South Seattle College

BIO241 – Anatomy and Physiology I

Fall Quarter 2021
Written by C.Schroyer

Laboratory 6: Membrane Potentials and Action Potential Generation

(Chapter 11 Section 11.3 – 11.8)

Due Date: See Course Calendar

Worth: 10 points
Project Type: Individual submission. The (submitted on the Lab 7 Answer Sheet).

LAB OBJECTIVES: Review the lab objectives listed below and summarize them in 1-3
sentences in your own words in the space provided (or in your lab notebook).

Learning Objectives:
1. Describe the functional and structural components of a neuron.
2. Define the resting membrane potential.
3. Compare and contrast voltage and ligand gated channels.
4. Identify and describe the components responsible for maintaining the resting membrane
potential.
5. Define an action potential and explain why it is essential for life.
6. Identify and describe the components of an action potential.
7. Explain how action potentials are generated. Be sure you can explain the locations and
movements of ions, changes in electrical potential and the membrane proteins involved.
8. Identify and explain how drugs can impair and/or facilitate action potential generation.

Note to Students: These learning objectives are designed to help prepare you for lecture exams
and/or lab practical exams.
South Seattle College
BIO241 – Anatomy and Physiology I
Fall Quarter 2021
Written by C.Schroyer

Background Information on The Nervous System

Background:
When a nerve cell is stimulated, it triggers what is known as an action potential. An action
potential is the change in electrical potential that propagates (travels) along the membrane of a
nerve cell. This is how information moves through our nerves!

In this activity, you’ll be investigating the changes that take place in a nerve cell during an
action potential. Pay close attention to the locations and movement of ions, changes in
electrical potential, and membrane proteins that are involved.

Generation of Action Potentials

During the resting state, the membrane potential arises because the membrane is
predominantly permeable to K+. An action potential begins at the axon hillock as a result of
depolarization. During depolarization voltage-gated sodium ion channels open due to an
electrical stimulus. As the sodium ions rush back into the cell, their positive charge changes
potential inside the cell from negative to more positive.

The Resting Membrane Potential

The resting membrane potential of cells varies depending on the cell type. For neurons, it
typically sits between -50 and -75mV. This value depends on the types of ion channels that are
open and the concentrations of different ions in the intracellular and extracellular fluids during
the resting state. In neurons, K+ and organic anions are typically found at a higher
concentration within the cell than outside, whereas Na+ and Cl- are typically found in higher
concentrations outside the cell.

This difference in concentrations provides a concentration gradient for ions to flow down when
their respective channels are open. Hence, K+ ions would be moving out of the cells, while Na+
and Cl- ions would be moving into the cell. At the resting state, the cell is mostly permeable to
K+, as such this exerts the greatest influence on the resting membrane potential out of the
three ions. These concentration gradients are maintained by the action of the Na+/K+
ATPase via active transport, which in turn allows the membrane potential to be maintained.

Generation of Action Potentials

During the resting state, the membrane potential arises because the membrane is
predominantly permeable to K+. An action potential begins at the axon hillock as a result of
depolarization. During depolarization voltage-gated sodium ion channels open due to an
electrical stimulus. As the sodium ions rush back into the cell, their positive charge changes
potential inside the cell from negative to more positive.
South Seattle College
BIO241 – Anatomy and Physiology I
Fall Quarter 2021
Written by C.Schroyer

If a threshold potential is reached, then an action potential is produced. Action potentials will
only occur if a threshold is reached. Additionally, if the threshold is reached, then the response
of the same magnitude is always elicited, irrespective of the strength of the stimulus. Hence,
action potentials are described as “all-or-nothing“.

Once the cell has been depolarized the voltage-gated sodium ion channels begin to close. The
positive potential inside the cell causes voltage-gated potassium channels to open and K+ ions
now move down their electrochemical gradient out of the cell. As the K+ moves out of the cell,
the membrane potential becomes more negative and starts to approach the resting potential.

Typically, repolarization overshoots the resting membrane potential, making the membrane
potential more negative. This is known as hyperpolarization. It is important to note that the
Na+/K+ ATPase is not involved in the repolarization process following an action potential.
Every action potential is followed by a refractory period. This period can be further divided into:
• the absolute refractory period which occurs once the sodium channels close after an AP.
Sodium channels then enter an inactive state during which they cannot be reopened,
regardless of the membrane potential.

