Small-conductance chloride channels activated by calcium on cultured endocrine cells from mammalian pars intermedia

Abstract

Porcine intermediate lobe (IL) endocrine cells maintained in primary culture have been studied using patch-clamp derived configurations to record unitary activity on outside-out vesicles. Solutions were devised so as to record Cl current in isolation and to fix cytoplasmic Ca concentration [Ca]i between 0.1 microM and 3 microM. Between [Ca]i 0.5 and 1 microM, the chloride permeability was restricted to single events with a small amplitude, that varied linearly with the membrane potential. Mean slope conductance of this chloride channel was 2.5 pS. Single channel analysis yielded two mean open time values of 10 and 55 ms at -80 mV. Relaxations of chloride currents on outside-out patches was examined at different [Ca]i. Relaxation was negligible at 0.15 microM [Ca]i, whereas at higher [Ca]i, the current exhibited relaxation in response to voltage jumps the kinetic of which could be fitted by two exponentials. At 0.5 microM [Ca]i, the fast relaxation time constant was shown to be voltage insensitive with a value of about 10 ms. The slow relaxation time constant had a mean value of 75 ms at -60 mV and increased with membrane depolarization with a twofold change over 120 mV. Another voltage effect was to favour the slow opening mode at the more depolarized potentials: the ratio of fast to slow relaxations being 5:1 at -60 mV as compared to 1:1 at +80 mV). Finally the estimated probability of opening (po) linearly increased with voltage. po displayed a bell-shaped dependence on [Ca]i, so that full activation of the channels was not achieved.