Essential Concepts: Mechanisms of Membrane Transport and Force GenerationJerome W. Breslin, PhDJerome W. Breslin, PhD

IDP/DPT GI Course, Fall 2011IDP/DPT GI Course, Fall 2011

Dept. of Physiology, LSUHSC-NODept. of Physiology, LSUHSC-NO

MEB 7208, MEB 7208, jbresl@lsuhsc.edu, x2669, x2669

OutlineOutlineComposition of Cell MembranesComposition of Cell Membranes

Transport Across MembranesTransport Across Membranes

Passive Diffusion, Facilitated Passive Diffusion, Facilitated Diffusion, and Active TransportDiffusion, and Active Transport

Membrane Potentials and Action Membrane Potentials and Action PotentialsPotentials

Smooth Muscle and Excitation Contraction Smooth Muscle and Excitation Contraction CouplingCoupling

Suggested ReadingSuggested Reading

William Ganong, William Ganong, Review of Medical Review of Medical PhysiologyPhysiology

Chapters 1 and 2Chapters 1 and 2

This textbook is available free to This textbook is available free to LSUHSC students online at LSUHSC students online at www.accessmedicine.com and also and also through our library’s website.through our library’s website.

Distribution of Distribution of body fluids:body fluids:

Important forImportant for1. tissue 1. tissue

homeostasishomeostasis2. nutrient 2. nutrient deliverydelivery

3. drug delivery3. drug delivery4. performance4. performance

5. immune 5. immune functionfunction

Transport:Transport:Across tissue barriersAcross tissue barriers

Gut Wall (absorption of nutrients)Gut Wall (absorption of nutrients)

Lung Alveoli (exchange of gases)Lung Alveoli (exchange of gases)

Capillary Beds (blood-tissue exchange of Capillary Beds (blood-tissue exchange of nutrients, gases, and waste products)nutrients, gases, and waste products)

Nephrons in kidneys (urine formation)Nephrons in kidneys (urine formation)

Transcellular versus ParacellularTranscellular versus Paracellular

Across cell membranesAcross cell membranes

Regulation of cell sizeRegulation of cell size

Regulation of cell electrical activityRegulation of cell electrical activity

Passive vs. Active TransportPassive vs. Active Transport

Example of transport across a tissue:Example of transport across a tissue:Epithelium of small intestineEpithelium of small intestine

Ganong,Ganong,Fig. 25-7Fig. 25-7

Find: trancellular transport, Find: trancellular transport, paracellular transportparacellular transport

Fig. 1-5Fig. 1-5GanongGanong

Cell Membranes: Fluid Mosaic ModelCell Membranes: Fluid Mosaic ModelPhospholipid bilayer, and integral membrane proteinsPhospholipid bilayer, and integral membrane proteins

Cell membranes: Selective Cell membranes: Selective BarriersBarriers

Lipophilic compounds can pass through.Lipophilic compounds can pass through.

Hydrophilic compounds do not pass through.Hydrophilic compounds do not pass through.

Specialized transporters and channels Specialized transporters and channels allow transport of hydrophilic compounds.allow transport of hydrophilic compounds.

Channels - selective pores for ions, Channels - selective pores for ions, water, and very small molecules. water, and very small molecules. Constructed from proteins.Constructed from proteins.

Transporters - membrane proteins that Transporters - membrane proteins that move particular ions or small molecules move particular ions or small molecules across membranes.across membranes.

Transport across membranes:Transport across membranes:Passive TransportPassive Transport

Diffusion, OsmosisDiffusion, Osmosis

Solutes or water cross a membrane Solutes or water cross a membrane by following a concentration by following a concentration gradient.gradient.

Facilitated DiffusionFacilitated Diffusion

Carrier Molecule binds a solute Carrier Molecule binds a solute and brings it across the membrane and brings it across the membrane with no energy expended.with no energy expended.

Active TransportActive Transport

Requires energy expenditure, and Requires energy expenditure, and can go against a gradient.can go against a gradient.

