We have previously reported that the positive inotropic response to β-adrenoceptor stimulation was markedly diminished in diabetic rat papillary muscles. This finding prompted us to characterize the properties of cardiac β-adrenoceptrors in diabetes by means of radioligand binding with ICYP. Male Wistar rats received 45 mg/kg streptozotocin to induce diabetes , and after a lapse of 4 weeks ventricular membrane fractions were prepared. Binding of ICYP to control and diabetic membranes was a saturable process of high affinity with very low non-specific binding. Scatchard analysis resulted in linear plots for both groups (K_D ：60-100 pM), indicating interaction with a single class of binding sites. The maximal number of ICYP binding sites, however, was about 40% less in diabetic group. The isoproterenol (ISO) displacement curve for ICYP showed two classes binding sites that have high and low affinity states for ISO. The proportion and dissociation constant of high affinity states were not different between both groups. GppNHp converted the heterogenous binding into a homogenous one of low affinity. If β-adrenoceptors expressing high, GppNHp-sensitive affinity for ISO have functionally an important role, their decrease under diabetes may explain in part the diminished inotropic response via β-adrenoceptors.