VOLTAGE-DEPENDENCE OF CONDUCTANCE CHANGES EVOKED BY GLYCINE RELEASE IN THE ZEBRAFISH BRAIN

1. The kinetics and mechanisms underlying the voltage dependence of in hibitory postsynaptic currents (IPSCs) recorded in the Mauthner cell ( M cell) were investigated in the isolated medulla of 52-h-old zebrafis h larvae, with the use of whole cell and outside-out patch-clamp recor dings. 2. Spontaneous miniature IPSCs (mIPSCs) were recorded in the pr esence of 10(-6) M tetrodotoxin (TTX), 10 mM MgCl2, and 0.1 mM [CaCl2] (o). Depolarizing the cell from -50 to +50 mV did not evoke any signif icant change in the distribution of mIPSC amplitudes, whereas synaptic currents were prolonged at positive voltages. The average decay time constant was increased twofold at +50 mV. 3. The voltage dependence of the kinetics of glycine-activated channels was first investigated dur ing whole cell recording experiments. Currents evoked by voltage steps in the presence of glycine (50 mu M) were compared with those obtaine d without glycine. The increase in chloride conductance (g(Cl)-) evoke d by glycine was time and voltage dependent. Inactivation and reactiva tion of the chloride current were observed during voltage pulses from 0 to -50 mV and from -50 to 0 mV, respectively, and they occurred with similar time constants (2-3 s). During glycine application, voltage-r amp analysis revealed a shift in the reversal potential (E(Cl)-) occur ring at all [Cl-](i) tested. 4. The basis of the voltage sensitivity o f glycine-evoked g(Cl)- was first analyzed by measuring the relative c hanges in the total open probability (NPo) of glycine-activated channe ls with voltage. This study focused on patches displaying a single con ductance level of 40 pS. NPo increased steadily from -60 to 0 mV and t hen remained stable for higher holding potential values. As for g(Cl)- , measured during whole cell recordings, changes in NPo with voltage w ere not instantaneous. 5. The relative contribution of opening frequen cy and mean open time and/or burst duration to the voltage dependence of NPo, were analyzed by comparing closed time and burst duration hist ograms at -50 and +50 mV. Three mean burst durations (0.6, 3, and 30 m s; V-h = -50 mV) characterized glycine channel activity in the M cell. Short gaps (0.6 ms) within a burst and a short burst component (0.9 m s) were voltage independent, whereas medium (tau(b2)) and long bursts (tau(b3)) were lengthened when the patch was depolarized to +50 mV. At positive voltages, burst frequency was also increased, whereas the re lative proportion of tau(b1) tau(b2), and tau(b3) remained unchanged. 6. Slow and fast changes in channel kinetics with membrane potential s uggest a complex channel behavior. A model is proposed in which an ina ctivated state is linked to the liganded closed state with voltage-dep endent inactivation and closing rate constants. 7. Our results suggest that a change in the mean open time of the glycine-gated channel esse ntially accounts for voltage-dependent properties of the inhibitory sy naptic currents. Voltage-dependent desensitization will only occur dur ing long-lasting epileptic activity of the inhibitory presynaptic cell s.