Evidence for interactions between intracellular calcium stores during methylmercury-induced intracellular calcium dysregulation in rat cerebellar granule neurons

摘要

Acute exposure to methylmercury (MeHg) causes severe disruption o fintracellular Ca~(2+) ([Ca~(2+)]_i} regulation,which apparently contriubtes to neuronal death.Activation of the mitochonently contributes to this effect.We examined in more detailthe contribution of mitochondrial Ca~(2+) ([Ca~(2+)]_m) to elevations of [Ca~(2+)]_i caused by acute exposure to a low concentration of MeHg in primary cultures of rat cerebellar granule neurons.In particular,we sought to determine whether interactions occurred between Ca~(2+)_i pools in response to MeHg.Prior depletion of Ca~(2+)_m using carbonyl cyanide m-chlorophenylhydrazone (CCCP) and oligomycin significantly decreased the amplitude of [Ca~(2+)]_i release from intracellular stores,and delayed the onset of whole-cell [Ca~(2+)]_i elevations,caused by 0.5 muM MeHg.CCCP alone hastened the MeHg-induced release of Ca~(2+) within the cell,whereas oligomycin alone delayed the MeHg-induced influx of extracellular Ca~(2+).In granule cells loaded with rhod-2 acetoxymethylester to measure changes in [Ca~(2+)]_m,MeHg exposure caused a biphasic increase in fluorescence.The initial increase in fluorescence occurred in the absence of extracellular Ca~(2+) and was abolished by mitochondrial depolarization.The secondary increase was associated with spreading of the dye from punctate staining to whole-cell distriubtion,and was delayed significantly by the MTP inhibitor cyclosporin A and the smooth endoplasmic reticulum Ca~(2+) ATPase inhibitor thapsigargin.We conclude that MeHg causes release of Ca~(2+) from the mitochondria through opening of the MTP,which contributes the bulk of the elevated [Ca~(2+)]_i observed during MeHg neurotoxicity.Additionally,the Ca~(2+) that enters the mitochondria seems to originate in the smooth endoplasmic reticulum,providing a mechanism for the observed mitochondrial Ca~(2+) overload.