Glutamate-induced calcium transient triggers delayed calcium overload and neurotoxicity in rat hippocampal neurons.
Glutamate-induced changes in intracellular free Ca2+ concentration ([Ca2+]i) were recorded in single rat hippocampal neurons grown in primary culture by employing the Ca2+ indicator indo-1 and a dual-emission microfluorimeter. The [Ca2+]i was monitored in neurons exposed to 100 microM glutamate for 5 min and for an ensuing 3 hr period. Ninety-two percent (n = 64) of these neurons buffered the glutamate-induced Ca2+ load back to basal levels after removal of the agonist; thus, the majority of cells had not lost the ability to regulate [Ca2+]i at this time. However, following a variable delay, in 44% (n = 26) of the neurons that buffered glutamate-induced Ca2+ loads to basal levels, [Ca2+]i rose again to a sustained plateau and failed to recover. The changes in [Ca2+]i that occur during glutamate-induced delayed neuronal death can be divided into three phases: (1) a triggering phase during which the neuron is exposed to glutamate and the [Ca2+]i increases to micromolar levels, followed by (2) a latent phase during which the [Ca2+]i recovers to a basal level, and (3) a final phase that begins with a gradual rise in the [Ca2+]i that reaches a sustained plateau from which the neuron does not recover. This delayed Ca2+ overload phase correlated significantly with cell death. The same sequence of events was also observed in recordings from neuronal processes. The delayed Ca2+ increase and subsequent death were dependent upon the presence of extracellular Ca2+ during glutamate exposure. Calcium influx during the triggering phase resulted from the activation of both NMDA and non-NMDA receptors as indicated by studies using receptor antagonists and ion substitution. Treatment with TTX (1 microM) or removal of extracellular Ca2+ for a 30 min window following agonist exposure failed to prevent the delayed Ca2+ overload. The delayed [Ca2+]i increase could be reversed by removing extracellular Ca2+, indicating that it resulted from Ca2+ influx. The three phases defined by changes in the [Ca2+]i during glutamate-induced neuronal toxicity suggest three distinct targets to which neuroprotective agents may be directed.
Randall
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RD
Department of Pharmacology, University of Minnesota Medical School, Minneapolis 55455.
Thayer
S A
SA
D000596:Amino Acids / Q000378:metabolism; D000818:Animals; D002118:Calcium / Q000378:metabolism / Q000502:physiology*; D002470:Cell Survival / Q000187:drug effects; D002478:Cells, Cultured; D005971:Glutamates / Q000494:pharmacology*; D018698:Glutamic Acid; D006624:Hippocampus / Q000166:cytology / Q000187:drug effects* / Q000502:physiology; D007477:Ions; D009474:Neurons / Q000187:drug effects* / Q000378:metabolism / Q000502:physiology; D009498:Neurotoxins / Q000494:pharmacology*; D051381:Rats; D011930:Reaction Time / Q000187:drug effects; D017459:Receptors, Amino Acid; D011956:Receptors, Cell Surface / Q000037:antagonists & inhibitors / Q000502:physiology; D013779:Tetrodotoxin / Q000494:pharmacology
D000596:Amino Acids; D005971:Glutamates; D007477:Ions; D009498:Neurotoxins; D017459:Receptors, Amino Acid; D011956:Receptors, Cell Surface; D018698:Glutamic Acid; D013779:Tetrodotoxin; D002118:Calcium