STIMULATION OF CA2-ACTIVATED NONSPECIFIC CATIONIC CHANNELS BY PHOSPHOLIPASE C-LINKED GLUTAMATE RECEPTORS IN SYNAPTONEUROSOMES()

The regulation of intracellular Ca2+ concentration ([Ca2+](i)) by glut amate metabotropic receptors (mGluR) was studied in 8-day-old rat fore brain synaptoneurosomes using spectrofluorimetric methods. Here we dem onstrate that metabotropic glutamate agonists induce in rat brain syna ptoneurosomes a Ca2+ influx largely dependent upon the presence of Ca2 + in the external medium. The pharmacological profile of this influx i s strongly correlated with the pharmacological profile of the activati on of phosphoinositide hydrolysis, i.e. quisqualic acid >> 1S,3R-amino -1-dicarboxylate-1,3 cyclopentane congruent to glutamate. This metabot ropic glutamate receptor-induced Ca2+ influx is insensitive to voltage -dependent Ca2+ channel antagonists and occurs through a Mn2+ impermea nt pathway. The study of the rapid kinetics shows that this influx is triggered after a 300 ms delay compared with that elicited by depolari zing agents and Ca2+ ionophore A23187. In order to assess further if m GluR stimulate this influx through the recruitment of inositol triphos phate (IP3)-sensitive intracellular Ca2+ stores, we have tested the ef fect of thapsigargin on membrane potential and intracellular Ca2+ simu ltaneously. Thapsigargin induces a depolarization of the synaptoneuros omal membrane followed by a massive Ca2+ influx, occurring via a Mn2nonpermeant route. This depolarizing effect is sensitive to the presen ce of the intracellular Ca2+ chelator 2-aminophenoxy)ethane-N,N,N',N'- tetraacetoxymethyl ester], and partially sensitive to extracellular Na +, but insensitive to the presence of extracellular Ca2+. Taken togeth er, our data suggest that mGluR stimulate self-maintained increases of [Ca2+](i) in rat forebrain synaptoneurosomes via the activation of a multistep mechanism, sequenced in the following steps: (i) mGluR-induc ed IP3 synthesis; (ii) IP3-stimulated intracellular Ca2+ release; (iii ) Ca2+-activated non-specific cation channel, leading to local depolar ization and a Ca2+ influx; and (iv) activation of Ca2+-sensitive phosp holipase C.