished from one another histologically, the precursor cells can be distinguished with a trained and practiced eye.

General Features: Understanding the general process of hematopoiesis will be extremely helpful in distinguishing and identifying the different cells in a bone marrow smear or in an intact bone marrow preparation. Basically an immature, precursor cell goes from a cell that is making lots of protein to a cell that is making much less protein.

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Since structure is (always) related to function, the structure of the precursor cell changes as it goes from making more protein to making less protein. Thus, a cell that is making a lot of protein will have a nucleus containing dispersed or active chromatin, i.e. that is being transcribed actively. When this cell is making less protein, the chromatin is condensed or clumped because it is not being transcribed. Likewise, a cell that is making a lot of protein will have many large nucleoli, the site of ribosomal RNA synthesis and assembly; as protein secretion decreases there are smaller and fewer nucleoli. Cells with high protein synthetic activity have more ribosomes in their cytoplasm and consequently the cytoplasm stains more basophilic (hematoxylin staining of the RNA in ribosomes). Cells with lower protein synthetic activity have fewer ribosomes, thus less basophilic staining with hematoxylin leaving the cytoplasm appearing more acidophilic due to the eosin staining of cytoplasmic proteins. In cells with high protein synthetic activity, the Golgi apparatus is highly developed, occupies much of the cytoplasm thus pushing the nucleus off to one side (acentric nucleus). Cells with low protein synthetic activity have a smaller Golgi and the nucleus tends to be more centrally located.

The chart below summarized these features.

Cell Organelle Making lots of Protein Making less Protein

Nucleus chromatin is dispersed

chromatin is clumped

Nucleoli more

fewer

Cytoplasm more ribosomes; basophilic

fewer ribosomes; acidophilic

Golgi apparatus *large; nucleus off center *smaller; nucleus more centered

* except in RBC precursors where the nucleus becomes more off centered until it is extruded from the cell at the last stage before maturity

ERYTHROPOIESIS

As the cells are maturing in the erythrocytic series, the cells are usually getting smaller, the nucleus is becoming smaller and more condensed and is eventually lost, and the cytoplasm is becoming pinker rather than blue.

The cells in the developing erythrocyte series are as follows:

• Unipotent stem cell: cannot be distinguished from other unipotent stem cells by histology

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• Proerythroblast: nucleus still rather large, taking up most of the cell; nucleus not condensed; cytoplasm still very blue or basophilic

• Basophilic erythroblast: very difficult to distinguish from the proerythroblast

• Polychromatophilic erythroblast: nucleus is more condensed than that of the proerythroblast; cytoplasm less blue, more grayish

• Orthochromatophilic erythroblast: nucleus more condensed, smaller than that of previous cells and looks pyknotic by comparison; cytoplasm beginning to take on a more pinkish cast

• Reticulocyte: no nucleus; cytoplasm still stains somewhat bluish due to presence of remnants of polyribosomes

• Erythrocyte: mature erythrocyte has no nucleus (in mammals); cytoplasm stains very pink due to lack of ribosomes and presence of high amounts of protein, i.e., hemoglobin

GRANULOCYTE DEVELOPMENT

• Unipotent stem cell: cannot be distinguished from other unipotent stem cells by histology

• Myeloblast: large cell with blue-staining cytoplasm; large nucleus; often has a clear area near the nucleus - this is where the rather large Golgi is located

• Promyelocyte: still a rather large cell with azurophilic (not specifically stained) granules

• Myelocyte: overall cell still rather large; nucleus still round without indentation; granules staining appropriately for the series, i.e., pink for eosinophilic, blue for basophilic, neutral for neutrophilic

• Metamyelocyte: cell is about the size of a mature granulocyte; nucleus with slight indentation; granules present that stain appropriately for the series, i.e., pink for eosinophilic, blue for basophilic, neutral for neutrophilic

• Band cell: cell is about the size of a mature granulocyte; nucleus with definite indentation - looks like a horseshoe; prominent granules that stain appropriately for the series

• Mature (segmented) granulocyte: cell is mature and looks like normal, mature granulocytes in the blood with lobed nucleus and prominent granules that stain appropriately for the series

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MONOCYTES

Not responsible for knowing the sequence of development of monocytes.

PLATELETS

Platelets, also called thrombocytes, play an important role in hemostasis by:

• plugging holes in blood vessels to prevent bleeding • promoting formation of clots to further prevent bleeding • helping to repair damaged blood vessels

Platelet granules contain (1) the secretory material that platelets produce to help repair damaged blood vessels, (2) growth factor, and (3)many other proteins. Some of these are:

• platelet factor 4 - regulates vascular permeability, calcium mobilization from bone, chemotaxis of monocytes and neutrophils

• beta thromboglobulin - function unknown; used to monitor activation of platelets in some diseases

• coagulation factors - fibrinogen, factor V, factor VIII • fibronectin, thrombospondin, platelet-derived growth factor - all may be involved in

repair of damaged blood vessels • serotonin (taken up from plasma and stored in granules) • lysosomal enzymes such as hydrolases

Platelets appear as round, oval or biconcave disks and have a diameter of about 1.5 to 3.5 µm. They are somewhat difficult to see in blood smears because of their small size and because they are often clumped together. Despite their small size, they contain all of the normal organelles and are rich in granules that are difficult to resolve with the light microscope but can be easily seen with the electron microscope.

Platelets are formed in the bone marrow from megakaryocytes (30-100 µm diameter), very large cells with a polyploid, multilobed nucleus. Platelets are released from fragmenting megakaryocytes in at least two ways:

• extension of pseudopodia through the wall of the sinuses; pseudopodia contain "strings" of platelets that are pinched off and released into the circulation

• passage of mature megakaryocyte into circulation and fragmenation in the pulmonary vascular bed

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Immune System

This system consists of cells and tissues that have as their main function the protection of the body from the invasion by microorganisms and disease-producing entities foreign to the animal. To achieve this goal this system has components spread widely throughout the body with concentrations in specific places. Components of the system may be single lymphocytes located strategically in the epithelium of mucous membranes, aggregations of lymphocytes associated with the mucosa of strategically placed organs, or entire organs highly organized and strategically located in reference to lymph and blood flow patterns.

The components are:

• Lymphocytes T cells B cells

• Plasma cells • Bone marrow • Thymus • Lymph Node • Mucosa-associated lymphoid tissue (MALT) • Spleen

NOTE: The bone marrow and thymus are considered as primary immune/lymphoid components because they contain the stem cells that will develop into T cells, B cells and natural killer cells of the functioning immune and lymphatic systems. Lymphocytes and Plasma Cells

There are basically two different types of lymphocytes, T lymphocytes (T cells) that are involved in cell-mediated immunity and B lymphocytes (B cells) that are involved in humoral immunity. Both types of cells originate from stem cells in bone marrow. In addition there are many types of T lymphocytes depending on their specific role in the immune response. In circulating blood, lymphocytes may be either small lymphocytes (6-9 µm) or large lymphocytes (9-15 µm) with the latter representing only about 3% of circulating lymphocytes. Although it is not possible by routine histological methods to differentiate the various types of small lymphocytes found in blood, they are of several different types that are in the process of migrating through the circulation to take up residence in an organ or they are "searching" for foreign antigen. Large lymphocytes are mostly activated B lymphocytes.

Immature T lymphocytes move from the bone marrow into the thymus, take up residence and become thymus-dependent or mature T lymphocytes. These mature T cells then pass through the circulation to find homes in lymph nodes, mucosa-associated lymphoid tissue or the spleen. There are several types of T lymphocytes, i.e., T helper cells, cytotoxic T cells and suppressor T cells.

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B lymphocytes originate and mature in the bone marrow then move through the circulation to various sites throughout the body. Upon interaction with foreign antigen and usually with the assistance of T helper cells, B lymphocytes become mature antibody secreting cells called plasma cells. Clones of plasma cells making specific immunoglobulins are produced thus providing the large numbers of plasma cells needed to mount a good antibody (humoral immune) response. Plasma cells are rarely found in the circulation but reside mostly in connective tissue (lamina propria) beneath epithelia, in the medullary cords of lymph nodes and in the white pulp of the spleen.

These immune cells are strategically located in areas that come in close contact with foreign substances. They represent one of the first lines of defense against invading microorganisms, viruses and parasites. A good example is the small intestine. In these types of locations, they are perfectly positioned to interact with invading foreign substances and they recognize these substances as non-self or foreign. Upon such "recognition" lymphocytes are activated and function to neutralize or destroy the invading foreign substance.

Plasma cells are derived from activated B lymphocytes that have left the blood stream and taken up residence in connective tissue. They are easily identified in histological sections due to their unique morphology which reflects their high protein synthetic activity. Usually the round to oval nucleus is eccentrically located in the cell due to the presence of a large Golgi apparatus where immunoblobulin synthesis is completed and the molecules packaged for secretion. The predominant staining pattern of the cytoplasm is bluish to purple (basophilic) due to the large amount of rough endoplasmic reticulum and associated ribosomes. Usually the cytoplasm is packed with rough ER. In a very well stained, relatively thin seciton, the nucleus has the appearance of being "spoked" or having a "clock face".

Thymus

Located posterior to the sternum in the anterior part of the mediastinum, the thymus is a bi-lobed nodular organ that is very large in the first year or two of life reaching maximum size at puberty then becoming smaller in a process called involution. During this degenerative process connective tissue fibers and fat cells replace the previously functional tissue (parenchyma) of the organ and even though only a few pieces of functional tissue remain, it is enough to continue to supply the organism with sufficient mature lymphocytes. Immature T lymphocytes move from the bone marrow into the thymus where they become immunocompetent T cells. These T cells then leave the thymus, go into the circulation and eventually find their way to lymph nodes, mucosa-associated lymphoid tissue or the spleen.

