Oligodendrocyte MCT1 Modulates Neuronal Viability

BAC-MCT1 mice were developed and MCT1 expression localized to specific cells by crossing with cell specific reporter lines, immunostaining for cell-specific markers, or isolating mRNA by FACS and BacTRAP. Critical function of oligodendrocyte MCT1 was evaluated in vitro in organotypic spinal cord cultures, in vivo in MCT1^+/− mice or wild-type mice injected with lentiviral vectors. Neuronal toxicity, measured by loss of neurofilament-containing neurons and incorporation of PI, was provoked in organotypic cultures by treating with ASO or MCT1i. MCT1^+/− mice were evaluated by histology, immunohistochemistry, and EM, and compared to littermate controls. For lentiviral experiments, MCT1shRNA was subcloned into lentivirus plasmid along with 3 different promoters (i.e., H1, myelin basic protein (MBP), and Cre-dependent H1). H1-MCT1shRNA lentivirus was injected into the spinal cord of C57Bl6 wild-type mice and motoneurons in the vicinity of virus were counted and compared to control virus injections. MBP-MCT1shRNA was injected into the optic nerve of Sprague –Dawley rats and degenerating axons quantified by EM and compared to the contralateral optic nerve injected with control virus. Cre-dependent H1-MCT1shRNA lentivirus was injected into the corpus callosum of PLP-Cre mice and axon pathology by non-phosphorylated neurofilament immunostaining. Finally, MCT1 expression was evaluated by Western blots of cortex from ALS patients and non-ALS controls; and MCT1 expression in SOD1^G93A transgenic mice, obtained from Jackson laboratories, was evaluating by crossing these mice to MCT1 BAC reporter mice.