The endocannabinoid system in appetite regulation and treatment of obesity.

The endocannabinoid system (ECS) is a complex cell-signaling system that is responsible for maintaining homeostasis by modulating various regulatory reactions in response to internal and environmental changes. The influence of ECS on appetite regulation has been a subject of much recent research, however, the full extent of its impact remains unknown. Current evidence links human obesity to ECS activation, increased endocannabinoid levels in both central and peripheral tissues, along with cannabinoid receptor type 1 (CBR1) up-regulation. These findings imply the potential pharmacological use of the ECS in the treatment of obesity. Here, we present various pathophysiological processes in obesity involving the ECS, highlighting different pharmacological options for modulating endocannabinoid activity to treat obesity. However, the potential of those pharmacological possibilities remains under investigation and requires further research.

Pharmacological inhibition of the CB1 cannabinoid receptor restores abnormal brain mitochondrial CB1 receptor expression and rescues bioenergetic and cognitive defects in a female mouse model of Rett syndrome.

BACKGROUND: Defective mitochondria and aberrant brain mitochondrial bioenergetics are consistent features in syndromic intellectual disability disorders, such as Rett syndrome (RTT), a rare neurologic disorder that severely affects mainly females carrying mutations in the X-linked MECP2 gene. A pool of CB1 cannabinoid receptors (CB1R), the primary receptor subtype of the endocannabinoid system in the brain, is located on brain mitochondrial membranes (mtCB1R), where it can locally regulate energy production, synaptic transmission and memory abilities through the inhibition of the intra-mitochondrial protein kinase A (mtPKA). In the present study, we asked whether an overactive mtCB1R-mtPKA signaling might underlie the brain mitochondrial alterations in RTT and whether its modulation by systemic administration of the CB1R inverse agonist rimonabant might improve bioenergetics and cognitive defects in mice modeling RTT. METHODS: Rimonabant (0.3 mg/kg/day, intraperitoneal injections) was administered daily to symptomatic female mice carrying a truncating mutation of the Mecp2 gene and its effects on brain mitochondria functionality, systemic oxidative status, and memory function were assessed. RESULTS: mtCB1R is overexpressed in the RTT mouse brain. Subchronic treatment with rimonabant normalizes mtCB1R expression in RTT mouse brains, boosts mtPKA signaling, and restores the defective brain mitochondrial bioenergetics, abnormal peripheral redox homeostasis, and impaired cognitive abilities in RTT mice. LIMITATIONS: The lack of selectivity of the rimonabant treatment towards mtCB1R does not allow us to exclude that the beneficial effects exerted by the treatment in the RTT mouse model may be ascribed more broadly to the modulation of CB1R activity and distribution among intracellular compartments, rather than to a selective effect on mtCB1R-mediated signaling. The low sample size of few experiments is a further limitation that has been addressed replicating the main findings under different experimental conditions. CONCLUSIONS: The present data identify mtCB1R overexpression as a novel molecular alteration in the RTT mouse brain that may underlie defective brain mitochondrial bioenergetics and cognitive dysfunction.

Actomyosin-mediated inhibition of synaptic vesicle release under CB1R activation.

Long-term synaptic plasticity is critical for adaptive function of the brain, but presynaptic mechanisms of functional plasticity remain poorly understood. Here, we show that changes in synaptic efficacy induced by activation of the cannabinoid type-1 receptor (CB1R), one of the most widespread G-protein coupled receptors in the brain, requires contractility of the neuronal actomyosin cytoskeleton. Specifically, using a synaptophysin-pHluorin probe (sypH2), we show that inhibitors of non-muscle myosin II (NMII) ATPase as well as one of its upstream effectors Rho-associated kinase (ROCK) prevent the reduction of synaptic vesicle release induced by CB1R activation. Using 3D STORM super-resolution microscopy, we find that activation of CB1R induces a redistribution of synaptic vesicles within presynaptic boutons in an actomyosin dependent manner, leading to vesicle clustering within the bouton and depletion of synaptic vesicles from the active zone. We further show, using sypH2, that inhibitors of NMII and ROCK specifically restore the release of the readily releasable pool of synaptic vesicles from the inhibition induced by CB1R activation. Finally, using slice electrophysiology, we find that activation of both NMII and ROCK is necessary for the long-term, but not the short-term, form of CB1R induced synaptic plasticity at excitatory cortico-striatal synapses. We thus propose a novel mechanism underlying CB1R-induced plasticity, whereby CB1R activation leads to a contraction of the actomyosin cytoskeleton inducing a reorganization of the functional presynaptic vesicle pool, preventing vesicle release and inducing long-term depression.

