Guanine nucleotide and cation regulation of radioligand binding to R i adenosine receptors of rat fat cells

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Abstract
The modulation of radioligand binding at R i adenosine receptors of rat fat cells by guanine nucleotides and cations was investigated. Guanine nucleotides (in the order of potency: GTP=GDP>Gpp(NH)p>5′-GMP) decreased the binding of the R i receptor agonist (−)N6-phenylisopropyl[3H]adenosine ([3H]PIA), but did not affect binding of the antagonist 1,3-diethyl-8[3H]phenylxanthine ([3H]DPX). Saturation of [3H]PIA binding revealed that GTP (100 μmol/l) converts the high affinity form of the R i receptor into a low affinity form. This effect was confirmed in kinetic experiments. GTP decreased the potency of agonists in competing for [3H]DPX binding, as shown by a 50-fold shift of the K i-value for (−)PIA, whereas antagonist-induced inhibition of binding remained unchanged. The divalent cations Mg2+ and Ca2+ produced a slight increase in [3H]PIA binding but did not affect [3H]DPX binding. Mn2+ markedly decreased both agonist and antagonist binding at R i adenosine receptors. Divalent cations reversed the guanine nucleotide-induced decrease of affinity of the R i receptor. Na+ did not significantly affect agonist or antagonist binding but abolished the stimulatory effect of Mg2+ on agonist binding in the presence of GTP. Our data indicate that guanine nucleotides convert the R i adenosine receptor of rat fat cells from a high to a low agonist affinity state and that the modulation of radioligand binding by mono-and divalent cations differs from that of R i receptors of other tissues.