• the relative refractory period which occurs when sodium channels slowly come out of
the inactivation. During this period the neuron can be excited with stimuli stronger than
the one normally needed to initiate an AP. Early on in the relative refractory period, the
strength of the stimulus required is very high. Gradually, it becomes smaller throughout
the relative refractory period as more sodium channels recover from the inactivation
stage.

Source of the above information: https://teachmephysiology.com/nervous-system/synapses/action-potential/

Please review the diagram below outlining action potential generation.

South Seattle College
BIO241 – Anatomy and Physiology I
Fall Quarter 2021
Written by C.Schroyer

Source: https://vlab.amrita.edu/?sub=3&brch=212&sim=742&cnt=1

Exercise 1: Comparing and Contrasting the Resting Membrane

Potential and Action Potential

Directions:
1. Carefully read the information below.
2. To complete this activity you will need online access and you will need to register for the
phet website so you can complete the interactive activity.
3. All Exercise 1 questions will be graded. Answers should be written and submitted on the Lab
6 Homework answer sheet (download from the Lab HW 6 submission link) or write directly
on the lab document in the space provided.

Getting Started:
Go to the following website: https://phet.colorado.edu/sims/html/neuron/latest/neuron_en.html
Name________________________________________________ Period_________ Date_____________ ​Action Potential Simulation  
 
Background​:
When a nerve cell is stimulated, it triggers what is known as an ​action potential​. An action potential is the change in electrical potential
that propagates (travels) along the membrane of a nerve cell. This is how information moves through our nerves!

In this activity, you’ll be investigating the changes that take place in a nerve cell during an action potential. Pay close attention to the
locations and movement of ions, changes in electrical potential, and membrane proteins that are involved​.

Getting Started​:
Go to the following website: ​https://phet.colorado.edu/sims/html/neuron/latest/neuron_en.html

The image shown represents a cross-section of the nerve axon. (What’s a cross-section? Imagine laying a nerve cell lengthwise along an
x-axis and slicing through it along the y-axis: that’s how you get a cross-section!) The yellow represents the plasma membrane of the
axon. The myelin sheath is not shown here.

When you click “Stimulate Neuron,” you’ll notice a purple and yellow thing moving down the length of the axon: this represents the action
potential. What you’ll be observing in this activity is what happens when the action potential reaches this cross-section of axon. You are
not observing the ​entire​ nerve cell, just a tiny fraction of its membrane! The changes you observe here are what happen down the entire
length of the neuron.

In the box labeled “show,” start by checking all of the boxes. This will allow you to see everything that is going on. Hit the “Stimulate
Neuron” button on the lower right corner of the simulation to simulate an action potential. You can pause the simulation at any time, scroll
back on the potential chart to rewind, zoom in/out, and speed up or slow down the animation.
 
Take several minutes to play around with this simulation and get comfortable with it. What happens when you check or uncheck boxes or
click different buttons? You’re not going to break it, so go ahead and click everything!

After you’re comfortable with the various settings, use the simulation to answer the questions below. ​Pro tip: You’re going to need to 
zoom in and change the speed to really understand what’s happening as you work through these questions. 

When you’re ready to start answering questions, check ALL of the boxes in the section labeled “Show.”
 
1. Observe the membrane closely while the axon is at rest.
a. Which membrane channels are open? _______________________________________________________________________
b. Which are closed? _______________________________________________________________________________________

2. Are there more open or closed channels present in the membrane when the axon is at rest?

3. The concentrations of sodium and potassium ions are different inside and outside the membrane. Which direction will sodium ions
move as a result of facilitated diffusion through the “leak” channels? Potassium ions?

4. Which side of the neuron is negatively charged - inside the neuron or outside? __________________________
a. This simulation only shows positively charged ions. How can one side of the membrane have a net negative charge in this
scenario? What must be present but not shown in the image?