Functions

• production of immunocompetent T lymphocytes • production of mature but naïve T cells for peripheral tissues and circulation • immunological self-tolerance • regulation of T cell maturation, proliferation and function via secretion of hormones

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Histology of the Thymus

Each lobule has an outer, darker staining cortex and an inner, paler staining medulla. High concentrations of T lymphocytes in the cortex are the basis for the intense basophilia of this region and this is the site of precursor cell proliferation and maturation. Mature, immunocompetent T cells then move from the cortex toward the medulla where they enter the bloodstream to be taken out of the thymus.

The thymus has two tissue components: parenchyma and stroma. The parenchyma is composed mostly of T lymphocytes in various stages of development into mature T cells whereas the stroma is composed of special thymic epithelial cells.

The parenchyma and stroma have different appearances depending on whether you are looking at the cortex or the medulla.

Cortex

In the cortex, the parenchyma consists mostly of the developing T lymphocytes. It is here that the T cell receptor (TCR) genes are rearranged so that the mature T cells obtain their specific surface markers. The stroma consists of sparse, delicate epithelial cells obscured by all of the lymphocytes. These epithelial cells form the support structure for the developing T cells but also play an important role in isolating the T cells from foreign anitgens during their development.

Medulla

In the medulla, the stroma consists of prominent epithelial cells that have large, pale-staining nuclei and substantial amounts of eosinophilic (pink-staining) cytoplasm. There are fewer T cells because most of them have entered the blood stream via vessels at the corticomedullary junction. Antigen presenting cells (APC) are also found in the medulla where they are called thymic interdigitating cells. These cells are thought to present self-antigens to the matured T cells. T cells that recognize these self-antigens are removed by a process called apotosis. This process helps to prevent autoimmune diseases.

One prominent and identifying feature of the medulla is the presence of Hassall's corpuscles thought to represent degenerating epithelial cells. These impressive structures begin to form in the fetus and increase in number and size as the animal ages.

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Structural basis for function of the thymus

In performing its major functions of producing immunocompetent but naïve T cells and in achieving immunological self-tolerance, the thymus has some special structural arrangements unlike those found in other organs.

• First, to keep the developing T lymphocytes "protected" so that they can develop their surface receptors in a "climate" that is not influenced by antigens, the thymic epithelial cells form a continuous layer along the inner surface of the capsule extending into the thymus along the septa and along blood vessels. These cells actually provide a cellular framework for a space that is kept separate from other spaces such as the bloodstream. This separation is maintained by desmosomes between adjacent epithelial cells and their close contact with endothelial cells of capillaries. The "barrier" that results is called the blood-thymus barrier; it is similar in structure to the blood-brain barrier. It is within this confined and protected space that the T lymphocytes develop into immunocompetent yet naïve T cells. The integrity of the space within the epithelial cell framework is extremely important because it prevents the premature stimulation of T cells by antigens. Blood-thymus barrier - components - tight junctions between endothelial cells - basal lamina of endothelium - small connective tissue space - basal lamina of epithelial cell - continuous sheet of epithelial cell

• Second, to provide a mechanism by which the newly developed immunocompetent and naïve T cells can be added back to the circulation, the blood supply of the thymus also has some peculiarities. Most arteries enter the thymus through the capsule, course via connective septae through the cortex down to the level of the corticomedullary junction where they then actually enter the parenchyma of the organ. Capillaries from these arterial branches return to the region of the cortex within the parenchyma. These capillaries are special in that they are not permeable to macromolecules, thus preventing any antigenic contact with developing T cells in the cortex. Postcapillary venules that derive from these same capillaries are permeable to macromolecules and lymphocytes. The new immunocompetent T cells move into these postcapillary venules to eventually join the general circulation and move to the other tissues and organs that are part of the immune system. Some capillaries from the arterial branches entering the thymus from the capsule extend down directly into the medulla to supply the tissue with oxygen and nutrients then reconvene as postcapillary venules that join the postcapillary venules coming from capillaries in the cortex. Thus, blood draining the cortex and the medulla combine in the postcapillary venules and exit the thymus through typical venous pathways.

• Third, to ensure that self-tolerance is acheived, the medulla of the thymus has antigen presenting cells (APC) that are thought to present self-antigens to the matured T cells.

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Any T cells that recognize these self-antigens are removed thus preventing development of autoimmune diseases.

Lymph Nodes

After maturing in the thymus, T cells move through the circulation to other organs, including lymph nodes. Lymph nodes are small lima-bean shaped organs that are spread throughout the body but occur in groups in areas where lymphatic vessels come together to form larger vessels such as in the groins, neck and axilla. Lymph nodes are also part of the lymphatic system that includes the lymphatic vessels, lymphoid tissue and lymphoid organs. Lymphatic vessels drain fluid (lymph) from peripheral tissues and bring it to the venous system. Lymph consists of interstitial fluid that is similar to blood plasma but with a lower protein concentration, lymphocytes and macrophages. Lymph nodes filter and purify the lymph before it flows into the venous system.

Functions

• filter debris and microorganisms via phagocytosis by fixed macrophages • facilitate the interaction between antigen presenting cells and circulating lymphocytes to

initiate an immune response • B lymphocytes: activation and proliferation; plasma cell formation and antibody

production in response to antigens • T lymphocytes: activation to become T helper and T cytotoxic cells

The location and structural organization of lymph nodes makes them perfect for the above functions. They are positioned so that all lymphatic vessels draining back to the venous circulation from the tissues pass through a lymph node. The afferent lymphatic vessels branch outside the organ, penetrate the capsule and empty into the subcapsular sinus. From here the lymph flows into and through cortical sinuses enabling the lymph to come in close contact with cells in the cortex of the node. In the medulla there are also sinuses (medullary sinuses) that enable the lymph to flow toward the hilum and enter efferent lymphatic vessels. Eventually the filtered lymph enters the bloodstream through the thoracic duct or right lymphatic duct.

Lymph nodes are surrounded by a fibrous connective tissue capsule that enters the organ as trabeculae which define a cortex and medulla. The capsule and trabeculae are the source of reticulin fibers that are found throughout the node and form the main supporting network of the organ. These fibers serve to keep the sinuses open and to support the massive number of lymphocytes and macrophages. Beneath the capsule is a subcapsular sinus into which lymph flows from the afferent lymphatic vessels.

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Histological organization of Lymph Nodes

The Cortex

The cortex is composed of the cortical sinuses surrounded by dense accumulations of lymphocytes. In the more superficial cortex the lymphocytes are arranged into spherical follicles, lymphoid follicles. It is here that B lymphocytes are activated and undergo proliferation.

Germinal Center (GC)

The open, pale-staining nature of the nuclei of these cells indicate that they are B lymphocytes undergoing active proliferation. Other cells include:

• follicular dendritic cells that present antigen to the B cells • tingible body macrophages that engulfed dead B cells that have died by apotosis

Resting B cells enter the lymph node parenchyma though the high endothelial venules and if they encounter an antigen with which they can react, they then enter the cycle of blast transformation to produce clones of plasma cells and B memory cells. This production of clones occurs in the germinal centers of lymphoid follicles. Paracortical zone Deeper regions of the cortex contain primarily T lymphocytes that do not form into follicles. T lymphocytes (helper and cytotoxic/suppresor) arrive through the circulation, enter the lymph node parenchyma through the high endothelial venules and take up residence in the paracortical zone. If activated, the T lymphocytes undergo active proliferation to produce expanded clones of activated T lymphocytes.

T lymphocytes that arrive at the lymph node via the arterial blood stream gain access to the parenchyma of the lymph node through the wall of the high endothelial venules located in the paracortical zone. These blood vessels contain endothelial cells that are expressing specific lymphocyte binding molecules called addressins. These surface molecules are available to bind to lymphocytes that recognize them, the lymphoctyes bind to the surface of the endothelium, then cross the vessel wall and enter the lymph node parenchyma.

Mantle zone (corona)

The germinal center is surrounded by a ring of darker-staining cells. The condensed nature of their nuclei indicates that these are resting B cells. Also present in the mantle zone are T helper cells, macrophages and dendritic cells.

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In a T cell-dominated response, the paracortical zone of the lymph nodes may be greatly enlarged. Interdigitating dendritic cells are the main antigen presenting cell in the paracortical zone.

The Medulla

The medulla of a lymph node is composed of medullary cords interspersed between medullary sinuses.

The medullary cords are composed of plasma cells producing antibodies, their precursors, macrophages and T helper cells. The most prominent cell in the cord is the precursor to plasma cells or immunoblasts that came from the germinal centers of the lymphoid follicles in the cortex of the node.

In the medullary cords, the plasma cells undergo final maturation and secrete antibodies into the lymph that is collected by efferent lymphatic vessels in the node and eventually carried to the general circulation. Plasma cells may also get into the general circulation in this manner.

The medullary sinuses are composed primarily of reticular fibers (RF) providing the support framework, reticular cells (fibroblast-like cells that secret the reticulin) (RC) and macrophages.

Overview of the Blood Flow Pattern in a Lymph Node

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Mucosa-associated lymphoid tissue (MALT)

MALT is really connective tissue located beneath mucous membranes in which the lymphocyte is the predominant cell type. Examples occur in the respiratory, gastrointestinal, urinary and reproductive tracts. The exact extent of these aggregations of lymphocytes is not easily discernible because they have no distinct capsule like that of lymph nodes. However, they are like lymph nodes in that they often have a pale-staining germinal center containing actively dividing lymphocytes like the germinal centers in lymph nodes. The larger aggregations contain B and T cell zones and antigen processing cells; the smaller, more scattered MALT components such as those in the intestines and respiratory tract are mostly T lymphocytes. Some B cells and plasma cells are also present.

