The effects of chlorethylclonidine, WB 4101 and nifedipine on norepinephrine-induced contractions of rat, guinea-pig, rabbit and dog aortae were investigated in order to characterize the α1-adrenoceptor subtype(s) present in the aortae of these different species. The putative α1A-adrenoceptor antagonist, WB 4101, was significantly more potent in the rat aorta compared to the rabbit, guinea-pig and dog aortae which were not significantly different from each other. The calcium channel antagonist, nifedipine (1 µM), had little or no effect on norepinephrine-induced contractions in aortic segments from the rabbit, guinea pig and dog; whereas in the rat aorta, nifedipine significantly inhibited the response to norepinephrine. Based on the studies with WB 4101 and nifedipine, α1-adrenoceptors in rat aorta would be tentatively classified as α1A-adrenoceptors, whereas those in the guinea-pig, rabbit and dog aortae would be of the α1B-adrenoceptor subtype. The putative irreversible α1B-adrenoceptor antagonist, chlorethylclonidine, inhibited the response to norepinephrine in aortae from all species, but to dramatically different degrees. The response to norepinephrine was inhibited by 500-fold and 450-fold by chlorethylclonidine in the rat and dog aortae, respectively, whereas in the guinea-pig and rabbit aortae, the potency of norepinephrine was reduced by only 3- and 20-fold, respectively. Thus, based on studies with chlorethylclonidine, α1-adreno-ceptors in the rat and dog aortae would be classified as α1B-adrenoceptors (i.e., chlorethyl-clonidine-sensitive), whereas α1A-adrenoceptors (chlorethylclonidine-insensitive) would predominate in the guinea-pig aorta, and possibly both α1A- and α1B-adrenoceptors would coexist in the rabbit aorta. It is apparent that the subclassification of α1-adrenoceptors based on chlorethylclonidine does not agree with the subclassification based on WB 4101 and nifedipine in these tissues. Specifically, studies with WB 4101 and nifedipine indicate that the rat aorta contains primarily α1A-adrenoceptors, whereas the high sensitivity to chlorethylclonidine in the rat aorta would indicate the existence of α1B-adrenoceptors. Similar discrepancies exist in the aortae from the other species. The results indicate clearly that significant heterogeneity exists in α1-adrenoceptors in mammalian aortae. Furthermore, our findings suggest that chlorethylclonidine and WB 4101 are not sufficiently reliable tools to use in subclassifying α1-adrenoceptors in vascular smooth muscle, and the results obtained in the present study do not allow α1-adrenoceptors in mammalian aortae to be subclassified reliably as either α1A-or α1B<