RABBIT RENAL EPITHELIAL ANGIOTENSIN-II RECEPTORS

The characteristics of angiotensin II (ANG II) receptors were compared in basolateral (BLM) and brush-border membranes (BBM) of rabbit kidney cortex. BLM showed high-affinity binding of I-125-labeled [Sar(1)]ANG II [dissociation constant (K-d) = 0.31 +/- 0.04 nM and maximum binding capacity (B-max) = 136 +/- 17 fmol/mg protein]. Losartan inhibited I-125-[Sar(1)]ANG II in a heterogeneous manner, with 73 +/- 4% of the sites showing high affinity [inhibition constant (K-i) = 2.63 +/- 0.82 nM] and 27 +/- 4% of sites having low affinity (K-i = 125 +/- 18 nM). Masking studies confirmed the presence of two different binding sites. The sites with high affinity for losartan resemble the AT(1A) receptor, previously described and cloned. The sites with low affinity resemble the pharmacological AT(1B) subtype, previously identified as the minority subtype in mesangial cells. In BBM, two sites could be distinguished on the basis of I-125-[Sar(1),Ile(8)]ANG II affinity, with K-d values of 1.43 +/- 0.26 and 782 +/- 90 nM for ANG II. The low-affinity site, however, comprised the main population of binding sites. ANG II binding to BBM was not sensitive to losartan. PD-123319 and CGP-42112A competed for binding with a one-half inhibitory concentration (IC50) of similar to 100 and 19.0 +/- 1.0 mu M, respectively. I-125-labeled AT(2)-selective antagonist PD-122979 bound specifically and with high magnitude to both BLM and BBM. A very high rate of ANG II degradation was observed, despite the presence of protease inhibitors, suggesting the possibility of the presence of receptors for ANG II fragments in BLM and BBM. These observations substantiate significant differences in ANG II binding in BBM and BLM consistent with at least three pharmacologically distinct receptor subtypes for angiotensin peptides.