Regulation of Neurotransmitters in Neuropsychiatric SystemAgnes KwenangDepartment of BiochemistryFac. of Medicine UNHAS

IntroductionNerve impulse transmission between cells is mediated by chemical mechanism, occurs by difussion of specific chemical compounds, which act as signals. Many of amino acids have been studied intensively because of their function in the nervous system. As prerequisite knowledge, we shall assume that the student has a degree of familiarity with the basic concepts of neuroanatomy and neurophysiology.

We shall mainly discuss about the regulation of neurotransmitter in neuropsychiatric system related to metabolism and function of amino acids group ( glutamate, and GABA) and biogenic amines group which amino acid act as precursor (catecholamines and serotonin).

General Learning Objective.Student should be able to understand the regulation of Neurotransmitters in Neuropsychiatric System.

Specific learning objectivesWhen student have mastered this topic, student should be able to :Define a neurotransmitter.List the neurotransmitters commonly found in brain and nerve.Describe the metabolism and function of amino acids act as a neurotransmitter (glutamate, and - amino butyric acid = GABA,).

Describe the metabolism and function of biogenic amines act as a neurotransmitter (catecholamines, and serotonin).Know the acetylcholine receptor, glutamate receptor, and dopamine receptor.

1. NEUROTRANSMITTERS definitionChemical compound is an established neurotransmitters must be met five important criteria :A synthetic mechanism exists within the presynaptic neuron;A mechanism of storage (in vesicles) in evident;

The transmitter is released in proportion to the strength of the stimulus (frequently of firing).Postsynaptic action of the transmitter has been demonstrated directly by microiontophoresis ; andAn efficient means for inactivation of the transmitter is present.

2. Classification of NeurotransmitterBiochemical Communication Hormones and NeurotransmittersTable 1.

(Voet: 2nd Ed. 1263)

HormoneOriginMajor effectsPolypeptides met-enkephalin Leu-enkephalin -endorphin

Aminoacid derivatives Epinephrine

NorepinephrineAdenohypophysis

Adrenal medulla

Adrenal medullaOpioid effects on CNS

Stimulates contraction of some smooth muscles and relaxes others.Increases heart rate and blood pressure.Stimulates glycogenolysis in liver, muscleStimulates lipolysis in adipose tissue.

Stimulates arteriole contractionDecreases peripheral circulationStimulates lipolysis in adipose tissue

Fig. 1

Substances that can be considered possibleNeurotransmitters or NeuromodulatorsTable 2.

Biogenic amines acetyl choline Catecholamines : Dopamine* Norepinephrine* Epinephrine* 5-hydroxytryptamine= serotonin* Other primary aminesHistamineOctopaminePhenylethylaminePhenyletahanolamine

PolyaminesPutrescineSpermineSpermidine

Amino acidsGlutamic acid*Aspartic acidGlycine-aminobutyric acid* = GABATaurineProlinePurinesAdenosineATP, etc

Regarded asEstablishedtransmittersPossible candidatetransmitters

NeuropeptidesSubstance PCarnosineThyrotropin-releasing hormone (TRH)NeurotensinSomatostatin-endorphin EnkephalinAngiotensin IAngiotensin IIOxytocinVasopressinCholecystokininBradykinin

(De Robertis: 690).AcetylcholineGlutamateGABAGlycineNorepinephrineDopamineSerotoninAspartateTaurineA large number of peptides

Nor epinephrineDopamineSerotoninVarious neuro peptidesSometimes called Neuromodulators rather than Neurotransmitters

This compound may not initiate a nerve impulse but may act on adenylate cyclase to increase or decrease cAMP levels and protein kinase activityThey may also diffuse through the extracellular space to influence a region of the brain greater than a single synaptic cleft.However, the distinction between transmitters and modulators is not exact.

Enzymes in neuromuscular junctions synthesize not only acetylcholine but also catecholamines, taurine and GABA.Some synapses in CNS release both Glycine and GABA. (Metzler 2nd ed. 1782).