Diffusion: Fick’s Diffusion: Fick’s First LawFirst Law

JJss = -PS(C = -PS(Coo - C - Cii))“Outside” “Inside”

JJss = solute flux; P = permeability = solute flux; P = permeability coefficient;coefficient;

S = surface area for diffusion;S = surface area for diffusion;(C(C00 - C - Cii) = concentration gradient of ) = concentration gradient of

the solutethe solute

What determines permeability (P)?What determines permeability (P)?

P = kT/6πrηxP = kT/6πrηx

k = Boltzmann’s constantk = Boltzmann’s constantT = absolute temperatureT = absolute temperaturer = radius of the soluter = radius of the solute

η = viscosity of the mediumη = viscosity of the mediumx = thickness of membranex = thickness of membrane

Osmosis: Transport of WaterOsmosis: Transport of WaterWater also moves down its concentration gradient.Water also moves down its concentration gradient.

Osmotic pressure = amount of pressure to keep Osmotic pressure = amount of pressure to keep water from entering side B in this diagram.water from entering side B in this diagram.

B AA B

OsmosisOsmosisDescribed by Described by van’t Hoff Equationvan’t Hoff Equation::

π = RT(π = RT(ϕϕic)ic)π = osmotic pressureπ = osmotic pressure

R = ideal gas constantR = ideal gas constant

T = absolute temperatureT = absolute temperature

ϕϕ = osmotic coefficient = osmotic coefficient

i = number of ions formed by i = number of ions formed by dissociation of a solute moleculedissociation of a solute molecule

c = molar concentration of a solutec = molar concentration of a solute

((ϕϕic) = osmolarity of the solutionic) = osmolarity of the solution

Molarity and Molarity and OsmolarityOsmolarityMolarity = number of molecules in solution Molarity = number of molecules in solution (moles/L).(moles/L).

Osmolarity = number of particles in Osmolarity = number of particles in solution (osmoles/L).solution (osmoles/L).

So, for molecules that don’t dissociate in So, for molecules that don’t dissociate in solution, molarity = osmolaritysolution, molarity = osmolarity

For molecules that do dissociate in For molecules that do dissociate in solution, osmolarity will be higher than solution, osmolarity will be higher than molarity.molarity.

Osmolarity vs. Osmolarity vs. MolarityMolarityExamples:Examples:

GlucoseGlucose - doesn’t dissociate in solution. If one - doesn’t dissociate in solution. If one mole is dissolved in one liter of water, you get mole is dissolved in one liter of water, you get 1 mol/L or 1 M. Osmolarity = 1 osmoles/L.1 mol/L or 1 M. Osmolarity = 1 osmoles/L.

NaClNaCl - dissociates into Na and Cl ions in - dissociates into Na and Cl ions in solution, so, a 1 M solution has an osmolarity solution, so, a 1 M solution has an osmolarity of 2 osmoles/L.of 2 osmoles/L.

CaClCaCl22 - dissociates into Ca and 2 Cl ions in - dissociates into Ca and 2 Cl ions in solution, so a 1 M solution has an osmolarity of solution, so a 1 M solution has an osmolarity of 3 osmoles/L.3 osmoles/L.

How do permeating How do permeating solutes affect solutes affect osmotic flow?osmotic flow?Flow water (V) = “Permeability” of Water Flow water (V) = “Permeability” of Water (L) x Pressure Gradient(L) x Pressure Gradient

L = hydraulic conductivityL = hydraulic conductivity

The pressure gradient is directly The pressure gradient is directly proportional to π. (V = Lπ) when solutes proportional to π. (V = Lπ) when solutes are impermeable.are impermeable.

Correction with permeant solute:Correction with permeant solute: V = V = σσLLΔΔππ

σσ = reflection coefficient, a value from 0 = reflection coefficient, a value from 0 to 1to 1

0 = freely permeable; 1 = impermeable0 = freely permeable; 1 = impermeable

Modes of transport for ions Modes of transport for ions and small molecules across and small molecules across biological membranesbiological membranes

ChannelsChannels

TransportersTransporters

ATPasesATPases

Other - endocytosis (not Other - endocytosis (not really across membrane, but really across membrane, but into vesicle inside cell)into vesicle inside cell)

Fig. 1-30Fig. 1-30

Ion Channels -- “gated pores” for particular ionsIon Channels -- “gated pores” for particular ions

Ligand-gated ion channelLigand-gated ion channel

Voltage-Voltage-gated ion gated ion channelchannel

Electrical recording of gatingElectrical recording of gating

Fig. 1-28Fig. 1-28

Patch Clamping is an experimental Patch Clamping is an experimental technique to measure gating of ion technique to measure gating of ion

channels.channels.

