pnb 2274 lab practical 2 review questions

Events of an AP

- Ligand-gated Na+ channels open - Voltage gated Na+ channels open and Na+ floods into the neuron - Na+ channels inactivate and voltage gated K+ channels open - Voltage gated K+ channels close and membrane potential approaches the resting membrane potential

Events of an action potential

- Stimulation of AP in motor neuron - Influx of Ca2+ into presynaptic axon terminal - Vesicle docking and releasing - Diffusion of ACh across the synaptic cell - Binding of ACh to receptors on the muscle membrane - Sodium influx through receptor channels - Action potential propagating along muscle membrane

Which of the following could result in no signal or poor signal?

- The worms were drowned - The recording pan is filled with Worm's Ringers when ensuring the worm is fully anesthetized. - The contact between the recording pins and the worm is poor. (e.g., dirty pins, electrodes are not pinned through the worm, etc.) - The head of the earthworm is not touching both stimulating electrodes.

The cockroach experimental setup is an __________ recording. A recording collected from this setup contains action potential from __________ sensory axon(s).

extracellular; multiple

To avoid excessive amounts of background noise, you need to make sure that the cockroach leg is:

leg is elevated from the surface of the Petri dish.

reciprocal activation

simultaneous activation of an agonist muscle and inhibition of an antagonist muslce

co-contraction

simultaneous contraction of both the agonist and the antagonist muscles around a joint

Given a typical neuron with ENA+ = +50mV and Vm = -70mV, determine the direction of Na+ flow when the membrane is forcibly set to +120mV.

Flow out of the cell

Which of the following is an acceptable scientific hypothesis?

Genetically modified tomatoes contain more fructose than conventional tomatoes.

If tetrodotoxin's effect is on neurons, why did Dr. Westwood experience both of these symptoms?

He experienced both symptoms because TTX blocks the sodium channels so an action potential will not be generated. Signals will not be transmitted from neuron to neuron and to muscle cells, which is the cause of the numbness and the paralysis.

What are the structural differences between the sheep's mammillary body and the human mammillary body?

Humans have two mammillary bodies while sheep only have one.

Describe your hypotheses that explains how batrachotoxin affects action potential generation.

If a neuron is exposed to batrachotoxin, the blocking of voltage-gated sodium channels will become inhibited, allowing for a constant influx of sodium ions, which will not allow the cell to repolarize.

Frame your question in the form of an experimentally testable hypothesis.

If an overhead triceps extension is performed using a heavier weight, the RMS triceps will be higher than the RMS triceps without the weight because the heavier weight provides a greater resistance.

What is the relationship between stimulus strength and response amplitude in a single axon?

In an axon, stimulus strength and response amplitude are related by the all-or-none principle. This means that an action potential will occur only if the stimulus is strong enough for the response amplitude to reach the threshold.

Design the experiment you will do to examine the hypothesis you formulated above. Make sure you include all the equipment you will need, the number of volunteers required, and that you list all the measurements you are going to make.

In this experiment, 2 people are required. The first person will be connected to the electrodes and will be performing the exercise with a 5 pound weight while the other person will hit the start and stop button. First we will perform a control by recording the muscle activity while the arm is performing 3 repetitions of the exercise without a weight. After a resting period of 2 minutes, we will record three repetitions of the overhead triceps extension using a two pound weight. After resting period of 2 minutes, perform three repetitions of the overhead triceps extension using a five pound weight. Compare sets of data.

What is the optimal sarcomere length for maximal force generation in muscle?

Intermediate length

The purpose of this week's lab is to:

Investigate the sensory responses by recording action potentials from a cockroach.

An earthworm giant axon:

Is composed of individual cells, one in each segment, linked through gap junctions to its neighbors in adjacent segments.

During reciprocal activation, why is the antagonist muscle not completely silent?

It is an antagonist pairing and the both work to stabilize the arm and joint

For the first experiment (spontaneous recording and puff stimuli), please copy your collected data into the Excel sheet below this question. In the Excel sheet, you can select different bin sizes. What did you choose as the bin number for the histogram? Why did you choose this bin number?

