Developmental effects of cannabinoids on zebrafish larvae

Cannabinoids are natural or synthetic compounds related chemically to (-)-(6aR,10aR)-6,6,9-trimethyl-3-pentyl-6a,7,8,10a-tetrahydro-6H-benzo[c]chromen-1-ol (Δ(9)-THC), the principle psychotropic constituent of the hemp plant, Cannabis sativa L. Here we examine the effects of the cannabinoids Δ(9)-THC, (R)-(+)-[2,3-dihydro-5-methyl-3-(4-morpholinylmethyl)pyrrolo [1,2,3-de]-1,4-benzoxazin-6-yl]-1-napthalenylmethanone and 2-[(1R,2R,5R)-5-hydroxy-2-(3-hydroxypropyl) cyclohexyl]-5-(2-methyloctan-2-yl)phenol, and the cannabinoid antagonist (AM 251). Exposures were either acute (1-12-h exposure at 108 hours of postfertilization [hpf]) or chronic (96-h exposure starting at 24 hpf). Geometric range finding was used to determine the experimental concentrations. The concentration of the chemical that kills 50% of the test animals in a given time (LC50) was determined based on cumulative mortality at 5 days of postfertilization. At day 5, behavioral analysis (visual motor response test) was carried out in which movement of individual larvae was analysed using automated video-tracking. With acute exposure, embryos showed a biphasic response to the dark challenge with all three cannabinoids tested. This response consisted of stimulation of the locomotor activity at low concentrations, suppression at high doses. With chronic exposure, embryos habituated to the effects of all three cannabinoids when assayed with the dark challenge phase. Further, the excitation was ameliorated when the antagonist AM 251 was coadministered with the cannabinoid. When AM 251 was administered on its own (chronically or acutely), the locomotor activity was suppressed at high concentrations. We examined the embryos for a range of malformations after chronic exposure to cannabinoid. Only Δ(9)-THC was associated with a significant increase in malformations at 5d (yolk sac and pericardial edema, bent tail/body axis). We conclude that cannabinoids have behavioral effects in zebrafish that are comparable to some of those reported in the literature for mammals. In particular, the acute exposure response resembles behavioral effects reported for adult rodents. Our data are consistent with these behavioral effects being mediated, at least in part, by the CB1 receptor.