Bioelectronic sensing platform emulating the human endocannabinoid system for assessing and modulating of cannabinoid activity.

Cannabinoids are involved in physiological and neuromodulatory processes through their interactions with the human cannabinoid receptor-based endocannabinoid system. Their association with neurodegenerative diseases and brain reward pathways underscores the importance of evaluating and modulating cannabinoid activity for both understanding physiological mechanisms and developing therapeutic drugs. The use of agonists and antagonists could be strategic approaches for modulation. In this study, we introduce a bioelectronic sensor designed to monitor cannabinoid binding to receptors and assess their agonistic and antagonistic properties. We produced human cannabinoid receptor 1 (hCB1R) via an Escherichia coli expression system and incorporated it into nanodiscs (NDs). These hCB1R-NDs were then immobilized on a single-walled carbon nanotube field-effect transistor (swCNT-FET) to construct a bioelectronic sensing platform. This novel system can sensitively detect the cannabinoid ligand anandamide (AEA) at concentrations as low as 1 fM, demonstrating high selectivity and real-time response. It also successfully identified the hCB1R agonist Δ9-tetrahydrocannabinol and observed that the hCB1R antagonist rimonabant diminished the sensor signal upon AEA binding, indicating the antagonism-based modulation of ligand interaction. Consequently, our bioelectronic sensing platform holds potential for ligand detection and analysis of agonism and antagonism.

Acute rimonabant treatment prevents anhedonia and memory loss in rats submitted to mild restraint stress.

Stress-related disorders are becoming increasingly common and are often associated with cognitive impairments. Within this context, the endocannabinoid (ECB) system, particularly the type 1 cannabinoid (CB1) receptor, seems to play a decisive role in restoring body homeostasis. There is consistent evidence in the literature that disrupted CB1-mediated neurotransmission can ultimately contribute to stress-related diseases. Therefore, the present study aimed to evaluate the participation of CB1 receptors in the integrity of stress-induced peripheral and behavioral responses. For this purpose, male adult Wistar rats underwent physical restraint (1 h/day, for 7 days), followed by a single administration of rimonabant (CB1 receptor antagonist, 3 mg/Kg, intraperitonial) at the end of stress protocol. Animals were then subjected to evaluation of neuroendocrine responses, behavioral tests and quantification of Iba-1 (microglial) immunoreactivity in the parvocellular subdivisions of the paraventricular nucleus of the hypothalamus (PVN). No effects of restraint stress or rimonabant administration were detected on body mass variation. However, stress significantly increased adrenal relative mass and corticosterone secretion, and reduced thymus relative size. The stress effects on adrenal size and corticosterone plasma levels were absent in rimonabant-treated rats, but the thymus size was further reduced in the restraint-rimonabant group. Restraint stress also induced anhedonia, a depression-like behavior, and reduced object recognition index, indicating memory recovery impairment. Treatment with the CB1 antagonist significantly reversed stress-induced anhedonia and memory deficit. In the PVN, restraint stress reduced the number of Iba-1 positive cells in the medial parvocellular region of vehicle- but not rimonabant-treated animals. Taken together, these results indicate that the acute inhibition of the CB1-mediated endogenous pathway restores stress-induced depression-like behaviors and memory loss, suggesting a role for endocannabinoids in the neuro-immune-endocrine interplay at both peripheral and hypothalamic levels.

The molecular signature of the peripheral cannabinoid receptor 1 antagonist AM6545 in adipose, liver and muscle tissue.

