Inhibitory effects of verapamil, prenylamine and D 600 on Ca2+-dependent noradrenaline release from the sympathetic nerves of isolated rabbit hearts.

Abstract

In isolated rabbit hearts we investigated the effects of verapamil, D 600 and prenylamine on noradrenaline release from the terminal sympathetic nerves. Noradrenaline in the perfusate was determined spectrofluorimetrically. 1. Perfusion of hearts with Ca2+-free low Na+ (high K+) solution caused an increase in noradrenaline output. Verapamil (1–100 μM) did not affect this Ca2+-independent output evoked by Na+ deprivation. 2. Verapamil inhibited the noradrenaline output evoked by introduction of 1.8 mM CaCl2 after perfusion of hearts with Ca2+-free, K+-rich solution with either a low or a normal Na+ concentration. Double reciprocal plot of noradrenaline output against the Ca2+ concentration used for stimulation (0.9–7.2 mM; α-adrenoceptors and neuronal uptake blocked by phentolamine and cocaine, respectively) yielded a straight line (K m =1.5 mM Ca2+), and the inhibition by verapamil proved to be competitive (K i =8.8 μM verapamil). By contrast, tetracaine and tetrodotoxin did not alter the Ca2+-induced noradrenaline output. 3. In hearts perfused with Tyrode's solution noradrenaline output evoked by increasing the KCl concentration was inhibited by verapamil, D 600 or prenylamine. When the CaCl2 concentration in the Tyrode's solution was decreased, the verapamil concentration-response curve was shifted to lower concentrations. 4. Verapamil also inhibited noradrenaline output evoked by electrical stimulation of the postganglionic sympathetic nerves (IC50: 73.3 μM). 5. Considerably lower concentrations of verapamil, D 600 or prenylamine decreased the noradrenaline output evoked by 72 μM or 180 μM acetylcholine (in the presence of 3.5 μM atropine; IC50: 1.2, 1.1 and 1.0 μM, respectively). The verapamil concentration necessary for the inhibition of acetylcholine-induced output was not affected by alterations of the Ca2+ concentration in the perfusion fluid (0.58–5.8 mM Ca2+). Double reciprocal plot of acetylcholine-induced noradrenaline output against the Ca2+ concentration in the Tyrode's solution (α-adrenoceptors and neuronal noradrenaline uptake blocked) revealed no straight line.