It showed a better trend of the data, while giving a good distribution of the data: it showed enough detail without losing track of the trend and losing the point of making a graph in the first place. A smaller bin size (larger number of bins) also allows you to tell how many spines are stimulated.

Describe how the isolated muscle responded to high frequency stimulation.

The high frequency stimulation caused the muscle to fatigue and slowly contract with less and less force.

What are your conclusions based on the recorded data?

When the weight is increased while performing an overhead tricep extension, the triceps have a larger EMG amplitude because of increased number of motor units needed for the contraction.

As the mouse breathes inside the metabolic chamber, the soap bubble seal moves because CO2 is generated as a result and absorbed by the soda lime placed at the bottom of the chamber. Which of the following does NOT affect the rate of this volume change?

Air consumption rate DOES AFFECT: CO2 generation and absorption rate and O2 consumption rate

During your recording, you see a flat line being recorded. What should you do?

Adjust the scale immediately.

Did your MetaNeuron simulation support your hypothesis about the effects of TTX you formulated in Exercise 1?

No. When TTX was checked, there was no real peak, meaning that there was no action potential.

Calculate the sampling rate in the image. It shows a rate of:

10 samples per second

Which of the following correctly describe bin number?

A large bin number will allow us to see more detailed information about the data. (large bin number = smaller bin size)

During the falling phase of the action potential, the membrane hyperpolarizes beyond the resting membrane voltage. The phenomenon is due to:

A large number of open potassium channels

Brett ate poisonous wild mushroom, which partially paralyzed his motor unit. His doctor performed an EMG test on him and the EMG trace is shown above. Predict what normal EMG from Brett could look like? 1 / 1

A normal EMG would have more regular intervals and a higher amplitude. Because Brett's motor units are paralyzed, his muscles are not able to fully relax before the next contraction, which doesn't allow the peaks to be as large in comparison to the dips as they should be. The inhibition of the motor unit leads to less electrical signals being generated.

What conclusion can you draw based on the graph showed above?

A single spine can give a set of APs with similar amplitudes. Different spines give APs with different amplitudes. All or none principle

In light of the "all or none" law of muscle contraction, how can you explain the graded response?

All of none is the required threshold a neuron needs to fire an action potential. A graded response is the adding up of all the individual responses from the motor units. Feedback from grader Just remember that "all or none" applies to single muscle fibers, but when graded response, multiple muscle fibers are activated.

Describe how the isolated muscle responded as the stimulus interval was decreased progressively.

As the stimulus interval was decreased, the two peaks of contraction became one larger peak.

As you increase voltage to the muscle, describe how it responds to the increased stimulus.

As the voltage increases, the contractile force increases until it peaks. After the voltage that corresponds with the maximum contractile force is increased, the contractile force starts to return back down.

In your own words, explain how the EMG trace changed when you added weights to your arm? Based on the data you collected what can you infer is happening to the muscles as weight is added?

As you increase the weight, more motor units need to be recruited in order to contract the muscle. This was seen in the data as the electrical activity of the muscles increased as weight was added to the arm.

Describe what happened to the recorded action potentials when you applied the press-and-hold stimulus to each spine. Consider amplitude, density, frequency, and any other differences in the action potentials. What is the evidence showing the spines became adapted to the stimulus?

At the start, the amplitude, density, and frequency were all high. As time progressed it became clear that the spine was adapting to the stimulus: the density and frequency started to decrease until they leveled out.

Take a screenshot of your histogram generated in the Excel file, with the optimal bin size, and upload it to Kuracloud. Based on the histogram, how many spines are likely contributing to the spontaneous firing?

Based on the histogram, there are 5 spines contributing to spontaneous firing.

Formulate two different hypotheses that explain how batrachotoxin might work.

Batrachotoxin prevents the sodium channels from closing which allows for an influx of sodium ions that keeps the membrane depolarized and prevents repolarization.

