GROUP 1

ANAPHY POST - LAB #1

Introduction

(Summary of the situation)

Terminologies

Sensory Neurons

Chemoreceptors-are sensory neurons that detect changes in chemicals that bind to a receptor in the
neuron’s membrane, transducing them into action potentials.Then receptors can be specific to one
chemical or respond to a variety of chemicals with similar molecular structures. Based on their location,
they are classified as either central or peripheral.

Mechanoreceptors- are sensory neurons that are activated by mechanical pressure. They are located in
internal tissues like joints, ligaments, tendons or muscles,or external ones like the skin.Deformation or
stimulation of the tissues in which the mechanoreceptors lie produces the inflow of sodium via voltage-
gated channels,which triggers an action potential.

Mechanoreceptors allow an organism to recognize and respond to physical pressure that are being
acted upon by the organism.

Thermoreceptors - sensory neurons that are activated by changes in temperature. They reside in the
skin, liver, skeletal muscles, and in the hypothalamus. Some of them are activated by heat and others
activated by cold.

Photoreceptors - considered as very special neurons that convert incoming light into an electrical signal
that can be carried to the brain to create conscious vision.

TRPV1 Channel - The TRPV1 receptor (transient receptor potential cation channel subfamily V member
1) is a non-selective ligand-gated ion channel found in certain types of sensory neurons in the peripheral
nervous system (thermoreceptors and nociceptors).
Exogenous substances, such as capsaicin (the unpleasant molecule found in hot chili peppers) or allyl
isothiocyanate (the pungent compound found in mustard and wasabi), as well as heat (greater than 43
degrees Celsius) and low pH, can activate TRPV1.

TRPV1 opens when activated, allowing cations to flow inward. A fast increase in intracellular Ca2+
concentrations frequently occurs, leading in signal transduction (action potential) and the response to
pain or heat.

Dorsal Root Ganglia

The dorsal root ganglia are a collection of cell bodies of the afferent sensory fibers in a dorsal root of a
spinal nerve, lying in the intervertebral foramina.

PATCH CLAMP BUFFERS

Cell membranes of excitable cells contain many different kinds of ion channels, some are voltage-gated.
In order for us to know one type of voltage-gated channel we will be using the voltage clamp method
where it is using a inactivation method of the rest of the channels unneeded. To achieve that, a buffer
blocking is used for the rest of the channels in the cell membrane. Example of the used of patch clamp
buffers is in studying sodium is one can block potassium and calcium channels.

VOLTAGE CLAMP

The voltage clamp is an experimental method used by electrophysiologists to measure the ion currents
through the membranes of excitable cells, such as neurons, while holding the membrane voltage at a set
level. It is a method where it locks the voltage automatically. VOLTAGE CLAMP is very useful for
understanding what is happening on the cellular level.

Threshold Potentials

is the minimum potential difference that must be reached in order to fire an action potential.

The resting membrane potential of neurons is about -70 mV. However, when a membrane potential
neuron hits the threshold value that is not less than -55 mV, action potential will be a result.

Lidocaine
Is a lipid-soluble anaesthetic that can be used as a pain killer.

The sensation of pain decreases which occurs in nociceptors due to the transmission in neurons by
prolonged deactivation of the voltage-gated sodium channels in the neuronal membrane which are in
charge for action potential propagation.

It can also inhibit the propagation of action potentials in other transmitting cells, such as motor neurons,
which results in a typical feeling of numbness in the body.

Sensory transduction - Learn why you feel pain when you get hit by a rock

Questions and Answers

1. What is the effect of the TRPV1 channel opening?

Answer: Positive ions flow inwards

Explanation

Positive ions (most importantly calcium) enter the cell by flowing down their concentration gradient.
This causes a graded depolarisation.

2. What is an important characteristic of the buffer used for studying sodium in patch clamp
experiments?

Answer: One that blocks potassium and calcium channels

Explanation

We want to block potassium and calcium channels so that we are only observing the current flowing
through sodium channels. This effectively means we will be observing only the influx of sodium into the
neuron.

3. If this cell is a neuron, using the voltage clamp to change the cell membrane potential from -70.0 mV
to -5.0 mV should cause:

Answer: A current that indicates a large amount of sodium briefly entering the cell

Explanation
Depolarization of a neuron beyond the threshold should cause the voltage-gated sodium channels to
briefly open, with an influx of sodium and then close.

4. Looking at the graph, these data are consistent with which statement below?

Answer: Opens the TRPV1 channels with no effect on sodium channels

Explanation

Action potential still occurs when the membrane potential is shifted from -70 mV to -5 mV, capsaicin
likely does not affect the voltage-gated sodium channels. There’s no action potential at -70 mV. If
capsaicin opened potassium channels, current would go in the opposite direction.

5. What action does the drug Lidocaine produce?

Answer: Lidocaine inhibits the voltage-gated sodium channels

Explanation:

Due to buffer choice, the current only represents the inward flow of sodium ions through the voltage-
gated sodium channels.