5. Although facilitated diffusion is clearly happening, the indicated ion concentrations are not changing. That’s because this image is
actually missing a very important protein called the ​sodium-potassium pump​! Think about what you know about the concentration
gradients. What must this pump be doing to maintain the concentration gradient? What kind of transport is it doing? Which direction must
each ion be moving through the pump?

Now it’s time to stimulate your neurons!

6. On the axes provided, sketch the graph that is generated when you click “stimulate neuron.” Include ALL necessary titles, labels, and
units! (This does not need to be perfect, so don’t bother writing every number or filling in grid lines.) Then, complete the table below by
writing the answer or circling the correct word. Pay close attention to +/- signs on the membrane potential values!

  At Rest (0-2 ms)  Peak of Action Potential  Immediately After Action 

(~2.7 ms)  Potential (4 ms) 

Membrane potential (mV)

Which side of the membrane has a net negative

Inside / Outside Inside / Outside Inside / Outside
charge?

Which side of the membrane has a greater

Inside / Outside Inside / Outside Inside / Outside
concentration of sodium ions?

Which side of the membrane has a greater

Inside / Outside Inside / Outside Inside / Outside
concentration of potassium ions?

State of the gated channels? Open / Closed Open / Closed Open / Closed / Closing

7. When membrane potential is negative, which side of the membrane is negatively charged? What about when the membrane potential
is positive?

8. What about the role of relative permeability of the ions? Before looking at that, let’s calculate the equilibrium (Nernst) potential of the
two ions.

9. Plugging in the concentrations of sodium and potassium, calculate the equilibrium potentials for sodium and potassium.

E​Na+​ = E​K+​ =

If the membrane potential is at the equilibrium potential for a specific ion, there is no net tendency for that ion to move in or out of the cell.
But what if the membrane potential is not at the equilibrium potential for a specific ion, and there are open ion channels that will allow the
ion to cross? The ion will tend to move across the membrane.* If the ion is moving across the membrane (i.e. the membrane is permeable
to it), then the membrane potential should get closer and closer to the equilibrium potential for the ion. So, if the cell is more permeable to
sodium, the membrane potential will become closer to the equilibrium potential of sodium (E​Na​). If the cell is more permeable to potassium,
the membrane potential will become closer to the equilibrium potential of potassium (E​K​).
10. When the cell is at rest, is the membrane potential closer to the equilibrium potential of potassium or sodium ions? What about at the
peak of the action potential?

11. Based on this information, predict the membrane’s relative permeability to sodium and potassium at rest, during the upstroke of the
action potential, and during the down stroke of the action potential. (Think about when the potential is approaching E​Na+​ and when it is
approaching E​K+​.) Explain your prediction.

Observe the sodium and potassium ions passing across the membrane during the simulation. You might want to slow down the simulation
speed and use the zoom feature to get a closer view.
12. Which gated channel opens first (during the upstroke)?

13. Which opens second (during the peak/downstroke)?

14. What direction are sodium ions moving through the sodium gated channel? What direction is potassium moving?

15. What is happening to the membrane potential as each of these channels open?

16. Do these observations agree with your prediction in question 11? Explain.

17. Look carefully at the graph of the membrane potential. At the bottom of the downstroke, what is the approximate membrane potential?
Is this higher, lower, or the same as the resting membrane potential?

18. Observe the “Stimulate Neuron” button throughout the course of the action potential. It turns gray, indicating that it is not available to
be clicked. This actually represents an important concept in neuron function, the ​refractory period​. This is a period after the action
potential occurs during which a second action potential cannot be propagated. What is the membrane potential when you can stimulate
the neuron again? How does this compare to the resting membrane potential?

19. The drug ouabain inhibits the function of the ​sodium-potassium pump ​(Explained in question 5). Predict the short-term and
long-term effects of ouabain on the excitability (ability to be stimulated) of a neuron. Think about the effect this would have on the resting
membrane potential.
Activity written by Kara Reichert, M.Ed. for AP Biology classes at Jones College Prep HS, Chicago (2019).
Includes work from “AP Sim” by Hwa Tsu of North Central High School.
Additional contributions made by Nichole Lowery of Jones College Prep.

*https://courses.washington.edu/conj/membpot/equilpot.htm