OBJECTIVE The subtype specificity, localization and distribution of urethral alpha1-adrenoceptors were studied in the male rabbit urethra. METHODS The properties of the urethral alpha1-adrenoceptors were investigated using radioligand receptor binding and light microscopic autoradiography with [125I]iodo-2-[b-(4-hydroxyphenyl)-ethylaminomethyl]tetralone (HEAT), and immunohistochemistry with monoclonal anti-alpha smooth muscle actin and anti-alpha sarcomeric actin antibodies. RESULTS Saturation experiments with [125I]HEAT demonstrated the presence of significant amounts of a single high affinity binding site for alpha1 adrenoceptors in the male rabbit urethra. The pharmacological profile of the alpha1 adrenoceptors in rabbit urethra, determined by inhibition experiments with subtype selective alpha1 adrenoceptor antagonists, was characterized by the following rank order of potency of inhibition constants (Ki values): prazosin < or = WB 4101 < spiperone < 5-methylurapidil < BMY 7378. The pKi values for the rabbit urethra were correlated with the pKi values for rat spleen, submaxillary glands, and vas deferens and for those reported for cloned alpha1d receptors with correlation coefficients of 0.68, 0.929, 0.909, and 0.523, respectively. CONCLUSIONS The pharmacological characterization demonstrates the predominance of alpha1A or alpha1A + alpha1B adrenoceptor subtype(s) in male rabbit urethral smooth muscle. Furthermore, the autoradiographic and immunohistochemical studies show a heterogeneous distribution of alpha1 adrenoceptors along the longitudinal axis of the urethra, within the smooth muscle fibers, with the receptors being localized more densely in the proximal than in the distal urethra.