Patch-clamp analysis of voltage-gated currents in intermediate lobe cells from rat pituitary thin slices.

Abstract

Ionic currents of hypophyseal intermediate lobe cells were studied using a thin-slice preparation of the rat pituitary in conjunction with conventional and perforated whole-cell patch-clamp recording techniques. A majority (89%) of the cells studied generated Na+, Ca2+ and K+ currents upon depolarizing voltage steps and responded to bath application of gamma-aminobutyric acid (GABA; 20-50 microM) with inward currents (in symmetrical chloride, holding potential -80 mV). A small percentage of cells (11%) did not display inward membrane currents upon depolarization and was unresponsive to GABA. In the first type of cells, Ca2+ and K+ currents were further studied in isolation. Ca2+ tail currents showed a biphasic time course upon repolarization, with time constants and amplitudes of 2.07 +/- 0.29 ms, 123 +/- 22 pA (for the slowly deactivating component) and 0.14 +/- 0.06 ms, 437 +/- 33 pA (for the fast-deactivating component; means +/- SD of n = 4 cells). Slowly and fast-deactivating conductances were half-maximally activated at around -10 mV and +10 mV respectively. Depolarizing voltage steps elicited two types of K+ current, which were separated using a prepulse protocol. A fast-activating, transient component showed half-maximal steady-state inactivation between -65 mV and -45 mV depending on the divalent cation composition of the external solution. Its decay was fitted by single-exponential functions with time constants of 36 +/- 11 ms and 3.9 +/- 0.9 ms at -20 mV and +40 mV respectively (mean +/- SD; n = 4 cells). Whereas the peak current amplitudes of the transient K+ current component remained stable, the amplitude of the second, delayed component increased progressively throughout the course of whole-cell experiments. In cells recorded with the perforated whole-cell technique, bath application of dopamine (10 nM-1 microM) induced large hyperpolarizations from a spontaneous membrane potential of -40 mV, but did not consistently affect the amplitude of the voltage-gated K+ conductances. These data are compared to previous studies using other preparations of the intermediate lobe, and differences are discussed, thus helping to extend our knowledge of electrical excitability of hypophyseal cells.