Glomeruli and microvessels of the rabbit kidney contain both A₁- and A₂-adenosine receptors

Summary

Rabbit renal cortices were fractionated by collagenase dispersion and glomeruli, microvessels and tubuli purified on a discontinuous sucrose gradient. Binding experiments with (−)[¹²⁵I]N⁶-(4-hydroxyphenylisopropyl)-adenosine ([¹²⁵I]HPIA) provided evidence for the presence of A₁-adenosine receptors in the glomerular and microvascular fraction. With glomeruli, saturation isotherms for specific [¹²⁵I]HPIA binding were mono-phasic with a K _D of 1.3 nmol/l and a B _maxof 7.7 fmol/mg protein. In kinetic experiments, an association rate constant of 4.9 × 105 (mol/ 1)⁻¹ s⁻¹ and a dissociation rate constant of 4.3 × 10⁻⁴ s⁻¹ were obtained, yielding a K _D of 0.9 nmol/l. Adenosine analogs displaced [¹²⁵I]HPIA binding with a rank order of potency typical of A₁-adenosine receptors; furthermore, binding was inhibited by methylxanthines and modulated by GTP. Saturation experiments with the microvessels revealed a K _D of 1.9 nmol/l and a B _max of 13.4 fmol/mg protein. However, no inhibition of glomerular and microvascular adenylate cyclase activity could be demonstrated, but instead both 5′-N-ethylcarboxamido-adenosine (NECA) and N⁶-(R-phenylisopropyl)-adenosine (R-PIA) stimulated enzyme activity, with EC₅₀ values of 0.14 μmol/l and 1.5 μmol/l, respectively. The concentration-response curve for NECA was shifted to the right (factor 9) by 10 μmol/l 8-phenyltheophylline. On the other hand, computer simulation of biphasic curves (adenylate cyclase inhibition in the presence of activation via a stimulatory receptor) indicates that the failure to observe an A₁-adenosine receptor-mediated inhibition of adenylate cyclase activity in the presence of stimulatory adenosine receptors may be attributable to methodological constraints. The results demonstrate that both A₁- and A₂-adenosine receptors are present in rabbit glomeruli and microvessels. It is suggested that both receptors are involved in the control of renin secretion.