Sodium and potassium conductances in somatic membranes of rat Purkinje cells from organotypic cerebellar cultures.

Abstract

1. The somatic voltage-gated conductances of Purkinje cells in organotypic cultures (Gahwiler, 1981) were studied using the outside-out patch recording configuration of the patch-clamp technique (Hamill, Marty, Neher, Sakmann and Sigworth, 1981). 2. When activated by step depolarizations, the tetrodotoxin-sensitive voltage-dependent Na+ current presented two distinct phases: an initial surge of inward current fluctuations which activates rapidly upon pulse onset and decays within 20-40 ms, and a later phase in which discrete bursts of single-channel activity are interspersed with silent periods. 3. Ensemble fluctuation analysis of the current fluctuations during the early phase of the Na+ current and measurements of single channels during both early and late phases indicate that a single type of Na+ channel can account for both phases of the Na+ current. This channel has an elementary current amplitude of -2 pA at -40 mV. This amplitude did not vary significantly between -60 and -20 mV. The mean open time depended on membrane potential, increasing by a factor of three between -60 and -20 mV. 4. The early component of the Na+ current activated at a threshold of -60 mV and reached its maximum amplitude at -20, mid-point for the activation curve being -40 mV. Times-to-peak current decreased with membrane potential, from 3.5 ms at -60 mV to 0.3 ms at 0 mV. The decay phase of the current presented two exponential components, with time constants of 1.5 and 10 ms at -40 mV. The steady state inactivation curve had a mid-point at -75 mV. 5. The late component of the Na+ current was observed in the voltage range from -60 to -20 mV, with a maximum at -40 mV. Its maximum amplitude corresponded to approximately 1.7% of the peak amplitude of the early component. 6. Macroscopic potassium currents were observed upon step depolarizations above a threshold of -30 mV. The currents activated in a voltage-dependent fashion, times-to-peak decreasing with depolarization, and partially inactivated during 40 ms depolarizing steps. Peak current amplitudes at any given membrane potential were decreased by depolarizing the holding potential. The macroscopic properties of the K+ current varied from patch to patch. 7. Two types of single-channel K+ currents were observed during steady-state depolarizations. The unitary current amplitudes were 2.7 and 10.4 pA at 30 mV, corresponding to chord conductances of 28 and 90 pS respectively. The 28 pS channel was observed as well upon step depolarizations, the time course of activation for the resulting averaged currents being voltage dependent. Both 28 and 90 pS channels were completely inhibited by external application of 1 mM-tetraethylammonium chloride.