Interneurones, spike timing, perception & synchronous clapping R Miles, INSERM EMI 0224, Paris.

1. Interneurones, spike timing, perception & synchronous clapping R Miles, INSERM EMI 0224, Paris.

2. The message for perception, action potentials must be generated with precise timing. for precise timing, a precise stimulus isn’t enough. for precise timing, the stimulus must be biphasic. positive - negative. depolarising - hyperpolarising

3. Important article on synchronous clapping Self organising processes: the sound of many hands clapping. Neda et al Nature (2000) Global sound intensity Local sound intensity Index of synchrony 0 10 20 30 Time sec After a performance, audiences clap at first randomly and then synchronously. Can we compare synchronous clapping to neuronal synchronisation…

4. Gamma frequency oscillations synchronous oscillations at 30 – 70 Hz cortex, hippocampus perception, attention, and sensorimotor coordination the binding problem – how to link the activity of different neurones engaged in the same cognitive task. Gamma oscillation evoked by visual stimulation in area 17 of the visual cortex Of the awake cat (Gray, 1994)

5. Synchronous clapping I cells in gamma oscillations Must be excited Must be excited Interaction between members Synaptic interactions between of audience interneurones Must not clap at different rhythms Interneurones must have similar properties Clapping and interneurons: common mechanisms for synchrony 1 2 3

6. Excitatory drive to interneurones in gamma oscillations In vivo – glutamatergic excitation and liberation of ACh. 20 mV Slow, excitation of layer 5 cortical P cell Initiated by stim of cholinergic pedunculopontine tegmental nucleus cat - ketamine / xylazine (Steriade & Amzica, PNAS, 1996) Control mAChR blocked 0.2 s Stim In vitro « models » of gamma oscillations tetanic stimulation – glutamate plus? agonists at muscarinic AChR agonists at kainate receptors agonists at mGluR Intra Spikes Power Field Mouse somatosensory cortex. 0.3 µM kainate + 20 µM carbachol -20 0 20 1 10 100 Freq Hz Time ms Buhl, Tamas & Fisahn J Physiol, 1998

7. Interneurone connectivity • Interneurons that inhibit any target cell • Interneurons that inhibit exclusively interneurons • Interneurons that excite interneurons What molecular and developmental mechanisms underly the formation of distinct connectivities?

8. Interneurons that inhibit any target cell I-cell intra 20 mV extra 20 µV 20 ms Popln Activity Sp / s 0 50 100 Time ms (Cohen & Miles, 2000) In a network, these interneurons generate an inverse synchrony in principal cells – they tell them when not to clap…

9. Interneurons that inhibit interneurons Interneurons containing Calretinin contact selectively other I-cells 200 µm 20 mV I cell IPSP 2 mV Random firing 20 mV Synchrony 10 µm Reconstruction: Red – axons Black – soma dendrites Black – CR+ I-cell axon Brown –CB+ I cell soma 20 ms Synchrony occurs when IPSPs cohere with intrinsic cellular AHP Gulyas, Hajos & Freund, 1996 These cells generate a rhythm by synchronising IPSPs within the population of I-cells. They synchronise and spread an anti-clapping message. Lytton & Sejnowski, 1991 Wang & Rinzel, 1993

10. 20 µm 200µm Interneurons that excite interneurons. Gap junctions occur between subsets of cortical and hippocampal interneurons. Mediate electrotrotonic coupling Formed by the connexin family of proteins Transmit electrical signals rapidly between coupled cells Gibson Bierlein & Connors, 1999 Galaretta & Hestrin, 1999 0.1 µm Tamas, Buhl, Lorincz & Somogyi, Nat Neurosci, 2000

11. Gap junctional coupling between cortical interneurons Gap junctions transmit signals rapidly They act as low pass electrical filters. Slow events (AHP) are better transmitted than fast events such as action potentials. A presynaptic action potential induces a postsynaptic « spikelet » of 0.5 – 2 mV. Gap junctions and GABAergic synapses may exist between two interneurones 1 nA Current 30 mV Cell 1 0.5 mV Cell 2 CR 0.5 mV 10 mV 25 ms Freq Hz 0.5 mV Galaretta & Hestrin, Nature 1999 Gibson, Beierlein & Connors, Nature 1999 5 ms

12. Interneurons should have similar properties… Interneurones form ~10% of cortical nerve cells. Diversity in co-transmitter peptides – VIP, CCK, SS Ca-binding proteins – PV, CB, CR. Morphological diversity Dendritic arborisation – which fibres can excite? Axonal arborisation – site of inhibition: somatic, dendritic, axonal. Parra, Gulyas & Miles, Neuron, 1998