The two lateral giant nerve fibers:

Behave as a single fiber because of extensive cross-connections between them.

Which of the following are mechanisms of fatigue?

Conduction failure - constant AP and K channel opening causes the T-tubule to be filled with K causing constant depolarization and an inability to regenerate an AP Lactic acid buildup - constant conversion of glucose to lactate in glycolysis --> acidification of the muscle fiber, decreasing the pH and causing proteins like actin/myosin to degrade/function improperly Inhibition of cross-bridge cycling - hydrolysis of ATP causes ADP and P buildup, directly inhibits the cross-bridge cycling by delaying cross-bridge detachment from actin

The PowerLab hardware is NOT designed to:

Display a signal

Describe in detail the cellular events responsible for the absolute refractory period AND the relative refractory period. (Hint: Discuss the activity of the voltage-gated channels).

During the absolute refractory period, sodium channels are open but inactivated. Since sodium influx isn't possible, depolarization doesn't occur. During the relative refractory period, some sodium channels are open; this allows some sodium to enter the cell which starts the depolarization process and allows for more voltage-gated channels to open up. This has the potential to become an action potential.

Which of the following statements about electromyography (EMG) recording is NOT true?

EMG is a technique that measures the contractile force of the muscles. EMG signals are recorded from multiple muscle fibers.

In the anterior pituitary region, do you see multiple cell types in each follicle or does each follicle contain a single cell type? What is the most abundant cell type? Which is the least prevalent?

Each follicle in the anterior pituitary has multiple cell types. The most abundant are chromophobes and the least abundant are gonadotropes.

Unlike the discrete waveform from an electrocardiogram, the electromyogram waveform is irregular. Why do you suppose this is?

Electrocardiograms record cardiac muscle which all contract at the same time but electromyograms record skeletal muscle which do not contract at the same time. These asynchronously contracting muscle fibers cause an irregular waveform.

When placing recording electrodes on the biceps and triceps:

Ensure that the electrodes are in good contact with the skin.

From your data, the theoretical maximum frequency at which nerve impulses can be generated by the median giant fiber was calculated. Do you think that the nerve fiber would be able to generate impulses continuously at this rate?

Eventually the muscle would fatigue and there wouldn't be enough ATP to keep having full action potentials. At first the nerve fiber would be able to generate impulses, but it soon would not be able to.

Propose the experiments you will design to test your hypotheses.

Examine the action potentials of normal neurons and the action potential of neurons when they become exposed to the toxin. Carefully compare the area of repolarization, or what should be repolarization in the neurons exposed to the toxin.

A single motor neuron innervates one muscle fiber.

False

During the refractory period experiment, Action Potential generated from the first stimulus always appears before the stimulus artifact generated by the second stimulus in your recording.

False

One motor unit can respond with multiple twitches per stimulus

False

Spines found on cockroaches' leg contain receptors for tactile and audio-vibratory stimuli. Each spine contains 2-3 sensory neurons.

False

To anesthetize the worm, you can place it in a beaker filled with Worm Ringer's with 10% ethanol.

False

In this week's lab, we will be using a transducer

False (transducer converts a mechanical signal into an analog voltage

Given a typical neuron with ENA+ = +50mV and Vm = -70mV, determine the direction of Na+ flow when the membrane is forcibly set to 0mV.

Flow inside the cell.

What stretch resulted in the highest contraction force? What happens to the muscle at the highest stretch levels?

Length 6 had the highest contraction force where it showed maximum contraction. At this point, the muscle had the highest force for its amount stretched. Feedback from grader At the highest stretch levels, the contractile force should be low because cross-bridges between myosin and actin filaments are limited.

Refer to the image above. What does Point C on the earthworm action potential recording indicate?

Median axon action potential

During lethal injection, K+ is added to the extracellular fluid. What would you predict the resting membrane potential to be?

More depolarized

The intracellular and extracellular concentrations are the same for cation A. The resting membrane potential is -70mV. What is the net driving force for cation A during the resting stage?

