The effects of vanadate on rabbit ventricular muscle adenylate cyclase and sodium pump activities

Vanadate in the +5 oxidation state has been reported to have a positive inotropic action on cardiac ventricular muscle. We have investigated the biochemical actions of vanadate on ventricular muscle adenylate cyclase and sodium pump activities in both intact or disrupted cell systems in an attempt to elucidate the mechanism(s) responsible for the physiological response. Vanadate at concentrations up to 100 microM (Ka = 2 microM) stimulated adenylate cyclase activity in sarcolemmal membrane preparations or disrupted myocytes isolated from rabbit ventricular muscle by 2-3-fold. Increasing the vanadate concentrations above 100 microM resulted in a progressive inhibition of basal or hormone-stimulated adenylate cyclase activity (Ki = 5 mM) which was similar to that found by the reaction product, pyrophosphate (Ki = 0.5 mM). Both activation and inhibition by vanadate was fully reversible. Maximum activation of adenylate cyclase by vanadate and isoprenaline were not additive whereas maximum fluoride activation was decreased (18%) and the forskolin-stimulated response was slightly potentiated. Vanadate reversibly inhibited ouabain-sensitive p-nitrophenylphosphatase activity (Ki = 60 nM) in sarcolemmal membrane preparations and disrupted myocytes. Complete inactivation was found at 1 microM vanadate. Acute or chronic incubation of intact myocytes with vanadate at concentrations up to 0.5 mM had no measurable affect on ouabain-sensitive 86Rb influx or isobutylmethylxanthine, isoprenaline or forskolin-stimulated accumulation of intracellular cAMP concentration. Inhibition of 86Rb influx and cAMP accumulation was found at higher concentrations of vanadate; however, this accompanied the progressive decrease in cell viability as measured by the decrease in percentage of rod-shaped cells. It is concluded that vanadate, at concentrations which have been reported to induce a positive inotropic action on mammalian ventricular muscle, does not increase adenylate cyclase activity or inhibit the sodium pump activity in intact myocytes. These results show that caution must be applied when extrapolating the actions found with vanadate in broken cell systems to intact tissues.