Stimulation of canine cardiac sarcoplasmic reticulum Ca 2+ uptake by dihydropyridine Ca 2+ antagonists

Abstract

We examined the effects of four Ca 2+ antagonists that possess the ability to bind to calmodulin—felodipine, nitrendipine, prenylamine, and verapamil—as well as the effect of the calmodulin antagonist trifluoperazine on Ca 2+ uptake and Ca 2+ + Mg 2+/ATPase activity in canine cardiac sarcoplasmic reticulum. In the presence of 20–30 μM felodipine and 100–200 μM nitrendipine, Ca 2+ uptake increased from 69 nmoles · mg −1 · min −1 to 107 and 108 nmoles · mg −1 · min −1, respectively, with half-maximal stimulation occurring at 7.5 and 28 μM respectively. Ca 2+ + Mg 2+/ATPase activity was unchanged over the same concentration ranges. In contrast, both Ca 2+ uptake and Ca 2+ + Mg 2+/ ATPase activities were inhibited in the presence of 10–100 μM trifluoperazine ( ic 50 = 25 μ M) , 10–100 μM prenylamine ( ic 50 = 35 μ M) and 100–200 μM verapamil (inhibition insufficient for ic 50 determination). None of the drugs affected membrane permeability to Ca 2+ as determined by passive 45Ca 2+ efflux in the presence of ethyleneglycol bis (β-amenoethyl ether) N, N, N 1-tetraacetic acid (EGTA). Drug inhibition of calmodulin-dependent turkey gizzard myosin light chain kinase activation in a purified protein system was used as a direct measure of calmodulin antagonism, and felodipine, nitrendipine, trifluoperazine, prenylamine, and verapamil blocked this activation at ic 50 values of 9.8, 55, 6.4,31, and 93 μM respectively. None of the drugs studied, however, had any effect upon endogenous phospholamban phosphorylation in our cardiac sarcoplasmic reticulum preparations. These observations indicate that di-hydropyridine Ca 2+ antagonists stimulate cardiac sarcoplasmic reticulum Ca 2+ uptake in vitro either by increasing the efficiency of the transport process or by inhibiting Ca 2+-dependent Ca 2+ release, and suggest that these effects do not result from interference with calmodulin-mediated processes.