Resting membrane potential

Lab 3: Action Potentials

Objectives

By completion of this lab, you will be able to…

• Understand and describe the resting membrane potential
• Outline the steps of an action potential
• Describe the membrane dynamics during an action potential

Neurons rely on action potentials to transmit information down their axons. You can think of it as the “on switch” for a neuron. What action potentials really are though, are movements of ions across the membrane. More specifically, an action potential alters the permeability of the axolemma (the plasma membrane of the axon) to certain ions. These changes in permeability propagate all the way to the axon terminal. Changes in permeability alter the neurons electrochemical gradient that is present at rest– its resting membrane potential.

In this lab exercise, you will use the PhET Neuron Simulator. This simulation provides you with a visual representation of the axon as an action potential travels down it. You will explore this simulation in order to understand how action potentials work.

Activity 1: Resting Membrane Potential Overview

Setup:

1. Open up the simulation.
2. Zoom out (- button) so that you can see the whole axon (i.e., one full circle)
3. Click the options to show the Charges and Concentrations

1. What two ions are involved with the neuron at rest? List them in the table below. What are the concentrations of each inside and outside of the cell? Note: the simulation uses the units mM. What is the charge for the ion (e.g., +2, -1, etc.)

Ion	Extracellular Concentration	Intracellular Concentration	Charge

2. List all 4 of the transport proteins embedded in the axolemma. Which 2 are open while neuron is at rest?
3. For the 2 transport proteins you identified in the previous question, how many of each are embedded in the axolemma? Based off of that, which ion do you think the axolemma is more permeable to?
4. Observe a potassium leak channel. Does potassium go only one way, or both? Hint: Try speeding up the simulation. Do not zoom in.
5. Fill in the blank: In terms of electrical charges (and maybe romantic relationships, too) opposites_______________
6. Leak channels are facilitated diffusion channels. Where is the highest concentration of potassium (intracellular or extracellular)? Which way would potassium diffuse according to this concentration gradient?
7. Does the axon have a positive or negative charge at rest according to the simulation? How does this relate to potassium’s occasional inward flow? Hint: see your answer to question 5.

Activity 2: Resting Membrane Potential Math

Using the concentration of an ion both inside and outside the cell, the membrane potential (more accurately, the equilibrium potential) can be calculated by the Nernst equation.

𝑉𝑒𝑞= 𝑅𝑇𝑧𝐹 ln ([𝐼𝑜𝑛𝐸𝑥𝑡𝑟𝑎𝑐𝑒𝑙𝑙𝑢𝑙𝑎𝑟][𝐼𝑜𝑛𝐼𝑛𝑡𝑟𝑎𝑐𝑒𝑙𝑙𝑢𝑙𝑎𝑟]

R is the Universal Gas Constant and F is Faraday’s Constant. We will assume the neuron is a mammalian neuron, so its temperature is 37oC (or 310 K in the case of this equation). z is charge of the ion in question (e.g., +1, -2, etc.). Since both sodium and potassium ions have the same charge, and we want our answer in mV, not volts, we can use this simplified equation:

𝑉𝑒𝑞= 26.7 ln ([𝐼𝑜𝑛𝐸𝑥𝑡𝑟𝑎𝑐𝑒𝑙𝑙𝑢𝑙𝑎𝑟][𝐼𝑜𝑛𝐼𝑛𝑡𝑟𝑎𝑐𝑒𝑙𝑙𝑢𝑙𝑎𝑟]

For each of the ions you will enter its concentration (the brackets mean concentration) for its concentration inside and outside of the cell. Note, the membrane potential of only one ion can be calculated at a time.

8. What is the membrane potential if only potassium is taken into account?
9. What is the membrane potential if only sodium is taken into account?
10. Click Potential Chart and then Stimulate Neuron. What is the voltage of a neuron before the action potential has occurred?
11. What might account for the discrepancy between the voltages from the previous 3
    3-4
12. A more complex equation is the Goldman-Hodgkin-Katz equation. Use the link to read about this equation. What extra parameters does it take into account compared to the Nernst equation?
13. Using the link in the previous question, scroll to the bottom of the page to find a calculator for the Goldman-Hodgkin-Katz equation. Click the button that says Fill Cells with Sample Data. This will enter data for a “standard” neuron into the calculator. Rank the ions by their permeability.
14. Change the voltage units to mV and hit calculate. What is the number it gives you? What is this number very close to from earlier in this worksheet?

Resting membrane potential cannot be explained entirely by the above work. In particular, it is important to remember the presence of the sodium-potassium pump. At the cost of ATP, 3 sodium are moved to outside the cell, and 2 potassium are brought in. This manifests as the concentration gradients we see for sodium and potassium.

Activity 3: Action Potentials

Setup:

1. Refresh the page with the simulation.
2. Click the options to show the Charges, Concentrations, and Potential Chart
3. You may have to slow down the simulation, zoom in, start/pause the simulation, and/or repeat steps to answer the following questions.

15. Hit Stimulate Neuron. Screenshot the graph and paste it here.
    3-5
16. What is the peak voltage? What is the lowest voltage?
17. What channels are open during the “uphill” phase (depolarization phase)? What happens to this channel during the downhill phase (pay attention to the ball and chain)? What ions are moving and in what direction?
18. What channels are open specifically in the “downhill” phase (repolarization phase)? At what approximate voltage do these channels open? What ions are moving and in what direction?
19. After the downhill phase is a hyperpolarization phase? Why do you think it gets this name? Hint: compare the resting membrane potential and the hyperpolarization phase.
20. After hitting Stimulate Neuron it becomes grey for a while. What physiological process of the neuron does this represent?