Ionotropic receptors and synapses

Question 1

What is a neuromuscular junction?

Answer 1

Motor neurone to muscle fibre.
Acetylcholine neurotransmitter acts on nicotinic receptors.
Principles of central nervous system synapses are similar.

Question 2

What is the mechanism of a neuromuscular junction?

Answer 2

ACh is released from the neurone and acts on the nicotinic receptors of the membrane of the muscle fibre.
This causes the opening of ligand gated sodium channels and influx of Na+, which causes depolarisation.
The end plate potential is the excitation of the muscle fibre due to activation of the neuromuscular junction.

Question 3

What is the voltage and current of neuromuscular junctions?

Answer 3

Voltage, in mV is about 10mV change, positive deflection.
Current, in nano amps, recorded by voltage clamp.
The voltage clamp response is a mirror image of the voltage, so when there is depolarisation, it shows a compensatory negative current to balance and maintain the voltage clamp.
See picture.

Question 4

What are excitatory neuronal synapses in the central nervous system?

Answer 4

The presynaptic neurone contains vesicles filled with glutamate neurotransmitter.
An action potential depolarises the membrane and releases glutamate.
Glutamate activates ligand gated channels, which increases permeability to ions.
Na+ move into the cell due to electrochemical driving force, and depolarises the membrane, which produces an excitatory postsynaptic potential (EPSP).

Question 5

What does the EPSP look like?

Answer 5

EPSP is positive deflection of around 2 or 3mV.
This is smaller than the 10mV of the EPP in NMJ.
See picture.

Question 6

What are inhibitory neuronal synapses in the central nervous system?

Answer 6

The presynaptic interneuron synthesises glycine neurotransmitter in the spinal cord, and GABA (gamma aminobutyric acid) in the brain.
These neurotransmitters have different receptors, but are both permeable to Cl-, which hyperpolarises the membrane and causes an inhibitory postsynaptic potential (IPSP).

Question 7

What does the IPSP look like?

Answer 7

The IPSP happens soon after the action potential fires.
The IPSP is a downward deflection, as voltage hyperpolarises in response to Cl- influx.
IPSC is inhibitory postsynaptic current.
See picture.

Question 8

What is voltage clamping inhibitory synapses?

Answer 8

Looks at the response of the inhibitory synapse when Cl- channels - glycine and GABA receptors are activated using a stimulating electrode.
Voltage clamped at different levels -100mV, -80mV, -70mV, -65mV and -40mV.
See picture.

Question 9

What is voltage clamping at -65mV for inhibitory synapses?

Answer 9

-65mV is resting membrane potential.
There is a small IPSP in response to activation.
So there is a small upward deflection in voltage clamp recording, as compensating for negative charge.
See picture.

Question 10

What is voltage clamping at -40mV for inhibitory synapses?

Answer 10

The IPSP and IPSC are increasing in size, due to a larger Cl- influx.
From -65 to -40mV, the electrical driving force has increased as voltage in the cell is more positive, and negative Cl- is attracted to positive force.
Therefore there is a larger postsynaptic compensatory current.
See picture.

Question 11

What is voltage clamping at -70mV for inhibitory synapses?

Answer 11

There is no IPSP or IPSC.
The synapse is activated and neurotransmitter released, but there is no current.
This is because -70mV is the equilibrium potential for Cl-, so there is no net Cl- movement.
See picture.

Question 12

What is voltage clamping at -80mV for inhibitory synapses?

Answer 12

When hyperpolarised beyond RMP, there is a current in the opposite direction.
There is a stronger driving force for Cl- to leave the cell, so removes its negative charge, which depolarises the cell and causes a upwards deflection.
This causes a downward deflection in current.
See picture.

Question 13

What is voltage clamping at -100mV for inhibitory synapses?

Answer 13

There is a greater upwards deflection in voltage, and greater downwards deflection in current.
See picture.

Question 14

How does the driving force affect the IPSP?

Answer 14

The equilibrium potential of Cl- is -70mV.
When more depolarised, Cl- enter.
When hyperpolarised at Ecl-, no driving force so no movement,
When hyperpolarised more than Ecl-, more Cl- leaves due to electrical driving force.

Question 15

What is the effect of changing the input on excitatory neurones?

Answer 15

If there is light presynaptic input on excitatory neurone, have an EPSP.
If there is strong input, there is an EPSP and a spike, as the membrane is sufficiently depolarised to open the VGNaC and generate action potentials.
See picture.

Question 16

What happens when you activate both excitatory and inhibitory synapses?

Answer 16

When just activate inhibitory neurone, have an IPSP, downward deflection due to Cl- entry which hyperpolarises.
When activate excitatory interneuron strongly, and normal inhibitory interneuron, there is a small EPSP, with no spike.
See picture.

Question 17

What is the mechanism behind activating both excitatory and inhibitory synapses?

