Subtypes of alpha-1 adrenergic receptors on rat parotid gland acinar cell membranes were characterized using subtype selective alpha adrenergic receptor antagonists. The alpha-1 adrenergic receptor antagonist beta-iodo-[125I]-4-hydroxyphenyl-ethyl-aminomethyl-tetralone (125IBE) had an equilibrium dissociation constant for specific binding sites on these membranes of 0.241 +/- 0.03 nM and a total number of specific radioligand binding sites of 41 +/- 4 fmol bound/mg of protein. Displacement of 125IBE binding by subtype-selective alpha-1 adrenergic receptor antagonists 2-(2,6-dimethoxyphenoxyethyl)-aminomethyl-1,4-benzodioxane HCl (WB4101) and 5-methylurapidil fit best to biphasic competition curves. The high- and low-affinity inhibition equilibrium dissociation constant for WB4101 were 0.45 +/- 0.1 and 27 +/- 6 nM, respectively. Similarly, the high- and low-affinity inhibition equilibrium dissociation constants for 5-methylurapidil were 0.16 +/- 0.03 and 71 +/- 20 nM, respectively. These affinities for 125IBE binding sites suggest the presence of alpha-1A and alpha-1B adrenergic receptor subtypes on acinar cell membranes. The irreversible alpha-1 adrenergic receptor antagonist chloroethylclonidine was used to inactivate alpha-1B adrenergic receptors on acinar cell membranes. After treatment with chloroethylclonidine, saturation binding analysis demonstrated no change in the total number of 125IBE binding sites. In addition, competition curves for WB4101 and 5-methylurapidil again showed two sites of 125IBE displacement, with no change in antagonist affinities in membranes treated with chloroethylclonidine.(ABSTRACT TRUNCATED AT 250 WORDS)