Properties of Angiotensin II Receptors of Isolated Rat Glomeruli: Factors Influencing Binding Affinity and Comparative Binding of Angiotensin Analogs*

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Abstract
Previous studies by other investigators have suggested that angiotensin II binds to three receptors of rat renal glomeruli with different affinities. In contrast, angiotensin II binds to a single, high affinity receptor in rat adrenal glomerulosa and uterine smooth muscle. These studies were designed to evaluate the binding of [125I]angiotensin II to isolated rat glomeruli, to characterize the receptors with the use of angiotensin analogs, and to determine whether glomerular receptors share properties in common with adrenal or uterine angiotensin II receptors. Glomeruli were isolated from the outer 1 mm of rat kidney cortex by a sieving technique and were employed in an angiotensin II radioreceptor assay. Linear Scatchard plots were obtained, indicating the presence of a single class of high affinity binding sites. The equilibrium association constants (Kn) and receptor concentrations were similar for glomeruli (Kn = 0.82 ± 0.05 × 109 M–1; 1127 ± 55 fmol/mg protein; n = 18) and adrenal glomerulosa (Ka = 0.89 ± 0.04 × 109 M–1; 1804 ± 285 fmol/mg protein; n = 6), and both were significantly higher than uterine smooth muscle (Ka = 0.23 ± 0.02 × 109 M–1; 194 ± 46 fmol/mg protein; n = 9). The Kn of glomeruli was significantly influenced by the protein concentration employed in receptor-binding studies. There was an inverse relationship between protein concentration and Kn (r = –0.77; n = 33; P < 0.001). The Kn was independent of glomerular surface area over a wide range (7,000–18,000 μm2). For glomeruli, the order of binding inhibition potencies of angiotensin analogs was: [Sar1,Ile8]angiotensin II > [Sar1.Thr8]-angiotensin II ≫ [Sar1,Thr(Me)8]angiotensin II ≫ angiotensin II ≫ [Sar1,Ala8]angiotensin II = [Sar1,Ser8]angiotensin = des-Asp1[Ile8]angiotensin II s des-Asp1-angiotensin II >>> [Val5,Ser9]angiotensin I. The correlation between dose ratios of binding inhibition potencies of angiotensin analogs for glomeruli and adrenal glomerulosa (r = 0.83) was significant (P ≪ 0.02), whereas that for glomeruli and uterine smooth muscle (r = 0.72) was not (P < 0.05). des-Asp1-angiotensin II (angiotensin III) was significantly less potent in binding inhibition in uterine smooth muscle (21%) than in glomeruli (75%) and adrenal glomerulosa (87%). We conclude that there is a single class of specific angiotensin II receptors on isolated rat glomeruli of a high affinity equal to that noted for adrenal glomerulosa. Glomerular receptors share more properties in common with adrenal glomerulosa than with uterine smooth muscle, suggesting that intrarenal angiotensin II receptors may differ from extrarenal smooth muscle angiotensin II receptors. Differences in glomerular protein concentration were noted to be a major determinant of the Kn. This finding is of practical significance, since artifactual differences in affinity related to receptor concentration could account for differences in affinity when comparing experimental groups. (Endocrinology106: 1923, 1980)