What is a membrane potential? How do we know a cell is “alive”? 10/24 10/27

How do we calculate Vm when many different ions and permeabilities exist? GHK-Equation

How do environmental changes affect cellular Vm?Why are patch clamps useful for studying Vm?What are the properties of voltage-gated channels?What is “self-propagation” and why is this property important

with regards to a cellular membrane potential?What is saltatory conduction and why is it so fast?How do gap junctions create an electric syncitium in the heart?How do action potentials use ligand-gated channels to cross

synaptic clefts between cells and create new APs in adjacent cells?

Good practice problems: 13-1, 2, 4, 5, 6, 7 and 8

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It is important to remember that membranes are “selectively” permeable and that this property generates “gradients” which represent potential energy and even voltage (ions).

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Remember that there are considerable differences within organs and across species, these are “ballpark” figures. There will be a TQ asking for typical intra and extracellular ion concentrations for a mammalian cell.

Because membranes are permeable to many different ions at different relative permeabilities, we need a more complicated equation to determine the true membrane potential!Goldman-Hodgkin-Katz Eq. Predicts Membrane

Potential (voltage)!Equation lets us predict how different ions and permeabilities

interact for a predictable Memb.Pot!Applications: Toxicity, Environmental Quality, Nutrition,

Human/Vet.Med…uses are innumerable!

• Vm=2.303RT/F log Pk(K)o+Pna(Na)o+Pcl(Cl)i Pk(K)i+Pna(Na)i+Pcl(Cl)o

F=FaradayConstant= 23,062 cal/Volt-moleRT as before:R= GasConstant=1.987 cal/mole-degrees Temp=298K (20oC)P=permeability relative to most permeable ion (K+)

100%=1.0 10%=0.1….not used as an absolute valueP values: K=100% Pk=1.0; Cl=45% Kcl=.45; Na=4% Pna=.04

• Ca++: negligible perm. at cellular rest, so folks generally ignore its contribution!

Example: How to calculate Vm given ion concentrations

What is the Vm for a normal neuron in your brain?

• Vm=2.303RT/F log Pk(K)o+Pna(Na)o+Pcl(Cl)i• Pk(K)i+Pna(Na)i+Pcl(Cl)o

i=intracellular (inside) o=extracellular (outside)

• Vm=2.303RT/F log 1(.005)o+0.04(0.14)o+0.45(.01)i• 1(.14)i+0.04(0.01)i+0.45(.125)o• Vm=_____volts or _____millivolts• Vm= Try the math for the test! What happens to Vm if

the Na+ permeability changed to from 0.04 to 1??

• Why does [K+]normally matter most for Vm at rest?Why does [Na+] determine Vm during action potential

when a temporarily OPENING of Na-channels occurs?• Why is the permeability for chloride ions ONE (1)?

What happens to Vm values of a cardiac muscle cell if you eat a banana (rich in KCl) or some

salty chips (NaCl)? Which causes a heart attack Na+ or K+?• Banana: assume plasma K+ goes to 15 mM/Cl goes to 135mM

• Vm=2.303RT/F log 1.0( )o+0.04( )o+0.45( )i• 1.0( )i+0.04( )i+0.45( )o• Salty Chips: assume plasma Na+ goes to 160mM/Cl=145 mM• Vm=2.303RT/F log 1.0( )o+0.04( )o+0.45( )i• 1.0( )i+0.04( )i+0.45( )o• What happens to salt if a toxin increases Pna to 0.40?• Vm=2.303RT/F log 1.0( )o+0.40( )o+0.45( )i• 1.0( )i+0.40( )i+0.45( )o

• What are some environmental conditions that could cause these effects?

• Why does plasma calcium have little effect on Vm for a healthy cell?

• What happens to Vm when a toxin changes the permeability?

How are Transmembrane Channel types characterized?

• Some Physical Properties to consider:– Membrane potential: mV difference across membrane– Channel Current: Amperes..means ions are moving!

• Channel type?• Ion type?• Ions/second passing through?

– Conductance: movement of ions when a specific voltage is applied across the membrane

• 4 Membrane Ion Channels: “4 ways to open”– 1) Ligand Gated Channel:– 2) Mechanosensitive Channel:– 3) Signal Gated Channel:

• Any of these three can initiate an action potential!– 4) Voltage Gated channel (VGC): V.I.P.!!

• This is the channel that “Propogates” an action potential!

These Channels can be in OPEN or CLOSED stateQuestion: is channel open or closed and for how

long?

There are four different types of channel (mechanosenitive not shown below).

Remember that pores and porins are always open!

What are some properties specific to the VGC?

Function of VGC: Convert a change in Vm into a change in ion permeability! Create Action Potential!

Three Main Types of VGC: Na, K, and CaVGCs have an amino acid segment that changes

conformation if the membrane potential changes!

VGCs have excellent ion selectivity:– Related to size of charge and atomic mass!– Related to size of sphere of hydration? MOST IMPORTANT

VGCs have channel gate(s):VGCs can be opened(activated)/closed (deactivated)VGCs modulate ACUTE CHANGES in permeability!

