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A Prototypic Intracellular Calcium Antagonist, TMB‐8, Protects Cultured Cerebellar Granule Cells Against the Delayed, Calcium‐Dependent Component of Glutamate Neurotoxicity
Author(s) -
Malcolm Craig S.,
Ritchie Lyndsay,
Grieve Angus,
Griffiths Roger
Publication year - 1996
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.1471-4159.1996.66062350.x
Subject(s) - glutamate receptor , neurotoxicity , excitotoxicity , extracellular , calcium , granule (geology) , intracellular , viability assay , nmda receptor , calcium in biology , fura 2 , chemistry , cytosol , biochemistry , biology , biophysics , microbiology and biotechnology , toxicity , cell , receptor , paleontology , organic chemistry , enzyme
The effect(s) of a prototypic intracellular Ca 2+ antagonist, 8‐( N,N ‐diethylamino)octyl‐3,4,5‐trimethoxybenzoate (TMB‐8), on glutamate‐induced neurotoxicity was investigated in primary cultures of mouse cerebellar granule cells. Glutamate evoked an increase in cytosolic free‐Ca 2+ levels ([Ca 2+ ] i ) that was dependent on the extracellular concentration of Ca 2+ ([Ca 2+ ] o ). In addition, this increase in [Ca 2+ ] i correlated with a decrease in cell viability that was also dependent on [Ca 2+ ] o . Glutamate‐induced toxicity, quantified by 3‐(4,5‐dimethylthiazol‐2‐yl)‐2,5‐diphenyltetrazolium bromide (MTT) staining, was shown to comprise two distinct components, an “early” Na + /Cl − ‐dependent component observed within minutes of glutamate exposure, and a “delayed” Ca 2+ ‐dependent component (ED 50 ∼50 µ M ) that coincided with progressive degeneration of granule cells 4–24 h after a brief (5–15 min) exposure to 100 µ M glutamate. Quantitative analysis of cell viability and morphological observations identify a “window” in which TMB‐8 (at >100 µ M ) protects granule cells from the Ca 2+ ‐dependent, but not the Na + /Cl − ‐dependent, component of glutamate‐induced neurotoxic damage, and furthermore, where TMB‐8 inhibits glutamate‐evoked increases in [Ca 2+ ] i . These findings suggest that Ca 2+ release from a TMB‐8‐sensitive intracellular store may be a necessary step in the onset of glutamate‐induced excitotoxicity in granule cells. However, these conclusions are compromised by additional observations that show that TMB‐8 (1) exhibits intrinsic toxicity and (2) is able to reverse its initial inhibitory action on glutamate‐evoked increases in [Ca 2+ ] i and subsequently effect a pronounced time‐dependent potentiation of glutamate responses. Dantrolene, another putative intracellular Ca 2+ antagonist, was completely without effect in this system with regard to both glutamate‐evoked increases in [Ca 2+ ] i and glutamate‐induced neurotoxicity.