Bax Regulates Neuronal Ca2+ Homeostasis

Excessive Ca2+ entry during glutamate receptor overactivation (“excitotoxicity”) induces acute or delayed neuronal death. We report here that deficiency in bax exerted broad neuroprotection against excitotoxic injury and oxygen/glucose deprivation in mouse neocortical neuron cultures and reduced infarct size, necrotic injury, and cerebral edema formation after middle cerebral artery occlusion in mice. Neuronal Ca2+ and mitochondrial membrane potential (Δψm) analysis during excitotoxic injury revealed that bax-deficient neurons showed significantly reduced Ca2+ transients during the NMDA excitation period and did not exhibit the deregulation of Δψm that was observed in their wild-type (WT) counterparts. Reintroduction of bax or a bax mutant incapable of proapoptotic oligomerization equally restored neuronal Ca2+ dynamics during NMDA excitation, suggesting that Bax controlled Ca2+ signaling independently of its role in apoptosis execution. Quantitative confocal imaging of intracellular ATP or mitochondrial Ca2+ levels using FRET-based sensors indicated that the effects of bax deficiency on Ca2+ handling were not due to enhanced cellular bioenergetics or increased Ca2+ uptake into mitochondria. We also observed that mitochondria isolated from WT or bax-deficient cells similarly underwent Ca2+-induced permeability transition. However, when Ca2+ uptake into the sarco/endoplasmic reticulum was blocked with the Ca2+-ATPase inhibitor thapsigargin, bax-deficient neurons showed strongly elevated cytosolic Ca2+ levels during NMDA excitation, suggesting that the ability of Bax to support dynamic ER Ca2+ handling is critical for cell death signaling during periods of neuronal overexcitation.