Answer 17

The inhibitory interneuron fires, Cl- enters the postsynaptic cell and tries to hyperpolarise it to Ecl, while the excitatory interneuron tries to depolarise it.
This forms a middle point which prevents the cell from reaching threshold, so VGNaC are not opened and action potentials are not fired.
So inhibitory interneurones regulate the excitability of the postsynaptic cell.

Question 18

What does postsynaptic activity look like with IPSPs?

Answer 18

Without inhibitory input, there is steady frequency of firing in response to sensory input.
If there is occasional activation of the inhibitory neurone, there are IPSPs, and a pause in firing when there is an IPSP.
The length of time of the pause depends on how much activation there is of the inhibitory neurone.
See picture.

Question 19

What is the importance of inhibitory neurones?

Answer 19

Inhibitory neurones alter the pattern and frequency of the postsynaptic cell.
This is important as information in the nervous system is encoded by a pattern and frequency of neuronal cells in a circuit.

Question 20

What are the glutamate receptors?

Answer 20

Ionotropic glutamate receptors are AMPA and NMDA.
These form an open channel in response to glutamate that is permeable to Na+ or K+, and causes IPSP.

Question 21

What are AMPA channels?

Answer 21

At resting membrane potential -65mV, it is typically Na+ influx.
There is some efflux of K+ due to chemical driving force (higher concentration inside), but the efflux is small compared to Na+ influx.

Question 22

What are NMDA channels?

Answer 22

These are also permeable to Ca2+.
These channels can be blocked by Mg2+, and APV drug.
When channel is open, Ca2+ influx, due to higher extracellular concentration, and depolarises the membrane.

Question 23

What is voltage clamping of excitatory synapses?

Answer 23

Clamp the cell at -60mV.
Stimulate axons with stimulating electrode.
Sharp rapid influx of Na+ through activated channels.
There is a negative deflection due to voltage clamp compensating a positive voltage.
There is then rapid decay of the synaptic current.
See picture.

Question 24

What is the hippocampus?

Answer 24

Strongly associated with learning and memory, and spatial navigation in humans.
Lesions prevents new memory formation.
Contains well-organised neuronal circuitry.

Question 25

What is the anatomical structure of the hippocampus?

Answer 25

There is a hippocampus on the left hemisphere, and a hippocampus on the right hemisphere - paired. There is a circuit consisting of 3 sets of neurones which connect to each other, then output to other regions of the brain.

Question 26

What is the circuit in the hippocampus?

Answer 26

The entorhinal cortex inputs into the dentate gyrus. The perforin pathway activates the granule cells in the dentate gyrus, which fire action potentials along axons through the mossy fibres to the CA3 regions. The CA3 regions has pyramidal cells, which are big neurones. These are excited and project along the Schaffers Collateral to the dendrites in the CA1 region.

Question 27

What do CA1 regions do?

Answer 27

The CA1 regions the send action potentials via axons to the subiculum, which connects to the cortex. The thalamus is the relay station of sensory input into the brain, it processes and sends information to the cortex as well.

Question 28

What is voltage clamping of glutamate receptors in hippocampal synapses?

Answer 28

The changes in voltage show the different glutamate receptors - AMPAR and NMDAR. Schaffer collaterals are stimulated, and causes an artefact on the recording. There is then a voltage clamp response.

Question 29

What is voltage clamping at -80mV in hippocampal synapses?

Answer 29

There is a sharp synaptic current - rapid onset and rapid decay. The current is exclusively carried by AMPAR. These are not blocked by Mg2+, so open and close in response to glutamate. See picture.

Question 30

What is voltage clamping at -40mV in hippocampal synapses?

Answer 30

The amplitude of synaptic response is reduced, and there are slower kinetics. The voltage is depolarised, which is sufficient to remove some of the Mg2+ block from some NMDAR, so both AMPAR and NMDAR are mediating the current. AMPAR are rapid, NMDAR are slow, so there is a slower decay of current. There is a decrease in current amplitude, due to voltage being closer to Ena, so driving force is reduced. See picture.

Question 31

What is voltage clamping at +20mV in hippocampal synapses?

Answer 31

The current moves through the channel at reversed polarity - upwards deflection. The cell is fully depolarised, so the Mg2+ block is fully removed, and there is full contribution of NMDAR, and AMPAR. There is K+ efflux rather than Na+ influx, which hyperpolarises. So voltage clamp has positive compensatory current. See picture.

Question 32

What is the patch clamp technique?

Answer 32

This uses a single electrode to measure the voltage and pass the current. This is useful as using two electrodes can be difficult as neurones can be small. The electrode has a large blunt opening, which removes a small patch of membrane, and forms a tight seal between the glass and membrane. So can record the activity of a population of the channels.

Question 33

How many channels does the patch clamp measure?

