A novel method for recording whole-cell and single-channel currents from differentiating cerebellar granule cells in situ

Abstract

A preparation procedure is described yielding very thin tissue layers (1-3 cells) of the rat cerebellum. One-week-old rat cerebellum was embedded in agarose and cut in 1 mm slices, which then were mounted by a fresh fibrin preparation on the bottom of a culture dish. After coagulation of the fibrin the slice was gently lifted, leaving a thin fingerprint-like layer of cells on the bottom of the dish. Individual cells could be identified by their topographical positions. From these preparations patch-clamp recordings were made of cells in the external and the internal granule layer and of migrating cells. Whole-cell currents of cells in the external granule layer, recorded using the perforated-patch method, show that these neuroblasts already possess a full complement of ionic currents, consisting of a transient sodium current and a transient and a sustained potassium current. Whereas the potassium currents are predominant, the maximum sodium peak currents are minute (42 +/- 5 pA, mean +/- S.E.M., n = 21), i.e. too small for generating action potentials. There were no significant differences between cells of the external and the internal granule layer. The ionic channels passing these currents were identified and characterized in single-channel studies on cell-attached patches. Two types of potassium channel were found: a non-inactivating channel with a single-channel conductance of 21 pS and an inactivating channel with a conductance of 6 pS.