The endocannabinoid system plays an important role in the regulation of metabolism, growth and regeneration of peripheral tissues, including liver, adipose and muscle tissue. Studies in cells, rodents and humans showed that cannabinoid receptor 1 (CB1) antagonist treatment is an effective strategy to improve features of metabolic health such as substrate metabolism, at least in models of metabolic dysregulation. However, acute signaling events that might induce these metabolic adaptations are not understood. It is not clear whether, and to which extent, a single treatment with a CB1 antagonist induces acute effects in peripheral, metabolic tissues. Therefore, the present study compared the phosphorylation status of signaling pathways and metabolic markers in liver, adipose and muscle tissue of mice treated with the peripherally restricted CB1 antagonist AM6545 and vehicle-treated mice. Protein kinase A phosphorylation was downregulated in white and brown adipose tissue, whereas the mitogen-activated protein kinase, phospho-extracellular signal-regulated kinase, was higher in liver, white adipose and muscle tissue of AM6545-treated mice. Additionally, Akt-mammalian target of rapamycin activation was higher in all tissues of AM6545-treated mice, whereas the phosphorylation status of metabolic markers remained unaffected. These data indicate that acute CB1 antagonism is effective to induce phosphorylation events of signaling cascades and metabolic markers in metabolic tissues of healthy, lean mice within a 90-min time window. The observed adaptations to AM6545 treatment do not fully align with earlier in vitro and in vivo findings, which could be ascribed to differences in cell type, exposure intensity (dose and time), health status and species.

Local activation of CB1 receptors by synthetic and endogenous cannabinoids dampens burst firing mode of reticular thalamic nucleus neurons in rats under ketamine anesthesia.

The reticular thalamic nucleus (RTN) is a thin shell that covers the dorsal thalamus and controls the overall information flow from the thalamus to the cerebral cortex through GABAergic projections that contact thalamo-cortical neurons (TC). RTN neurons receive glutamatergic afferents fibers from neurons of the sixth layer of the cerebral cortex and from TC collaterals. The firing mode of RTN neurons facilitates the generation of sleep-wake cycles; a tonic mode or desynchronized mode occurs during wake and REM sleep and a burst-firing mode or synchronized mode is associated with deep sleep. Despite the presence of cannabinoid receptors CB1 (CB1Rs) and mRNA that encodes these receptors in RTN neurons, there are few works that have analyzed the participation of endocannabinoid-mediated transmission on the electrical activity of RTN. Here, we locally blocked or activated CB1Rs in ketamine anesthetized rats to analyze the spontaneous extracellular spiking activity of RTN neurons. Our results show the presence of a tonic endocannabinoid input, since local infusion of AM 251, an antagonist/inverse agonist, modifies RTN neurons electrical activity; furthermore, local activation of CB1Rs by anandamide or WIN 55212-2 produces heterogeneous effects in the basal spontaneous spiking activity, where the main effect is an increase in the spiking rate accompanied by a decrease in bursting activity in a dose-dependent manner; this effect is inhibited by AM 251. In addition, previous activation of GABA-A receptors suppresses the effects of CB1Rs on reticular neurons. Our results show that local activation of CB1Rs primarily diminishes the burst firing mode of RTn neurons.

CBD Versus CBDP: Comparing In Vitro Receptor-Binding Activities.

Phytocannabinoids with seven-carbon alkyl chains (phorols) have gained a lot of attention, as they are commonly believed to be more potent versions of typical cannabinoids with shorter alkyl chains. At the time of this article, cannabidiphorol (CBDP) and tetrahydrocannabiphorol (THCP) can both be purchased in the North American market, even though their biological activities are nearly unknown. To investigate their relative potency, we conducted in vitro receptor-binding experiments with CBDP (cannabinoid CB1/CB2 receptor antagonism, serotonin 5HT-1A agonism, dopamine D2S (short form) agonism, and mu-opioid negative allosteric modulation) and compared the observed activity with that of CBD. To our knowledge, this is the first publication to investigate CBDP's receptor activity in vitro. A similar activity profile was observed for both CBD and CBDP, with the only notable difference at the CB2 receptor. Contrary to common expectations, CBD was found to be a slightly more potent CB2 antagonist than CBDP (p < 0.05). At the highest tested concentration, CBD demonstrated antagonist activity with a 33% maximum response of SR144528 (selective CB2 antagonist/inverse agonist). CBDP at the same concentration produced a weaker antagonist activity. A radioligand binding assay revealed that among cannabinoid and serotonin receptors, CB2 is likely the main biological target of CBDP. However, both CBD and CBDP were found to be significantly less potent than SR144528. The interaction of CBDP with the mu-opioid receptor (MOR) produced unexpected results. Although the cannabidiol family is considered to be a set of negative allosteric modulators (NAMs) of opioid receptors, we observed a significant increase in met-enkephalin-induced mu-opioid internalization when cells were incubated with 3 µM of CBDP and 1 µM met-enkephalin, a type of activity expected from positive allosteric modulators (PAMs). To provide a structural explanation for the observed PAM effect, we conducted molecular docking simulations. These simulations revealed the co-binding potential of CBDP (or CBD) and met-enkephalin to the MOR.

