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Abstract:
1. Postnatal rat retinal ganglion cells (RGCs) can be maintained and identified in dissociated long-term culture. After 4-7 days in vitro they form glutamatergic synapses with other RGCs or putative amacrine cells. Here we intended to characterize the postsynaptic features of these in vitro synapses. 2. Pair patch-clamp recordings in the whole cell configuration were performed to study the properties of synaptic glutamate receptors. Immunohistochemically and physiologically identified RGCs were activated by short depolarizing voltage steps. This elicited glutamatergic excitatory postsynaptic currents (EPSCs) in coupled neurons. At room temperature, evoked EPSCs (eEPSCs) had latencies between 3 and 7 ms and amplitudes between 36.4 and 792.6 pA. 3. Postsynaptic neurons were electrotonically compact and therefore well suited for analysis of fast synaptic events. All cells were responsive to exogenous glutamate, alpha-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid (AMPA) and N-methyl-D-aspartate (NMDA). The current-voltage relationships of AMPA-activated currents were linear, whereas NMDA-induced whole cell currents displayed the typical characteristics including a negative slope conductance in the presence of Mg2+. In contrast to AMPA-activated currents, NMDA-activated currents had the usual slow onset and decay. 4. RGCs obviously failed to generate NMDA-receptor-mediated EPSCs, because all postsynaptic cells lacked a slow current component even in the absence of added Mg2+ and in the presence of glycine. Retinal eEPSCs were completely blocked by 6,7-dinitroquinoxaline-2,3-dione (DNQX). 5. eEPSCs as well as spontaneous EPSCs (sEPSCs) were characterized by a very rapid time course. In eEPSCs, 20-80% rise times and time constants of decay (tau DS) were on average 0.64 and 1.96 ms, respectively. eEPSCs were extremely fast, with average rise times of 0.34 ms and tau DS of 1.20 ms. The latter numbers closely correspond to the values obtained for DNQX-sensitive miniature EPSC (mEPSC) in postnatal day 5 rat RGCs in situ. 6. To clarify whether the decay of fast AMPA-receptor-mediated EPSCs of retinal neurons was determined by the onset of glutamate receptor desensitization, we compared the decay of sEPSCs with the decay of the glutamate response of excised out-side-out membrane patches. Glutamate-activated currents were elicited by a rapid superfusion device (time constant of rise = 0.7 ms). The response to 1 mM of glutamate decayed 2 to 4 times more slowly than the sEPSCs. 7. These results suggest that desensitization did not limit the rate of decay of purely AMPA-mediated EPSCs in response to ganglion cell activation.(ABSTRACT TRUNCATED AT 400 WORDS)
