³⁶Cl fluxes in dispersed rat submandibular acini: Effects of Ca²⁺ omission and of the ionophore A23187

Abstract

Transmembrane fluxes of³⁶Cl were investigated in dispersed acini of the rat submandibular gland in Ca²⁺-containing and Ca²⁺-free media and also in the presence of the divalent cation ionophore A23187. In Ca²⁺-replete medium, a time-dependent uptake of tracer resulted in a steady state³⁶Cl content of 8.5±0.3 nmol/mg protein in 3–5 min. This uptake was reduced 32% by 1 mM furosemide and 27% by 1 μM acetylcholine. In the presence of Ca²⁺, the ionophore (10⁻⁵ M) reduced tracer uptake 36% and prevented further effects of either acetylcholine or furosemide. Both acetylcholine and A23187 caused a rapid net efflux of³⁶Cl from tracer-preloaded acini in Ca²⁺-containing medium (37% and 20%, respectively). When Ca²⁺ was omitted from the incubation medium, basal³⁶Cl uptake in the absence of added test substance was the same as in Ca²⁺-containing medium but was not affected by acetylcholine, while it was still reduced 29% by furosemide. Addition of acetylcholine to preloaded acini in Ca²⁺-free medium caused only a transient and unsustained³⁶Cl efflux but subsequent addition of Ca²⁺ produced a 36% reduction in tracer content. The ionophore caused a net³⁶Cl efflux in Ca²⁺-containing medium (24% decrease in³⁶Cl content) but had no effect in Ca²⁺-free medium. Subsequent addition of Ca²⁺ resulted in a 27% net efflux of tracer. The calmodulin inhibitor triflouperazine caused a 14% increase in³⁶Cl uptake but did not cause³⁶Cl efflux from preloaded cells or modify acetylcholine-induced efflux of tracer from these cells. These results suggest that intracellular Ca²⁺ regulates the efflux but not the uptake of Cl into salivary cells. Since this efflux is thought to occur by way of luminal membrane channels in salivary acini, the role of Ca²⁺ is likely to be to control the opening of these channels. Ca²⁺ does not seem to regulate Cl entry into salivary acini directly, but, after binding to calmodulin, may modulate some aspects of Cl entry into salivary cells.