Relatively flat line of the current in the bottom graph around 0 = lidocaine inhibits voltage-gated sodium
channels from opening.

6. What action does the drug Rebine produce?

Answer: Rebine seems to have no impact on the cell

Explanation:

Action potential still occurs when membrane potential goes from -70 mV to -5 mV. Rebine seems to not
affect the voltage-gated sodium channels.

7. All the experiments so far are consistent with:

Answer: Rebine requires open TRPV1 channels in order to inhibit voltage-gated channels

Explanation:

- If Rebine opened sodium channels from the outside = induce action potential on its own

- Binding of refine and capsaicin together to the TRPV1 channel would not inhibit the action
potential, nor would blocking the capsaicin binding tot the TRVP1 channel
8. What would be the effect on the membrane voltage of tripling the clamped current while applying
Rebine and capsaicin together to the nociceptor?

Answer: Membrane voltage will increase without an action potential occurring

Explanation:

- Even increasing the current to 3 times higher while applying capsaicin and Rebine to nociceptor
does not trigger an action potential.

- Increasing current flow so more positive ions enter the nociceptor does raise the membrane
voltage, even to above the neurones activation threshold. However, an action potential requires the
opening of the voltage-gated sodium channels which are inhibited

9. What does a longer latency to withdrawal time reflect?

Answer: Slower reflex due to being less sensitive to pain

Explanation:

Lidocaine inhibit the activation of nociceptors

10. What is the significance of Rebine entering the cell through the TRPV1 channel?

Answer: Rebine only enters and inhibits nociceptors

Explanation:

- Rebine only enters nociceptors and only when the TRPV1 channel is already open

- Rebine only affects nociceptors that already sending painful signals to the brain -- does not
inhibit other cells, such as muscles.

11. What results can you conclude from this expirement

Answer: Lidocaine inhibits motor function while rebine does not

Summary/Conclusion:

Why do you feel pain when you get hit by a rock?

- The sensory neurons that we have on our body allows us to react with the sensation of pain.
Pain is transmitted via nociceptors. People feel pain when specific nerves called nociceptors detect
tissue damage and transmit information about the damage along the spinal cord to the brain. The
stimulus caused by the sensation triggers the brain leading to the feeling of pain.
What drug will help to reduce the pain?

- Between Lidocaine and Rebine, Rebine will be the best option because it allows motor function.
This would prevent the pain and could lead them to go home safe and painless.

That’s the end of our discussion, this is the Group 1. Thankyou for Listening.

GROUP 1

ANAPHY POST - LAB #1

Introduction

(Summary of the situation)

Terminologies

Sensory Neurons

Chemoreceptors-are sensory neurons that detect changes in chemicals that bind to a receptor in the
neuron’s membrane, transducing them into action potentials.Then receptors can be specific to one
chemical or respond to a variety of chemicals with similar molecular structures. Based on their location,
they are classified as either central or peripheral.

Mechanoreceptors- are sensory neurons that are activated by mechanical pressure. They are located in
internal tissues like joints, ligaments, tendons or muscles,or external ones like the skin.Deformation or
stimulation of the tissues in which the mechanoreceptors lie produces the inflow of sodium via voltage-
gated channels,which triggers an action potential.

Mechanoreceptors allow an organism to recognize and respond to physical pressure that are being
acted upon by the organism.

Thermoreceptors - sensory neurons that are activated by changes in temperature. They reside in the
skin, liver, skeletal muscles, and in the hypothalamus. Some of them are activated by heat and others
activated by cold.

Photoreceptors - considered as very special neurons that convert incoming light into an electrical signal
that can be carried to the brain to create conscious vision.
TRPV1 Channel - The TRPV1 receptor (transient receptor potential cation channel subfamily V member
1) is a non-selective ligand-gated ion channel found in certain types of sensory neurons in the peripheral
nervous system (thermoreceptors and nociceptors).

Exogenous substances, such as capsaicin (the unpleasant molecule found in hot chili peppers) or allyl
isothiocyanate (the pungent compound found in mustard and wasabi), as well as heat (greater than 43
degrees Celsius) and low pH, can activate TRPV1.

TRPV1 opens when activated, allowing cations to flow inward. A fast increase in intracellular Ca2+
concentrations frequently occurs, leading in signal transduction (action potential) and the response to
pain or heat.

Dorsal Root Ganglia

The dorsal root ganglia are a collection of cell bodies of the afferent sensory fibers in a dorsal root of a
spinal nerve, lying in the intervertebral foramina.

PATCH CLAMP BUFFERS

Cell membranes of excitable cells contain many different kinds of ion channels, some are voltage-gated.
In order for us to know one type of voltage-gated channel we will be using the voltage clamp method
where it is using a inactivation method of the rest of the channels unneeded. To achieve that, a buffer
blocking is used for the rest of the channels in the cell membrane. Example of the used of patch clamp
buffers is in studying sodium is one can block potassium and calcium channels.