Fig. 1-32Fig. 1-32

Active TransportActive TransportExample: Sodium-Potassium ATPase (Na/K Pump)Example: Sodium-Potassium ATPase (Na/K Pump)

Costs 1 ATP to transport 3 Na Costs 1 ATP to transport 3 Na and 2 Kand 2 K

Transporters (Facilitated Diffusion)Transporters (Facilitated Diffusion)Selective for particular solutesSelective for particular solutes

Can be ReversibleCan be Reversible

Example of transport across a tissue:Example of transport across a tissue:Epithelium of small intestineEpithelium of small intestine

Ganong,Ganong,Fig. 25-7Fig. 25-7Find: trancellular transport, Find: trancellular transport,

paracellular transport, passive paracellular transport, passive diffusion, facilitated diffusion, active diffusion, facilitated diffusion, active

transporttransport

Transporters are important for absorption in the GI tract.Transporters are important for absorption in the GI tract.

Fig. 1-13Fig. 1-13

Gap Junctions (Connexons) are specialized Gap Junctions (Connexons) are specialized channels that allow direct movement of channels that allow direct movement of

ions and water between cellsions and water between cells

Fig. 1-25Fig. 1-25

Cells also can pick up and release water Cells also can pick up and release water and solutes using vesicles.and solutes using vesicles.

ExocytosisExocytosis

EndocytosisEndocytosis

Fig. 1-34Fig. 1-34

Different modes of communication between cellsDifferent modes of communication between cells

How are signals spread inHow are signals spread in neurons and muscle cells?neurons and muscle cells?

Fig. 1-33Fig. 1-33

The composition of various transporters The composition of various transporters and channels on a cell determines and channels on a cell determines

membrane potential (Vmembrane potential (Vmm).).

Concentration (mM)Concentration (mM)

IonIon Inside CellInside Cell Outside Outside CellCell

Equilibrium Equilibrium Potential Potential

(mV)(mV)

NaNa++ 1414 142142 +61+61

KK++ 140140 44 -94-94

ClCl-- 99 125125 -70-70

Typical Concentrations of ions inside Typical Concentrations of ions inside and outside human nerve cells:and outside human nerve cells:

Ionic equilibria can produce Ionic equilibria can produce electrochemical potential electrochemical potential

differences.differences.A B

100 mM K+ 10 mM K+

Δμ(KΔμ(K++) = RTln([K) = RTln([K++]]AA/[K/[K++]]BB) + zF(E) + zF(EAA - E - EBB))

= “diffusive” + “electrical” influences = “diffusive” + “electrical” influences

Electrochemical equilibrium can be Electrochemical equilibrium can be described by the Nernst Equation:described by the Nernst Equation:

At equilibrium, Δμ(KAt equilibrium, Δμ(K++) = 0,) = 0,soso

at equilibrium:at equilibrium:RTln([KRTln([K++]]AA/[K/[K++]]BB) = zF(E) = zF(EAA - E - EBB),),

oror

EEAA - E - EB B = (RT/zF)ln([K= (RT/zF)ln([K++]]AA/[K/[K++]]BB))

solving for RT/F and converting ln to solving for RT/F and converting ln to log,log,

EEAA - E - EB B = (-60mV/+1) x log([K= (-60mV/+1) x log([K++]]AA/[K/[K++]]BB))

A B

100 mM K+ 10 mM K+

So for the above example,So for the above example,EEAA - E - EB B = (-60mV/+1) x log([0.1 = (-60mV/+1) x log([0.1

M]/[0.01 M])M]/[0.01 M])

EEAA - E - EB B = -60 mV x log 10= -60 mV x log 10

EEAA - E - EB B = -60 mV= -60 mV

What does this mean?What does this mean?

To hold a 10-fold concentration To hold a 10-fold concentration difference of K+difference of K+

across a membrane, an electrical across a membrane, an electrical potential differencepotential difference

of about 60 mV is required.of about 60 mV is required.

