Na+-Ca2+ exchange in cultured vascular smooth muscle cells

Abstract

Vascular smooth muscle cells (VSMC) contract as intracellular free calcium ([Ca2+](i) rises. While Na+-Ca2+ exchange has been proposed to contribute to transmembrane Ca2+ flux, its role in cultured VSMC is unknown. Accordingly, we have investigated the role of Na+- Ca2+ exchange in unidirectional and net transmembrane Ca2+ fluxes in cultured rat aortic VSMC under basal conditions and following agonist-mediated stimulation. Transmembrane Ca2+ uptake was significantly increased in response to a low external Na+ concentration ([Na+]0) compared with 140 mM [Na+]0. Na+-dependent Ca2+ uptake in response to low [Na+]0 was further increased by intracellular Na+ loading by preincubation of the VSMC with 1 mM ouabain. Under steady-state conditions, Ca2+ content varied inversely with [Na+]0, increasing from 1.0 nmol Ca2+/mg protein at 140 mM [Na+]0 to 4.0 nmol Ca2+/mg protein at 20 mM [Na+]0. Increasing [K+]0 to 55 mM also enhanced Na+-dependent Ca2+ influx. Augmentation of Ca2+ uptake with K+ depolarization was not significantly inhibited by the calcium channel antagonist verapamil. Transmembrane Ca2+ efflux was increased in response to 130 mM [Na+]0 compared with zero [Na+]0 (iso-osmotic substitution with choline+), and was further stimulated by the vasoconstrictor angiotensin II, which is known to elevate [Ca2+](i). These changes in [Ca2+](i) were studied directly using fura-2 fluorescence measurements. Elevated [Ca2+](i) levels returned to baseline more rapidly in the presence of normal (130 mM) [Na+]0 compared with zero [Na+]0 (iso-osmotic substitution with choline+). These findings suggest that a bidirectional Na+- Ca2+ exchange mechanism is present in cultured rat aortic VSMC. Na+- Ca2+ exchange appears to play a part in Ca2+ homeostasis, particularly under conditions of altered intracellular Na+ or increased [Ca2+](i) following agonist stimulation.