Kainate receptors coupled to G(i)/G(o) proteins in the rat hippocampus

Kainate receptors are a subtype of ionotropic glutamate receptors, permeable to cations and thus expected to have an excitatory depolarizing action on neurons. However, kainate receptor activation inhibits gamma-aminobutyric acid release in the hippocampus through activation of protein kinase C in a pertussis toxin-dependent manner, suggesting a coupling of kainate receptors to G proteins. Thus, we directly investigated the G protein coupling of kainate receptors in the rat hippocampus by using a selective kainate receptor agonist, [(3)H](2S,4R)-4-methylglutamate ([(3)H]MGA). [(3)H]MGA bound to a single site to hippocampal membranes with a K(D) value of 32 nM and a B(max) value of 1024 fmol/mg protein. This binding likely represents kainate receptors because it was displaced by domoate (K(i) = 4 nM), kainate (K(i) = 11 nM), and 6-cyano-7-nitroquinoxaline-2,3-dione (K(i) = 1.4 microM), but not by alpha-amino-3-hydroxy-5-methyl-4-isoxazole-propionate (K(i) > 10 microM), (RS)-alpha-methyl-4-phosphonophenylglycine (K(i) > 10 microM), or (+/-)-1-aminocyclopentane-trans-1,3-dicarboxylic acid (K(i) > 10 microM). Guanylylimidodiphosphate (30 microM), which uncouples all G protein-coupled receptors, shifted to the right the saturation curve of [(3)H]MGA (K(D) = 133 nM). This effect was mimicked by pretreatment of hippocampal membranes with modifiers of G(i)/G(o) proteins [30 microM N-ethylmaleimide (K(D) = 98 nM) or 25 microgram/ml pertussis toxin (K(D) = 95 nM)] but not by a modifier of G(s) proteins [50 microgram/ml cholera toxin (K(D) = 32 nM)]. Treatment of solubilized hippocampal membranes with pertussis toxin (25 microgram/ml) decreased [(3)H]MGA affinity (K(D) = 105-113 nM), which was recovered by reconstitution of these pretreated solubilized hippocampal membranes with G(i)/G(o) proteins (K(D) = 41-76 nM). These results indicate that hippocampal kainate receptors are coupled to G(i)/G(o) proteins.