Distribution of MALT

In the digestive system:

• in the wall of the pharynx - tonsils (palatine, lingual, pharyngeal) • in the wall of the small intestine - aggregate lymphoid nodules • in the wall of the colon-aggregate lymphoid nodules • in the walls of the appendix

In the reproductive system:

• in the wall of the vagina

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Types of MALT and Functional aspects

• larger aggregates function much like lymph nodes • smaller, scattered MALT are mostly T lymphocytes but also have B cells and plasma

cells

-- mostly IgA in the intestines and respiratory tract to protect against pathogens that may gain access to underlying tissues -- IgG and IgM secreted into lamina propria to counteract pathogens that have gained access to connective tissue -- IgE secreted into lamina propria; mediates the release of histamine from mast cells

• single lymphocytes found within the lamina epithelialis are mostly T lymphocytes • MALT is drained by efferent lymphatics but there are no afferent lymphatics • lymphocytes exposed in MALT regions go through regional lymph nodes then return to

the MALT region after activation

Spleen

Located between the stomach, left kidney and diaphragm, the spleen is the largest lymphoid organ in the body, performing functions for the blood similar to those performed by the lymph nodes for the lymph. It is a soft organ, conforming to the contours of the organs and structures surrounding it. At the hilus on the visceral surface, the splenic artery brings blood into the spleen, the splenic vein takes blood from the spleen to the hepatic portal system, and lymphatics drain lymph from the spleen. In some domestic species such as the horse and dog, the spleen functions as a reservoir from which blood can be mobilized when needed and in these species, smooth muscle is a prominent feature of the capsule and trabeculae of the spleen.

Functions

• removal of abnormal blood cells and particulate matter via phagocytosis • storage of iron from recycled red blood cells • initiation of the immune responses by B cells and T cells in response to antigens

circulating in the blood • hematopoiesis in fetus and sometimes in adult

Histology

The exterior surface of the spleen consists of a capsule containing collagen and elastic fibers; the interior components are collectively called the "pulp". Upon gross examinagtion of a slice of the spleen, the pulp has two very different appearances: red and white. The organ appears as a large expanse of red pulp dotted with white pulp. Histologically,red pulp is "red" due to the presence of large numbers of erythrocytes in blood vessels called sinuses and white pulp is "white" due to lack of these sinuses and consequently fewer erythrocytes. The red pulp surrounds the white pulp while the latter looks like lymphatic nodules. Closer inspection of the white pulp indicates

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that there is a "central arteriole", sometimes called a central artery, close to the center of each area of white pulp.

Red Pulp

The red pulp of the spleen is characterized by a parenchyma (PN) that consists of macrophages of the sheathed capillaries as well as other macrophages and blood cells that have not yet entered the venous sinuses. The rest of the red pulp is occupied by numerous venous sinuses (VS). The walls of the sinuses are very open and can be easily traversed by blood cells. Their lining consists of long endothelial cells (arrows) oriented along the longitudinal axis of the vessel. Large spaces occur between adjacent endothelial cells and the underlying basement membrane is discontinuous, thus blood cells can easily pass between the endothelial cells and gain access to the bloodstream on the venous side. A continuous reticulin network forms the framework that supports the macrophages and a few fibroblasts responsible for producing the reticulin fibers; special stains are required to visualize the reticular network.

White Pulp

The white pulp of the spleen is characterized by a parenchyma that consists of two types of lymphocytes, i.e., B cells and T cells located in two different areas of the spleen. B cells are located in the lymphoid follicles scattered throughout the organ. In younger animals, a germinal center can be seen as seen in lymph nodes. In fact, this type of white pulp functions much in the manner that lymphoid follicles of lymph nodes function, i.e., initiation of immune responses by B cells to foreign antigens in the blood. T cells are located around the central arteries and form a kind of sheath. This site is called the periarteriolar lymphoid sheath.

Blood flow in the spleen

To properly understand the histology of the spleen, it is necessary to understand its blood supply. The splenic artery enters the spleen at the hilus, then branches into numerous arterioles that run through the parenchyma or pulp of the spleen. When these arterioles acquire a coating of T cells, the arterioles are called central arteries and the surrounding lymphoid tissue is called the PALS, i.e., the periarteriolar lymphoid sheath.

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Smaller penicillary arteries branch off of the central arteries and end in sheathed capillaries that are special capillaries which actually have no endothelial cells and end blindly. These unique capillaries are surrounded by macrophages which serve to filter materials from the blood. After the blood flows through these sheathed capillaries, it flows into a complicated system of sinuses that drain into larger and larger sinuses which eventually drain into the splenic vein which joins with the hepatic portal vein. Blood flows out of the sheathed capillaries into a space that is not considered part of any "true" blood vessel, then the blood cells re-enter the bloodstream through

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the walls of the sinuses. This particular arrangement of "blood flow" in the spleen is considered to be an open circulation pattern.

Integument

The Integument - the skin and all of its derivatives

Components

• skin (epidermis, dermis, hypodermis) • derivatives (sweat glands, sebaceous glands, mammary glands, hair, nails, claws,

hooves, horns, antlers, combs, wattles, and feathers)

Functions

• protection - from drying out, from invasion by microorganisms, from UV light and from insults (mechanical, chemical or thermal)

• sensation - for touch, pressure, pain and temperature • thermoregulation - decreases heat loss in cold temperatures; increases heat loss in hot

temperatures • metabolic functions - energy stored in fat deposits; synthesis of vitamin D

Structure of the Skin

Three distinct layers can be seen in the skin

• Epidermis - consists of keratinizing stratified squamous epithelium • Dermis - consists of fibroelastic connective tissue • Hypodermis - consists mostly of white adipose tissue (sometimes referred to as the

subcutis)

The thickness of these layers varies depending on the specific location in/on the body and in a given location on the amount of exposure to wear and tear. For example, in the buccal cavity, the epidermis consists of a moist stratified squamous epithelium which is relatively think and not highly keratinized whereas the epidermis of skin on the ball of the foot is thick and highly keratinized. The skin covering the dorsum of the hand has a rather thin hypodermis whereas the skin over the buttocks has a very thick hypodermis containing numerous fat cells.

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Epidermis

Layers of the Epidermis

- in order from outermost (surface) to innermost (deepest)

• Stratum corneum - consists of the remains of keratinocytes; mostly composed of the protein, keratin

• Stratum lucidum - present only in very thick skin; pale-staining layer of cells between the stratum corneum and stratum granulosum in which the dying keratinocytes contain a lot of keratin but are not completely replaced by it

• Stratum granulosum - consists of keratinocytes containing large numbers of granules that contribute to the process of keratinization

• Stratum spinosum- consists of large, polyhedral keratinocytes that are actively synthesizing keratin which is inserted as tonofibrils into the area of the plasma membrane beneath desmosomes that connect adjacent cells together. These "connections" or desmosomes between cells in this layer help hold them together and result in the "spiny" appearance of the cells that gives this layer its name.

• Stratum basale - consists of keratinocytes undergoing mitosis to produce the constant supply of keratinocytes needed for replacement of the dead and dying cells in the more superficial layers of the epidermis

Dermis

Two zones of the dermis:

• papillary zone - consists of loose areolar connective tissue containing collagen and fine elastic fibers; connects the epidermis to the thicker and denser reticular zone of the dermis

• reticular zone - contains dense, irregular and coarse collagen fibers and thick elastic fibers interspersed with fibroblasts and blood vessels and nerves

Glands in the skin.

Several different types of glands are located in the dermis of the skin serving a variety of functions.

• Sebaceous glands

• Apocrine sweat glands

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• Merocrine (= eccrine) sweat glands

Sebaceous Glands: The epithelium of this gland is an outgrowth of the external root sheath of the hair follicle and the gland empties its oily product directly into the follicle itself. The glands are of a branched acinar type and produce a lipid product called sebum that serves to reduce the entry of microorganisms into the body through the skin, to lubricate the hair and preventing it from drying out. The secretory cells die and become part of the product; a holocrine mode of secretion. These glands are not found in hooves, foot pads, claws or horns.

Apocrine Sweat Glands: These glands are coiled, tubular glands with a large lumen and a duct connecting it to an adjacent hair follicle. These glands secrete a viscid, milky product and are analogous to odiferous glands of many mammals. Once thought to use the apocrine mode of secretion, it is now known that their mode of secretion is more like that of the merocrine sweat glands. These glands are the primary sweat gland of domestic animals and are especially prominent in the horse.

Merocrine Sweat Glands: These glands are unbranched tubular in form and appear as a mass of tubules in cross section. They are plentiful in the upper regions of the fatty hypodermis. They secrete a watery product that is hypotonic to the plasma. It is the evaporation of this secretion on the surface of the skin that aids in thermoregulation. These are sometimes called eccrine sweat glands.

Hair

General structure of hair and associated structures:

• Hair shaft: the part of the hair above the surface of the skin • Hair root: the part of the hair below the surface • Bulb: an enlarged, hollow portion at the base of the root • Hair papilla: projection of dermis into center of the bulb • Follicle: the indentation in the skin within which the root lies

Hair has three layers:

• Cuticle: the outermost layer. Single layer of flattened, keratinized cells. Overlap like shingles, with free edge distally.

• Cortex: the thickest, intermediate layer. Consists of several layers of keratinized cells containing hard keratin. If hair is colored, these cells contain pigment. Cells held together by desmosomes.

• Medulla: central core; loosely packed cuboidal cells.

The structure and organization of the cuticle and medulla cells are species-specific.

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Hair follicles

The hair follicle is the structure that anchors the hair in the dermis and produces the hair itself. It is composed of five layers of epithelial cells arranged concentrically. The inner three layers form the hair shaft through a process of keratinzation while the outer two layers form the hair sheath.

1. cells in the innermost layer form the medulla of the hair or core of the hair shaft 2. cells in the next layer form the cortex that makes up most of the hair 3. cells in the third layer form the cuticle on the surface of the hair 4. cells in the fourth layer make up the internal root sheath 5. cells in the fifth or outermost layer form a layer called the external root sheath that does

not take part in hair formation

The external root sheath is separated from the surrounding connective tissue by a thick basement membrane known as the glassy membrane.

Types of follicles

Hair follicles can be classified in two ways: based on their size, i.e., diameter and based on their organization.