Some of the Neurotransmitters found in Nervous Tissue.EXCITATORY :Acetylcholine, Aspartate, Dopamine, Histamine, Nor epinephrine, Epinephrine, ATP, glutamate, 5-hydroxytryptamine.INHIBITORY :GABA, Glycine

(Devlin : p.931)

Fig. 2. Involvement of the astrocytes in the metabolism of GABAand Glutamate

3. Metabolism and function of amino acids (GABA and Glutamate)GABA can be made and degraded through a series of reactions commonly known as the GABA SHUNT. In this reaction, -KG of the TCA cycle is transaminated to produce glutamate. Gludecarboxylase removes the 1-carboxyl group to produce GABA. The GABA is catabolized by deamination and oxidation reactions to produce succinate that can enter the TCA cycle.

GABA the inhibitory neurotransmitter and glutamate, the excitatory neurotransmitter, may share some common routes of metabolism involving the astrocytes. Both glutamate and GABA are taken up by astrocytes and converted to glutamate, which is then transported back into the presynaptic vesicles.

In the excitatory nerve, glutamine is converted to glutamate. In the inhibitory nerve, the glutamine is converted to GABA with glutamate as an intermediate.(Devlin :937-938)

Fig. 3

Fig. 4

GABA CycleIn the GABA cycle acetyl and OAA are converted into Citrate (step a) in the usual way and the citrate is then converted into 2-oxoglutarate (ketoglutarate). The latter is transformed to L-glutamate either by direct amination (b) and transamination (c), the amino donor being GABA.

GABA is formed by decarboxylation (d) of glutamate and is catabolized via transamination (e) to succinic semialdehyde, which is oxidation to succinate and OAA.The two transamination steps in the pathways may be linked, as indicated in Fig.3, to form a complete cycle that paralles the TCA but in which 2-oxoglutarate in oxidized to succinate via glutamate and GABA.No TPP is required, but 2-oxoglutarate, is reductively aminated to glutamate.

The Role of Glutamine in brain.As precursor of Glutamate in neuronal cells.Removed of ammonia to blood.Brain tissue can form urea, although this does not play a significant role in ammonia removal. Formation of Glutamine must be preceded by synthesis of Glutamate in the brain itself because the supply of blood glutamate is adequate to account for the increase blood glutamine formed in brain in the presence of high levels of blood ammonia.

The immediate source of glutamate for this purpose is -ketoglutarate.This would rapidly deplete the supply of TCA intermediates unless they could be replaced by CO2 fixation with conversion of pyruvate to oxaloacetic acid = OAA.

Fig. 6

4. Metabolism and function of Biogenic aminesRegulation of catecholamines.The rate limiting step in catecholamine synthesis is the conversion of Tyrosine to form dihydroxyphenylalanine by tyrosine hydroxylase. This enzyme is controlled in neurons, but not in adrenal medulla, by a calmodulin-sensitive protein kinase. The rise of calcium ion concentration cause by a neural impulse stimulates the kinase to phosphorylate an activator protein. The phosphorylated protein than combines with the tyrosine hydroxylase causing to initiate the replenishment of the catecholamines.

Defective regulation leading to excessively high concentrations of noradrenaline may be a factor in the development of schizophrenia.Fate of catecholamines.After discharge into the synaptic cleft as transmitters, catecholamines are recovered by an Na+-coupled symporter into the cytosol, from which they can be again concentrated into vesicles. However, catecholamines in the cytosol are subject to attack by a monoamine oxidase in the outer membrane of mitochondria.

This enzyme is an important protective device; its activity is high enough to remove amines rapidly.It is a flavoprotein that uses dioxygen and forms hydrogen peroxide as a product :

R-CH2-NH3+ + O2 R-CH=O + NH4+ + H2O2 AmineAldehydeA second enzyme in the cytosol methylates the 3-hydroxy group in the deaminated catechol compounds.

MAO

The methylated aldehyde that is formed may either be oxidized to an acid or to reduced to an alcohol. The alcohol (3-methoxy-4-hydroxy-phenylethylene glycol) is the principal product in the CNS. It is slowly transported into the blood and excreted in the urine. However, the principal catechol derivateve in the urine is the acid as its anion 3-methoxy-4-hydroxy mandelate, formed by the liver from catecholamines released by the adrenal medulla and other tissues.

