Autoradiography of receptor-activated G-proteins in post mortem human brain

The agonist-stimulated guanosine 5'-(gamma-[S-35]thio)triphosphate binding assay was used to anatomically localize receptor-activated G-proteins by au toradiography in post mortem human brain. The optimal conditions for guanos ine 5'-(gamma-S-35]thio)triphosphate binding to human brain sections were e stablished in post mortem samples of the prefrontal cortex, hippocampus, ba sal ganglia, brainstem and cerebellar cortex. An excess of GDP (2 mM) was r equired to decrease basal activity and obtain effective stimulation by spec ific agonists. guanosine 5'-(gamma-[S-35]Thio)triphosphate binding was incr eased after stimulation with specific agonists of different G-protein-coupl ed receptors. They include cannabinoid (WIN55212-2), mu-opioid ([D-Ala(2)N- MePh4,Gly(5)-ol]enkephalin), serotonin-1A [(+/-)-8-hydroxy-2-(di-n-propylam ino)tetralin and serotonin-1B/1D (sumatriptan), cholinergic muscarinic rece ptors (carbachol) and alpha(2)-adrenoceptors (UK14304). Such stimulation re ached 1458%, 440%, 188%, 219%, 61% and 339%, respectively, over the basal l evels. In tissue sections, the use of the above-mentioned agonists (10(-4) M) showed patterns of anatomical distribution similar to those already desc ribed by receptor autoradiography, with high densities over the hippocampus (serotonin-1A receptors), cortex (alpha(2)-adrenoceptors) and striatum (mu -opioid receptors). The highest binding levels were reached with the cannab inoid receptor agonist in most of the analysed brain regions. Carbachol pro duced only moderate stimulation of those same regions. The blockage of agon ist-stimulated guanosine 5'-(mu-[S-35]thio)triphosphate binding by selectiv e antagonists verified that the effect was receptor mediated. This technique provides a method to identify modifications of the receptor- mediated activation of G-proteins in post mortem human brain with anatomica l resolution. It also provides valuable information on the level of drug ef ficacy in the human species. (C) 2000 IBRO. Published by Elsevier Science L td.