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Brevetoxin‐Induced Autocrine Excitotoxicity Is Associated with Manifold Routes of Ca 2+ Influx
Author(s) -
Berman Frederick W.,
Murray Thomas F.
Publication year - 2000
Publication title -
journal of neurochemistry
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.75
H-Index - 229
eISSN - 1471-4159
pISSN - 0022-3042
DOI - 10.1046/j.1471-4159.2000.0741443.x
Subject(s) - chemistry , nmda receptor , excitotoxicity , population , neurotoxicity , channel blocker , biophysics , calcium , biochemistry , receptor , biology , demography , organic chemistry , toxicity , sociology
Real‐time alterations in intracellular Ca 2+ ([Ca 2+ ] i ) were monitored in fluo‐3‐loaded cerebellargranule neurons (CGNs) exposed to the brevetoxin PbTx‐1.[Ca 2+ ] i was measured using a fluorescent plate reader(FLIPR), which measures simultaneously the mean intracellular Ca 2+ change in a population of cultured cells in each well of a 96‐well plate.PbTx‐1 produced rapid and concentration‐dependent increases in neuronal[Ca 2+ ] i with a potency nearly identical to thatdetermined previously for PbTx‐1‐induced neurotoxicity. The NMDA receptorantagonists MK‐801, dextrorphan, and D(‐)‐2‐amino‐5‐phosphonopentanoic acid,and tetanus toxin, an inhibitor of Ca 2+ ‐dependent exocytoticneurotransmitter release, effected significant reductions in both theintegrated fluo‐3 fluorescence response and excitatory amino acid release andprotected CGNs against PbTx‐1 neurotoxicity. The L‐type Ca 2+ channel antagonist nifedipine produced a modest reduction in the fluo‐3response but reduced substantially the plateau phase of the PbTx‐1 incrementin [Ca 2+ ] i when combined with MK‐801. When nifedipineand MK‐801 were combined with the Na + /Ca 2+ exchanger(reversed mode) inhibitor KB‐R7943, the PbTx‐1 increment in[Ca 2+ ] i was nearly completely attenuated. These datashow that Ca 2+ entry into PbTx‐1‐exposed CGNs occurs through threeprimary routes: NMDA receptor ion channels, L‐type Ca 2+ channels,and reversal of the Na + /Ca 2+ exchanger. There was aclose correlation between reduction of the integrated fluo‐3 fluorescenceresponse and the level of neuroprotection afforded by blockers of eachCa 2+ entry pathway; however, simultaneous blockade of L‐typeCa 2+ channels and the Na + /Ca 2+ exchanger,although reducing the integrated [Ca 2+ ] i response to alevel below that provided by NMDA receptor blockade alone, failed tocompletely attenuate PbTx‐1 neurotoxicity. This finding suggests that inaddition to total [Ca 2+ ] i load, neuronal vulnerabilityis governed principally by the NMDA receptor Ca 2+ influx pathway.