What do K+-VGC look like? Why is the sphere of hydration important? Which is bigger with respect to its atomic mass and sphere of

hydration? Na+ or K+

The sodium VGC actually has two gates! Some books simplify it a bit by calling it one gate. Often times environmental toxins work by disabling these VGC! Some classic Na-VGC neurotoxins from pufferfish(tetrodotoxin or TTX) and red tide(dinoflagellates) work at this channel (fatal).What would these toxins do to action potential formation and “breathing”?

Patch clamps are fantastic tools for isolating channels and evaluating each type of membrane channel for its contribution to total membrane ion permeability. If the scientist is “lucky” they hope to get a patch with just one channel protein (voltage and amps current).

Patch Clamps measure “flickers” created by the opening and closing of an individual channel or channels! We often times determine the nature of a toxin by examining effects on membrane potentials with patch clamps.In the example below, the patch contains TWO channels, each opening at different times, and occasionally at the same time (doubling the current)!

VGC have a unique amino acid sequence (letters): some parts are variable between different VGCs (not shaded below), and some (in pink below) give each type of VGC a specificity for a single ion: Na+, K+ or Ca++Why does similarity/dissimilarity explain why some drugs affect only certain ion permeabilities, people or different species of animal?

What does an action potential look like as it starts self propagating?What happens to Na, K, Na-VGC, K-VGC and Vm over time?Can you draw/describe this for a test?

• Lets sketch this on the membrane below:

• Extracellular• ---------------------------------------------------------------------------------------• ---------------------------------------------------------------------------------------• Intracellular• Side

• The key is that when the VGC open at “A”, it changes the ions overlying the VGC at “B” causing them to open too, this causes the VGC at “C” to open and the AP can self-propagate on down the membrane!

Dendrites carry the AP towards the cell body and axons carry the AP away from cell body to the next cell! If the stimulus occurs

artificially in the middle of an axon, the AP can travel both ways!

Once the membrane is depolarized how do we get its depolarized membrane potential back to its original hyperpolarized state?Solution:1) We limit the amount of time the VGC stay open! 2) Hyperpolarize Na-VGC before they can open again!

What are the terms used to describe the events/changes in membrane potential (mV) and time (msec) on the last diagram?

• #1Resting Membrane Potential• “Sub-threshold Depolarization”

• #2“Depolarization Phase”• “Overshoot”• #3“Repolarization Phase”• #4“Undershoot/Hyperpolarization Phase”

• Refractory Period: V.I.P.• “Absolute” Refractory Period- can channels open?• “Relative” Refractory Period- will high P-K+ let membrane

depolarize? Would an electric shock open channels?

The changes in Vm occur because the many many gates open or close. In the end the Na/K-ATPase cleans up the rest to get back to the original 140 mM/10mM concentrations! Conductance describes permeability

Once a local depolarization that exceeds threshold is created the AP spreads in either direction from the origin. Normally this is from the cell body towards the synapse!

What happens to the AP-velocity if we wrap layers of electrical insulation (sphingomyelin) in a sheath between pockets of VGC?Extremely Rapid Saltatory Conduction Results!

• Unmyelinated Velocity is about 0.5 meters/sec

• Myelination increases velocity to up to 70-100 meters/sec

Blue Whale: How long will it take to go from brain to tail?

30 Meters: With Myelination:__

Without Myelinationation:__

How tall are you head to tail?

Why does sphingomyelin help?

What is a Node of Ranvier?

Why no VGC between nodes?

Why does Saltatory Conduction use less Glucose/ATP?

The myelin sheath is created by cells to GREATLY increases Action Potential velocity! Oligodendrites in the central nervous systemSchwann Cells in the peripheral nervous systemGaps between cells are called Nodes of Ranvier

Synaptic Gap Junctions (if present) permit the incoming Na+ from the action potential in one cell to diffuse INTO the next cell creating a threshold potential opening its Na-VGC: new APcreated!

This creates an Electrical Syncitium! (T.Q.)This synaptic device isn’t too common!Poor regulation!Importance to heart?Why can the heart fibrillate?

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Acetylcholine is a ubiquitous ligand that opens a sodium ligand gated channels. How does ACH do this? When is ACH released at a synapse? Why is acetylcholinesterase important in regards to limiting the duration of the Increase in Permeability? Why do some toxins work by modifying the activity of ACHase?

The action potential can also cross between cells by causing chemicals to be released at the pre-synaptic cell that find/open Ligand Gated Channels on the post synaptic cell and change the Na-permeability of the second cell to create a new AP on (or change in) the second cell!

Costs of membrane potential with respect to ATP use and production. What organelles “could” facilitate this and why are neurons so sensitive to hypoxia?

Neurons maintain a “FINE” balance between ATP Supply and Use, why is this so?

• Perfusion of the brain

• Na+/K+-ATPase function at rest and when cell is “active”

• Long-term potentiation in neuronsmemory

• Neuron packing and space for mitochondria

• Glycolysis and the cytosol (lactate?) ATP yield

• Mitochondria ATP yield

• Phosphocreatine ATP backup

• If blood supply stops: Why do you pass out in 20 second and why do brain cells begin to undergo infarct in 5 minutes?