Cytosolic free calcium regulation in response to acute changes in intracellular pH in vascular smooth muscle

This study examined the mechanisms whereby alterations of intracellular pH (pH(i)) impact on free cytosolic calcium (Ca(i)/²⁺) in cultured rat aortic vascular smooth muscle cells (VSMC) assayed in the presence of HCO₃/CO₂. Rapid cell alkalinization, effected by the exposure to NH₄Cl or removal of CO₂ from the superfusate, produced a rapid increase in Ca(i)/²⁺. The rise in Ca(i)/²⁺ was markedly diminished when sarcoplasmic reticulum (SR) Ca²⁺ stores had been depleted by prior exposure to arginine vasopressin (AVP) in Ca²⁺-free media or when SR release and reuptake of Ca²⁺ were blocked by the addition of 3,4,5-trimethoxybenzoic acid 8-(diethylamino)octyl ester (TMB-8), but was unaffected by the removal of external Ca²⁺ or inhibition of Ca²⁺ entry using NiCl₂. Cell acidification also resulted in a rapid increase in Ca(i)/²⁺. This Ca(i)/²⁺ increase was most apparent when pH(i) was very low (<6.6) and was unaffected by removal of external Ca²⁺ or NiCl₂ addition. Unlike the effect of cell alkalinization, the increase in Ca(i)/²⁺ associated with cell acidification was not prevented by pretreatment with AVP or TMB-8. We conclude that, in cultured VSMC, acute intracellular alkalinization and, to a lesser extent, acidification result in release of Ca²⁺ from internal stores. Alkalinization increases Ca(i)/²⁺ by promoting its release from a store which is AVP and TMB-8 sensitive, most likely the SR. Cell acidification increases Ca(i)/²⁺ from an intracellular store(s) that is neither AVP nor TMB-8 sensitive. The increase in Ca(i)/²⁺ produced by cell acidification may be explained on the basis of cell buffering such that, as cytosolic H⁺ increases, it displaces Ca(i)/²⁺ from internal buffers with similar affinities for Ca²⁺ and H⁺.