Interference with the Cannabinoid Receptor CB1R Results in Miswiring of GnRH3 and AgRP1 Axons in Zebrafish Embryos

Abstract

The G protein-coupled cannabinoid receptors type 1 (CB1R) and type 2 (CB2R), and their endocannabinoid (eCBs) ligands, have been implicated in several aspects of brain wiring during development. Here we aim to assess whether interfering with CB1R affects development, neuritogenesis and pathfinding of GnRH and AgRP neurons, forebrain neurons that control respectively reproduction and appetite. We pharmacologically and genetically interfered with CB1R in zebrafish strains with fluorescently labeled GnRH3 and the AgRP1 neurons. By applying CB1R antagonists we observed a reduced number of GnRH3 neurons, fiber misrouting and altered fasciculation. Similar phenotypes were observed by CB1R knockdown. Interfering with CB1R also resulted in a reduced number, misrouting and poor fasciculation of the AgRP1 neuron’s axonal projections. Using a bioinformatic approach followed by qPCR validation, we have attempted to link CB1R functions with known guidance and fasciculation proteins. The search identified stathmin-2, a protein controlling microtubule dynamics, previously demonstrated to be coexpressed with CB1R and now shown to be downregulated upon interference with CB1R in zebrafish. Together, these results raise the likely possibility that embryonic exposure to low doses of CB1R-interfering compounds could impact on the development of the neuroendocrine systems controlling sexual maturation, reproduction and food intake.

Anosmin-1-Like Effect of UMODL1/Olfactorin on the Chemomigration of Mouse GnRH Neurons and Zebrafish Olfactory Axons Development

“…The fish strain gnrh3:EGFP, used to visualize the early GnRH3 neurons (Abraham et al, 2009(Abraham et al, , 2010Garaffo et al, 2015;Zuccarini et al, 2019), was obtained from Dr. Y. Zohar (Univ. Maryland Biotechnology Institute, Baltimore, United States).…”

Marijuana and Opioid Use during Pregnancy: Using Zebrafish to Gain Understanding of Congenital Anomalies Caused by Drug Exposure during Development

“…THC and CBD interact with both CNR1 and CNR2 receptors (THC binds to both types of receptors and CBD modifies the receptors’ ability to bind to cannabinoids). Continuous exposure to different CNR1 receptor ligands WIN55,212-2 (1 nM-1 µM), AM251 (100 nM-5 µM) or rimonabant (SR141716A:1 nM-1 µM) at various concentrations from 0–72 hpf or 0–96 hpf, without the replacement of solution for the entire exposure period, did not impact hatching [ 88 ]. Although the highest concentration of WIN55,212-2 (1 µM; CNR1 agonist) caused complete embryonic lethality, lower concentrations had no impact on the survival of the embryos.…”

“…Although the highest concentration of WIN55,212-2 (1 µM; CNR1 agonist) caused complete embryonic lethality, lower concentrations had no impact on the survival of the embryos. Exposure to CNR1 antagonists AM251 or rimonabant also did not change the survival rate [ 88 ]. In another study, Migliarini et.…”

CB1 antagonism increases excitatory synaptogenesis in a cortical spheroid model of fetal brain development

“…Global CB 1 knockouts in mice, as well as specific interneuron and pyramidal cell CB 1 knockouts, are viable but show axonal guidance errors and impaired postsynaptic target selection 6 , 36 , 37 . Interestingly, pharmacological treatment with both CB 1 agonists and antagonists creates axon fasciculation errors during development in mice 7 , 37 , 40 , 41 . It is clear from this research that the ECS is necessary for correct axonal targeting and the subsequent establishment of synapses in the developing brain.…”