MODULATION OF ADENOSINE AGONIST [H-3] N6-(R)-PHENYLISOPROPYLADENOSINE BINDING TO PIG BRAIN CORTICAL MEMBRANES BY CHANGES OF MEMBRANE FLUIDITY AND OF MEDIUM PHYSICOCHEMICAL CHARACTERISTICS

The binding of [H-3]N6-(R)-phenylisopropyladenosine ([H-3]R-PIA) to pig brain cortical membranes was investigated as a function of membrane fluidity and of the physicochemical characteristics of the medium. The two affinity states of the A1 adenosine receptor behave differently in the experiments performed. Increases (up to 6.7 poises) or reduction (up to 1.3 poises) in membrane microviscosity did not lead to significant variation of equilibrium parameter values except for a slight modification of the low-affinity K(d) value at 1.3 poises. Addition of cholesterol to raise the microviscosity up to 9.0 poises led to a marked decrease of binding to the high-affinity state without modifying the K(d). Furthermore, the K(d) value for the low-affinity state rose markedly. Addition of sucrose, which modifies the conductance, viscosity and density of the aqueous medium, did not lead to significant changes when used at a concentration of 0.25 M, either in isolation or in radioligand binding assay medium. The presence of 0.32 M sucrose in the isolation medium led to a 30% fall in the total binding without affecting the distribution and K(d) values for either affinity state very much. However, the presence of greater-than-or-equal-to 0.32 M sucrose in the binding assay medium produced the disappearance of the low-affinity states and the appearance of high-affinity states, only one affinity state was found to have a somewhat increased K(d) value. On the other hand, increases of the medium conductance did not lead to the disappearance of the low-affinity state although some decrease of the binding was observed at high Tris concentrations. These results strongly suggest a modulation of the agonist binding to A1 adenosine receptors by changes in membrane fluidity and the physicochemical characteristics of the medium.