Regional differences of the inhibition of GABAB ligand binding by the GTP analogue Gpp(NH)p.

In order to determine whether the interactions between GABAB receptors and G-proteins differ in several brain areas, we have used the reduction in high-affinity GABAB binding by the GTP analogue Gpp(NH)p as an internal assay marker for G-protein linkage to GABAB receptors. The results indicate that Gpp(NH)p inhibits the binding of the GABAB receptor agonist [3H]CGP 27492 (80 to 95%) in a biphasic manner between 0.1 nM and 1 mM. The IC50 for high-affinity sites is significantly higher in cerebellum (70 nM, 53% of binding sites) than in cortex, hippocampus, corpus striatum and thalamus (15–30 nM, 63–73% of binding sites). The IC50s of the low-affinity sites in hippocampus and cortex (170 μM and 210 μM, respectively) were significantly higher than the IC50s in cerebellum, thalamus and corpus striatum (18–39 μM). All these binding sites are sensitive to pertussis toxin (PTX; 7–15 μg/mg protein), implicating that they are linked either to G1 or to G0 proteins. The two binding sites observed (high affinity, nM and low affinity, μM for Gpp(NH)p) and the regional dependence in affinity of these sites may originate either from different GABAB receptor subtypes, different G-proteins or different coupling mechanisms between G-proteins and GABAB receptors. Whereas the PTX site of G-protein linked to GABAB receptors changes with age [24], the GTP binding site does not differ between peripubertal rats (5–6 weeks) and adults rats (10–12 weeks).