Based on size (diameter):

• Primary hair follicle: large having sweat gland, sebaceous gland and arrector pili muscle; ex. overcoat or guard hairs in dogs

• Secondary hair follicle: smaller, lacking sweat glands and arrector pili muscle; ex. underhair

Based on Organization:

• Simple follicle: a single hair from one follicle • Compound follicle: cluster of several follicles with several hairs emerging from one

opening onto surface of skin

The Hoof

The equine foot includes the hoof, dermis, first, second and third phalanges and associated structures. The hoof itself is the cornified layer of the epidermis, lacking the stratum granulosum and stratum lucidum. It is important to understand the histology of the hoof because a disease involving the epithelium of the foot, called laminitis, is the most devastating clinical disease of the foot.

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The peculiar histology of the hoof is formed from special relationships between the dermis (or corium) and the overlying epidermis. In some places, the dermal papillae and epidermal pegs are confluent forming apparent layers, i.e., they are laminar or consist of lamellae; in other places they are more typical. It is this lamellar interaction between the epidermis and dermis that gives the hoof its strength.

The wall of the hoof is that part of the hoof which is visible when the foot is on the ground, and it can be divided into three layers. From outside to inside, they are the stratum externum (tectorium), the stratum medium, and the stratum internum (lamellatum).

The Wall of the Hoof

• The stratum externum or tectorium is an extension of the perioplic epidermis and is composed of cornified eipithelial cells which appear as a soft, white, shiny material. This tissue attaches the hoof to the epidermis of the skin of the foot.

• The stratum medium or coronary epidermis is composed of prominent tubular and intertubular horn, and this layer comprises the bulk of the wall of the hoof.

• The stratum internum or stratum lamellatum is the epidermis in the laminar region.

Stratum Lamellum

This layer is made of nontubular horn which fuses with the stratum medium and helps hold the wall to the foot. In this region, the dermal papillae and epidermal pegs form elongated ridges oriented perpendicular to the ground. These ridges are formed from primary and secondary laminae - the secondary laminae being oriented at close to a right angle to the primary laminae. There are about 600 primary laminae and about 100-200 secondary laminae for each primary lamina. This system of interdigitating primary and secondary laminae provides the tight bond between the wall of the hoof and the underlying dermis. Thus, damage to the laminae leads to disruption of this interdigitating system which results in separation of the hoof wall from the dermis and phalanx beneath it.

Laminitis (acute laminar degeneration) is an inflammation of the laminae within the hoof. Many pathophysiologic mechanisms are thought to cause laminitis, among them vasoconstriction within the digit, perivascular edema, arteriovenous shunting of blood at the level of the coronary band, venoconstriction and microthombosis. These lead to less than normal perfusion of blood to the digit resulting in ischemia, edema and eventually necrosis of the laminae.

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Male Reproductive System

The primary function of the male reproductive system is reproduction, which includes the production of spermatozoa, the transportation of spermatozoa from the testes out of the male body, the secretion by glands, and the placement of spermatozoa in the female reproductive tract. Spermatozoa are produced in the testes then transported from the testes by a series of ducts which become gradually larger and connect with the urethra of the penis. Various accessory glands in the male system secrete materials which together with the spermatozoa constitutes the semen. A secondary function of the male reproductive system is the production of the male hormones which are responsible for the secondary sex characteristics of the male animal.

The male reproductive system is composed of several distinct organs. These include the testes, epididymis, deferent ducts, accessory glands, and the penis. The testis (plural, testes) is both an exocrine organ (compound, coiled, tubular gland) producing cells, i.e., spermatozoa, and an endocrine organ, secreting hormones, i.e., testosterone. Accessory glands (not all are present in all species) include the ampullary glands, vesicular glands, prostate gland, bulbourethral gland and urethral glands.

The testes are paired organs, and each one is enclosed in a fibrous white capsule of dense connective tissue (tunica albuginea) containing blood vessels (the stratum vasculare). A layer of peritoneum is tightly adhered to the tunica albuginea of each testis. The stallion has obvious smooth muscle fibers in the capsule. The connective tissue of the capsule continues into the testis on the posterior aspect as the mediastinum testis.

The dense connective tissue of the tunica albuginea is continuous with the loose areolar connective tissue of the septuli testis (septa) which extend through the parenchyma of the testis and divide it into lobules. Each lobule is composed of several seminiferous tubules (tubuli contorti) and the surrounding connective tissue. Spermatogenesis (formation of spermatozoa) occurs in the epithelial lining of the seminiferous tubules. The interstitium is composed of loose connective tissue containing fibroblasts and Leydig cells (interstitial cells). Spermatozoa produced in the seminiferous epithelium move through the lumen of the tubules to the tubuli recti (straight tubes) which extend to a network of spaces in the mediastinum, the rete testis (except in the stallion). Efferent ductules (ductuli efferentes) carry the spermatozoa from the rete testis, then converge to form the ductus epididymis, a convoluted duct. The ductus epididymis straightens and becomes the ductus deferens. In domestic mammals, testes are not in a major body cavity, but are enclosed in the scrotum. Each testis is suspended at the end of a tissue called the spermatic cord which contains the ductus deferens, the blood vessels, and the nerves supplying the testis.

Testis. Each testis is composed of an exocrine part (seminiferous tubules) and an endocrine part (interstitial or Leydig cells). The testis is divided into lobules by septa consisting of loose areolar connective tissue. Several seminiferous tubules are found in each lobule, and interstitial cells are found in the connective tissue septa surrounding the seminiferous tubules. The seminiferous tubules are the exocrine portion of the testis producing and "excreting"

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spermatozoa. These tubules are lined by a stratified epithelium that consists of the developing spermatozoa and supporting cells (Sertoli cells).

Seminiferous tubules. The stratified epithelium of the seminiferous tubules is composed of different stages of developing sperm cells. Spermatogonia are stem cells located near the basement membrane of the tubule which proliferate by mitosis. Some of the progeny cells differentiate into sperm and move away from the basement membrane toward the lumen of the tubule. These differentiating cells first undergo meiosis then undergo a morphological change to become spermatozoa. Some of the progeny cells undergo mitosis again to produce more progeny cells providing a continuous source of stem cells for the production of spermatozoa.

Interstitium. The interstitial tissue of the testis consists of loose areolar connective tissue containing numerous reticular fibers which serves to support the seminiferous tubules. The interstitial cells (Leydig cells), located in this connective tissue, are responsible for the synthesis and secretion of the steroid hormone testosterone.

Spermatogenesis: the process by which stem cells develop into mature spermatozoa. There are three phases: (1) Spermatocytogenesis (Mitosis), (2) Meiosis, and (3) Spermiogenesis.

1. Spermatocytogenesis (also called Mitosis): Stem cells (Type A spermatogonia; singular = spermatogonium) divide mitotically to replace themselves and to produce cells that begin differentiation (Type B spermatogonia). Spermatogonia have spherical or oval nuclei, and rest on the basement membrane. (You are not responsible for distinguishing between Type A and Type B spermatogonia in lab.)

2. Meiosis: Cells in prophase of the first meiotic division are primary spermatocytes. They are characterized by highly condensed chromosomes giving the nucleus a coarse chromatin pattern and an intermediate position in the seminiferous epithelium. This is a long stage, so many primary spermatocytes can be seen. Primary spermatocytes go through the first meiotic division and become secondary spermatocytes. The cells quickly proceed through this stage and complete the second meiotic division. Because this stage is short there are few secondary spermatocytes to be seen in sections. You are not responsible for identifying secondary spermatocytes in lab. Meiosis is the process by which the diploid number of chromosomes present in spermatogonia (the stem cells) is reduced to the haploid number present in mature spermatozoa.

The products of the second meiotic division are called spermatids. They are spherical cells with interphase nuclei, positioned high in the epithelium. Since spermatids go through a metamorphosis into spermatozoa, they occur in early through late stages. You are not responsible for distinguishing the different stages of spermatids, but you are required to identify a spermatid.

All of these progeny cells remain attached to each other by cytoplasmic bridges. The bridges remain until sperm are fully differentiated.

3. Spermiogenesis: This is the metamorphosis of spherical spermatids into elongated spermatozoa. No further mitosis or meiosis occurs. During spermiogenesis, the acrosome forms, the flagellar apparatus

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forms, and most excess cytoplasm (the residual body) is separated and left in the Sertoli cell. Spermatozoa are released into the lumen of the seminiferous tubule. A small amount of excess cytoplasm (the cytoplasmic droplet) is shed later in the epididymis.

Sertoli Cell & Developing Sperm Cells: an interaction

The Interaction At all stages of differentiation, the spermatogenic cells are in close contact with Sertoli cells which are thought to provide structural and metabolic support to the developing sperm cells. A single Sertoli cell extends from the basement membrane to the lumen of the seminiferous tubule although its cytoplasm is difficult to distinguish at the light microscopic level. They are characterized by the presence of a vesicular, oval, basally positioned nucleus which contains a prominent nucleolus. The nuclear envelope often contains a definite fold. The significance of the very close association of the two types of cells is unknown. Sertoli cells are endocrine cells - they secrete the polypeptide hormone, inhibin. Inhibin acts at the level of the pituitary to reduce the secretion of follicle stimulating hormone.

Blood-testis barrier. Large molecules cannot pass from the blood into the lumen of a seminiferous tubule due to the presence of tight junctions between adjacent Sertoli cells. The spermatogonia are in the basal compartment (deep to the level of the tight junctions) and the more mature forms such as primary and secondary spermatocytes and spermatids are in the adluminal compartment. The function of the blood-testis barrier (red highlight in diagram above) may be to prevent an auto-immune reaction. Mature sperm (and their antigens) arise long after immune tolerance is established; therefore, a male animal is capable of making antibodies against his own sperm. Injection of sperm antigens causes inflammation of the testis (autoimmune orchitis) and reduced fertility. Thus, the blood-testis barrier may reduce the likelihood that sperm proteins will induce an immune response.

