Cannabinoid CB1 receptors are expressed in a sub-set of dopamine neurons and underlie cannabinoid modulation of DA release and DA-related behavior in mice / 中国药理学与毒理学杂志

OBJECTIVE Cannabinoids modulate do-pamine(DA)transmission and DA-related behavior,which has been thought to be mediated initially by acti-vation of cannabinoid CB1 receptors(CB1Rs)on GABA neurons.However,the cellular and receptor mechanisms underlying cannabinoids' psychoactive effects are not fully understood.The present study is to explore the pos-sible expression character of CB1Rs and elucidated the underlying mechanism of them.METHODS We took advantage of RNAscope in situ hybridization(ISH)assays and triple-staining assays to detect the CB1R-expressing neurons.We established an optical intracranial self-stimulation(OICSS)behavioral model by using opto-genetics to study dopaminergic reinforcement function.Natural and synthetic cannabinoids were used to study the function of CB1Rs.Conditional genetic depletion of CB1Rs and behavioral assay were performed to study the modulatory role of CB1Rs in DA-related behaviors.RESULTS We found that CB1Rs are also expressed in a subset of DA neurons and functionally underlie cannabi-noid action in male and female mice.ISH assays demon-strated CB1 mRNA in tyrosine hydroxylase(TH)-posi-tive DA neurons in the ventral tegmental area(VTA)and glutamate decarboxylase 1(GAD1)-positive GABA neu-rons.The CB1R expressing DA neurons were located mainly in the middle portion of the VTA with the number of CB1-TH colocalization progressively decreasing from the medial to the lateral VTA.Triple-staining assays indi-cated CB1R mRNA colocalization with both TH and vesicular glutamate transporter 2(VgluT2,a glutamate neuronal marker)in the medial VTA close to the midline of the brain.Optogenetic activation of this population of DA neurons was rewarding as assessed by OICSS.D9-tetrahydrocannabinol(D9-THC)or ACEA(a selective CB1R agonist)dose-dependently inhibited optical intra-cranial self-stimulation in DAT-Cre control mice,but not in conditional knockout mice with the CB1R gene absent in DA neurons.In addition,deletion of CB1Rs from DA neurons attenuated D9-THC-induced reduction in DA release in the NAc,locomotion,and anxiety.CONCLU-SION Our results indicated that CB1Rs are expressed in a subset of DA neurons that corelease DA and gluta-mate,and functionally underlie cannabinoid modulation of DA release and DA-related behavior.

Endocannabinoid system and periodontitis: mechanisms and therapeutic implications / Sistema endocanabinoide e periodontite: mecanismos e implicações terapêuticas

Introdução: A periodontite é um importante problema de saúde pública. Embora o princípio da terapia da periodontite esteja focado principalmente na remoção do biofilme dental e dos fatores associados, sua fisiopatologia registra diferentes eventos moleculares e inflamatórios relacionados ao sistema imunológico do hospedeiro, como a participação do sistema endocanabinoide. Objetivo: Esta revisão teve como objetivo explorar e elucidar os mecanismos e papéis do sistema endocanabinoide na fisiopatologia da periodontite e suas possibilidades para futuras terapias relacionadas. Material e método: Realizou-se uma busca eletrônica na plataforma PubMed por estudos envolvendo a ação do sistema endocanabinoide sobre a periodontite. Resultado: Dezenove estudos clínicos e pré-clínicos foram incluídos nesta revisão narrativa. Conclusão: Os receptores canabinoides tipo 1 e 2 são componentes integrais do sistema endocanabinoide e manifestam-se de várias formas nos tecidos periodontais. As ações e mecanismos através dos quais os receptores canabinoides são ativados em locais saudáveis ou inflamados continuam a ser o foco de investigações em curso. Além disso, os fitocanabinoides e canabinoides sintéticos apresentam potencial como tratamentos, com estudos pré-clínicos indicando benefícios na redução da inflamação e na facilitação da reparação dos tecidos.

