RESUMEN
BACKGROUND: Cannabis usage is increasing with its widespread legalization. Cannabis use by mothers during lactation transfers active cannabinoids to the developing offspring during this critical period and alters postnatal neurodevelopment. A key neurodevelopmental landmark is the excitatory to inhibitory gamma-aminobutyric acid (GABA) switch caused by reciprocal changes in expression ratios of the K+/Cl- transporters potassium-chloride cotransporter 2 (KCC2) and sodium-potassium-chloride transporter (NKCC1). METHODS: Rat dams were treated with Δ9-tetrahydrocannabinol or a synthetic cannabinoid during the first 10 days of postnatal development, and experiments were then conducted in the offspring exposed to these drugs via lactation. The network influence of GABA transmission was analyzed using cell-attached recordings. KCC2 and NKCC1 levels were determined using Western blot and quantitative polymerase chain reaction analyses. Ultrasonic vocalization and homing behavioral experiments were carried out at relevant time points. RESULTS: Treating rat dams with cannabinoids during early lactation retards transcriptional upregulation and expression of KCC2, thereby delaying the GABA switch in pups of both sexes. This perturbed trajectory was corrected by the NKCC1 antagonist bumetanide and accompanied by alterations in ultrasonic vocalization without changes in homing behavior. Neurobehavioral deficits were prevented by CB1 receptor antagonism during maternal exposure, showing that the CB1 receptor underlies the cannabinoid-induced alterations. CONCLUSIONS: These results reveal how perinatal cannabinoid exposure retards an early milestone of development, delaying the trajectory of GABA's polarity transition and altering early-life communication.
Asunto(s)
Cannabinoides , Alucinógenos , Animales , Dronabinol , Femenino , Lactancia , Masculino , Embarazo , Ratas , Ácido gamma-AminobutíricoRESUMEN
Relative to Δ9-tetrahydrocannabinol (THC), the synthetic cannabinoid CP 55,940 (CP) is significantly more potent and efficacious at cannabinoid receptors, the primary targets for endogenous cannabinoids (eCBs). eCBs belong to a large, interconnected lipidome of bioactive signaling molecules with a myriad of effects in optimal and pathological function. Recreational use of highly potent and efficacious synthetic cannabinoids is common amongst adolescents, potentially impacting brain development. Knowledge of the molecular outcomes of synthetic cannabinoid use will be important to develop more targeted therapies for synthetic cannabinoid intoxication and to prevent long-term disruption to the CNS. Here, we test the hypothesis that CP has age and region-dependent effects on the brain lipidome. Adolescent [post-natal day (PND) 35 and PND 50] and young adult female mice were given either an acute dose of CP or vehicle and brains were collected 2 h later. Eight brain regions were dissected and levels of â¼80 lipids were screened from each region using HPLC/MS/MS. CP had widespread effects on the brain lipidome in all age groups. Interestingly, more changes were observed in the PND 35 mice and more were reductions in a lipid's concentration, including region-dependent lowering of eCB levels. CP levels were highest in the cortex at PND 35, the hippocampus at PND 50, and in the cerebellum in the adult. These data provide novel insights into how high-potency, synthetic cannabinoids drive different, age-dependent, cellular signaling effects in the brain.
RESUMEN
Numerous studies have been carried out in the mouse model, investigating the role of the cannabinoid receptor type 1 (CB1). However, mouse CB1 (mCB1) receptor differs from human CB1 (hCB1) receptor in 13 amino acid residues. Two splice variants, hCB1a and hCB1b, diverging in their amino-termini, have been reported to be unique for hCB1 and, via different signaling properties, contribute to CB1 receptor physiology and pathophysiology. We hypothesized that splice variants also exist for the mCB1 receptor and have different signaling properties. On murine hippocampal cDNA, we identified two novel mCB1 receptor splice variants generated by splicing of introns with 117 bp and 186 bp in the N-terminal domain, corresponding to deletions of 39 or 62 amino acids, respectively. The mRNAs for the splice variants mCB1a and mCB1b are expressed at low levels in different brain regions. Western blot analysis of protein extracts from stably transfected HEK293 cells indicates a strongly reduced glycosylation because of the absence of two glycosylation sites in mCB1b. On-cell western analysis in these stable lines revealed increased internalization of mCB1a and mCB1b upon stimulation with the agonist WIN55,212-2 as compared to mCB1. Results also point toward an increased affinity to SR141716 for mCB1a, as well as slightly enhanced inhibition of neurotransmission compared to mCB1. In mCB1b, agonist-induced MAPK phosphorylation was decreased compared to mCB1 and mCB1a. Identification of mouse CB1 receptor splice variants may help to explain differences found between human and mouse endocannabinoid systems and improve the understanding of CB1 receptor signaling and trafficking in different species.