Intracellular alkalinization by NH4Cl increases cytosolic Ca2+ level and tension in the rat aortic smooth muscle

Intracellular pH (pHi) is elucidated to be an important regulator of various cell functions, but the role of pHi in smooth muscle contraction remains to be clarified. The purpose of the present study is to examine the effects of cell alkalinization by exposure to NH4Cl on cytosolic Ca2+ level (Ca2+]i) and on muscle tone. We attempted simultaneous measurements of both Ca2+]i and contractile force in rat isolated thoracic aorta from which the endothelium was removed. NH4Cl (10-80 mM) increased both Ca2+]i and muscle tone in the presence of external Ca2+. These responses were reproducible. The removal of Ca2+ from the nutrient solution partially inhibited the rise in Ca2+]i and the smooth muscle contraction induced by NH4Cl. In addition, the Ca2+ channel blocker verapamil also partially attenuated the responses to NH4Cl. The NH4Cl-induced responses were gradually reduced as NH4Cl was repeatedly added in a Ca(2+)-free solution. Norepinephrine (NE, 1 microM) induced a transient increase in Ca2+]i and sustained contraction in the absence of external Ca2+, and the subsequent application of NE had little effect on Ca2+]i. After internal Ca2+ stores were depleted by exposure to NE, the subsequent application of NH4Cl induced increases in Ca2+]i and tension of the aorta in a Ca(2+)-free solution. These results suggest that NH4Cl mainly evokes Ca2+ release from the internal Ca2+ stores that are not linked with adrenergic alpha-receptor and causes Ca2+ influx through voltage-dependent Ca2+ channels in the vascular smooth muscle.