Pertussis toxin inhibits signal transduction at a specific metabolotropic glutamate receptor in primary cultures of cerebellar granule cells

In primary cultures of cerebellar granule cells, glutamate receptors have been classified into metabolotroplc (G_P1 and G_P2) and ionotroplc (G_C1 and G_C2). The G_P1 and G_C1 receptors are negatively modulated by magnesium and noncompetitively inhibited by phencyclidine; G_P2 and G_C2 receptors are insensitive to inhibition by magnesium and phencyclidine (Costa, Fadda, Kozikowski, Nicoletti and Wroblewski, 1988). Exposure of cultured cerebellar granule cells to pertussis toxin (PTX, 1 μg/ml for 14-16 hr) reduced the stimulation of the hydrolysis of inositol phospholipids (PI) by the G_P2 receptor agonists, glutamate and quisqualate in the presence of magnesium, but did not inhibit the stimulation of the hydrolysis of PI by G_P1, receptor agonists. The stimulation of the hydrolysis of PI by the muscarinic cholinergic receptor agonist, carbamylcholine, remained unchanged after pretreatment with pertussis toxin. In membranes prepared from cerebellar granule cells in primary culture, the addition of guanosine 5′-0-(3-thiotriphosphate) (GTP-γ-s), a nonhydrolyzable analogue of GTP, enhanced the hydrolysis of PI and reduced the B_max of quisqualate-displaceable binding of [³H]glutamate. These results indicate that, in primary cultures of cerebellar granule cells, a specific class of metabolotroplc glutamate receptors (the G_P2 receptor) is coupled with the hydrolysis of PI through a pertussis toxin-sensitive GTP-binding protein.