Move from extracellular to intracellular

A motor unit includes:

Multiple muscle fibers innervated by one motor neuron

What causes the bubbles to move through the pipette during the experiment?

O2 is consumed by the mouse and the generated CO2 is absorbed in the setup.

When the concentration increases inside the cell, Vrest becomes more negative. When the concentration decreases inside the cell (or increases outside the cell) Vrest becomes more positive. Refer to your calculations in Table 2, does ion [K] or [Na] have a bigger effect on Vrest? Furthermore, mathematically justify your answer with the GHK equation.

Potassium has a larger effect on Vrest because it has a higher conductance than the sodium ion (0.37 > 0.02).

In addition to the overall size difference, describe the major structural differences you observed using a sheep brain and a human brain model. (Include at least 4 structural differences.)

Sheep brains have proportionally larger olfactory bulbs because they rely on smell more than humans do. Humans have two mammillary bodies but sheep just have one larger mammillary body. The human brain has more sulci and gyri because of its greater surface area. The fornix of the sheep brain is thicker than the fornix of the human brain.

How does the size of sheep olfactory bulbs compare with those of humans? Is the sense of smell more important as a protective and a food-getting sense in sheeps or in humans?

Sheep olfactory bulbs are larger than those of humans because they use smell more for survival than humans do.

Is there any relationship between the position of the spine and the amplitude of the stimulated action potentials? Suggest factors other than the position on the leg that could influence the amplitude of action potentials.

Some factors that could influence the relationship are the diameter of the axon, the type of stimulus applied, and the duration of the stimulus.

Refer to the image above. What does Point A on the earthworm action potential recording indicate?

Stimulus Artifact

Refer to the image above. What does Point B on the earthworm action potential recording indicate?

Stimulus Artifact

Referring to your hypothesis in Exercise 1, your simulation results in Exercise 2, and your membrane drawing in Exercise 3, predict the mechanism of action of tetrodotoxin (TTX) on a neuron.

TTX blocks the opening of voltage-gated sodium channels which doesn't allow for the depolarization of the action potential to occur.

Based on table 1 and table 2, what conclusions can you make? Make sure to include the factors that influence the membrane potential.

Table 1 shows that the ratio of ions in the cell has an effect on the membrane potential. Table 2 shows that an influx of sodium ions causes depolarization (more negative) and an eflux of potassium ions causes hyperpolarization (more positive) of the neuron.

Define tetanus. At which stimulus interval did you observe tetanus? Explain the mechanism behind this phenomenon.

Tetanus is sustained muscle contraction because of a rapid series of nerve impulses. We saw complete tetanus at 0.2 seconds (20 ms) and this is when the action potentials were conducted at a very high rate and there was sustained muscle contraction. Tetanus happens when the applied stimulus is really high and muscle does not have time to relax before the second stimulus arrives. This results in multiple contractions added together to form a single and smooth contraction trace.

What happened to the recorded action potentials when you applied the press-and-hold stimulus to each spine? What is the evidence showing the spines became adapted to the stimulus?

The AP decreases and the stimulation is not as strong.

The EMG is a recording of:

The action potentials of the innervated muscle fibers

Is there any relationship between the position of the spine and the amplitude of the stimulated action potentials? Suggest factors other than the position on the leg that could influence the amplitude of action potentials.

The amplitude of the stimulated action potential is dependent on the size of the nerve, but more importantly, the location of the spine in relation to the recording electrode.

Based on your recorded EMG trace and the values you measured in Exercise 1, describe this phenomenon qualitatively.

The biceps and triceps muscles work by reciprocal activation, meaning that while one is contracted the other is relaxed, because they are antagonistic muscles. It is shows through the data from the table on the left. When the biceps are contracting by flexing the arm, the biceps have a larger EMG amplitude than the triceps. When the arm is extended, the triceps are contracting while the biceps are relaxing, which is shown by the larger EMG amplitude in the triceps. The table on the right shows that as you increase the weight or force that the muscles have to exert on, the EMG amplitude of the muscles contracting together increases, as expected. Feedback: The agonist muscle is mainly activated, while the antagonist is partially inhibited.

