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Endocannabinoids (eCBs) mediate their effects through actions on several receptors including the cannabinoid receptors CB1R and CB2R. The role played by eCBs in the development of locomotor systems is not fully understood. In this study we investigated the roles of the eCB system in zebrafish development by pharmacologically inhibiting the CB1R and CB2Rs (with AM251 and AM630 respectively) in either the first or second day of development. We examined the morphology of motor neurons and we determined neuromuscular outputs by quantifying the amount of swimming in 5 dpf larva. Blocking CB2R during the first day of development resulted in gross morphological deficits and reductions in heart rate that were greater than that following treatment with the CB1R blocker AM251. Blocking CB1Rs from 0 to 24 hpf resulted in an increase in the number of secondary and tertiary branches of primary motor neurons, whereas blocking CB2Rs had the opposite effect. Both treatments manifested in reduced levels of swimming. Additionally, blocking CB1Rs resulted in greater instances of non-inflated and partially inflated swim bladders compared with AM630, suggesting that at least some of the deficits in locomotion may result from an inability to adjust buoyancy. Together these findings indicate that the endocannabinoid system is pivotal to the development of the locomotor system in zebrafish, and that perturbations of the eCB system early in life may have detrimental effects. [ABSTRACT FROM AUTHOR]

The fatty acid amide hydrolase (FAAH) and monoacylglycerol lipase (MAGL) enzymes are the predominant catabolic regulators of the major endocannabinoids (eCBs) anadamide (AEA) and 2-arachidonoylglycerol (2-AG), respectively. The expression and roles of eCBs during early embryogenesis remain to be fully investigated. Here, we inhibited FAAH and MAGL in zebrafish embryos during the first 24 h of life and examined motor neuron and locomotor development at 2 and 5 days post fertilization (dpf). Application of the dual FAAH/MAGL inhibitor, JZL195 (2 µmol l-1), resulted in a reduction in primary and secondary motor neuron axonal branching. JZL195 also reduced nicotinic acetylcholine receptor (nAChR) expression at neuromuscular junctions. Application of URB597 (5 µmol l-1), a specific inhibitor of the FAAH enzyme, also decreased primary motor neuron branching but did not affect secondary motor neuron branching and nAChR expression. Interestingly, JZL184 (5 µmol l-1), a specific inhibitor of MAGL, showed no effects on motor neuron branching or nAChR expression. Co-treatment of the enzyme inhibitors with the CB1R inhibitor AM251 confirmed the involvement of CB1R in motor neuron branching. Disruption of FAAH or MAGL reduced larval swimming activity, and AM251 attenuated the JZL195- and URB597-induced locomotor changes, but not the effects of JZL184. Together, these findings indicate that inhibition of FAAH, or augmentation of AEA acting through CB1R during early development, may be responsible for locomotor deficiencies., Competing Interests: Competing interests The authors declare no competing or financial interests., (© 2021. Published by The Company of Biologists Ltd.)