Group I mGluRs coupled to G proteins are regulated by tyrosine kinase in dopamine neurons of the rat midbrain

Metabotropic glutamate receptors (mGluRs) modulate neuronal function via di fferent transduction mechanisms that are either dependent or independent on G-protein function. Here we investigated, using whole cell patch-clamp rec ordings in combination with fluorimetric measurements of intracellular calc ium concentration ([Ca2+],), the metabolic pathways involved in the respons es induced by group I mGluRs in dopamine neurons of the rat midbrain. The i nward current and the [Ca2+](i) increase caused by the group I mGluR agonis t (S)-3,5-dihydroxyphenylglycine (DHPG, 100 muM) were permanently activated and subsequently abolished in cells loaded with the nonhydrolizable GTP-an alogue GTP-gamma -S (600 muM). In addition, when GDP-beta -S (600 muM) was dialyzed into the cells to produce the blockade of the G proteins, the DHPG -dependent responses were reduced. When the tissue was bathed with the phos pholipase C inhibitor 1-[6[[(17 beta)-3-methoxyestra-1,3,5(10)-trien-17-yl] amino]exyl]-1H-pyrrole-2,5-dione (10 muM), the DHPG-induced calcium transie nts slightly diminished but the associated inward currents were not affecte d. Interestingly, a substantial depression of the DHPG-induced inward curre nt and transient increase of [Ca2+](i) was caused by the protein tyrosine k inase inhibitors tyrphostin B52 (40 muM) and 4',5,7-trihydroxyisoflavone (g enistein; 40 muM), whereas genistein's inactive analogue 4',5,7-trihydroxyi soflavone-7-glucoside (40 muM) was ineffective. The blockade of the Src fam ily of tyrosine kinase by 4-amino-5-(4-methylphenyl)-7-(t-butyl)pyrazolo [3 ,4-d]pyrimidine (20 muM), mitogen-activated protein kinase by 2'-amino-3' m ethoxyflavone (50 muM), and protein kinase C by staurosporine (1 muM) had n o effect on the cellular responses caused by DHPG. The mGluR5-selective ant agonist 2-methyl-6-(phenylethynyl)-pyridine (10-100 muM) did not affect the actions of DHPG. Thus our results indicate that the responses, mainly medi ated by mGluRs1 in dopamine neurons, are activated by intracellular mechani sms coupled to G proteins and regulated by tyrosine kinases.