Sodium and calcium fluxes in a clonal nerve cell line.

1. 22Na+ and 45Ca2+ fluxes were studied in the clonal nerve cell line PC12. Three distinct types of ion channels were found: (a) voltage‐dependent Na+ channels, (b) voltage‐dependent Ca2+ channels, and (c) acetylcholine‐activated channels permeable to both ions. 2. 22Na+ uptake through voltage‐dependent Na+ channels is induced by veratridine and scorpion venom, and is inhibited 50% by 5 X 10(‐7) M‐tetrodotoxin and greater than 98% by 5 X 10(‐6) M‐tetrodotoxin. 3. 45Ca2+ uptake through voltage‐dependent Ca2+ channels is induced by depolarizing the cells in 50 mM‐KCl. This flux is not dependent on the presence of Na+ in the medium and is insensitive to 5 X 10(‐6) M‐tetrodotoxin. However, 1 mM‐Mn2+ causes a 95% inhibition of K+‐induced 45Ca2+ uptake. 4. Veratridine and scorpion venom also induce voltage‐dependent 45Ca2+ uptake which can be blocked by 1mM‐Mn2+. In contrast to KCl‐induced 45Ca2+ uptake, this flux is completely blocked by 5 X 10(‐6) M‐tetrodotoxin and is abolished by removal of Na+ from the medium. Thus the depolarizing stimulus for Ca2+ uptake in this case is Na+ from the medium. Thus the depolarizing stimulus for Ca2+ uptake in this case is Na+ influx through voltage‐dependent Na+ channels. 5. Carbamylcholine induces both 22Na+ and 45Ca2+ fluxes which are blocked by nicotinic cholinergic antagonists with the exception of alpha‐bungarotoxin. The 22Na+ flux occurs exclusively via acetylcholine receptor channels, as evidenced by the lack of effect of 5 X 10(‐6) M‐tetrodotoxin. In the presence of Na+, almost all of the 45Ca2+ uptake can be blocked by 1 mM‐Mn2+ and thus occurs via voltage‐dependent Ca2+ channels which are activated by the depolarizing Na+ influx. 6‐‐8% of the total 45Ca2+ flux, however, is insensitive to 1 mM‐Mn2+, suggesting that this portion of the uptake occurs via the acetylcholine receptor channels. In Na+‐free medium, the Mn2+‐resistant 45Ca2+ component increases to 40% of the total uptake, apparently due to lack of competition from Na+ for the acetylcholine receptor channels. This receptor‐linked flux still causes sufficient depolarization to induce the additional 60% of the Ca2+ flux through voltage‐dependent, Mn2+ sensitive Ca2+ channels. 6. Mn2+ inhibits Ca2+ flux through voltage‐dependent Ca2+ channels by competing for entry through these channels. 50 mM‐KCl induces 54Mn2+ fluxes in PC12 cells that are comparable in magnitude to 45Ca2+ fluxes. 7. In normal saline 45Ca2+ efflux from PC12 cells is several times more rapid than in Na+‐free medium, indicating the presence of a Ca2+‐Na+ exchange mechanism.