Calcium channel blocker inhibition of the calmodulin-dependent effects of thyroid hormone and milrinone on rabbit myocardial membrane Ca 2+-ATPase activity

Abstract

The Ca 2+-ATPase activity of rabbit myocardial membranes is stimulated in vitro by l-thyroxine and by milrinone, a bipyridine. These effects are concentration dependent and calmodulin requiring. The calcium channel blockers nifedipine and verapamil have been reported to have anticalmodulin effects in other assay systems. In this study we have examined the effects of nifedipine and verapamil on rabbit myocardial membrane Ca 2+-ATPase activity, in the absence (basal activity) and presence of exogenous l-thyroxine (T4), 10 −10M, and milrinone, 10 −7 M. Basal enzyme activity was inhibited by a minimum of 10 −6 M nifedipine ( ic 50 of 3.4 × 10 −5 M) and 10 −5 M verapamil ( ic 50 of 1.5 × 10 −4 M). Both calcium antagonists inhibited enzyme stimulation by T 4 and milrinone, with halfmaximal inhibition of T 4 and milrinone effects, respectively, at 2.9 × 10 −5 M and 9.0 × 10 −6 M nifedipine and 3.0 × 10 −5 M and 5.2 × 10 −5 M verapamil. The addition of exogenous purified calmodulin, 40 ng/μg, membrane protein, in the presence of 10 −5 M nifedipine or verapamil restored T 4-stimulated enzyme activity. Nifedipine and verapamil, each at a concentration of 10 −6 M, significantly inhibited binding of radioiodinated calmodulin to rabbit heart membranes in vitro. These studies provide evidence that nifedipine and verapamil have an anti-calmodulin effect in this myocardial enzyme system. Through interaction with calmodulin, the channel blockers inhibit thyroid hormone and milrinone stimulation of myocardial membrane Ca 2+-ATPase.