What is the cause of the blip (?) in the [Na+] current trace? (Circled in purple in the image above)

The blip is when the potassium channels open up and the sodium influx compensates for that.

The image directly above shows an extracellular recording. Which of the statements regarding the pointed part from a biphasic waveform is correct?

The charge at the positive recording electrode is more negative compared to the negative recording electrode.

Does your data support your hypothesis? Please provide some explanations in the following space, if your data do not support your hypothesis.

The data supports our hypothesis. When performing the exercise with no weight, the average EMG amplitude was 0.113 mV. The average amplitude when performing the exercise with 2 pounds was 0.16 mV. The average amplitude when performing the exercise with 5 pounds was 0.46 mV. As the weight was increased, the EMG amplitude also increased.

Describe how the isolated muscle responded as it was stretched progressively.

The force continued to increase as the isolated muscle was stretched progressively.

How does the relative size of the fornix in the sheep brain compare with the human fornix? Why do you suppose this is so? (Hint: What is the function of this band of fibers?)

The fornix is made up of afferent neurons which is why it is larger in sheep than humans, because sheep rely more on their senses, especially smell, for survival and gathering food.

Review the data generated in your 3D graph. What is the maximum temperature (C) to generate an action potential under the default cellular ion settings? What trends in the action potential do you notice as the temperature begins to decrease?

The maximum temperature is 22 degrees Celsius. As temperature begins to decrease, the action potential is longer.

In theory, how do the normalized O2 consumption rates compare to each other? How do you explain this phenomenon?

The normalized consumption rate is highest for TSH and lowest for PTU. This is because TSH increases basal metabolic rate while PTU decreases it, so it makes sense that when a mouse is treated with TSH, it consumes more oxygen and the mouse treated with PTU consumes less oxygen than the control mouse.

What happens to your EMG signal when you increase the contraction strength? Explain the observation.

The peaks are larger

Provide a possible mechanism for why the muscle was unable to maintain a prolonged contraction in the fatigue experiment.

The prolonged contraction could cause a conduction failure as the opening of potassium channels into the t-tubules causes depolarization.

From the MetaNeuron program, what is the final cell resting membrane potential (R.P. or Vrest) in mV? From your calculations on the previous page, what is the final cell resting membrane potential? What are the reasons for the differences between the 2 values?

The resting membrane potential is -64.81 mV. Our calculations gave us -73.06 mV. A reason for the different resting potential could be the temperature difference. We assumed room temperature so the constant we used was 58 for cations but if the temperature used is different on MetaNeuron, the (RT/zF) constant would be different.

The biphasic nature of the action potential recorded extracellularly is a consequence of:

The use of two extracellular electrodes spaced some distance apart to record the potential.

Explain why there is a difference in the conduction velocity values produced by the two calculation methods for the median nerve. Which method is more accurate?

There is a difference in calculations because one used the difference method and the other used the absolute method. The difference method is more accurate because it uses multiple data points which gives it more accuracy and precision, rather than relying on two points.

The typical shape of the action potential is biphasic with two peaks of opposite sign. Can you explain why this is?

This because during an action potential, the outside of the cell becomes more negative while the inside becomes more positive. The negative electrode measures the first peak and picks up the negative charge because it is extracellular. The double negative charge from the electrode and the earthworm becomes a positive. The positive electrode picks up the negative charge as the action potential travels down the axon and shows a negative signal, opposite from the positive.

Critique the following hypothesis: "Aphid-infected plants that are exposed to ladybugs will have fewer aphids after a week than aphid-infected plants which are left untreated. "

This is a good hypothesis

Critique the following hypothesis: "The potential of our brain is unlimited, with which we can achieve any goals we set"

This is a poor hypothesis because it is not testable.

You have seen that an action potential generated very soon after a preceding one is not full-size. Explain this observation.

This is because the previous action potential was in a refractory period still. This means that some of the sodium channels are still inactivated and cannot contribute to the depolarization of the action potential, which is why it is not full size.

