Treatment with cannabidiol (CBD) or KLS-13019 (novel CBD analog), has previously been shown to prevent paclitaxel-induced mechanical allodynia in a mouse model of chemotherapy-induced peripheral neuropathy (CIPN). The mechanism of action for CBD- and KLS-13019-mediated protection now has been explored with dissociated dorsal root ganglion (DRG) cultures using small interfering RNA (siRNA) to the mitochondrial Na+ Ca2+ exchanger-1 (mNCX-1). Treatment with this siRNA produced a 50–55% decrease in the immunoreactive (IR) area for mNCX-1 in neuronal cell bodies and a 72–80% decrease in neuritic IR area as determined with high-content image analysis. After treatment with 100 nM KLS-13019 and siRNA, DRG cultures exhibited a 75 ± 5% decrease in protection from paclitaxel-induced toxicity; whereas siRNA studies with 10 μM CBD produced a 74 ± 3% decrease in protection. Treatment with mNCX-1 siRNA alone did not produce toxicity. The protective action of cannabidiol and KLS-13019 against paclitaxel-induced toxicity during a 5-h test period was significantly attenuated after a 4-day knockdown of mNCX-1 that was not attributable to toxicity. These data indicate that decreases in neuritic mNCX-1 corresponded closely with decreased protection after siRNA treatment. Pharmacological blockade of mNCX-1 with CGP-37157 produced complete inhibition of cannabinoid-mediated protection from paclitaxel in DRG cultures, supporting the observed siRNA effects on mechanism.
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Based on the high-content analysis of decreases in mNCX-1 immunoreactive area in DRG neurons, the siRNA approach confirmed the importance of this target in producing acute (5 h) protective actions of CBD and KLS-13019. Coupled with the pharmacological studies, the target knockdown observations suggested that with both cannabinoids, the mNCX-1 target was sufficient to mediate the protection from toxicity produced by paclitaxel in sensory neurons. An unexpected finding was the identification of neuritic mitochondria as a target for the protective action of these cannabinoids. A working hypothesis emerging from these studies is that the cellular location of a target may be of equal importance to the identity of that target, thereby completing or helping to define the mechanistic concept.
These studies were supported by a Grant from the National Institute on Drug Abuse of the National Institutes of Health (R41DA044898).
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Brenneman, D.E., Kinney, W.A. & Ward, S.J. Knockdown siRNA Targeting the Mitochondrial Sodium-Calcium Exchanger-1 Inhibits the Protective Effects of Two Cannabinoids Against Acute Paclitaxel Toxicity. J Mol Neurosci 68, 603–619 (2019). https://doi.org/10.1007/s12031-019-01321-z
- Sodium-calcium exchanger-1 (NCX-1)
- Dorsal root ganglion