(+)-Niguldipine inhibited specific 125I-BE 2254 binding more potently in membrane preparations from rat tissues enriched in the alpha 1A subtype (hippocampus and vas deferens) than those with the alpha 1B subtype (liver and spleen). Inhibition curves for (+)-niguldipine were better fit by a two-site model in most tissues, although Kl values for each site varied markedly between tissues. The potency of this lipophilic drug was highly dependent on tissue concentration, probably accounting for most of this variability. Pretreatment of membranes with chloroethylclonidine (CEC) to inactivate the alpha 1B subtype did not completely eliminate the low affinity sites for (+)-niguldipine, particularly in heart. Saturation analysis showed that (+)-niguldipine competitively inhibited both alpha 1A and alpha 1B subtypes. However, substantial non-competitive inhibition was also observed in several tissues. Analysis of inhibition curves for 5-methylurapidil gave similar proportions of alpha 1A and alpha 1B receptor sites as were calculated for (+)-niguldipine in various tissues. Although (+)-niguldipine and 5-methylurapidil revealed variable proportions of low affinity sites in CEC-pretreated hippocampus and heart, this was not observed with inhibition curves for WB 4101 and phentolamine. These results are generally consistent with the previously defined alpha 1A and alpha 1B subtypes. 5-Methylurapidil currently appears to be the best antagonist for discriminating these subtypes; (+)-niguldipine shows similar selectivity but is complicated by a high lipophilicity. However, the persistence of low affinity sites for 5-methylurapidil and (+)-niguldipine after CEC pretreatment and the noncompetitive effects of (+)-niguldipine in some tissues raise the possibility of an additional subtype(s) of alpha 1-adrenergic receptors in rat tissues.