Schild plots for (-)-propranolol, timolol and IPS 339, but not penbutolol or carazolol, as antagonists of isoproterenol in the isolated, field-stimulated rat vas deferens exhibited biphasic curves indicating the presence of both high and low affinity sites for these agents in this preparation. Schild plots determined for (-)-propranolol in the presence of prazosin (0.3 micron) or using the prostatic end of the rat vas deferens yielded two similar affinity sites suggesting the lack of involvement of postsynaptic alpha 1-adrenergic mechanisms in the observed response. In the presence of Gpp(NH)p(4 micron), (-)-propranolol, timolol and IPS 339 exhibited only a single affinity site which corresponded to their high affinity site in antagonizing isoproterenol in the absence of Gpp(NH)p. Similarly, only a single high affinity site was observed for (-)-propranolol when salbutamol, rather than isoproterenol, was used as the agonist. The data suggest the two affinity sites observed with some beta-adrenergic blockers in this preparation reflects a possible action of isoproterenol and the antagonists upon two subpopulations of beta 2-adrenergic receptors, one of which is sensitive to Gpp(NH)p.