Proceedings of The Physiological Society

University College London (2003) J Physiol 547P, PC29

Poster Communications

Regulation of cerebellar inhibitory synaptic plasticity by presynaptic NMDA receptors

I.C. Duguid and T.G. Smart

Department of Pharmacology, University College London, Gower Street, London WC1E 6BT, UK

Inhibitory synaptic transmission mediated by GABAA receptors on cerebellar Purkinje neurones (PNs) exhibits a long-lasting 'rebound potentiation' (RP) subsequent to repetitive heterosynaptic climbing fibre activation (Kano et al. 1992). We report that during rebound potentiation a novel, enhancement in GABA release termed depolarization-induced potentiation of inhibition (DPI) that involves the retrograde release of a glutamate-like transmitter and subsequent activation of presynaptic N-methyl-D-aspartate (NMDA) receptors.

To induce DPI, cultured PNs (16-21 DIV) from Sprague-Dawley rats were voltage clamped at -70mV in normal ACSF containing both tetrodotoxin (TTX) and 6-cyano-7-nitroquinoxaline-2,3-dione (CNQX) to block action potentials and non-NMDA receptor-mediated currents. Cells were then subjected to a single depolarising pulse protocol that involved rapid depolarisation of the cell eight times from -70 to 0 mV (2 s intervals). RP (Llano et al. 1991; Kano et al. 1992) was associated with an increase in the mean mIPSC amplitude (146 ± 9 % of control; n = 10; P < 0.01; t test) while the mean mIPSC frequency displayed a clear biphasic profile. Initially, a rapid reduction in the mean mIPSC frequency was observed (79 ± 5% of control; lasting ~60 s), before developing into a novel, transient increase in the mean mIPSC frequency observed at 3 min post-stimulus (149 ± 8% of control; n = 10; P < 0.01). We examined whether the coincident presynaptic component to RP occurred also in immature (P6-8) slice preparations, during a stage in development when morphology and synaptic connectivity closely resembles that of cultured PNs. Induction of RP resulted in a similar increase in the mean mIPSC amplitude (154 ± 21% of control; n = 7; P < 0.02) and frequency (160 ± 30% of control; n = 7; P < 0.02) measured 5 min post-stimulus. The involvement of presynaptic NMDA receptors during the increase in GABA release was implicated by three main findings. Firstly, the antagonist, D-(2)-amino-5-phosphovalerate (D-APV; 50 µM), completely abolished the mIPSC frequency potentiation in both cultured and immature (P6-8) PNs (98 ± 11 % of control and 101 ± 13% of control, respectively). Secondly, brief (4 s), focal applications of NMDA (100 µM) induced a sustained increase in the release of GABA from basket/stellate cells persisting for > 13 min (no postsynaptic NMDA currents were observed). Finally, triple immunocolocalisation of NMDA receptor subunits (NR1, NR2A,C&D) provided further evidence for the existence of presynaptic NMDA receptors at putative basket/stellate cell axon terminals.

This presynaptic regulation of transmitter release, during the induction of cerebellar synaptic plasticity, will ensure control over PN excitability and subsequent sensorimotor co-ordination in the cerebellar cortex.

This work was supported by the MRC.

Where applicable, experiments conform with Society ethical requirements