Introduction: Periodontitis is a major public health problem. Although the principle of periodontitis therapy is mainly focused on removing dental biofilm and associated factors, its physiopathology enrolls different molecular and inflammatory events related to the host immune system, as the participation of the endocannabinoid system. Objective: This review aimed to explore and elucidate the mechanisms and roles of the endocannabinoid system on periodontitis physiopathology and its possibilities for future related therapies. Material and method: An electronic search was carried out on the PubMed platform for studies involving the action of the endocannabinoid system on periodontitis. Result: Nineteen clinical and preclinical studies were included in this narrative review. Conclusion: Cannabinoid receptors type 1 and 2 are integral components of the endocannabinoid system, manifesting in various forms in the periodontal tissues. The actions and mechanisms through which cannabinoid receptors are activated in healthy or inflamed sites remain the focus of ongoing investigations. Moreover, phytocannabinoids and synthetic cannabinoids show therapeutic potential, with pre-clinical studies indicating benefits in reducing inflammation and facilitating tissue repair

Pharmacological characterizationof synthetic cannabinoid MAM-2201:radioligand binding and abuse-related effects / 中国药理学与毒理学杂志

OBJECTIVE Over 30% of all new psychoactive substances identified by the UN Office on Drugs and Crime in 2016 were synthetic cannabinoids. The recent emergence of MAM-2201 on the illicit market is troubling because this drug has no precedent in either the scientific or patent literature, and appears to be a novel compound developed specifically as a ″graymarket″ drug of abuse bystruc?turally combining the known synthetic cannabinoids JWH- 122 and AM- 2201. There is currently no published information regarding the pharmacology of MAM-2201. METHODS The present studies characterized cannabinoid-like effects of MAM-2201 in vitro (interactions with cannabinoid type 1 receptors [CB1Rs]) and in vivo (in mice and rats). RESULTS In a radioligand binding assay using [3H]CP55,940 in HEK cell membranes transfected with the CB1R, MAM-2201 (Ki=5.4 nmol·L- 1), had higher binding affinity than WIN 55,212-2 (Ki=80 nmol·L-1), and D9-THC (Ki=8.3 nmol·L-1). The Emax values for MAM-2201 and WIN 55,212-2 in an assay of agonist inhibition of forskolin-stimulated cAMP were 85% (EC50=0.45 nmol·L-1) and 95%, respectively, as compared with the D9-THC Emax of 74%. In mice, MAM-2201 (0.003-1.0 mg·kg-1, IP) produced dose-dependent cannabimimetic effects which were both more potent and more effective than those of D9-THC. MAM-2201 and D9-THC dose-dependently produced hypothermia:ED50=0.287 and 25.4 mg·kg-1, analgesia: ED50=0.125 and 29.4 mg·kg-1, and catalepsy: ED50=0.301 and 18.9 mg·kg-1 in adult male CD1 mice. Importantly, MAM-2201 also elicited convulsant effects at a dose of 1.0 mg·kg-1 in 8/8 murine subjects. In rats, MAM-2201 produced dose-dependent D9-THC-like intero?ceptive effects in subjects trained to discriminate 3.0 mg·kg-1 (IP) D9-THC from saline. CONCLUSION MAM-2201 binds CB1Rs with high affinity and agonist efficacy, and functions as a potent cannabinoid agonist in vivo across several complementary measures of cannabinoid activity in two rodent species.

A spider derived peptide, PnPP-19, induces central antinociception mediated by opioid and cannabinoid systems

Background: Some peptides purified from the venom of the spider Phoneutria nigriventer have been identified as potential sources of drugs for pain treatment. In this study, we characterized the antinociceptive effect of the peptide PnPP-19 on the central nervous system and investigated the possible involvement of opioid and cannabinoid systems in its action mechanism. Methods: Nociceptive threshold to thermal stimulation was measured according to the tail-flick test in Swiss mice. All drugs were administered by the intracerebroventricular route.Results: PnPP-19 induced central antinociception in mice in the doses of 0.5 and 1 µg. The non-selective opioid receptor antagonist naloxone (2.5 and 5 µg), µ-opioid receptor antagonist clocinnamox (2 and 4 µg), δ-opioid receptor antagonist naltrindole (6 and 12 µg) and CB1 receptor antagonist AM251 (2 and 4 µg) partially inhibited the antinociceptive effect of PnPP-19 (1 µg). Additionally, the anandamide amidase inhibitor MAFP (0.2 µg), the anandamide uptake inhibitor VDM11 (4 µg) and the aminopeptidase inhibitor bestatin (20 µg) significantly enhanced the antinociception induced by a low dose of PnPP-19 (0.5 µg). In contrast, the κ-opioid receptor antagonist nor-binaltorphimine (10 µg and 20 µg) and the CB2 receptor antagonist AM630 (2 and 4 µg) do not appear to be involved in this effect. Conclusions: PnPP-19-induced central antinociception involves the activation of CB1 cannabinoid, µ- and δ-opioid receptors. Mobilization of endogenous opioids and cannabinoids might be required for the activation of those receptors, since inhibitors of endogenous substances potentiate the effect of PnPP-19. Our results contribute to elucidating the action of the peptide PnPP-19 in the antinociceptive pathway.(AU)