13. Linkage GluR Markers Physiol Tout Cell number Interneurons should have similar properties … Diversity in firing patterns – fast firing cells – Kv3 channel (Lien & Jonas 2003) Diversity in expression of AMPA, NMDA and mGluRs. Receptors for modulating transmitters. Cluster analysis - Cortical I cells 20 mV Musc mGluR NA 5HT 2 m SLM SR SO Musc ACPD NA 5HT Musc ACPD NA 5HT Musc ACPD NA 5HT Cauli et al 2000 Parra et al 1998

14. tACPD 100 pA 10 s % Of Cells mGluR 1 5 1+5 1 5 1+5 Differences in expression of proteins associated with gamma Group 1 mGluRs Connexins Type I 20 mV Type II 4 mV Type III Electrical coupling 100 ms Type IV Cx36 Cx32 Expression of two distinct connexins in 6 interneurons from RT- PCR Single cell RT-PCR for mGluR1 & mGluR5 Venance et al PNAS, 2001 Van Hooft et al, J Neurosci 2000

15. So, as for synchronous clapping, inhibitory cells…. • Recieve a slow excitation • Interact between themselves • Have quite similar properties But to understand how the rhythm emerges, must think about interactions involving I cells … • Interactions between inhibitory cells • Synaptic excitation of inhibitory cells • Excitation of pyramidal cells

16. Three types of interaction between inhibitory cells GABA-mediated inhibition Excitation mediated by gap junction Chemical inhibition + gap junction 0.5 mV 50 ms 200 ms Biphasic signals generated by gap and GABAergic interactions give the best spike transmission at gamma frequencies 1 mV Tamas et al, Nat Neurosci, 2002

17. Synaptic excitation of inhibitory cells I cells have active dendrites Active post-synaptic properties modify EPSP shape -55 mV 0.25 mV 5 ms -75 mV hot spots Voltage-dependent activation of both Inward – peak of EPSP enhanced and Outward – decay of EPSP accelerated 50 µm Ca transients evoked by somatic firing Galarretta & Hestrin, Science, 2001 Kaiser et al J. Physiol, 2001

18. Inhibitory cell Pyramidal cell EPSP 2 mV 20 ms Firing 20 mV Temporal precision 20 0 0 50 100 ms 0 50 100 ms Temporal precision of EPSP – spike coupling in hippocampal neurones (Fricker & Miles Neuron 2000)

19. Mechanism of precise EPSP – spike coupling in I-cells V-clamp response to EPSP waveform EPSP voltage dependance Command -47 -55 -70 mV 200 pA 5 mV Inward plus outward 20 ms 40 pA 10 ms Outward (TTX) Integral Peak Inward (4AP+TEA) -80 -60 -40 Vm mV So precise spike timing depends on a biphasic signal EPSP initiates inward - outward current

20. High variance Low variance But is pyramidal cell spike generation precise or imprecise? Fricker & Miles say isolated events initiate spikes with variable timing but Mainen and Sejnowski say noisy stimuli initiate firing with millisecond precision Or maybe the precision of the timing depends on the variance / amplitude of the noisy stimulus? 200 pA 20 mV noisy stimulus 30 mV 200 pA square pulse 200 ms 200 ms Axmacher & Miles, soumis Mainen & Sejnowski, Science, 1995

21. Low variance High variance Noise amplitude – cellular currents – precision in P-cells Noise 20 mV • -80 -56 • 74 -50 • 70 -46 • HP PP mV • -96 -64 • 88 -55 • 83 -51 • HP PP mV Cellular current 20 pA Firing 20 ms Low variance noise elicits purely inward currents and give low temporal precision High variance noise elicit inward – outward currents and give precise firing (maybe voltage dependent inactivation of K current and enhanced persistant Na current) Axmacher & Miles

22. Precision in timing of Pcell firing: EPSP – IPSP sequence Current clamp Voltage clamp EPSP Di-synaptic IPSC sum Di-synaptic EPSP Afferent EPSC 2.5 ms 5 ms 2 ms Inhibition functional …blocked 10 ms Two puise – Inhibition functional - Inhibition blocked Precise spike timing depends on a biphasic EPSP – IPSP sequence Pouille & Scanziani, Science, 2001

23. Somatic vs Dendritic summation - precision Dual somatic –dendritic records Soma Dendrite afferent EPSP sum Dendrite sum Soma di-synaptic IPSP Summation Double pulse summation 200 0 -100 -20 0 20 Interval ms Feedforward inhibition terminates on the soma Restricted somatic window for summation. Pouille & Scanziani, Science, 2001

24. Biphasic signals generate precisely timed spikes Gap junctional excitation – chemical IPSP 1 ms EPSP initiates inward - outward current 10 ms Monosynaptic EPSP - di-synaptic IPSP 2.5 ms

25. But why do we need precise spike timing? Oscillations - binding problem LTP LTD depend crucially on pre- and post spike timing Binaural sound discrimination depend on resolving timing differences in 100µs. Olfactory discrimination depend on resolving signals from oscillations And is imprecise, or delayed, firing ever useful? Late firing can maintain persistent activity in a network (XJ Wang) Reverberating circuits.