The impact of piperazine and antipsychotic co-exposures and CB1 blockade on the effects elicited by AMB-FUBINACA, a synthetic cannabinoid, in mice.

BACKGROUND & PURPOSE: The constant emergence and broad toxicological effects of synthetic cannabinoids create a discernible public health threat. The synthetic cannabinoid AMB-FUBINACA (AMB-FUB) is a potent agonist at the CB1 receptor and has been associated with numerous fatalities. Synthetic cannabinoids are commonly abused alongside other drugs and medications, including a "party pill" drug, para-fluorophenylpiperazine (pFPP), and the antipsychotic risperidone. This research aimed to investigate the mechanisms underpinning AMB-FUB toxicity and the impact of clinically relevant co-exposures in vivo. EXPERIMENTAL APPROACH: Male and female C57Bl/6 mice received a single dose of AMB-FUB (3 or 6 mg kg-1), pFPP (10 or 20 mg kg-1) or vehicle intraperitoneally. Mice were co-exposed to AMB-FUB (3 mg kg-1) and pFPP (10 mg kg-1) or risperidone (0.5 mg kg-1) to investigate these drug combinations. To study receptor-dependency and potential rescue of AMB-FUB toxicity, rimonabant (3 mg kg-1) was administered both pre- and post-AMB-FUB. Adverse effects caused by drug administration, including hypothermia and convulsions, were recorded. KEY RESULTS: AMB-FUB induced CB1-dependent hypothermia and convulsions in mice. The combination of AMB-FUB and pFPP significantly potentiated hypothermia, as did risperidone pre-treatment. Interestingly, risperidone provided significant protection from AMB-FUB-induced convulsions in female mice. Pre- and post-treatment with rimonabant was able to significantly attenuate both hypothermia and convulsions in mice administered AMB-FUB. CONCLUSION & IMPLICATIONS: Factors such as dose, CB1 signalling, and substance co-exposure significantly contribute to the toxicity of AMB-FUBINACA. Mechanistic understanding of synthetic cannabinoid toxicity and fatality can help inform overdose treatment strategies and identify vulnerable populations of synthetic cannabinoid users.

Seventeen years since rimonabant's downfall: reassessing its suicidality risk profile.

Targeting the cannabinoid type 1 receptor (CB1) is a clinically validated antiobesity therapeutic approach. The only such drug approved, rimonabant, was launched in 2006 in Europe but subsequently rejected by the US Food and Drug Administration (FDA) in 2007. The FDA cited the increased risk of suicidality in its opposition to rimonabant's approval, leading to the drug's eventual worldwide withdrawal and the abandonment of this class of therapeutics. Seventeen years later, a new class of CB1-targeting drugs is emerging, but the impact of the 2007 FDA decision remains a formidable obstacle to its clinical development. We revisit the suicidality data presented by the FDA in light of the evolution of suicidality assessment and cross-reference this with the data in the subsequently published clinical trials. We conclude that the publicly available data do not support the FDA's conclusion that the use of rimonabant was associated with an increase in the risk of suicidality.

AM6527, a neutral CB1 receptor antagonist, suppresses opioid taking and seeking, as well as cocaine seeking in rodents without aversive effects.