The Duct System

After production in the testes, spermatozoa pass through a series of ducts in their journey out of the male system.

Tubuli recti, Rete Testis, Efferent ductules

The genital ducts are tubular organs in which the lamina epithelialis varies from the stratified epithelium in the testes to a transitional epithelium in the urethra. In the terminal part of the seminiferous tubules, the epithelium contains only Sertoli cells which gradually blends with the squamous, cuboidal or columnar epithelium (species variation) of the tubuli recti and the rete testis. These epithelial cells may actually represent a continuation of the Sertoli cells which line the seminiferous tubules.

Epididymis The ductus epididymis is lined with a pseudostratified stereociliated columnar epithelium. Stereocilia are actually nonmotile, long microvilli which serve to increase the absorptive and/or secretory surface of this epithelium. With its associated connective tissue and muscle, the ductus epididymis coils to form the head, body and tail of the epididymis which then continues into the

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ductus deferens. Spermatozoa are stored within the epididymis while they undergo maturation to become mature sperm.

Ductus Deferens The ductus epididymis continues as the ductus deferens which is also lined by a pseudostratified columnar epithelium. However, the lamina propria submucosa of the ductus deferens is areolar loose connective tissue, and the tunica muscularis is very thick and contains two layers of smooth muscle. The tunica serosa is present and typical.

Urethra The male urethra consists of two portions; the pelvic urethra and the penile urethra. Both portions are lined with transitional epithelium, both contain erectile tissue, and both contain (species variable) branched tubular mucous glands, the urethral glands. In the pelvic urethra, the three layers of smooth muscle in the tunica muscularis near the bladder are replaced (or joined in some species) by the striated urethral muscle. The tunica adventitia is present and typical.

The tunica muscularis is smooth muscle, and cavernous (corpus cavernosum urethra) tissue is present in the connective tissue beneath the epithelium. In the penile urethra, the corpus cavernosum penis is also present.

Accessory Glands

Ampullary, vesicular, prostate, bulbourethral, and urethral glands

The products of these glands serve to nourish and activate the spermatozoa, to clear the urethral tract prior to ejaculation, serve as the vehicle of transport of the spermatozoa in the female tract, and to plug the female tract after placement of spermatozoa to help ensure fertilization. Although the glands are ususally described as being branched tubular or branched tubuloalveolar, they vary in their organization and in their distribution in different species.

Ampullary Glands

This branched tubular gland lined by simple columnar epithelium is an enlargement of the ductus deferens in its terminal portion. This is a typical tubular gland in ruminants, horses and dogs; absent in the cat and poorly developed in boars. The function of the white serous secretion is not known.

Vesicular Glands

The secretory endpieces of this gland are lined with simple columnar epithelium; the main ducts are lined with stratified columnar epithelium. These glands do not occur in carnivores, but are present in some form in horses, ruminants and swine. Seminal fuid, the product of this gland, serves as a vehicle for the transport of spermatozoa.

Prostate Gland

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Grossly the prostate gland can be divided into two parts: the body and the disseminate part. Low cuboidal to low columnar epithelium provides the lining for this compound, tubuloalveolar gland which consists primarily of serous secretory end pieces. The secretion of this gland is more serous in dogs and more mucous in bulls. It serves to promote the movement of spermatozoa and to form a vaginal plug. Additionally, in bulls, the secretion contains high amounts of fructose and citric acid. Concretions may be present in the secretory end pieces as well as parts of the duct system.

Bulbourethral Glands

The lining of these paired, compound, tubuloalveloar glands is simple columnar epithelium. A capsule of dense connective tissue contains some smooth muscle as well as skeletal muscle of the bulbocavernous and urethral muscles. All domestic species have these glands except the dog, and their mucous secretion serves to clear the urethra of urine and to lubricate it and the vagina. The product may also serve as an energy source for the spermatozoa.

Urethral Glands

In some species, branched tubular mucous glands are found along the length of the urethra, especially dorsal to the lumen of the urethra. The exact function of their product is not clear.

Penis: the copulatory organ

The penis provides an outlet for both urine and the copulatory ejaculate (spermatozoa and semen). The histology and gross anatomy of the penis varies dramatically from species to species and from region to region within the same species. In general, the body of the penis consists of the urethra, erectile tissue (corpora cavernosa penis and corpora cavernosum urethra), smooth and skeletal muscle, touch and pressure receptors (Pacinian corpuscles) and a dense connective tissue capsule (tunica albuginea).

Erectile tissue and the erectile mechanism. The erectile tissue is composed of dense irregular connective tissue which contains numerous elastic fibers and sinuses. Under stimulation, the primary blood supply of the penis is directed through helicine arteries which open into the venous sinuses. During erection, these vessels and the sinuses become engorged with blood, and the thin-walled veins beneath the tunica albuginea are effectively closed, further increasing the rigidity of the organ. Because the capsule around the erectile tissue of the corpus cavernosum urethra is not as thick as that around the corpus cavernosum penis, the urethra is not occluded during erection. After ejaculation, the helicine arteries contract and regain their normal tone resulting in a relaxing of the pressure around the veins which leads to the restoration of normal bloo

d flow to the region.

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Female Reproductive System The primary function of the female reproductive system is reproduction, which includes

the production of ova

the transportation of ova from the ovary to the site of fertilization

transportation of spermatozoa from the point of deposition in the female tract to the site of fertilization

nourishment of the developing embryo and fetus

parturition and nourishment of the infant

Included in the reproductive function of this system is the production of the female hormones which are responsible for the secondary sex characteristics of the female animal as well as the development of follicles in the ovary, ovulation, preparation of the uterus for implantation of an embryo, maintenance of pregnancy, parturition and preparation of the mammary glands for milk production.

Structure

The female reproductive system is composed of several distinct organs. These include the paired ovaries, paired uterine tubes, uterus (uterine horns), cervix, vagina, and the mammary glands. The ovaries are both an exocrine organ producing cells, i.e., ova, and an endocrine organ, secreting hormones, i.e., estrogen and progesterone. Note: in domestic animals the oviducts are usually called uterine tubes and the uterus is called uterine horns due to the structure of these organs. Ova are produced in the ovaries then transported from the ovaries to the site of fertilization in the upper part of the uterine tube. Sperm are transported from the site of deposition near the vagina and uterus to the site of fertilization. If fertilization occurs, the uterus serves to nourish the developing embryo and fetus until the time of parturition. The vagina receives the male copulatory organ, the penis, during copulation and is the birth canal for the infant during parturition. Mammary glands serve to nourish the infant.

Although all mammals have the same basic organs, their individual structure and association with each other varies according to species. The structure of the uterine tubes and uterus are especially variable.

Function

The ovary, or female gonad, is:

1. an exocrine gland, producing ova 2. an endocrine gland, secreting

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a. the female hormones estrogen and progesterone androgens, typically considered male hormones

The surface of the ovary is covered with surface epithelium, a simple epithelium which changes from squamous to cuboidal with age. Immediately beneath this surface epithelium there is a dense connective tissue sheath, the tunica albuginea ovarii.

In most species, the ovaries are composed of an outer cortex and inner medulla (except in the mare where the cortical region is interior to the medulla). The cortex is composed of ovarian follicles (developing oocytes with their associated follicular cells), interstitial gland cells and stromal elements. Ovarian follicles are in different stages of development (least mature to most mature): primordial, primary, secondary, secondary-vesicular and mature.

The cortex usually also contains the remains of degenerated follicles called atretic follicles which may arise at any stage of follicular development.

Interstitial gland cells are also present in the cortex. Although the function of these cells is not known for sure, they are thought to secrete estrogen since they have the structure of steroid-secreting cells.

The atretic follicles and the interstitial gland cells, though not shown on this diagram, will be discussed later.

The medulla is composed of loose areolar connective tissue containing numerous elastic and reticular fibers, large blood vessels, nerves and lymphatics.

The hilus is the region through which blood vessels, lymphatics and nerves enter and leave the ovary. It is contiguous with and histologically similar to the medulla.

Oogenesis: the production of female gametes, the ova

1. Early in embryogenesis, primordial germ cells migrate from the yolk sac endoderm to the genital ridge (developing ovary) where they take up residence and are called oogonia.

2. These diploid oogonia undergo several mitotic divisions prior to or shortly after parturition, thus providing the developing ovary with a large supply of future ova (eggs).

3. When oogonia begin the first meiotic division, they are called primary oocytes.

4. Primary oocytes are arrested in prophase of Meiosis I until the female reaches sexual maturity. They grow in size during this arrested phase, but do not divide. A human female is born with about 2 million primary oocytes in her ovaries, but by the time of puberty only about 400,000 are left due to atresia (degeneration).

When the female reaches sexual maturity and under the influence of follicle stimulating hormone (FSH), a small number of primary oocytes are stimulated to continue through Meiosis I.

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6. During this process the number of chromosomes is reduced from the diploid number (2N) to the haploid number (1N).

7. This division is uneven in that although the chromosomes are divided equally, most of the cytoplasm stays with the oocyte. The smaller polar body contains half the chromosomes but only a small amount of cytoplasm and will eventually degenerate.

8. After a primary oocyte completes the first meiotic division, it is called a secondary oocyte (1N). In most species Meiosis I is completed just before ovulation (release of the ovum from the ovary). However, in horses and dogs Meiosis I is completed after ovulation.

9. If a secondary oocyte is not penetrated by a sperm, it will degenerate.

10. If fertilization and pregnancy do not occur, a new cycle will begin in which FSH from the pituitary gland will stimulate a few more primary oocytes to continue through Meiosis I. 11. The process is the same as previously described and a secondary oocyte is formed.

12. However, some of the time a sperm will penetrate the zona pellucida and the secondary oocyte is stimulated to continue through Meiosis II, forming a second polar body and a mature ovum (1N). Again, the polar body contains half of the chromosome material, but little cytoplasm, and it eventually degenerates.