There was an increase in the number of fibers responding as the stimulus strength was increased. What is this type of behavior called? Is this of physiological significance to the earthworm?

This is called recruitment, which is when the number of additional motor fibers are increased when the stimulus strength is greater. The median and lateral fibers of an earthworm have different functions and recruitment doesn't really pertain to the earthworm.

An intracellularly recorded nerve action potential approximates 80 mV. Why is your recorded action potential so much smaller?

This is smaller because our action potential was recorded extracellularly which means it was recorded under different conditions. The condition outside of the worm is different from the inside and the smaller voltage could also have been due to poor recording conditions.

Based on your hypotheses on batrachotoxin, propose experiments that can be used to test your hypotheses. (Please be clear but concise.)

To test these hypotheses, the action potential could be measured on the muscle before and after injecting it with batrachotoxin to see its duration and other qualities. If possible, the sodium and potassium levels of the muscles should be checked before and after injecting the toxin to see if the levels of each ion changed and how that impacted the action potential.

After placing recording electrodes as instructed, you should always be able to observe action potentials that are firing instantaneously.

True

Ground electrode should be pinned through the worm somewhere between the stimulating electrodes and the recording electrodes.

True

In general, large action potentials are produced by axons with large diameters.

True

Recruitment refers to:

Twitches in all the fibers of several motor units (each motor unit has one twitch)

At which stimulus interval did you observe tetanus?

We observed complete tetanus at 20 ms.

What did you observe when you mechanically stimulated your retina? What you learned from lecture explains the observation?

We saw a bright circle of light moving up and down as we moved our finger up and down.

How does Vrest change in response to concentration changes of [K] and [Na]?

When the concentration increases inside the cell, Vrest becomes more negative. When the concentration decreases inside the cell (or increases outside the cell) Vrest becomes more positive.

In Table 2, what trends do you notice to the Vrest as the internal/external ion concentrations change in reference to the same ion? (Hint: It's all about the ratio of ions in to out.)

When the internal/external ion concentration ratio is greater than 1, Vrest is more negative than the original Vrest. When the ratio is less than 1, Vrest is more positive than the original Vrest.

When conductance is shown on the graph, why are both ionic traces positive? Conversely, when ionic currents are shown, why is the [K+] ionic current trace positive while the [Na+] ionic current trace is negative?

With conductance, both ionic traces are positive because the ion channels for both are open which makes the membrane permeable to both ions. With ionic currents, the potassium trace is positive because the ions are flowing out of the cell and the sodium trace is negative because the ions are flowing in.

Describe the improvements you have made towards your Hypothesis, Method, or Conclusions result from the in-class discussions with other groups.

With the in-class discussions, we were able to see what other groups decided to test and we were able to compare our results with groups that did a similar exercise as us. We found the difference in our results to be because of the way we performed the exercise and what data we specifically chose to record. Improvements that can be made from the discussion for next time are to do more trials or to average the data to have an easier graphical depiction.

Does your trace roughly match with a typical length tension curve? If not, could you provide some explanations?

Yes

The lightly stained region in the pituitary gland is rich in ______. It ______ hormones such as oxytocin and ADH.

capillary network; stores and releases

The figure shown above is an example of:

co-contraction

PTU (Propylthiouracil), a molecule that interferes with the iodination process during the production of thyroglobulin, will decrease the plasma thyroid hormone levels. After being subjected to PTU for a period of time, the metabolic rate of a mouse will ______.

decrease

The bubbles that are placed into the pipette are to ______.

indicate the oxygen/air consumption

The primary function of three stainless steel pins serve as the ______.

recording electrodes

The amplitude of action potential recorded in this week's lab can be affected by:

the size of the nerve and the proximity of the nerve to the recording electrodes.

Given that Neuron X has equilibrium potential values of ENA = 50mv, EK = -90mV, and ECl = -55mV, if Neuron X had a GNa that was 100x as large as either GK or GCl, what would be the approximate value of the resting membrane potential (Vm)?

~ -50 mV