Fluorescence measurement of changes in intracellular calcium induced by excitatory amino acids in cultured cortical astrocytes

Abstract

Population response of [Ca2+], in cultured cortical astrocytes to excitatory amino acids was measured at room temperature using the calcium-sensitive dye fura-2. Quisqualic acid (QA), glutamate (Glu), and kainic acid (KA) caused a peak increase in [Ca2+], in the order QA > Glu > KA. No response to N-methyl-D-aspartic acid (NMDA) was observed whether or not Mg2+ was present externally. Both QA and Glu (100 μM) frequently elicited a decaying oscillatory [Ca2+], response during sustained agonist application; the period of oscillations initially was 23.5 sec and increased as the response was damped. Comparatively, the [Ca2+], response to KA was nonoscillatory. Both responses to Glu and KA were reduced slightly by antagonist 7-D-glutamylaminomethyl-sulfonic acid (1 mM), but virtually were abolished by kynurenic acid (3 mM). Replacement of external Na+ by choline had no significant effect on the Glu response. Removal of external Ca2+ reduced the peak response to QA, Glu, and KA to 40, 34, and 18%, respectively; and markedly reduced the degree of QA- and Glu-induced [Ca2+], oscillations. Pretreatment with phorbol esters, a potent activator of protein kinase C, blocked the [Ca2+], response to Glu but not KA. It is concluded that cortical astrocytes express Glu receptors of the non-NMDA type in culture and that receptor activation leads to Ca2+ influx and release of internal Ca2+. Mobilization of Ca2+ apparently occurs via the known Glu-mediated hydrolysis of inositol lipids, which may come under negative-feedback control by protein kinase C activation. Oscillatory [Ca2+] signaling offers the possibility of a dynamic population response in an electrically coupled glial network.