Preclinical research has demonstrated the efficacy of CB1 receptor (CB1R) antagonists in reducing drug-taking behavior. However, clinical trials with rimonabant, a CB1R antagonist with inverse agonist profile, failed due to severe adverse effects, such as depression and suicidality. As a result, efforts have shifted towards developing novel neutral CB1R antagonists without an inverse agonist profile for treating substance use disorders. Here, we assessed AM6527, a CB1R neutral antagonist, in addiction animal models. Our findings revealed that AM6527 did not affect cocaine self-administration under fixed-ratio reinforcement schedules but dose-dependently inhibited it under progressive-ratio reinforcement schedules. Additionally, AM6527 dose-dependently inhibited heroin self-administration under both fixed-ratio and progressive-ratio reinforcement schedules and oral sucrose self-administration under a fixed-ratio reinforcement schedule, as well as cocaine- or heroin-triggered reinstatement of drug-seeking behavior in rats. However, chronic AM6527 administration for five consecutive days significantly inhibited heroin self-administration only during the initial two days, indicating tolerance development. Notably, AM6527 did not produce rewarding or aversive effects by itself in classical electrical intracranial self-stimulation and conditioned place preference tests. However, in optical intracranial self-stimulation (oICSS) maintained by optogenetic stimulation of midbrain dopamine neurons in DAT-cre mice, both AM6527 and rimonabant dose-dependently inhibited dopamine-dependent oICSS behavior. Together, these findings suggest that AM6527 effectively reduces drug-taking and seeking behaviors without rimonabant-like adverse effects. Thus, AM6527 warrants further investigation as a potential pharmacotherapy for opioid and cocaine use disorders.

The CB1 negative allosteric modulator PSNCBAM-1 reduces ethanol self-administration via a nonspecific hypophagic effect.

Alcohol use disorder (AUD) affects >15 million people in the United States. Current pharmacotherapeutic treatments for AUD are only modestly effective, necessitating the identification of new targets for medications development. The cannabinoid receptor type 1 (CB1) has been a target of interest for the development of medications for substance use disorders and other compulsive disorders. However, CB1 antagonists/inverse agonists (e.g., rimonabant) have severe side effects that limit their clinical utility, including anxiety, depression, and suicide. Recent development of CB1 negative allosteric modulators (NAMs), including PSNCBAM-1, may provide an alternative mechanism of attenuating CB1 signaling with reduced side effects. PSNCBAM-1 has not yet been evaluated for effects in models of AUD. In this study, we investigated the effects of the CB1 NAM, PSNCBAM-1, in rodent models of AUD using adult male mice. PSNCBAM-1 dose-dependently attenuated oral ethanol self-administration (8 % w/v ethanol in water), significantly reducing ethanol rewards at a dose of 30 mg/kg, but not at 10 or 18 mg/kg. PSNCBAM-1 also dose-dependently attenuated palatable food self-administration (diluted vanilla Ensure), significantly reducing food rewards at 18 and 30 mg/kg PSNCBAM-1. PSNCBAM-1 did not affect conditioned place preference for 2 g/kg ethanol. These results suggest PSNCBAM-1 reduces ethanol-taking behavior via a nonspecific hypophagic effect and does not reduce the rewarding effects of ethanol.

CB1 Receptor Negative Allosteric Modulators as a Potential Tool to Reverse Cannabinoid Toxicity.

While the opioid crisis has justifiably occupied news headlines, emergency rooms are seeing many thousands of visits for another cause: cannabinoid toxicity. This is partly due to the spread of cheap and extremely potent synthetic cannabinoids that can cause serious neurological and cardiovascular complications-and deaths-every year. While an opioid overdose can be reversed by naloxone, there is no analogous treatment for cannabis toxicity. Without an antidote, doctors rely on sedatives, with their own risks, or 'waiting it out' to treat these patients. We have shown that the canonical synthetic 'designer' cannabinoids are highly potent CB1 receptor agonists and, as a result, competitive antagonists may struggle to rapidly reverse an overdose due to synthetic cannabinoids. Negative allosteric modulators (NAMs) have the potential to attenuate the effects of synthetic cannabinoids without having to directly compete for binding. We tested a group of CB1 NAMs for their ability to reverse the effects of the canonical synthetic designer cannabinoid JWH018 in vitro in a neuronal model of endogenous cannabinoid signaling and also in vivo. We tested ABD1085, RTICBM189, and PSNCBAM1 in autaptic hippocampal neurons that endogenously express a retrograde CB1-dependent circuit that inhibits neurotransmission. We found that all of these compounds blocked/reversed JWH018, though some proved more potent than others. We then tested whether these compounds could block the effects of JWH018 in vivo, using a test of nociception in mice. We found that only two of these compounds-RTICBM189 and PSNCBAM1-blocked JWH018 when applied in advance. The in vitro potency of a compound did not predict its in vivo potency. PSNCBAM1 proved to be the more potent of the compounds and also reversed the effects of JWH018 when applied afterward, a condition that more closely mimics an overdose situation. Lastly, we found that PSNCBAM1 did not elicit withdrawal after chronic JWH018 treatment. In summary, CB1 NAMs can, in principle, reverse the effects of the canonical synthetic designer cannabinoid JWH018 both in vitro and in vivo, without inducing withdrawal. These findings suggest a novel pharmacological approach to at last provide a tool to counter cannabinoid toxicity.

