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 E Na Two microelectrodes in muscle fibre 2

Real synaptic current Result: they are not Na + channels - current reverses near 0 mV, far from E Na 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 Ca 2+ permeability too: but little Ca 2+ 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 transmission Glutamate receptor GABA/glycine receptors - Cell inside more positive than E cl - Inward Cl - flux - Equivalent to outward flow of positive charge - Cell inside more negative than E cl - 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 E Cl is usually close to resting potential E r 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 ); chapter 6 (up to page 125) Nicholls et al chapters 3 & 9 - sections on ACh, glutamate, GABA and glycine receptor channels; chapter 13 - pages Kandel et al chapter 11, chapter 12 (pages ) 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 ) Kandel et al chapter 13 (Note: All these readings go into a lot of depth, so read selectively based on examples in the lecture)