The determination of its excretion is a useful guide to the clinical diagnostic management of tumors such as neuroblastoma and pheochromacytoma that form adrenaline, noradrenaline and dopamine. The compound is known clinically as vanillmandelate or vanillyl mandelate, an example of bastard nomenclature is excruciating to chemists. Its origin lies in the widespread occurrence of fragrant methylated 3,4-dihydroxy phenyl derivatives in plants; among the is vanillin in vanilla(Goldstein, 650-651)

Degradation of DopamineFig. 7

Catecholamines are rapidly metabolized byCatechol-O-methyl transferase = COMTMonoamine oxidase = MAOMAO is found in neural tissue and deaminates serotonin, epinephrine, norepinephrine.MAO-B is found in extraneural tissuesNerve impulses regulate catecholamine synthesis. Nerve stimulation synthesis of catecholamines.

NeuroblastomaMalignant neoplasma, neural crest and sympathetic nervous system frequently in adrenal medulla15-50% of neonatal malignancies, 5% of all childhood cancer deathSome produce : -dopamine-dopamine and NorepinephrineSerum level dopamine hydroxylase correlate well of urinary vanillyl mandelic acidDopamine excreted as Homovanillic acid

Parkinsons Disease1% of the population over 50 years oldClinical manifestation related to deficiency of Dopamine in areas of the brain responsible for motor stimulationA lack of sufficient dopamine production in certain brain structures like the substancia nigra or the destruction of this structure by toxic compound leads to Parkinsons Disease or Parkinson Syndrome

The key pathologic characteristic is degeneration of the pigmented cells in the substansia nigraAdministration of large quantities of L-DOPA can reverse many of the symptoms in some casesDopamine itself cannot readily enter the neurons and is rapidly destroyedIts precursor DOPA, enter the neurons

Large doses are required because dopa decarboxylase in other tissues will rapidly convert it to DOPAMINE unless a decarboxylase inhibitor is also administratedCarbidopa is used for this purpose because it does not enter the neurons

Goldstein, p.652

Fig. 8

Serotonin (5-hydroxy tryptamine)Serotonin is a transmitter in some neurons of the CNS. These neurons affect many aspects of behavior: appetite, agression, sleep, sexual activity, and so on. The route of formation is like that of noradrenaline.It begin with hydroxylation of C5 in tryptophan by a mixed function oxidase using tetrahydrobiopterin as the electron donor. The 5-hydroxy tryptophan is decarboxylated to form serotonin by dihydroxyphenylalanine decarboxylase.

The tryptophan-5-monooxygenase is activated. As with the catecholamines, released serotonin is recovered by an Natrium ion coupled symporter. Any of the compound that escapes packaging in vesicles is removed by the action of MAO and aldehyde dehydrogenase, which convert it to the corresponding carboxylate for excretion in the urine. (Goldstein : 654)

NeuropeptidesAre involved in the mediate of sensory and emotional responses such as those associated with hunger, thirst, sex, pleasure, pain, etc.Included in this category are peptides such as enkephalins, endorphins, and substance P.Substance P is an excitatory neurotransmitter that has role in pain transmission, whereasendorphins have role in eliminating the sensation of painSome of the peptides found in brain tissues(Devlin: 937.938)

5. RECEPTORSFig. 9. Major signaling pathways from metabotropic and ionotropic receptors in neurons.

Various G proteins control the signaling from metabotropic receptors using phosphatidyl inositol-specific phospholipase C (PI-PLC) and adenylate cyclase or activity directly on K+ ion channels(Metzler 2nd p.1775)

Receptor-associated ion channelsMany neurotransmitters, including acetylcholine and glutamate, act to open ion channels that are part of the receptor protein or of a tight complete of proteins.Binding of Acetylcholine to its receptor in the neuromuscular junction causes the release of Ca2+ ions from the exterior into the muscle fiber.

Acetylcholine receptor

A diagram of the neuromuscular junctionThe junction consists of a single nerve terminal separated from the postsynaptic region by the synaptic cleft.The motor end-plate is the specialized portion of the muscle membrane involved in the junctionJunctional folds are prominent; they content a high density of AChRs in close proximity to the nerve terminal.

The overall process at the junction: 6 steps1. Synthesis of Ach (cytosol nerve terminal) Enzyme : Cholineacetyl transferase2.Ach is then incorporated into small membrane-boundd particles called synaptic vesicles and stored therein.3. Release of Ach from these vesicles into the synaptic cleft is the next step. This occurs by exocytosis,which involves fusion of the vesicles with the presynaptic membranes.