VOLTAGE CLAMP

The voltage clamp is an experimental method used by electrophysiologists to measure the ion currents
through the membranes of excitable cells, such as neurons, while holding the membrane voltage at a set
level. It is a method where it locks the voltage automatically. VOLTAGE CLAMP is very useful for
understanding what is happening on the cellular level.

Threshold Potentials

is the minimum potential difference that must be reached in order to fire an action potential.
The resting membrane potential of neurons is about -70 mV. However, when a membrane potential
neuron hits the threshold value that is not less than -55 mV, action potential will be a result.

Lidocaine

Is a lipid-soluble anaesthetic that can be used as a pain killer.

The sensation of pain decreases which occurs in nociceptors due to the transmission in neurons by
prolonged deactivation of the voltage-gated sodium channels in the neuronal membrane which are in
charge for action potential propagation.

It can also inhibit the propagation of action potentials in other transmitting cells, such as motor neurons,
which results in a typical feeling of numbness in the body.

Sensory transduction - Learn why you feel pain when you get hit by a rock

Questions and Answers

1. What is the effect of the TRPV1 channel opening?

Answer: Positive ions flow inwards

Explanation

Positive ions (most importantly calcium) enter the cell by flowing down their concentration gradient.
This causes a graded depolarisation.

2. What is an important characteristic of the buffer used for studying sodium in patch clamp
experiments?

Answer: One that blocks potassium and calcium channels

Explanation

We want to block potassium and calcium channels so that we are only observing the current flowing
through sodium channels. This effectively means we will be observing only the influx of sodium into the
neuron.
3. If this cell is a neuron, using the voltage clamp to change the cell membrane potential from -70.0 mV
to -5.0 mV should cause:

Answer: A current that indicates a large amount of sodium briefly entering the cell

Explanation

Depolarization of a neuron beyond the threshold should cause the voltage-gated sodium channels to
briefly open, with an influx of sodium and then close.

4. Looking at the graph, these data are consistent with which statement below?

Answer: Opens the TRPV1 channels with no effect on sodium channels

Explanation

Action potential still occurs when the membrane potential is shifted from -70 mV to -5 mV, capsaicin
likely does not affect the voltage-gated sodium channels. There’s no action potential at -70 mV. If
capsaicin opened potassium channels, current would go in the opposite direction.

5. What action does the drug Lidocaine produce?

Answer: Lidocaine inhibits the voltage-gated sodium channels

Explanation:

Due to buffer choice, the current only represents the inward flow of sodium ions through the voltage-
gated sodium channels.

Relatively flat line of the current in the bottom graph around 0 = lidocaine inhibits voltage-gated sodium
channels from opening.

6. What action does the drug Rebine produce?

Answer: Rebine seems to have no impact on the cell

Explanation:

Action potential still occurs when membrane potential goes from -70 mV to -5 mV. Rebine seems to not
affect the voltage-gated sodium channels.

7. All the experiments so far are consistent with:

Answer: Rebine requires open TRPV1 channels in order to inhibit voltage-gated channels

Explanation:
- If Rebine opened sodium channels from the outside = induce action potential on its own

- Binding of refine and capsaicin together to the TRPV1 channel would not inhibit the action
potential, nor would blocking the capsaicin binding tot the TRVP1 channel

8. What would be the effect on the membrane voltage of tripling the clamped current while applying
Rebine and capsaicin together to the nociceptor?

Answer: Membrane voltage will increase without an action potential occurring

Explanation:

- Even increasing the current to 3 times higher while applying capsaicin and Rebine to nociceptor
does not trigger an action potential.

- Increasing current flow so more positive ions enter the nociceptor does raise the membrane
voltage, even to above the neurones activation threshold. However, an action potential requires the
opening of the voltage-gated sodium channels which are inhibited

9. What does a longer latency to withdrawal time reflect?

Answer: Slower reflex due to being less sensitive to pain

Explanation:

Lidocaine inhibit the activation of nociceptors

10. What is the significance of Rebine entering the cell through the TRPV1 channel?

Answer: Rebine only enters and inhibits nociceptors

Explanation:

- Rebine only enters nociceptors and only when the TRPV1 channel is already open

- Rebine only affects nociceptors that already sending painful signals to the brain -- does not
inhibit other cells, such as muscles.

11. What results can you conclude from this expirement

Answer: Lidocaine inhibits motor function while rebine does not

Summary/Conclusion:
Why do you feel pain when you get hit by a rock?

- The sensory neurons that we have on our body allows us to react with the sensation of pain.
Pain is transmitted via nociceptors. People feel pain when specific nerves called nociceptors detect
tissue damage and transmit information about the damage along the spinal cord to the brain. The
stimulus caused by the sensation triggers the brain leading to the feeling of pain.

What drug will help to reduce the pain?

- Between Lidocaine and Rebine, Rebine will be the best option because it allows motor function.
This would prevent the pain and could lead them to go home safe and painless.

That’s the end of our discussion, this is the Group 1. Thankyou for Listening.