MUSCARINIC ACTIVATION INHIBITS T-TYPE CA2-CELLS( CURRENT IN HEN GRANULOSA)

The effect of the muscarinic agonist calbachol on Ca2+ currents in hen granulosa cells isolated from the largest ovarian Follicle was studie d. The major Ca2+ current observed using the perforated patch techniqu e with a recording solution containing 10 mM Ca2+, but no Na+ or K+, e xhibited characteristics typical of T-type Ca2+ current: maximal ampli tude at -20 mV, rapid inactivation (half-time of 42 +/- 3 msec at -20 mV), inhibition by 100 mu M Ni2+, and insensitivity to the dihydropyri dine Ca2+ channel antagonist, nifedipine. In all cells studied, carbac hol (0.5 mM) caused an inhibition of this current (elicited by depolar izing pulses from -70 to -20 mV) to an average maximal decrease of 90 +/- 2% below basal values. In some 50% of the cells, the Ca2+ current also partially recovered during the 10-min exposure to the muscarinic agonist. These effects were prevented by the muscarinic antagonist atr opine (1 mu M). To test whether this inhibition was due to increases i n intracellular free Ca2+ concentrations ([Ca2+](i)), [Ca2+](i) was si multaneously measured in fura-2-loaded cells. For cells Incubated in n ormal solution, [Ca2+](i) was 0.15 +/- 0.02 mu M, but increased to 0.2 5 +/- 0.6 mu M in cells exposed to the recording solution. Under these conditions, carbachol failed to produce the expected [Ca2+](i) transi ents, but, rather, caused a small decrease (8 +/- 2%) in basal [Ca2+]( i) attributable to its diminution of Ca2+ current. Thus, the results d emonstrated an important muscarinic inhibition of the T-type Ca2+ curr ent not related to [Ca2+](i) fluctuations. They indicate, on the other hand, that [Ca2+](i) can strongly modulate carbachol-induced mobiliza tion of Ca2+ from the intracellular stores.