Binding of [3H]angiotensin II and [3H]DuP 753 (Losartan) to rat liver homogenates reveals multiple sites. Relationship to AT1a- and AT1b-type angiotensin receptors and novel nonangiotensin binding sites.

The binding characteristics of radiolabeled angiotensin II and the nonpeptidergic angiotensin AT1 receptor antagonist, DuP 753 (Losartan), were studied in rat liver homogenates. Competition experiments with human angiotensin I, II, and III and with the angiotensin antagonists, CGP 42114A, saralasin, DuP 753, and PD123177, confirmed that the [3H]angiotensin II binding was to an AT1-type receptor. Computer analysis of the competition studies using the human angiotensins demonstrated that the data could be best fitted to a model that considers interaction at two sites. Angiotensin II, angiotensin III, and an angiotensin II analogue, [Sar1]angiotensin II, were calculated to have approximately one hundredfold selectivity at each of the two binding sites, but angiotensin I and the antagonists did not show a difference in affinity between the two sites. The addition of 120 mM NaCl and the nonhydrolyzable analogue of GTP, GppNHp (100 microM) to the buffer resulted in a reduction in [3H]angiotensin II binding at both sites. Thus, we suggest that the two sites may represent distinct angiotensin AT1-type receptors, possibly AT1a and AT1b subtypes. The addition of dithiothreitol (DTT) reduced [3H]angiotensin II binding, confirming the binding to AT1-type receptors. Binding studies using the selective AT1 angiotensin II receptor antagonist, [3H]DuP 753, were also performed on the rat liver homogenates. Saturation studies using both angiotensin II and DuP 753 to define nonspecific binding showed that [3H]DuP 753 bound to at least two types of site, one smaller population of receptors that was sensitive to both angiotensin II and DuP 753 and a second site that was sensitive to DuP 753 only.(ABSTRACT TRUNCATED AT 250 WORDS)