Glutamate receptor activation in cultured cerebellar granule cells increases cytosolic free Ca2+ by mobilization of cellular Ca2+ and activation of Ca2+ influx.
The Ca2+ sensitive fluorescent probe, fura-2 has been used to monitor cytosolic free calcium levels in mature primary cultures of cerebellar granule cells during exposure to L-glutamate and other excitatory amino acids: quisqualate (QA) kainate (KA) and N-methyl-D-aspartate (NMDA). Glutamate at micromolar concentrations produced a prompt and dose-related increase in the intracellular concentration of free Ca2+, ([Ca2+]i), whereas QA, KA and NMDA had no effect. This increase was also seen in the absence of extracellular Ca2+, suggesting that L-glutamate promotes mobilization of Ca2+ from intracellular stores. In the presence of extracellular calcium, the elevation of [Ca2+]i was, in part, mediated by an increase in the plasma membrane permeability to Ca2+. This Ca2+ influx was not affected by the Ca2+-channel antagonist l-Verapamil. However, L-Verapamil did block the increase in [Ca2+]i seen after depolarization of the cells with potassium. The Ca2+ response elicited by glutamate was partially blocked by the excitatory amino acid antagonist glutamate diethyl ester (GDEE). Furthermore, glutamate stimulated the formation of inositol mono-, bis-, tris- and tetrakisphosphates (IP1, IP2, IP3, and IP4) suggesting a role for these compounds for the increase in [Ca2+]i.
Bouchelouche
P
P
Department of Clinical Chemistry, Herlev Hospital, University of Copenhagen, Denmark.
Belhage
B
B
Frandsen
A
A
Drejer
J
J
Schousboe
A
A
D000818:Animals; D001224:Aspartic Acid / Q000031:analogs & derivatives; D001572:Benzofurans; D002118:Calcium / Q000493:pharmacokinetics* / Q000502:physiology; D002121:Calcium Channel Blockers / Q000494:pharmacology; D002478:Cells, Cultured; D002531:Cerebellum / Q000187:drug effects / Q000378:metabolism*; D003600:Cytosol / Q000378:metabolism*; D005456:Fluorescent Dyes; D016257:Fura-2; D005971:Glutamates / Q000494:pharmacology; D018698:Glutamic Acid; D007295:Inositol Phosphates / Q000378:metabolism; D007608:Kainic Acid / Q000494:pharmacology; D051379:Mice; D016202:N-Methylaspartate; D017470:Receptors, Glutamate; D017981:Receptors, Neurotransmitter / Q000187:drug effects / Q000502:physiology*; D013050:Spectrometry, Fluorescence
D001572:Benzofurans; D002121:Calcium Channel Blockers; D005456:Fluorescent Dyes; D005971:Glutamates; D007295:Inositol Phosphates; D017470:Receptors, Glutamate; D017981:Receptors, Neurotransmitter; D001224:Aspartic Acid; D018698:Glutamic Acid; D016202:N-Methylaspartate; D007608:Kainic Acid; D002118:Calcium; D016257:Fura-2