[Ca2+]i signaling between mitochondria and endoplasmic reticulum in neurons is regulated by microtubules -: From mitochondrial permeability transition pore to Ca2+-induced Ca2+ release

The positioning and dynamics of organelles depend on membrane-cytoskeleton interactions. Mitochondria relocate along microtubules (MT), but it is not clear whether MT have direct effects on mitochondrial function. Using two-photon microscopy and the mitochondrial fluorescent dyes rhodamine 123 and Rhod-2, we showed that Taxol and nocodazole, which correspondingly stabilize and disrupt MT, decreased potential and Ca2+ in the mitochondria of brain stem pre-Botzinger complex neurons. Without changing basal cytoplasmic Ca2+ ([Ca2+](i)), Taxol promoted the generation of [Ca2+](i) spikes in dendrites. These spikes were abolished after blockade of Ca2+ influx and after depletion of internal Ca2+ stores, indicating the involvement of Ca2+-induced Ca2+ release. Nocodazole decreased mitochondrial potential and [Ca2+] m and produced a long lasting increase in [Ca2+](i). MT-acting drugs depolarized single immobilized mitochondria and released previously stored Ca2+. All of these effects were inhibited by pretreatment with blockers of mitochondrial permeability transition pore (mPTP), cyclosporin A, and 2-aminoethoxydiphenyl borate. Induction of mPTP by Taxol and nocodazole was confirmed by using a calcein/Co2+ imaging technique. Electron and optical microscopy revealed tubulin bound to mitochondria. Mitochondria, MT, and endoplasmic reticulum ( ER) showed strong co-localization, the degree of which decreased after MT were disrupted. We propose that changes in the structure of MT by Taxol and nocodazole promote the induction of mPTP. Subsequent Ca2+ efflux stimulates the Ca2+ release from the ER that drives spontaneous [Ca2+](i) transients. Thus, close positioning of mitochondria to the ER as determined by MT can be essential for the local [Ca](i) signaling in neurons.