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

Acute exposure to methylmercury (MeHg) causes severe disruption of intracellular Ca(2+) (Ca(2+)) regulation, which apparently contributes to neuronal death. Activation of the mitochondrial permeability transition pore (MTP) evidently contributes to this effect. We examined in more detail the contribution of mitochondrial Ca(2+) (Ca(2+)) to elevations of Ca(2+) 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+) release from intracellular stores, and delayed the onset of whole-cell Ca(2+) elevations, caused by 0.5 microM 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+), 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 distribution, 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+) 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.