Cannabis use increases appetite; however, little is known about the molecular mechanisms that drive this behavior. In this context, a growing body of evidence supports a critical role for RNA biogenesis in the hypothalamus, the brain’s endogenous appetite center, as an environmentally adaptive mechanism that regulates appetite. We recently identified alternative polyadenylation (APA) as a novel post-transcriptional epigenetic mechanism that rapidly regulates information flow from genome to phenome in the context of DIO. Our preliminary data suggest that food intake increases 1 h after acute cannabis sativa exposure, and thus, we hypothesized that hypothalamic APAs underlie this behavioral response.
To address this contention, we employed a Whole Transcriptome Termini Site Sequencing (WTTS-Seq) approach to simultaneously measure differentially expressed APAs (DE-APAs) on multiple RNA biotypes in the hypothalamus of adult male Long Evans rats 1 h following acute vapor cannabis exposure. In silico validation was completed for all transcripts and qPCR was used as a functional validation for select transcripts.
Our WTTS-Seq analysis mapped approximately 59 unique hypothalamic DE-APAs in cannabis exposed rats relative to air controls 1 h after exposure. Notably, we detected DE-APAs on transcripts implicated in excitatory synaptic function such as Slc6a3, the dopamine transporter, and tyrosine hydroxylase, the rate-limiting enzyme for dopamine production. Importantly, we found that reduced DE-APA insertion led to decreased hypothalamic expression of Slc6a3, suggesting that reduced APA usage functionally alters mRNA abundance. Interestingly, we detected significant DE-APAs on Drosha, which is implicated in miRNA processing and addiction pathways.
Collectively, these data highlight DE-APAs as a novel genetic mechanism that directs hypothalamic RNA biogenesis following cannabis exposure. This insight may help compact both anorexia and obesity in a therapeutic context.