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(Abst. 1.309), 2017

Effects of Cannabidiol on Mitochondrial Bioenergetics and Homeostasis
Authors: Younghee Ahn, University of Calgary; Christopher Drummond-Main, University of Calgary; Nellie C. Yee, University of Calgary; Do Young Kim, Barrow Neurological Institute; and Jong M. Rho, University of Calgary
Content: Rationale: Cannabidiol (CBD) is one of the hundreds of cannabinoids identified in cannabis (medical marijuana) and has received much interest for the treatment of medically intractable epilepsy. Currently, there is abundant evidence that CBD possesses beneficial neuromodulatory properties, including anti-seizure effects in rodent models. However, the underlying mechanisms as well as both short- and long-term side effects of CBD remain poorly understood. In a previous study (Bernard et al., PNAS, 2005), chronic marijuana ingestion consumption significantly altered brain neurochemistry, morphology and physiology, in a manner that would be expected to impair cognition and behavior, especially during brain maturation. We hypothesized that CBD can disrupt brain mitochondrial bioenergetics and homeostasis at very low micromolar concentrations. Methods: Mitochondrial isolation and protein assay (Sullivan et al., Ann Neurol., 2003 and Pandya et al., Exp Neurol., 2009), spectrofluorophotometer assays for mPT (mitochondrial permeability transition) (Brown et al., J Biol Chem, 2006), bioenergetics profiling using Seahorse XF24 extracellular flux analyzer (Ahn et al, Dev Neurosci., 2014). Results: To assess whether CBD can impair mitochondrial function, we measured thresholds for calcium-induced activation of the mPT complex and respiration changes in acutely isolated brain mitochondria. CBD (3.75µM) reduced mPT in a manner similar to what was observed with the mitochondrial chloride channel inhibitor DIDS (4,4'-diisothiocyanatostilbene-2,2'-disulfonic acid). Using the same assay, we calculated mitochondrial calcium uptake. CBD at less than 1.875µM had no significant effect, but concentrations over 3.75µM led to significant reductions in mitochondrial calcium uptake. Next, we utilized the Seahorse XF24 analyzer to measure oxygen consumption rates (OCRs). CBD decreased OCR and ATP production at concentrations over 7.5µM. Given the impairment of mitochondrial functions, CBD would be predicted to induce neuronal injury. Indeed, we found that CBD (1µM or higher) significantly decreased cell viability in murine hippocampal HT22 cells. Conclusions: While CBD has been used in various formulations to treat epilepsy for several decades, and more recently for certain forms of childhood encephalopathic epilepsies (Devinsky et al, NEJM 2017), there is a dearth of information regarding both short-term and long-term side-effects. Our data indicate that at very low micromolar concentrations, CBD can impair several measures of mitochondrial function, lower the threshold for mPT, which collectively induces cell death. The results of our investigations may help establish safe dosing of CBD in clinical populations. Funding: Alberta Children's Hospital Research Institute