Postnatal hypofunction of N-methyl-D-aspartate receptors alters perforant path synaptic plasticity and filtering and impairs dentate gyrus-mediated spatial discrimination.

BACKGROUND AND PURPOSE: Transient hypofunction of the NMDA receptor represents a convergence point for the onset and further development of psychiatric disorders, including schizophrenia. Although the cumulative evidence indicates dysregulation of the hippocampal formation in schizophrenia, the integrity of the synaptic transmission and plasticity conveyed by the somatosensorial inputs to the dentate gyrus, the perforant pathway synapses, have barely been explored in this pathological condition. EXPERIMENTAL APPROACH: We identified a series of synaptic alterations of the lateral and medial perforant paths in animals postnatally treated with the NMDA antagonist MK-801. This dysregulation suggests decreased cognitive performance, for which the dentate gyrus is critical. KEY RESULTS: We identified alterations in the synaptic properties of the lateral and medial perforant paths to the dentate gyrus synapses in slices from MK-801-treated animals. Altered glutamate release and decreased synaptic strength precede an impairment in the induction and expression of long-term potentiation (LTP) and CB1 receptor-mediated long-term depression (LTD). Remarkably, by inhibiting the degradation of 2-arachidonoylglycerol (2-AG), an endogenous ligand of the CB1 receptor, we restored the LTD in animals treated with MK-801. Additionally, we showed for the first time, that spatial discrimination, a cognitive task that requires dentate gyrus integrity, is impaired in animals exposed to transient hypofunction of NMDA receptors. CONCLUSION AND IMPLICATIONS: Dysregulation of glutamatergic transmission and synaptic plasticity from the entorhinal cortex to the dentate gyrus has been demonstrated, which may explain the cellular dysregulations underlying the altered cognitive processing in the dentate gyrus associated with schizophrenia.

Effects of Repeated Treatment with the Monoacylglycerol Lipase Inhibitor MJN110 on Pain-Related Depression of Nesting and Cannabinoid 1 Receptor Function in Male and Female Mice.

MJN110 inhibits the enzyme monoacylglycerol lipase (MAGL) to increase levels of the endocannabinoid 2-arachidonoylglycerol , an endogenous high-efficacy agonist of cannabinoid 1 and 2 receptors (CB1/2R). MAGL inhibitors are under consideration as candidate analgesics, and we reported previously that acute MJN110 produced partial antinociception in an assay of pain-related behavioral depression in mice. Given the need for repeated analgesic administration in many pain patients and the potential for analgesic tolerance during repeated treatment, this study examined antinociceptive effects of repeated MJN110 on pain-related behavioral depression and CB1R-mediated G-protein function. Male and female ICR mice were treated daily for 7 days in a 2 × 2 design with (a) 1.0 mg/kg/d MJN110 or its vehicle followed by (b) intraperitoneal injection of dilute lactic acid (IP acid) or its vehicle as a visceral noxious stimulus to depress nesting behavior. After behavioral testing, G-protein activity was assessed in lumbar spinal cord (LSC) and five brain regions using an assay of CP55,940-stimulated [35S]GTPÉ£S activation. As reported previously, acute MJN110 produced partial but significant relief of IP acid-induced nesting depression on day 1. After 7 days, MJN110 continued to produce significant but partial antinociception in males, while antinociceptive tolerance developed in females. Repeated MJN110 also produced modest decreases in maximum levels of CP55,940-induced [35S]GTPÉ£S binding in spinal cord and most brain regions. These results indicate that repeated treatment with a relatively low antinociceptive MJN110 dose produces only partial and sex-dependent transient antinociception associated with the emergence of CB1R desensitization in this model of IP acid-induced nesting depression. SIGNIFICANCE STATEMENT: The drug MJN110 inhibits monoacylglycerol lipase (MAGL) to increase levels of the endogenous cannabinoid 2-arachidonoylglycerol and produce potentially useful therapeutic effects including analgesia. This study used an assay of pain-related behavioral depression in mice to show that repeated MJN110 treatment produced (1) weak but sustained antinociception in male mice, (2) antinociceptive tolerance in females, and (3) modest cannabinoid-receptor desensitization that varied by region and sex. Antinociceptive tolerance may limit the utility of MJN110 for treatment of pain.