In the resting state, single quanta (= 10.000 molecules of the transmitter, probably corresponding to the contents of one synaptic vesicle) are released spontaneously, resulting in small miniature end-plate potentials(MEPPs).When a nerve ending is depolarized by transmission of a nerve impuls, this process open voltage-sensitive Ca2+ channels, permitting an influx of Ca2+ from the synaptic space into the nerve terminal.This Ca2+ plays an essential role in the exocytosis that release Ach(contents of approximately 200 vspace.esicles) into the synaptic.

4.The released Ach diffuses rapidly across the synaptic cleft to its receptors in the junctional When 2 molecules of Ach bind to a receptor, it undergoes a conformatinal change, opening a channel in the receptor that permits a flux of cations across the membrane.The consequent entry of Na+ ions results in depolarization of the muscle mlate potentialembrane, forming the end-plate potential.This in turn depolarizes the adjacent muscle membrane, and action potentials are generated and transmitted along the fiber, resulting in contraction

5.When the channel closes, the Ach dissociates and is hydrolyzed by acetylcholinnesterase, which catalyzes the reaction:Ach + H2O Acetate + CholineThis important enzyme is present in high amounts in the basal lamina of the synaptic space.6. The choline recycled into the nerve terminal by an active transport mechanism, where it can be used again for synthesis of Ach.

The Acetylcholine Receptor of the Neuromuscular Junction is a Transmitter-gated ion Channel.The channel is closed in the absence of Ach.When 2 molecules of the transmitter bind to the receptor, one to each subunit, the protein undergoes a conformational change that results in opening of the ion channel for approximately 1 ms. During this time, Na+ flows in and K+ flows out. As mentioned above, it is the entry of Na+ that results in depolarization of the muscle membrane, generating the end-plate potential.

Because the presence or absence of Ach itself results in opening and closing of the channel, the Ach receptor is referred to as a transmitter-gated ion channelIn contrast to the 2 other types gated channels: voltage-gated and mechanical gated channels

Myasthenia GravisAutoantibodies damage Acetylcholine receptors and reduce their numbers.

Binding of glutamate to its ionotropic receptor in a synaptic ending of dendrite causes an influx of ions into the cytoplasm, initiating an action potential in the dendrite.In most instances the properties of the receptor channel favor the rapid flow of Ca2+ ions into the cytoplasm.Many other receptors are 7-helix transmembrane proteins, which activate guanine nucleotide G proteins.

The G proteins couple some receptors to adenylate cyclase and cyclic AMP-activated channel, and yet others via Phospholipase C to K+ channels and indirectly to Ca2+ channels. All of these G protein coupled receptors are referred to as metabotropic receptorsA single synapse often contain both ionotropic receptors and metabotropic receptors.The ionotropic receptors induce a rapid response, while the metabotropic receptors act more slowly.

However, in most cases the final effect is the release at Ca2+ ions into the cytoplasm.The rapid response may be initiation of an action potential, while the slow response may be activation of calmodulin-dependent kinases and phosphatases(Metzler 2nd 1774-1775)

Linked to intracellular production of diacylglycerol and inositol tri-polyphospo inositide

Glutamate receptorsSubdivided into 5 classes :NMDA (N-methyl-D-aspartate)AMPA (-amino-3hydroxy-5, methyl-4-isoxade propionic acid) = quisqualate receptorsKainate (a compund isolated from seaweed)L-AP4 (a synthetic agonist)Metabotropic receptors

Cation channels = ionotropic receptors

NMDA receptor.Excitotoxins : kainate, quinolinateStimulate NMDA res. causing release of excess glutamate

(depolarization plasma membrane) open ion channel. Allow accumulation of Ca2+ and Na+ into the cell osmotic swelling cell death

Fig. 10

It contains 5 distinct sites :A site binds the transmitter glutamateA regulatory site that binds glycineA voltage-dependent Mg2+ binding siteA site that binds phencyclidine and PCPA site that binds Zn2+(Murray : 24th ,798-799)

Alzheimers diseaseIs an incurable neuropsychiatric condition in which progressive impairment of cognitive functions occurs, usually accompanied by affective and behavioral disturbances.Is the commonest cause of dementiaLost of memory is often the first sign.The basic pathologic picture is of degenerative process characterized by the loss cells in certain areas of the brain (e.g the cortex and hippocampus).Amyloid protein is the major constituent of the plaques found in this disease.