13. After a sperm enters the cytoplasm of the ovum, two pronuclei form, containing genetic material from the ovum or the sperm.

14. Fertilization is complete when the two pronuclei fuse and restore the diploid chromosome number.

15. If fertilization is completed, the zygote undergoes several mitotic changes to become an embryo; otherwise it degenerates.

There are three stages in the development of follicles:

1. pre-ovulation 2. ovulation 3. post-ovulation.

The pars distalis of the pituitary gland (hypophysis) controls the process of follicular development and maturation by the secretion of:

1. gonadotropic hormones FSH (follicle stimulating hormone) 2. LH (luteinizing hormone) 3. prolactin (in some species).

The hormonal interactions will be explained in the discussion of the ovarian cycle.

Pre-ovulation: Development of Follicles in the Ovary.

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The primordial (quiescent) follicle consists of a primary oocyte and a single layer of flattened follicular cells. As the follicle develops, alterations occur in the primary oocyte and the surrounding follicular cells. The primary oocyte produces yolk granules and the follicular cells change from flattened to cuboidal or columnar.

The primary follicle consists of a primary oocyte with a single layer of cuboidal/columnar follicular cells. As development proceeds, the number of follicular cells increases by mitosis forming several layers around the primary oocyte. As these cells enlarge they release steroid hormones called estrogens of which estradiol is the dominant one prior to ovulation. During each cycle, a few primary follicles will continue to develop into secondary follicles.

The secondary follicle consists of several layers of cuboidal/columnar follicular cells, now collectively called the membrana granulosa which begin to secrete follicular fluid. A thick, amorphous layer, the zona pellucida, forms between the primary oocyte and the membrana granulosa. Previously undifferentiated stromal cells now develop into two distinct layers around the developing follicle: the theca interna and the theca externa . Cells in the theca interna are large, rounded and epithelial-like; cells in the theca externa are smaller, fibroblasts. Both layers of theca cells are separated from the membrana granulosa cells of the follicle by a basement membrane. As the follicular fluid secreted by the membrana graulosa cells accumulates, small pockets of fluid between granulosa cells begin to appear. Usually in human females only one secondary follicle will continue to develop.

The secondary-vesicular follicle is characterized by the presence of pockets of follicular fluid within the membrana granulosa. As the follicle continues to develop, the separate pockets fuse to form one large pocket of fluid called the follicular antrum.

During this development of the follicular antrum, the oocyte is still a primary oocyte, arrested in prophase of Meiosis I. It is still surrounded by granulosa cells which are contiguous with the membrana granulosa present around the periphery of the growing follicle.

Two regions of cells can be identified in the layer of granulosa cells surrounding the oocyte:

1. the corona radiata contains granulosa cells which remain attached to the oocyte after ovulation and are in close contact with the oocyte through cytoplasmic processes which pass through the zona pellucida and contact microvilli of the oocyte;

2. the cumulus oophorus contains granulosa cells which surround the oocyte and are continuous with the displaced cells of the membrana granulosa but remains in the ovary after ovulation.

The other granulosa cells form a layer around the periphery of the follicle and are separated from the theca interna cells by a distinct basement membrane.

The mature follicle, sometimes called the pre-ovulatory follicle, has all of the components of the secondary-vesicular follicle but is much larger and contains one single large antrum of follicular fluid. These follicles are very large and usually extend from the deepest parts of the cortex and protrude from the surface of the ovary. In some species just before ovulation, the primary oocyte in the mature follicle completes meiosis I producing a secondary oocyte and a polar body.

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Pre-ovulation: Development of Theca Cells in the Ovary.

As the oocyte and follicular (granulosa) cells are growing and developing in the ovary, the stromal cells differentiate and develop into the theca interna and theca externa cells. As a follicle goes from a primary to a secondary follicle, the stromal cells immediately surrounding the follicle differentiate into the theca folliculi. The cells closest to the follicle become the theca interna cells, round, foamy cells that secrete androgens, including testosterone. These two “male” hormones are converted by the granulosa cells to estrogens. The stomal cells farther away from the developing follicle become the theca externa cells, fibroblast-like cells arranged around the follicle outside the theca interna cells.

Post-ovulation

1. corpus hemorrhagicum After ovulation, hemorrhage into the remains of the follicle usually occurs resulting in a structure called a corpus hemorrhagicum. This transitory structure develops into a corpus luteum. 2. corpus luteum (yellow body) In most species LH from the pituitary gland initiates this luteinization and stimulates the granulosa cells to secrete progesterone. The granulosa cells undergo hyperplasia (proliferation), hypertrophy (enlargement) and are transformed into granulosa lutein cells. In several species, including the human, the accumulation of a yellow lipid pigment (lutein) and other lipids marks the transition to granulosa lutein cells. The cells of the theca interna are also transformed into lipid-forming cells called theca lutein cells. The resulting structure is highly vascular. If fertilization occurs, the corpus luteum persists and secretes progesterone. 3. orpus albicans If fertilization does not occur, the corpus luteum degenerates and is replaced by connective tissue forming a corpus albicans.

The Ovarian Cycle

Primordial follicles in the cortex of the ovary are stimulated by FSH (follicle stimulating hormone) secreted by the pars distalis of the pituitary gland. FSH stimulates the development of one or more primordial follicles in the ovary to begin the development into a mature (Graafian) follicle ready for ovulation. A surge of LH from the pars distalis initiates ovulation and induces luteinization of the granulosa and theca cells of the ruptured follicle. The oocyte with its surrounding corona radiata and cumulus cells moves into the uterine tubes while the ruptured follicle left behind becomes a corpus hemorrhagicum and under the influence of LH develops into a corpus luteum. If fertilization and implantation occur, the corpus luteum persists as the corpus luteum of pregnancy, but if fertilization does not occur, the corpus luteum degenerates into a corpus albicans which remains as a scar in the ovary.

The Uterine Tubes

Function The uterine tubes (also called Fallopian tubes or oviducts):

1. transport the ovum from the ovary to the site of fertilization

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2. help transport spermatozoa, the haploid male gametes, from the site of deposition to the site of fertilization

3. provide an appropriate environment for fertilization 4. transport the fertilized ovum (embryo) to the uterine horns where implantation and

further development may occur.

Structure The uterine tubes can be divided into three major parts:

1. the infundibulum 2. the ampulla 3. the isthmus

The infundibulum is the region most proximal to the ovary. It is funnel-shaped and has finger-like projections called fimbriae that extend into the pelvic cavity and make close contact with the ovaries. The tunica mucosa occupies most of the thickness of the wall of the organ.

The ampulla is the middle, one-third region in which fertilization usually occurs. Histologically it is very similar to the infundibulum having a very thick tunica mucosa and relatively thick tunica muscularis.

The thick-walled isthmus is the lower one-third region most proximal to the uterine horns. The smooth muscle in the wall of the isthmus helps propel (by peristalsis) the fertilized ovum toward the uterine horns and body of the uterus where implantation occurs. The tunica muscularis is the thickest part of the wall and the tunica submucosa is very thin as in the infundibulum and ampulla.

About the time of ovulation, the infundibulum, closest to the ovary, moves to cover the site of rupture of the mature (Graafian) follicle.

The ovum moves down the infundibulum of the uterine tube toward the ampulla, assisted by peristaltic contractions of the smooth muscle in the wall of the tube as well as fluid moved by ciliated epithelial cells in the mucosa of the tube. The ampulla is usually the site of fertilization.

After fertilization, the embryo moves down through the isthmus which connects the uterine tube with the uterine horns or uterus. The thick muscular wall of the isthmus of the uterine tube helps propel the embryo into the uterus where it can be nourished during further development.

Regional Variations:

The uterine tubes are paired tubular organs with the typical organization of a tubular organ, i.e., four tunics consisting of:

1. tunica mucosa 2. tunica submucosa 3. tunica muscularis 4. tunica serosa.

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The thickness and specific characteristics of these tunics varies with the region of the uterine tube. The tunica mucosa of the infundibulum helps capture the ovum from the surface of the ovary, bathes it in a supportive fluid and helps move it toward the uterus. The tunica mucosa of the ampulla provides the proper environment for fertilization. Consequently, in the infundibulum and ampulla the tunica mucosa is thick and highly developed.

It is the tunica muscularis of the isthmus that provides the strong contractions that at the right time propel the ovum or embryo into the uterine horns.

As a result, in the isthmus the tunica mucosa is reduced in thickness and the tunica muscularis is much thicker

Histology:

The lamina epithelialis of the tunica mucosa of the uterine tubes is an intermittently ciliated columnar epithelium that contains two types of cells: a ciliated cell and a non-ciliated, secretory cell. In the cow and sow the lamina epithelialis may be pseudostratified intermittently ciliated columnar. The secretory product of the non-ciliated, secretory cells is moved toward the uterine horns by the movement of the cilia on the ciliated cells. This secretion probably also protects and nourishes the ovum.

The lamina propria consists of a typical loose areolar connective tissue without glands, and it blends with the underlying, thin tunica submucosa. There is no lamina muscularis mucosae in the entire female reproductive tract. The tunica muscularis is sparse in the infundibulum and the ampulla but thick in the isthmus consisting of an inner circular and an outer longitudinal layer of smooth muscle. The tunica serosa is typical containing many blood vessels in a distinct vascular layer.

Cyclic Changes in the Epithelium : Under the influence of estrogen, the ciliated epithelial cells increase in height and in the number of cilia. Under the influence of progesterone, these cells decrease in height and in the number of cilia. These cells are at their tallest with the most numerous cilia at the time of ovulation. Their main function is to assist in the movement of the ovum toward the site of fertilization and the embryo toward the uterus. This action is secondary to the peristaltic movement of the isthmus region.

Clinical: The uterine tubes are the site of tubal ectopic pregnancies. They can also be the site of bacterial infection which can lead to Pelvic Inflammatory Disease, a major cause of infertility in women.

