The pro-, but not anti-nociceptive effects of Cannabidiol depend on Trpa1b in larval Zebrafish

The promiscuous ligand cannabidiol (CBD) shows promise as an analgesic, but its complex pharmacodynamics has made it difficult to identify its mechanism(s) of action. Numerous putative CBD receptors including cannabinoid receptors CB1 and CB2, as well as Trpv1 and Trpa1-the receptors for capsaicin and mustard-oil (AITC) respectively-have been proposed to contribute to CBD-mediated analgesia. Larval zebrafish have several attributes that lend themselves to inquiries into the biology of nociception. The neural circuits underlying nociception in zebrafish larvae are highly analogous to those found in higher vertebrates. Notably, the small size and optical clarity of zebrafish enable holistic evaluation of analgesic function utilizing behavioral and imaging platforms. Here we report that in larval zebrafish of either sex, CBD serves both anti- and pro-nociceptive functions. Utilizing place aversion assays as a proxy for nociception, we found that low concentrations of CBD inhibit aversion to noxious chemical stimuli including AITC and acetic acid. Counterintuitively, we found that higher concentrations of CBD potentiated nocifensive behavior as measured by enhanced thermal aversion and increased locomotion. Knockdown of Trpa1b eliminated the algogenic effects of CBD while having no effect on its analgesic properties, as it abolished <i>trpa1b+</i> sensory neuron responses to CBD, CBD-evoked thermal hypersensitivity, and increased locomotion in Trpa1b-null animals. Strikingly, CBD profoundly inhibited thermal aversion to noxious heat in Trpa1b-null animals but not in wildtype animals, indicating that CBD-mediated Trpa1b activation can oppose the analgesic properties of CBD. These studies provide a framework to investigate the genetic and neural substrates of CBD-mediated analgesia and nociception.<b>Significance statement</b> While Cannabis and its component compounds such as delta-9-tetrahydrocannabinol (THC) and cannabidiol (CBD) have long been used to alleviate pain, little is known about their mechanisms of action. Our study provides a framework for investigating the effects of cannabinoids on nociception in a vertebrate, by utilizing behavioral experiments in combination with neuroimaging and in-vitro studies. Together, these approaches allowed us to define the role of one molecular target, Trpa1b, involved in the effects of CBD upon nociception. Critically, activation of this receptor by CBD masks a separate, analgesic effect upon temperature sensation, which suggests a possible explanation for CBD's inconsistent efficacy across different models of pain.