A spider derived peptide, PnPP-19, induces central antinociception mediated by opioid and cannabinoid systems

Some peptides purified from the venom of the spider Phoneutria nigriventer have been identified as potential sources of drugs for pain treatment. In this study, we characterized the antinociceptive effect of the peptide PnPP-19 on the central nervous system and investigated the possible involvement of opioid and cannabinoid systems in its action mechanism. Methods Nociceptive threshold to thermal stimulation was measured according to the tail-flick test in Swiss mice. All drugs were administered by the intracerebroventricular route. Results PnPP-19 induced central antinociception in mice in the doses of 0.5 and 1 g. The non-selective opioid receptor antagonist naloxone (2.5 and 5 g), -opioid receptor antagonist clocinnamox (2 and 4 g), -opioid receptor antagonist naltrindole (6 and 12 g) and CB1 receptor antagonist AM251 (2 and 4 g) partially inhibited the antinociceptive effect of PnPP-19 (1 g). Additionally, the anandamide amidase inhibitor MAFP (0.2 g), the anandamide uptake inhibitor VDM11 (4 g) and the aminopeptidase inhibitor bestatin (20 g) significantly enhanced the antinociception induced by a low dose of PnPP-19 (0.5 g). In contrast, the -opioid receptor antagonist nor-binaltorphimine (10 g and 20 g) and the CB2 receptor antagonist AM630 (2 and 4 g) do not appear to be involved in this effect. Conclusions PnPP-19-induced central antinociception involves the activation of CB1 cannabinoid, - and -opioid receptors. Mobilization of endogenous opioids and cannabinoids might be required for the activation of those receptors, since inhibitors of endogenous substances potentiate the effect of PnPP-19. Our results contribute to elucidating the action of the peptide PnPP-19 in the antinociceptive pathway.

A Cannabinoid Receptor Agonist N-Arachidonoyl Dopamine Inhibits Adipocyte Differentiation in Human Mesenchymal Stem Cells

Endocannabinoids can affect multiple cellular targets, such as cannabinoid (CB) receptors, transient receptor potential cation channel, subfamily V, member 1 (TRPV1) and peroxisome proliferator-activated receptor gamma (PPARgamma). The stimuli to induce adipocyte differentiation in hBM-MSCs increase the gene transcription of the CB1 receptor, TRPV1 and PPARgamma. In this study, the effects of three endocannabinoids, N-arachidonoyl ethanolamine (AEA), N-arachidonoyl dopamine (NADA) and 2-arachidonoyl glycerol (2-AG), on adipogenesis in hBM-MSCs were evaluated. The adipocyte differentiation was promoted by AEA whereas inhibited by NADA. No change was observed by the treatment of non-cytotoxic concentrations of 2-AG. The difference between AEA and NADA in the regulation of adipogenesis is associated with their effects on PPARgamma transactivation. AEA can directly activate PPARgamma. The effect of AEA on PPARgamma in hBM-MSCs may prevail over that on the CB1 receptor mediated signal transduction, giving rise to the AEA-induced promotion of adipogenesis. In contrast, NADA had no effect on the PPARgamma activity in the PPARgamma transactivation assay. The inhibitory effect of NADA on adipogenesis in hBM-MSCs was reversed not by capsazepine, a TRPV1 antagonist, but by rimonabant, a CB1 antagonist/inverse agonist. Rimonabant by itself promoted adipogenesis in hBM-MSCs, which may be interpreted as the result of the inverse agonism of the CB1 receptor. This result suggests that the constantly active CB1 receptor may contribute to suppress the adipocyte differentiation of hBM-MSCs. Therefore, the selective CB1 agonists that are unable to affect cellular PPARgamma activity inhibit adipogenesis in hBM-MSCs.