The Uterus

Functions

1. serves to receive the sperm in mares 2. transports sperm from site of deposition to uterine tubes for fertilization 3. provides suitable environment for

a. implantation of the embryo b. nourishment of the embryo & fetus during pregnancy

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4. provides mechanical protection of the fetus 5. expels the mature fetus at the end of pregnancy

In the fundus and body of the uterus, the wall is divided into the

1. endometrium = tunica mucosa and tunica submucosa 2. myometrium = tunica muscularis 3. perimetrium= tunica serosa

These are terms specific to the uterus that apply to the typical tunics of a tubular organ.

The endometrium comprises the tunica mucosa and the tunica submucosa of the uterus. In the tunica mucosa the lamina epithelialis is usually simple columnar except in the sow and ruminants where it may be pseudostratified columnar. The lamina propria consists of loose connective tissue full of neutrophils and lymphocytes. It blends with the underlying tunica submucosa since there is no lamina muscularis mucosae in the entire female reproductive tract. Uterine glands are simple or branched tubular glands located in the lamina propria-tunica submucosa. Some regions of the endometrium in ruminants are void of glands and are highly vascular. It is in these regions, called caruncles, that contacts between the uterus and the extraembryonic membranes are made.

The myometrium is the tunica muscularis of the uterus. It is composed of a thick inner circular layer and a thinner outer longitudinal layer of smooth muscle. The region in between the two layers of smooth muscle contains large blood vessels.

The perimetrium is the tunica serosa of the uterus. It has the typical composition of loose connective tissue, but contains a large number of lymphatic vessels.

The stratum vasculare is a layer of large blood vessels located between the inner and outer layers of smooth muscle of the myometrium.

In the sow the stratum vasculare is indistinct and in the cow it may be located in the outer half of the circular muscle layer.

Glands

Uterine glands are simple or branched tubular glands, and may be coiled distally. Distal portions of the glands are in the lamina propria/tunica submucosa. The glands secrete mucus, glycogen, proteins, and lipids. The remainder of the endometrial tissue is loose connective tissue.

The Uterus: Changes in the Uterus during the Estrous Cycle

Domestic animals are either monestrous or polyestrous. Monestrous animals such as bitches have one estrous cycle per year; each cycle is followed by a long anestrous period. Polyestrous animals are either continuously cycling without an anestrous period such as cows and sows or seasonally cycling with an anestrous period such as mares, ewes, goats and queens. The uterine wall, especially the endometrium, undergoes greater changes in the monestrous animal than in the polyestrous animal but changes can be clearly seen in polyestrous animals.

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During the estrous cycle the uterus undergoes changes controlled by hormones secreted by the ovary. The changes are most pronounced in the glands in the tunica submucosa of the endometrium and in the smooth muscle and stratum vasculare located in the tunica muscularis.

The Chart and Diagrams below allow you to see the differences in the uterine wall between Proestrus, early in the estrous cycle, and Diestrus, later in the estrous cycle. First read through the text and view the associated images, then review the material by comparing the appearance of the uterus in proestrus versus diestrus by using the colored bars below the text.

Proestrus

1. endometrial glands at lowest level of size & secretion 2. estrogen rises from granulosa cells in the developing follicles of the ovary 3. uterine epithelial cells hypertrophy 4. uterine glands proliferate 5. vascular supply increases

Estrus

1. after ovulation progesterone rises from · granulosa & theca lutein cells in the corpus hemorrhagicum · granulosa & theca lutein cells in the corpus luteum

2. uterine epithelial cells continue to hypertrophy 3. uterine glands continue to proliferate

Metestrus

1. progesterone levels remain high 2. uterine glands undergo hyperplasia & coiling 3. uterine glandular secretory activity high

Diestrus

1. endometrial glands at maximum size, coiling & secretion 2. normal state (no fertilization)

· secretory activity is arrested · lining cells and glands begin to involute · vascularity of the entire wall decreases fertilized state · secretory activity is maintained (in order to nourish embryo/fetus during pregnancy)

3. the wall of the uterus returns to a state that resembles that of the Proestrus uterus

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Vagina

Histology:

The epithelium of the vagina is stratified squamous, usually nonglandular. It increases in thickness during proestrus and estrus. In some species (especially rodent and carnivores), the epithelium keratinizes during estrus.

The tunica mucosa and submucosa are highly folded.

Lymphocytes, and lymphatic nodules can be found in the connective tissue.

The cranial portion of the vagina has a tunica serosa; the larger caudal portion has a tunica adventitia.

Tunica mucosa: lamina epithelialis of stratified squamous epithelium which is nonglandular; highly folded; lamina propria is loose connective which blends with the denser connective tissue of the tunica submucosa since a lamina muscularis mucosae is not present; the tunica mucosa is thin prior to the onset of puberty and in old age; thickens under the influence of estrogens during the reproductive years; superficial cells accumulate glycogen which is maximum at the time of ovulation. The acid pH (app. 3.0) in the vagina is due to the breakdown of this glycogen by commensal lactobacilli which produce lactic acid. Thus, only acid-loving bacteria and fungi can exist in this low pH environment, thus detering bacterial pathogens and fungi such as Candida albicans.

Tunica submucosa: highly folded

Tunica muscularis: composed of two-three layers of smooth muscle

Tunica serosa: present cranially then turns into a tunica adventitia caudally

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Endocrine System

Endocrine organs are organs whose cells secrete their products, i.e., hormones, into the bloodstream whereas exocrine organs such as sweat glands, salivary glands and sebaceous glands secrete their products into a duct system. Hormones travel via the blood circulation and when they reach their "target organ" they exert their specific effect. Some organs are primarily exocrine while some are primarily endocrine and some contain elements of both. The ovary and testes are both exocrine organs, "secreting" ova and spermatozoa, respectively yet they are endocrine organs as well secreting hormones such as estrogen, progesterone and testosterone. In the pancreas, part of the organ is an exocrine gland (the acini) secreting digestive enzymes and part is endocrine , i.e., the islets of Langerhans which secrete various hormones such as insulin and glucagon. In the digestive system, some cells are endocrine cells, i.e., the enteroendocrine cells.

Other endocrine organs include the thyroid, parathyroid, adrenal, and pituitary gland.

The Thyroid Gland

Gross Anatomy

The thyroid gland is located dorsolateral to the trachea, close to the larynx. It has two lobes that are connected by a narrow isthmus.

II. Histology

The thyroid gland is composed of follicles and interfollicular connective tissue. The capsule, classified as loose areolar connective tissue, surrounds the mass of thyroid

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follicles and sends smaller pieces of connective tissue into the gland to surround the individual thyroid follicles.

Near the thyroid gland and embedded in the same connective tissue capsule is the parathyroid gland.

Sometimes patches of lymphocytes can be observed in the thyroid/parathyroid glands.

Thyroid follicles consist of a layer of simple epithelium surrounding a gel-like pinkish material called colloid.

The principal cell is the most numerous cell present in the simple epithelial layer and is responsible for secreting the thyroid hormones as well as thyroglobulin, a glycoprotein.

Thyroid hormones are stored extracellularly as part of the thyroglobulin which is the main component of the colloid.

The size of follicles and the height of principal cells varies even within one section of the gland. Squamous principal cells indicate a relatively inactive gland whereas cuboidal to columnar cells indicate more activity in removing the hormone from the stored form.

In addition to principal cells there is another type of functional cell in the thyroid gland. This is the parafollicular cell which may be found as single cells in the epithelial lining of the follicle or in groups in the connective tissue between follicles. They usually appear as large, clear cells since they do not stain well with hematoxylin and eosin. They are sometimes called parafollicular cells based on their location and clear cells (C cells) based on their appearance of their cytoplasm. Parafollicular cells secrete calcitonin, a hormone that lowers the level of calcium in the blood.

III. Function

• secretes the thyroid hormones tri-iodothyronine (T3 ) and tetra-iodothyronine (T4 or thyroxin) that help to regulate the metabolic rate

• also secretes calcitonin that helps control blood calcium concentration

IV. Mechanism of Secretion of T3 and T4 (thyroxin).

• Under the influence of increased TSH from the pituitary gland, principal cells concentrate iodine by active transport. At the same time they synthesize thyroglobulin and secrete it into the lumen of the thyroid follicle

• The iodination reaction, catalyzed by the enzyme peroxidase, is carried out on the large thyroglobulin molecule at the luminal surface of the principal cell. Various combinations of iodinated and non-iodinated tyrosine are possible. If the two molecules of tyrosine are both fully iodinated, the hormone resulting upon

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cleavage is T4 but if one of the tyrosines has only one iodine, then the hormone that results is T3. In the circulation T4 is converted to T3 which appears to be the active form of the hormone.

• Under the influence of rising TSH levels, the principal cells take up colloid by pinocytosis, the vesicles fuse with lysosomes which hydrolyze throglobulin releasing T3 and T4 (thyroxin) which diffuse into the blood and lymph

V. Parafollicular Cells

• Secrete calcitonin which inhibits osteoclasts from resorbing bone resulting in decrease in calcium in the blood

• Controlled by the level of calcium in the blood

PARATHYROID GLAND I. Gross Anatomy The parathyroid gland is difficult to see at the gross level. It is very close to and usually embedded within the capsule of the thryroid gland. II. Histology There are three types of cells in the parathyroid gland: adipocytes, chief cells and oxyphil cells. A reticular connective tissue framework surrounds and supports these cells. The main secretory cell is the chief cell. These cells secrete parathyroid hormone. Unfortunately these cells have no distinguishing features.

Another cell type present is the oxyphil cell in the human, ox and horse. These are large cells that contain numerous mitochondria. Their function is unknown.

III. Function

The parathryoid gland secretes parathyroid hormone which is essential for regulating the levels of calcium and phosphate in the blood. Parathyroid hormone acts on the following target organs.

