Characterization of Cannabinoid Withdrawal Using Pyruvate Dehydrogenase as a Marker of Inhibition of In Vivo Neuronal Activity

Abstract

Immediate early genes, such as c-fos, are used as molecular markers of neuronal activation; however, there are currently no available molecular markers of neuronal inhibition. Immunohistochemistry (IHC) is used to map activity in different brain regions in response to different stimuli, such as drug administration, exposure to anesthesia, or novel environments. Based on previous literature, we used phosphorylated pyruvate dehydrogenase (pPDH) as a potential marker of neural inhibition. This study used two female rats that were intravenously injected with either WIN 55,212-2 (WIN), a synthetic cannabinoid, or a vehicle solution via jugular catheters on a schedule with increasing doses for four days (0.2-0.8 mg/kg). On the fifth day, a final dose of 0.8 mg/kg of WIN or vehicle was administered. Then, four hours later, they were both injected intraperitoneally with 10 mg/kg of rimonabant, a selective antagonist/inverse agonist of the CB1 receptor, and were sacrificed 90 minutes after rimonabant administration. Using this model of precipitated withdrawal, pPDH immunohistochemistry was performed to detect possible neural inhibition in select brain regions. The regions of interest include various regions in the hippocampus and amygdaloid nucleus. There was a qualitative difference in expression between saline and WIN samples, and more samples would need to be analyzed to include cell counts. More development of this protocol needs to be done to result in more reliable and valid pPDH detection in conjunction with a bigger, more diverse sample size. Future directions should explore the validity and reliability of pPDH as a marker of neural inhibition as well as using c-fos alongside pPDH to investigate the bidirectional nature of neural activity.