Intracellular injection of brain extracts from Alzheimer’s disease patients trigger unregulated Ca2+ release from intracellular stores that hinders cellular bioenergetics

Strong evidence indicates that amyloid beta (Aβ) inflicts its toxicity in Alzheimer’s disease (AD) by promoting uncontrolled elevation of cytosolic Ca 2+ in neurons. We have previously shown that synthetic Aβ42 oligomers stimulate abnormal intracellular Ca 2+ release from the endoplasmic reticulum stores, suggesting that a similar mechanism of Ca 2+ toxicity may be common to the endogenous Aβs oligomers. To investigate this possibility, we use human postmortem brain extracts from control and AD-affected patients and test their ability to trigger Ca 2+ fluxes when injected intracellularly into Xenopus oocytes. Immunological characterization of samples from AD patients revealed elevated content of soluble Aβ oligomers, detected by the conformation-dependent OC-antibody, whereas no immunoreactivity was detected in the normal samples. Intracellular injection of brain extracts from control patients failed to trigger detectable changes in intracellular Ca 2+ . Conversely, brain extracts from AD patients triggered Ca 2+ events consisting of local and global Ca 2+ fluorescent transients rising within few seconds after injection and persisting for several seconds. Pre-incubation of brain extracts with the conformation specific OC antibody completely suppressed brain extract ability to trigger cytosolic Ca 2+ events. Comparison of the elementary events triggered by brain extracts and synthetic Aβ42 oligomer showed comparable temporal evolution and amplitudes to events triggered by direct injection of IP3. Moreover, bath application of caffeine reversibly inhibited local and global Ca 2+ signals in all the samples confirming the involvement of Ca 2+ release from the ER. Analysis of the recorded Ca 2+ fluorescence signals by computational modeling allowed quantification of the IP3 and Ca 2+ generated by each sample. The model further shows that the abnormal increase of Ca 2+ and IP3 may affect mitochondrial bioenergetics. These results, supports the hypothesis that endogenous amyloid oligomer contained in neurons of AD-affected brains may represent the toxic agents responsible for neurons malfunctioning and death, associated with the disruption of neuronal Ca 2+ homeostasis.