Intracellular survival pathways against glutamate receptor agonist excitotoxicity in cultured neurons. Intracellular calcium responses.
Cultured rat cerebellar granule cells are resistant to the excitotoxic effects of N-methyl-D-aspartate (NMDA) and non-NMDA receptor agonists under three conditions: 1) prior to day seven in vitro when cultured in depolarizing concentrations of potassium [25 mM]; 2) at any time in vitro when cultured in non-depolarizing concentrations of potassium 5 mM[; and 3) when neurons, cultured in depolarizing concentrations of potassium 25 mM[ for eight days in vitro, are pretreated with a subtoxic concentration of NMDA. The focus of this paper is to determine: a) whether the resistance to excitotoxicity by NMDA and non-NMDA receptor agonists is due to a decreased intracellular calcium Ca++[i response to glutamate receptor agonists in cultured rat cerebellar granule cells; or b) whether Ca++[i levels induced by the agonists are similar to those observed under excitotoxic conditions. Granule cells, matured in non-depolarizing growth medium, treated with glutamate resulted in an increase in Ca++[i followed by a plateau that remained above baseline in virtually all neurons that responded to glutamate. The response was rapid in onset (< 10 sec) and the pattern of response heterogeneous in that cells responsive to glutamate increased their Ca++[i to different extents; some cells did not respond to glutamate. Kainate also produced significant elevations in Ca++[i. The Ca++[i response to glutamate in neurons matured in depolarizing (25 mM K+) growth medium for three days was rapid, transient and heterogeneous, which reached a plateau that was elevated above baseline levels; removing the glutamate markedly reduced the Ca++[i concentration. Activation of the alpha-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid (AMPA)/kainate receptors by kainic acid produced similar changes in Ca++[i responses. At a time when cultured cerebellar granule cells become susceptible to the excitotoxic effects of glutamate acting at NMDA receptors (day in vitro (DIV) 8) in depolarizing growth medium, glutamate elicited Ca++[i responses similar to those observed at a culture time when the neurons are not susceptible to the excitotoxic effects of glutamate (DIV 3). Pretreatment of the cultured neurons with a subtoxic concentration of NMDA, which protects all neurons against the excitotoxic effects of glutamate, did not alter the maximal Ca++[i elicited by an excitotoxic concentration of glutamate.
Marini
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AM
Department of Neurology, Uniformed Services University of the Health Sciences, Bethesda, Maryland 20814, USA. Ann.Marini@NA.AMEDD.Army.Mil
Ueda
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June
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D000818:Animals; D000831:Animals, Newborn; D002118:Calcium / Q000378:metabolism*; D002478:Cells, Cultured; D002531:Cerebellum / Q000187:drug effects / Q000378:metabolism; D018690:Excitatory Amino Acid Agonists / Q000494:pharmacology*; D005089:Exocytosis; D018698:Glutamic Acid / Q000494:pharmacology*; D007608:Kainic Acid / Q000494:pharmacology*; D016202:N-Methylaspartate / Q000494:pharmacology*; D009474:Neurons / Q000187:drug effects / Q000378:metabolism; D011188:Potassium / Q000494:pharmacology; D051381:Rats; D017470:Receptors, Glutamate / Q000187:drug effects*; D018350:alpha-Amino-3-hydroxy-5-methyl-4-isoxazolepropionic Acid / Q000494:pharmacology
D018690:Excitatory Amino Acid Agonists; D017470:Receptors, Glutamate; D018698:Glutamic Acid; D016202:N-Methylaspartate; D018350:alpha-Amino-3-hydroxy-5-methyl-4-isoxazolepropionic Acid; D011188:Potassium; D007608:Kainic Acid; D002118:Calcium