Fig. 11. Schematic diagram illustrating the release of dopamine by a neuron in the substantia nigra and also showing the sites of action of drugs that ameliorate or induce parkinsonism.

The 6 steps described in the text are illustrated but are not numbered in the figure. The sites of actions of drugs (L-dopa, deprenyl, amantidine, and dopamine receptor agonists (eg. bromocriptine) that are used to treat parkinsonism are also shown; in general these drugs elevate local levels of dopamine. Thus countering its low level in parkinsonism, Sites of actio of certain drugs (reserpine and dopamine receptor antagonists, such as many neuroleptics) that induce parkinsonism by depleting dopamine or competing with it are also indicated. (Murray 24 th ed.805)

Dopamine acts as a neurotransmitterAs in the case of acetylcholine : divided into 6 steps.= process is involved in the action of monoamines neurotransmitters (norepi-nephrine, epinephrine).Synthesis of dopamineReaction is catalyzed by tyrosine hydroxylase is role-limiting

Storage of dopamine in synaptic vesicles.Entry of dopamine is driven by pit gradient established by a protein in vesicular membrane, that pump protons into the vesicle at the expense of ATP.Release of dopamine involves exocytosis.Binding of dopamine to its postsynaptic receptors.The amine reaches its receptors by diffusion across the synaptic cleft.

This produce their effector actions by affecting adenylyl cyclase positively or negatively or in at least one case by affecting another signaling system (Phospholipase C and Polyphospho inositide cycle).Reuptake of dopamineThis is achieved by a high-affinity transporter (which uses ATP) present in the presynaptic membrane. The recycled dopa-mine can again be incorporated into synaptic vesicles and reused as a transmitter.

Degradation of dopamine can occurWithin the synaptic cleft or following reuptake, within the presynaptic terminal.Dopamine DOPAC Homovanilic acid. Schizophrenia (homovanil acid in CSF)Deprenyl acts to inhibit MAO-B. (Murray 24th ed : 804-805)MAO-BCOMT

SchizophreniaGenetic, neurodevelopmental, dopaminergic factors may be involved in the causation of schizophrenia.Genetic linkage studies in schizophrenia suffer from a lack of replication.

Dopamine ReceptorGene cloning, at least five difference (D1 D5) with some subtypes formed by alternative splicing.They are membrane proteins, at least some of which are mic loops.Most appear to be coupled to G proteins.Some are positively coupled to adenylyl cyclase (eg. D1) at least one (D2) negatively.At least one subtype (of D1) appears to be coupled to Phospholipase C.

Some at least, are regulated by phosphorylation.Drug affinities of most neuroleptics for the D2 receptor reflect their potencies in treating Schizophrenia.The various show different anatomic distributions.The D4 receptor binds the atyprical neuroleptic clozapine with an affinity ten times higher than that of D2 sites.

Five distinct subtypes of the D4 receptor have been recognized, it being the first member of the catecholamine family of receptors to show polymorphic variation in the human population(Murray 24th , 811)

ConclusionWe have already discussed :Define a neurotransmitter List of the neurotransmitter.The metabolism and function of amino acids act as a neurotransmitter.The metabolism and function of biogenic amines act as a neurotransmitter.The receptors of glutamate and dopamine.

References :De Robertis EDP, De Robertis EMF Jr, 1987. Cell and Molecular Biology. 8th edition, Philadelphia : Lea and Febiger, 690.Devlin TH, 1993. Textbook of Biochemistry with Clinical Correlation 3rd edition. New York : Wiley-Liss Inc, 931, 937- 938Goldstein M, 1983. Biochemistry A Functional Approach 3rd edition. Tokyo : WB Saunders Company,650-652, 654, 648

Koolman J, Heinrich Rohm K, 2001. Color Atlas of Biochemistry. Terjemahan Jakarta : Penerbit Hipokrates, 316-317Metzler DE, 2003. Biochemistry the Clinical Reactions of Living Cells. 2nd edition. Vol 2, California, Elsevier Science, 958, 959, 1774, 1775, 1782Murray RK, Granner DK, Mayes PA, Rodwell VW, 1996. Harpers Biochemistry. 24th edition. Stamford : Appleton and Lange, 174, 303, 308, 563, 795-796, 798-799, 805.

Voet D, Voet JG, 1995. Biochemistry, 2nd edition. Canada : John Wiley and Sons Inc, 1263.

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