Next theme: What’s going on at the postsynaptic membrane? Ligand-gated ion channels: - ACh...

Next theme:

What’s going on at the postsynaptic membrane? Ligand-gated ion channels:- ACh receptors (excitatory)- glutamate receptors (excitatory)- GABA receptors (inhibitory)- glycine receptors (inhibitory)

1

How does ACh depolarise the neuromuscular junction?

Perhaps ACh opens Na+ channels? How to test:•Check the reversal potential of the current - in this case it should be near ENa

•Two microelectrodes in muscle fibre

2

Real synaptic current

•Result: they are not Na+ channels - current reverses near 0 mV, far from ENa

•So which ion conducts the current?

3

•Which ion conducts the current? - i.e.•Which ion has equilibrium potential near 0 mV? - i.e.•Which ion is at equal concentration outside and inside?

•None of them! - so the ACh receptor channel must conduct more than one ion•In fact both Na+ and K+ are conducted equally well•Substantial Ca2+ permeability too: but little Ca2+ available•So Na+ and K+ carry most of the current

4

Fluxes are equal here

5

Synaptic current and synaptic potential

•Current flows during the rising phase of the EPP•Inward current causes the depolarisation (“upstroke”) of the EPSP•Repolarisation is passive return to resting potential

6

Location of ACh receptors at the neuromuscular junction

7

How does ACh trigger an AP?

8

Recording single ACh receptors

9

Effect of a “puff” of ACh

•How to apply ACh briefly?•We need “outside-out” patches•ACh can then be added/removed very fast

10

How to apply a “puff” of ACh

ACh

No ACh

11

Effect of a “puff” of ACh

•Channels stay open as long as ACh is bound

•Unbinding of ACh is random: so channel open time is random

•Add single channel currents: we get the macroscopic endplate current

12

What does the ACh receptor look like?

•Very distant relative of Na+ channel•Hydropathy plot like this:

13

What does the ACh receptor look like?•Unlike the K+ channel - it has 5 subunits and not 4

14

How does the ACh receptor select cations?

Conserved negative charges in most subunits in the M2 helix

15

How does the ACh receptor select cations?

The negative charges are all on one side of the M2 helix

16

How does the ACh receptor select cations?

The M2 helices face inwards around the pore

17

How does the ACh receptor select cations?

So we would have 3 negatively charged rings around the pore

18

How does the ACh receptor select cations?

Mutating the rings alters ion conductance: so they are important in selecting ions to go through the pore

19

Central excitatory synaptic transmission•Introducing a new neurotransmitter: glutamate•Glutamate receptors: two types

20

•Introducing a new neurotransmitter: glutamate•Glutamate receptors: two types

•Non-NMDA receptors:- don’t respond to the glutamate analogue NMDA•Functionally like ACh receptor...but little sequence similarity•Opened by glutamate•Allow Na+ and K+ to pass

Central excitatory synaptic transmission

21

•NMDA receptors:- DO respond to the glutamate analogue NMDA•Opened by glutamate•Allow Na+ and K+ to pass•Similar sequence to non-NMDA receptors BUT very different functionally:- modulated by many substances- important in synaptic plasticity (maybe memory/learning: more later)

Central excitatory synaptic transmission

22

ACh receptor

AChR and GluR: functionally similar

Glutamate receptor

23

Synaptic inhibition

Inhibitory neurone

Excitatory neurone

24

Synaptic inhibition

Inhibitory neurone

Excitatory neurone

•What makes an excitatory neurone excitatory? (or an inhibitory neurone inhibitory)?- the kind of transmitter released from its terminal- the kind of receptor on the postsynaptic membrane

•Let’s look at inhibition

25

•Transmitters involved: GABA, glycine•Time to look at the transmitters

Central inhibitory synaptic transmission

26

•Transmitters involved: GABA, glycine•Receptor sequences closely related to ACh receptor

Central inhibitory synaptic transmission

•...but functionally they are the opposite of the AChR!27

Central inhibitory synaptic transmissionGlutamate receptor GABA/glycine receptors

- Cell insidemore positivethan Ecl

- Inward Cl- flux- Equivalent to outward flow of positive charge

- Cell insidemore negativethan Ecl

- Outward Cl- flux- Equivalent to inward flow of positive charge

28

Central inhibitory synaptic transmission

GABA/glycine receptors

Conclusion:•GABA and glycine receptors conduct chloride ions•ECl is usually close to resting potential Er

•So inhibitory postsynaptic potential (IPSP) is small: often negative but sometimes zero•Even if it’s zero it is still inhibitory (we will see why later)

29

Reading for this lecture:

•Purves et al chapter 5 (page 96 - 107); chapter 6 (up to page 125)•Nicholls et al chapters 3 & 9 - sections on ACh, glutamate, GABA and

glycine receptor channels; chapter 13 - pages 244-247•Kandel et al chapter 11, chapter 12 (pages 212-214)

Next lecture:Ion channel modulation by G proteins and second messengers:

slow synaptic transmission•Purves et al chapter 7 (up to page 153)

•Nicholls et al chapter 10 (especially pages 184-188)•Kandel et al chapter 13

(Note: All these readings go into a lot of depth, so read selectively based on examples in the lecture)