Dependence Potential of the Synthetic Cannabinoids JWH-073, JWH-081, and JWH-210: In Vivo and In Vitro Approaches

Synthetic cannabinoids (CBs) such as the JWH series have caused social problems concerning their abuse liability. Because the JWH series produces euphoric and hallucinogenic effects, they have been distributed illegally under street names such as "Spice" and "Smoke". Many countries including Korea have started to schedule some of the JWH series compounds as controlled substances, but there are a number of JWH series chemicals that remain uncontrolled by law. In this study, three synthetic CBs with different binding affinities to the CB1 receptor (JWH-073, 081, and 210) and Delta9-tetrahydrocannabinol (Delta9-THC) were evaluated for their potential for psychological dependence. The conditioned place preference test (unbiased method) and self-administration test (fixed ratio of 1) using rodents were conducted. Ki values of the three synthetic cannabinoids were calculated as supplementary data using a receptor binding assay and overexpressed CB1 protein membranes to compare dependence potential with CB1 receptor binding affinity. All mice administered JWH-073, 081, or 210 showed significantly increased time spent at unpreferred space in a dose-dependence manner in the conditioned place preference test. In contrast, all tested substances except Delta9-THC showed aversion phenomenon at high doses in the conditioned place preference test. The order of affinity to the CB1 receptor in the receptor binding assay was JWH-210 > JWH-081 >> JWH-073, which was in agreement with the results from the conditioned place preference test. However, no change in self-administration was observed. These findings suggest the possibility to predict dependence potential of synthetic CBs through a receptor binding assay at the screening level.

Inhibition of 5-HT3 Receptors-activated Currents by Cannabinoids in Rat Trigeminal Ganglion Neurons / 华中科技大学学报（医学）（英德文版）

This study investigated the modulatory effect of synthetic cannbinoids WIN55,212-2 on 5-HT3 receptor-activated currents (I5-Hr3) in cultured rat trigeminal ganglion (TG) neurons using whole-cell patch clamp technique.The results showed that:(!) The majority of examined neurons (78.70％) were sensitive to 5-HT (3-300 μmol/L).5-HT induced inward currents in a concentrationdependent manner and the currents were blocked by ICS 205-930 (1 μmol/L),a selective antagonist of the 5-HT3 receptor; (2) Pre-application of WIN55,212-2 (0.01-1 μmol/L) significantly inhibited I5-HT3 reversibly in concentration-dependent and voltage-independent manners.The concentration-response curve of 5-HT3 receptor was shifted downward by WIN55,212-2 without any change of the threshold value.The EC50values of two curves were very close (17.5±4.5) μmol/L vs.(15.2±4.5) μmol/L and WIN55,212-2 decreased the maximal amplitude of I5-HI3 by (48.65±4.15)％; (3) Neither AM281,a selective CBI receptor antagonist,nor AM630,a selective CB2 receptor antagonist reversed the inhibition of I5-HT3by WIN55,212-2; (4) When WIN55,212-2 was given from 15 to 120 s before 5-HT application,inhibitory effect was gradually increased and the maximal inhibition took place at 90 s,and the inhibition remained at the same level after 90 s.We are led to concluded thatWIN55,212-2 inhibited I5-Hr3 significantly and neither CB1 receptor antagonist nor CB2 receptor antagonist could reverse the inhibition of I5-HT3 by WIN55,212-2.Moreover,WIN55,212-2 is not an open channel blocker (OCB) of 5-HT3 receptor.WIN55,212-2 significantly inhibited 5-HT-activated currents in a non-competitive manner.The inhibition of I5-HT3 by WIN55,212-2 is probably new one of peripheral analgesic mechanisms of WIN55,212-2,but the mechanism by which WIN55,212-2 inhibits I5-HT3 warrants further investigation.

Participación del sistema endocanabinoide en el desarrollo de obesidad / A role for the endocannabinoid system in obesity

Endocannabinoids are the endogenous ligands for the cannabinoid receptors type 1 and 2. These membrane receptors are responsible for the psychotropic effects of Cannabis Sativa, when bound to its active component known as (-)-Δ9-tetrahydro-cannabinol. Cannabinoid receptors, endocannabinoids and the enzymes catalyzing their biosynthesis and degradation, constitute the endocannabinoid system (ECS), which has a remarkable role controlling energy balance, both at central nervous system and peripheral tissues. The ECS regulates food ingestion by stimulating a network of orexigenic neurons present in the hypothalamus and reinforcing motivation and reward to food consumption in the nucleus accumbens. Regarding peripheral tissues, this system controls lipid and glucose metabolism at different levels, reduces energy expenditure and leads energy balance to fat storage. Metabolic alterations, includ-ing excessive accumulation of abdominal fat, dyslipidaemia and hyperglicaemia, are suggested to be associated to a hyperactivated ECS. Since obesity is one of the major health problems in modern societies, in this review we discuss the role of the endocannabinoid system in metabolic pathways associated to control mechanisms of energy balance and its involvement in overweight and obesity. In addition, we also discuss therapeutic possibilities and emergent problems due to cannabinoid receptor type 1 antagonism utilized as treatment for such alterations.