Underlying mechanisms of long-term potentiation during the inhibition of the cannabinoid CB1 and GABAB receptors in the dentate gyrus of hippocampus.

BACKGROUND: The release of various neurotransmitters and thereby the excitability of neuronal circuits are regulated by the endocannabinoid system in an activity-dependent manner. Hippocampal long-term potentiation (LTP) is augmented in cannabinoid type 1 (CB1) receptor-deficient mice. CB1 receptors exist on GABAergic axon terminals in the hippocampus. In our previous work, we showed that CB1 antagonists increased the population spike (PS) amplitude, field excitatory post-synaptic potential (fEPSP), and the LTP induction in the dentate gyrus (DG) of the rat hippocampus while the GABAB antagonist decreased these parameters. Determining the underlying mechanisms of the pre- and/or postsynaptic locus of LTP expression is of great importance. In this study, we investigated whether LTP alteration acutely caused by CB1 and GABAB receptor antagonists (AM251 and CGP55845, respectively) happens at the postsynaptic or presynaptic regions, or at both. Therefore, the paired-pulse ratio (PPR) was assessed prior to and following the LTP induction in the studied groups. METHODS: Male Wistar rats were randomly assigned to the groups of control, AM251, CGP55845, CGP55845 + AM251. A high-frequency stimulation (HFS) of the perforant path (PP) was used to induce LTP in the DG region. RESULTS: Statistical analysis revealed that AM251 produced significant increase in excitatory postsynaptic potential (EPSP) slope and amplitude of PS. Conversely, administration of CGP55845 produced decrease in slope of EPSP. The current results indicated that the PPR was not influenced by LTP induction in the presence of AM251 or CGP55845 either alone or their combination. CONCLUSIONS: It can be concluded that the site causing LTP expression is, at least in part, the postsynaptic site because PPR was not influenced by LTP induction in the presence of AM251 or CGP55845 either alone or their combination.

Identification of a novel fatty acid binding protein-5-CB2 receptor-dependent mechanism regulating anxiety behaviors in the prefrontal cortex.

The endocannabinoid (eCB) system represents a promising neurobiological target for novel anxiolytic pharmacotherapies. Previous clinical and preclinical evidence has revealed that genetic and/or pharmacological manipulations altering eCB signaling modulate fear and anxiety behaviors. Water-insoluble eCB lipid anandamide requires chaperone proteins for its intracellular transport to degradation, a process that requires fatty acid-binding proteins (FABPs). Here, we investigated the effects of a novel FABP-5 inhibitor, SBFI-103, on fear and anxiety-related behaviors using rats. Acute intra-prelimbic cortex administration of SBFI-103 induced a dose-dependent anxiolytic response and reduced contextual fear expression. Surprisingly, both effects were reversed when a cannabinoid-2 receptor (CB2R) antagonist, AM630, was co-infused with SBFI-103. Co-infusion of the cannabinoid-1 receptor antagonist Rimonabant with SBFI-103 reversed the contextual fear response yet showed no reversal effect on anxiety. Furthermore, in vivo neuronal recordings revealed that intra-prelimbic region SBFI-103 infusion altered the activity of putative pyramidal neurons in the basolateral amygdala and ventral hippocampus, as well as oscillatory patterns within these regions in a CB2R-dependent fashion. Our findings identify a promising role for FABP5 inhibition as a potential target for anxiolytic pharmacotherapy. Furthermore, we identify a novel, CB2R-dependent FABP-5 signaling pathway in the PFC capable of strongly modulating anxiety-related behaviors and anxiety-related neuronal transmission patterns.