• Bone: increases blood calcium by inhibiting osteoblast deposition of calcium and stimulating osteoclast removal of calcium.

• Kidney: increases blood calcium by increasing calcium ion reabsorption by kidney tubular cells; inhibits reabsorption of phosphate ion from the glomerular filtrate

• Small intestine: increases the absorption of calcium from the small intestine

IV. Summary of hormonal control of blood calcium levels through action on bone.

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Calcitonin As the level of calcium in the blood rises, the amount of calcitonin secreted by the C cells of the thyroid increases. Calcitonin stimulates osteoblasts to form bone taking calcium out of the circulation. At the same time, calcitonin inhibits the mobilization of bone (and calcium) by osteoclasts. The end result is a decrease in the level of calcium in the blood thus helping to maintain proper blood calcium levels. Parathyroid Hormone A decrease in the normal levels of calcium in the blood causes the chief cells of the parathyroid gland to secrete more parathyroid hormone which stimulates osteoclasts to mobilize bone resulting in an increase in the level of calcium in the blood. Parathyroid hormone also increases Ca ion reabsorption in the kidney and decreases the reabsorption of phosphate ions.

Note: Whereas calcitonin is important in regulating the level of calcium in the blood, parathyroid hormone is essential!!

ADRENAL GLAND

I. Gross Anatomy The adrenal glands are located at the cranial end of the kidneys. They are flat organs embedded in fat. Each gland has an outer cortex that appears yellow in fresh tissue and an inner medulla that appears gray in fresh tissue.

II. Histology of the Adrenal Gland and Adrenal Cortex

The adrenal gland is surrounded on the surface by a connective tissue capsule. This capsule has projections into the cortex and through the cortex down into the medulla in some species.

In most species 4 cortical zones can be identified. From the zone nearest the capsule these are:

• Zona glomerulosa

o In ruminants this zone consists of cells in clusters.

o In carnivores, horses and pigs this zone consists of columnar cells in arches and is sometimes called a zona arcuata.

• Zona intermedia: This zone is relatively thin and contains mostly undifferentiated cells.

• Zona fasciculata This is the largest zone containing large, round, foamy-appearing cells arranged in cords that radiate from the zona glomerulosa down toward the medulla.

o Sinusoidal capillaries are located between cords of cells that are one cell thick.

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o Cells in the zona fasiculata have a foamy appearance due to the presence of many lipid droplets prior to processing for microscopy. These lipid droplets represent the precursors for steroid hormones.

• Zona reticularis

o Cells of the zona reticularis are arranged in a network of cords no longer arranged in parallel as in the zona fasiculata.

III. Function of the Adrenal Cortex

• Zona glomerulosa cells secrete mineralocorticoids (principal one is aldosterone). These steroid hormones act on the kidney to:

o Increase Na+ reabsorption o Increase K+ secretion

• Zona fasciculata and reticularis cells secrete glucocorticoids (e.g., cortisol, cortisone, corticosterone) which control glucose metabolism.

IV. Histology of the Adrenal Medulla

• Cells in the medulla are arranged in groups or cords, clustered around capillaries and venules.

• The cells have secretory granules which contain either epinephrine or norepinephrine.

• When fixed in potassium bichromate, the medullary cells become brown. Therefore, they are called chromaffin cells. The color is the result of a reaction between chromate and epinephrine or norepinephrine.

• Chromaffin cells are derived from neural crest cells. They are innervated by preganglionic sympathetic fibers. They release hormone by exocytosis when stimulated by those fibers.

V. Function of the Adrenal Medulla

• Secretes catecholamines (epinephrine and norepineprine).

VI. Vasculature of the adrenal gland.

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Blood is supplied to the adrenal gland via the suprarenal arteries. These arteries penetrate the capsule and form a plexus just beneath it. From this plexus, blood is further supplied via two different routes.

Cortical arteries of the subcapsular plexus branch into sinusoids in the cortex which intimately surround cells in the zona fasciculata and zona reticularis. These sinusoids drain into venules that empty into the central vein of the medulla. This blood supply provides a mechanism by which cells in the cortex can influence cells in the medulla.

The second route of blood supply is more direct to the medulla. In this case, cortical arteries run along trabecular branches of the connective tissue capsule directly into the medulla without forming capillaries or sinusoids in the cortex. These arteries then branch into capillaries in the medulla supplying the chromaffin cells with oxygenated blood. These capillaries join the same small medullary venules as the sinusoids of the cortex and all the blood flows into the central vein of the medulla. Because of this peculiar blood supply to the adrenal gland, the cortex has no veins.

PITUITARY GLAND (HYPOPHYSIS)

The pituitary gland or hypophysis is located at the base of the brain. Its function reflects its development from two different types of ectoderm, i.e., oral ectoderm which forms Rathke's pouch and neural ectoderm from the base of the diencephalon which forms the infundibulum. As developments proceeds, the ectoderm of the infundibulum grows downward and wraps around Rathke's pouch. In the adult the infundibulum forms the infundibular stalk and the pars nervosa; the ectoderm of Rathke's pouch forms the pars distalis and the pars intermedia. Some of the oral ectoderm remains associated with the infundibular stalk to form in the adult the pars tuberalis. A vestige of Rathke's pouch often remains visible in the adults of many species and forms a cleft that serves to distinguish the anterior lobe of the pituitary from the posterior lobe (useful in humans but not in domestic species).

The gland remains "connected" to the brain via both vascular and nervous routes (see below under Function). These relationships with the hypothalamus of the brain provide a basis for understanding the different functions of the pituitary gland.

Vascular Connection: The vascular connection provides the mechanism by which factors secreted by neuroendocrine cells in the hypothalamus reach and affect secretory cells in the pars distalis. These secretory cells respond to the "releasing factors" brought to them through the portal blood vessels.

Neural Connection: The neural connection provides the mechanism by which hormones secreted by neuroendocrine cells located in specific nuclei in the brain are actually released into the bloodstream at the level of the pars nervosa of the pituitary gland.

II. Histology

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The pituitary gland can be divided into various regions based on structure and function. One way to describe the pituitary gland is by the type of tissue present in different regions. If this system is used then regions are as follows:

Adenohypophysis - based on grouping of all regions composed of glandular tissue This includes the...

• pars distalis • pars intermedia • pars tuberalis

Neurohypophysis - based on grouping of all regions composed of neural or neurosecretory tissue This includes the . . .

• median eminence (not shown) • infundibular stalk • pars nervosa ( infundibular process)

Another way to describe the pituitary gland is using the terms anterior lobe and posterior lobe (in domestic species, ventral lobe and dorsal lobe) as follows . . . Anterior or Ventral Lobe This includes the . . .

• pars distalis • pars intermedia

Posterior or Dorsal Lobe This includes the . . .

• pars nervosa

The Adenohypophysis Pars distalis: This region of the pituitary gland is organized as cords or clusters of cells supported by a reticular connective tissue. With routine staining two types of cells can be observed: (1) chromophiles which stain readily and are either red (acidophiles), blue or purple (basophiles) depending on the type of secretory material present, and (2) chromophobes which do not take up the stain and thus appear unstained or rather clear. Chromophobes may be chromophiles that have lost their secretory granules or chromophiles that have not accumulated large numbers of secretory granules. Use of specific antibodies against the protein secretory products has allowed the identification of the different cells. The cells of the par distalis are:

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• Somatotrophs secrete growth hormone. • Mammotrophs secrete prolactin. • Corticotrophs secrete ACTH. • Thyrotrophs secrete TSH. • Gonadotrophs secrete FSH and LH.

Pars intermedia: With routine histological staining, the cells in the pars intermedia stain blue-purple and thus are basophilic.

• Cells secrete ACTH, MSH, endorphins and lipotrophins.

Pars tuberalis: This region is an extension of the glandular pituitary gland and its cells resemble those of the pars intermedia and pars distalis. The specific function of the cells in the pars tuberalis, however, is not clear. The Neurohypophysis Pars nervosa: This region consists of unmyelinated nerve axons (cell bodies are in the hypothalamus) and supportive cells called pituicytes.

• Secretes ADH (antidiuretic hormone) which is synthesized by neurons in the supraoptic nucleus of the hypothalamus.

• Secretes vasopressin which is synthesized by neurons in the paraventricular nucleus of the hypothalamus.

III. Function - Control of Secretion

Adenohypophysis.

The cells in the adenohypophysis secrete two classes of hormones: (1) direct acting and (2) trophic. Direct acting hormones include growth hormone (GH) and prolactin from the pars distalis, and melanocyte stimulating hormone (MSH) from the pars intermedia. Trophic hormones include adrenocorticotrophic hormone (ACTH), thyroid stimulating hormone (TSH), follicle stimulating hormone (FSH) and luteinizing hormone (LH).

Secretion of these hormones is controlled by specific releasing hormones in the hypothalamus. Most of the releasing hormones are stimulatory in their action except for the one for prolactin which is inhibitory and the one for growth hormone which has both inhibitory and stimulatory releasing hormones. Releasing hormones are produced in the median eminence of the hypothalamus and reach the adenohypophysis via a portal system of veins known as the pituitary portal system.

Neurohypophysis.

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The cells in the neurohypophysis secrete only direct acting hormones : (1) antidiuretic hormone (ADH) also known as vasopressin secreted by neurons in the supraoptic nucleus in the hypothalamus and (2) oxytocin secreted by neurons in the paraventricular nucleus in the hypothalamus. After synthesis in the hypothalamus, these hormones move down the axons of the hypothalamohypophyseal tract through the infundibular stalk and terminate near blood vessels in the pars nervosa. Accumulations of these hormones bound to specific glycoproteins can be observed along the axons of the hypothalamophypophyseal tract and in the pars nervosa. These "accumulations" often called Herring bodies represent a storage form of the hormone. Release of these hormone stores is determined by impulses in the axons of the hypothalamophypophyseal tract originating in the hypothalamus. Such a mechanism of secretion controlled by nerve impulses is called "neurosecretion".