Answer 33

The patch clamp has 100s of channels in the membrane in the electrode. The activity of a single channel can be isolated, due to the channels opening at different times.

Question 34

What are single NMDA channel currents?

Answer 34

Measures the NMDA currents of a single channel at different Mg2+ concentrations. Measured with Mg2+ (normal physiological Mg2+), and 0 Mg2+.

Question 35

What is the effect of the NMDA channel with Mg2+ at -60mV?

Answer 35

There is a small downward deflection due to brief opening of the channel, which lets in a small amount of Na+. This causes a small compensatory negative charge. -60mV is hyperpolarised, so NMDAR is blocked by Mg2+ and has no effect. See picture

Question 36

What is the effect of the NMDA channel with Mg2+ at -30mV?

Answer 36

The membrane is more depolarised, so some of the Mg2+ block is removed. See picture.

Question 37

What is the effect of the NMDA channel with Mg2+ at 0mV?

Answer 37

There is no channel activity, due to being in equilibrium of the channels. There is no driving force so no flux. See picture.

Question 38

What is the effect of the NMDA channel with Mg2+ at +30 and +60mV?

Answer 38

The current is reversed, there are large channel activations. The cell is completely depolarised so all the Mg2+ block is removed, and K+ can leave through the channels and drives the current. Positive charge is leaving, so hyperpolarises. See picture.

Question 39

What is the effect of NMDA channels with 0 Mg2+?

Answer 39

-60mV - big channel opening, NMDAR are available and activated. 0mV - no current due to 0 driving force. +30 and +60mV, large currents. This is evidence that NMDA receptors are dependent on Mg2+ block, when not present, there is no block and K+ hyperpolarises at all voltages. See picture.

Question 40

What is associative learning?

Answer 40

Co-incidence activity is important for association of information. The more often a connection is activated, the stronger the synapse becomes. Cells that fire together, wire together.

Question 41

How are NMDARs coincidence detectors?

Answer 41

NMDAR are both ligand gated and voltage gated. They require glutamate binding, and depolarisation, which removes the Mg2+ block, to open its channel. When both conditions are met simultaneously, the NMDAR opens and allows Na+ or Ca2+ flow. Depolarisation occurs by the AMPAR activation.

Question 42

What happens when there is Ca2+ influx through NMDAR?

Answer 42

Ca2+ depolarises the cell, but is also a strong intracellular signalling molecule. e.g. kinase activation which causes transient changes in membrane receptors by phosphorylation. Ca2+ can trigger changes in gene expression, which synthesises new proteins for somatic action.

Question 43

What is plasticity?

Answer 43

The brain's ability to change and adapt by strengthening or weakening synaptic connections between neurones. Summation of EPSPs increases depolarisation and can reach threshold.

Question 44

What is temporal summation?

Answer 44

NMDAR responding to repeated patterns of presynaptic activation. This triggers greater and greater amplitude of EPSPs. This unblocks NMDA and causes downstream signalling in plasticity. See picture.

Question 45

What is spatial summation?

Answer 45

Simultaneous activation of the pre synapse and post synapse. Input from different neurones at the same time. This is more important than temporal summation. See picture.

Question 46

What is long term depression plasticity?

Answer 46

Spatial summation that activates NMDA receptors but without excess Ca2+ current. This causes dephosphorylation of downstream receptors and therefore weakens synaptic signalling and affects memory. This is the opposite of long term potentiation, which improves memory. By Ca2+ movement through NMDA which activates signalling pathways such as protein kinase C, which phosphorylates proteins and strengthens synapses and memory.

Question 47

What does an outward or inward current mean?

Answer 47

The current reverses in direction around 0mV because the channel is permeable to both Na+ and K+. Therefore the current is mostly carried by K+ above 0mV and mostly carried by Na+ below 0mV. There is equal movement of Na+ and K+ in opposite directions at 0mV.

Question 48

A drug called APV is an antagonist of NMDA channels. What does the figure show about the voltage clamp before and after APV? (see picture)

Answer 48

NMDA channels do not contribute to EPSC at -60mV, contribution is seen by shaded part, but has increasing contribution at more positive values. This is because of the Mg2+ block at negative voltages. When the the cell is depolarised the Mg2+ block is lessened so NMDA channels contribute to the EPSC, by allowing Ca2+ and Na+ influx on top of the Na+ influx through AMPA channels.

Question 49

Why is there a difference in K+ movement at different voltages?

Answer 49

The equilibrium potential of K+ is -80mV, so at -60mV there is little drive for K+ efflux. At +20mV, there is a big driving force so a large amount of K+ efflux, and therefore the current is positive. For Na+, the equilibrium potential is +60mV, so as the voltage becomes more positive there is less driving force for Na+ influx, and the current is a positive deflection.

Question 50

What does the effect of pure AMPA look like?

Answer 50

APV removes the NMDA component and reveals the pure AMPA. See picture.