Why is this important?Why is this important?

These potential differences are an These potential differences are an energy sourceenergy source

transmission of signals in neurons and transmission of signals in neurons and muscle cells.muscle cells.

Gibbs-Donnan Gibbs-Donnan EquilibriumEquilibriumThe cytoplasm contains many large The cytoplasm contains many large proteins, which typically have a proteins, which typically have a negative charge. This influences how negative charge. This influences how cation-anion pairs distribute across cation-anion pairs distribute across the membrane, creating an uneven the membrane, creating an uneven distribution.distribution.

These nondiffusible ions influence how These nondiffusible ions influence how diffusible ions distribute across a diffusible ions distribute across a membrane, and is known as the membrane, and is known as the Gibbs-Gibbs-Donnan EffectDonnan Effect..

Fig. 1-33Fig. 1-33

The composition of various transporters The composition of various transporters and channels on a cell determines and channels on a cell determines

membrane potential (Vmembrane potential (Vmm).).

Resting Membrane Potential:Resting Membrane Potential:

Depends mainly on electrical potential Depends mainly on electrical potential differences generated by Na, K, and Cl differences generated by Na, K, and Cl

across cell membrane.across cell membrane.

EENaNa = (-60mV/+1) x log([14 mM]/[142 mM] = = (-60mV/+1) x log([14 mM]/[142 mM] = +60 mV+60 mV

EEK K = (-60mV/+1) x log([140 mM]/[4 mM] = -= (-60mV/+1) x log([140 mM]/[4 mM] = -93 mV93 mV

EEClCl = (-60mV/-1) x log([9 mM]/[125 mM] = = (-60mV/-1) x log([9 mM]/[125 mM] = -68 mV-68 mV

The resting potential is estimated by a The resting potential is estimated by a weightedweighted

average of the above potentials.average of the above potentials. Weight Weight is based on a parameter called is based on a parameter called

conductanceconductance (inverse of electrical (inverse of electrical resistance)resistance)

Actual resting potentials (EActual resting potentials (Emm) are about -70 mV.) are about -70 mV.

EENaNa = +60 mV, = +60 mV,which is a 130 mV difference from Ewhich is a 130 mV difference from Emm

This difference represents a This difference represents a potential driving force.potential driving force.

Nerve and muscle cells use this to Nerve and muscle cells use this to propagate signals.propagate signals.

Fig. 1-30Fig. 1-30

Voltage Gated Ion Channels - open or Voltage Gated Ion Channels - open or closed depending on membrane potentialclosed depending on membrane potential

At a “threshold” potential, voltage-At a “threshold” potential, voltage-gated sodium channels open, causing gated sodium channels open, causing rapid influx of sodium an driving rapid influx of sodium an driving

membrane potential up (membrane potential up (start of action start of action potentialpotential))

Action Potential in Nerve CellsAction Potential in Nerve Cells

Rhythmic waves of smooth muscle contraction in the gut are the result of waves of action potentials moving along via gap junctions.

Figure 15-23

Contraction of GI Smooth Muscle Cells

Regulation of Regulation of vascular smooth musclevascular smooth muscle tone: tone:Role of calcium in E-C couplingRole of calcium in E-C coupling

Fig. 21-2, Fig. 21-2, Berne & Berne & LevyLevy

MLC KinaseMLC Kinase

MLC-PPMLC-PP

MLCMLCCa-CalmodulinCa-Calmodulin

MLC PhosphataseMLC Phosphatase

Activation of Actin-Myosin ATPaseActivation of Actin-Myosin ATPase

Actin-myosin cross bridge reactionActin-myosin cross bridge reaction

ContractionContraction

??

Types of smooth Types of smooth muscle contraction:muscle contraction:

Tonic Contraction = Constrict or RelaxTonic Contraction = Constrict or Relax

All Smooth Muscle TypesAll Smooth Muscle Types

Phasic Contraction = Short contraction Phasic Contraction = Short contraction followed by period of relaxationfollowed by period of relaxation

GI smooth muscle - mixing GI smooth muscle - mixing contractions - will cover in more contractions - will cover in more detail in next lecture.detail in next lecture.