Comparison of the apparent irreversible beta-adrenoceptor antagonist Ro 03-7894 with propranolol in cardiac ventricular muscle by pharmacological and radioligand binding techniques.

Membrane fractions were prepared from guinea-pig ventricular muscle and the specific binding of [3H]dihydroalprenolol ([3H]DHA) assessed. The dissociation constant (Ki) of (+/-)-propranolol was determined (6.9 nM) from its ability to displace [3H]DHA binding. This compared with the pA2 value of propranolol of 8.32 (dissociation constant, 4.8 nM) determined for the antagonism of isoprenaline-induced positive inotropic responses of papillary muscles from guinea-pig hearts. Scatchard analysis of the saturation curves for specific [3H]DHA binding showed that in the presence of propranolol, the displacement was characteristic of competitive antagonism. That is, there was no change in the total beta-adrenoceptor was characteristic of competitive antagonism. That is, there was no change in the total beta-adrenoceptor binding sites (Bmax) but an apparent reduction of the dissociation constant (KD) of [3H]DHA. This antagonism was fully reversed by washing membranes that had been previously incubated with propranolol. In contrast, in the presence of the beta-adrenoceptor antagonist, Ro 03-7894, the Scatchard plots were displaced in a manner characteristic of irreversible antagonism. The Bmax was significantly reduced. This antagonism was resistant to washout, with the Scatchard plots still showing a reduced Bmax and no change in the dissociation constant (KD) of [3H]DHA. This apparent irreversible antagonism by Ro 03-7894 was also demonstrated in guinea-pig isolated papillary muscles. The maximum of the dose-response curve to isoprenaline, constructed after incubation with Ro 03-7894 and a 3 hr bath-washout, was depressed by 89.5 +/- 7.5%