Myoplasmic calcium, myosin phosphorylation, and regulation of the crossbridge cycle in swine arterial smooth muscle.

Abstract

Our objective was to test the hypothesis that changes in crossbridge phosphorylation in the swine carotid media are due to changes in the myoplasmic calcium concentration. The photoprotein aequorin was loaded intracellularly by incubation in a series of calcium-free solutions. This loading procedure did not affect subsequent stress development, myosin light chain phosphorylation, or ultrastructure. The time course of light production, myosin light chain phosphorylation, shortening velocity at zero load, and active stress were measured in three stimulus protocols: depolarization with 109 mM potassium chloride at 22 degrees C, 37 degrees C, and 37 degrees C, followed by a reduction in potassium chloride to 20 mM to induce stress maintenance with basal phosphorylation (latch). Light-predicted intracellular calcium concentration was found to correlate with myosin phosphorylation and unloaded shortening velocity. The calcium concentration required for half-maximal myosin phosphorylation was approximately twice that for stress maintenance. These estimates depend on many assumptions, but they compared favorably with the half-maximal myosin phosphorylation values obtained for the calcium-dependence of stress maintenance and phosphorylation in Triton X-100 skinned carotid media preparations. This supports the hypothesis that myoplasmic calcium is the determinant of myosin phosphorylation and mean crossbridge cycling rates in intact smooth muscle depolarized by potassium chloride.