Peripheral CB1 receptor blockade acts as a memory enhancer through a noradrenergic mechanism.

Peripheral inputs continuously shape brain function and can influence memory acquisition, but the underlying mechanisms have not been fully understood. Cannabinoid type-1 receptor (CB1R) is a well-recognized player in memory performance, and its systemic modulation significantly influences memory function. By assessing low arousal/non-emotional recognition memory in mice, we found a relevant role of peripheral CB1R in memory persistence. Indeed, the peripherally-restricted CB1R specific antagonist AM6545 showed significant mnemonic effects that were occluded in adrenalectomized mice, and after peripheral adrenergic blockade. AM6545 also transiently impaired contextual fear memory extinction. Vagus nerve chemogenetic inhibition reduced AM6545-induced mnemonic effect. Genetic CB1R deletion in dopamine ß-hydroxylase-expressing cells enhanced recognition memory persistence. These observations support a role of peripheral CB1R modulating adrenergic tone relevant for cognition. Furthermore, AM6545 acutely improved brain connectivity and enhanced extracellular hippocampal norepinephrine. In agreement, intra-hippocampal ß-adrenergic blockade prevented AM6545 mnemonic effects. Altogether, we disclose a novel CB1R-dependent peripheral mechanism with implications relevant for lengthening the duration of non-emotional memory.

Neutral CB1 Receptor Antagonists as Pharmacotherapies for Substance Use Disorders: Rationale, Evidence, and Challenge.

Cannabinoid receptor 1 (CB1R) has been one of the major targets in medication development for treating substance use disorders (SUDs). Early studies indicated that rimonabant, a selective CB1R antagonist with an inverse agonist profile, was highly promising as a therapeutic for SUDs. However, its adverse side effects, such as depression and suicidality, led to its withdrawal from clinical trials worldwide in 2008. Consequently, much research interest shifted to developing neutral CB1R antagonists based on the recognition that rimonabant's side effects may be related to its inverse agonist profile. In this article, we first review rimonabant's research background as a potential pharmacotherapy for SUDs. Then, we discuss the possible mechanisms underlying its therapeutic anti-addictive effects versus its adverse effects. Lastly, we discuss the rationale for developing neutral CB1R antagonists as potential treatments for SUDs, the supporting evidence in recent research, and the challenges of this strategy. We conclude that developing neutral CB1R antagonists without inverse agonist profile may represent attractive strategies for the treatment of SUDs.

Endocannabinoid-dependent formation of columnar axonal projection in the mouse cerebral cortex.

Columnar structure is one of the most fundamental morphological features of the cerebral cortex and is thought to be the basis of information processing in higher animals. Yet, how such a topographically precise structure is formed is largely unknown. Formation of columnar projection of layer 4 (L4) axons is preceded by thalamocortical formation, in which type 1 cannabinoid receptors (CB1R) play an important role in shaping barrel-specific targeted projection by operating spike timing-dependent plasticity during development (Itami et al., J. Neurosci. 36, 7039-7054 [2016]; Kimura & Itami, J. Neurosci. 39, 3784-3791 [2019]). Right after the formation of thalamocortical projections, CB1Rs start to function at L4 axon terminals (Itami & Kimura, J. Neurosci. 32, 15000-15011 [2012]), which coincides with the timing of columnar shaping of L4 axons. Here, we show that the endocannabinoid 2-arachidonoylglycerol (2-AG) plays a crucial role in columnar shaping. We found that L4 axon projections were less organized until P12 and then became columnar after CB1Rs became functional. By contrast, the columnar organization of L4 axons was collapsed in mice genetically lacking diacylglycerol lipase α, the major enzyme for 2-AG synthesis. Intraperitoneally administered CB1R agonists shortened axon length, whereas knockout of CB1R in L4 neurons impaired columnar projection of their axons. Our results suggest that endocannabinoid signaling is crucial for shaping columnar axonal projection in the cerebral cortex.