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In the brain, hippocampal circuits are crucial for cognitive performance (e.g., memory) and deeply affected in pathological conditions (e.g., epilepsy, Alzheimer). Specialized molecular mechanisms regulate different cell types underlying hippocampal circuitries functions. Among them, cannabinoid receptors exhibit various roles depending on the cell type (e.g., neuron, glial cell) or subcellular organelle (e.g., mitochondria). Determining the site of action and precise mechanisms triggered by cannabinoid receptor activation at a local cellular and subcellular level helps us understand hippocampal pathophysiological states. In doing so, past and current research have advanced our knowledge of cannabinoid functions and proposed novel routes for potential therapeutics. By outlining these data in this work, we aim to showcase current findings and highlight the pathophysiological impact of the cannabinoid receptor type 1 (CB1) localization/activation in hippocampal circuits. This work was supported by Ikerbasque, MINECO (Ministerio de Economia y Competitividad) PGC2018-093990-A-I00 (MICIU/AEI/FEDER, UE), and Endeavour Scholarship Scheme (Malta-EU) (to E.S.-G.); Basque Government (IT1230-19), Red de Trastornos Adictivos, Instituto de Salud Carlos III (ISC-III) and European Regional Development Funds-European Union (ERDF-EU; RD16/0017/0012), Ministry of Science and Innovation (PID2019-107548RB-I00) (to P.G.).

Epilepsy is one of the most common neurological conditions. Yearly, five million people are diagnosed with epileptic-related disorders. The neuroprotective and therapeutic effect of (endo)cannabinoid compounds has been extensively investigated in several models of epilepsy. Therefore, the study of specific cell-type-dependent mechanisms underlying cannabinoid effects is crucial to understanding epileptic disorders. It is estimated that about 100 billion neurons and a roughly equal number of glial cells co-exist in the human brain. The glial population is in charge of neuronal viability, and therefore, their participation in brain pathophysiology is crucial. Furthermore, glial malfunctioning occurs in a wide range of neurological disorders. However, little is known about the impact of the endocannabinoid system (ECS) regulation over glial cells, even less in pathological conditions such as epilepsy. In this review, we aim to compile the existing knowledge on the role of the ECS in different cell types, with a particular emphasis on glial cells and their impact on epilepsy. Thus, we propose that glial cells could be a novel target for cannabinoid agents for treating the etiology of epilepsy and managing seizure-like disorders. This work was supported by the Basque Government (IT1230-19, to P.G.); MINECO/FEDER, UE (SAF2015-65034-R, to P.G.); Ministry of Science and Innovation (PID2019-107548RBI00, to P.G.); Red de Trastornos Adictivos, Instituto de Salud Carlos III (ISC-III) and European Regional Development Funds-European Union (ERDF-EU, Investing in your future; RD16/0017/0012, to P.G.); J.E.-H. is a Postdoctoral Researcher contracted with funds of Red de Trastornos Adictivos, Instituto de Salud Carlos III (ISC-III) and European Regional Development Funds-European Union (ERDF-EU, Investing in your future; RD16/0017/0012), and the Basque Government (IT1230-19). E.S.-G. is funded by Ikerbasque and MINECO (PGC2018-093990-A-I00; MICIU/AEI/FEDER, UE).

Binge drinking (BD) is a serious health concern in adolescents as high ethanol (EtOH) consumption can have cognitive sequelae later in life. Remarkably, an enriched environment (EE) in adulthood significantly recovers memory in mice after adolescent BD, and the endocannabinoid, 2-arachydonoyl-glycerol (2-AG), rescues synaptic plasticity and memory impaired in adult rodents upon adolescent EtOH intake. However, the mechanisms by which EE improves memory are unknown. We investigated this in adolescent male C57BL/6J mice exposed to a drinking in the dark (DID) procedure four days per week for a duration of 4 weeks. After DID, the mice were nurtured under an EE for 2 weeks and were subjected to the Barnes Maze Test performed the last 5 days of withdrawal. The EE rescued memory and restored the EtOH-disrupted endocannabinoid (eCB)-dependent excitatory long-term depression at the dentate medial perforant path synapses (MPP-LTD). This recovery was dependent on both the cannabinoid CB1 receptor and group I metabotropic glutamate receptors (mGluRs) and required 2-AG. Also, the EE had a positive effect on mice exposed to water through the transient receptor potential vanilloid 1 (TRPV1) and anandamide (AEA)-dependent MPP long-term potentiation (MPP-LTP). Taken together, EE positively impacts different forms of excitatory synaptic plasticity in water- and EtOH-exposed brains. This research was funded by ISCIII (“RD16/0017/0012” to P.G.), co-funded by ERDF/ESF, “Investing in your future”; The Basque Government (IT1230-19 to P.G.); Ministry of Science and Innovation (PID2019-107548RB-I00 to P.G.); Ph.D. contract from MINECO (BES-2013-065057 to S.P.); Ph.D. contract from UPV/EHU (PIF 18/315 to L.L.), and Ph.D. contract from UPV/EHU (PIF 19/164 to M.S.).

The senses dictate how the brain represents the environment, and this representation is the basis of how we act in the world [...]

SUMMARYThe idea that the effects of drugs largely depend on subcellular target location is emerging as a novel predictive factor of their beneficial or adverse outcomes. G protein-coupled type-1 cannabinoid receptors (CB1) are regulators of several brain functions as well as the main targets of cannabinoid-based medicines.Besides their classical location at plasma membranes, CB1receptors are present at different locations within cells, including in association to mitochondrial membranes (mtCB1). Here we report the generation and characterization of a mutant mouse line, which lack mtCB1receptors.

Water intake is crucial for maintaining body fluid homeostasis and animals' survival [1-4]. In the brain, complex processes trigger thirst and drinking behavior [1-5]. The anterior wall of the third ventricle formed by the subfornical organ (SFO), the median preoptic nucleus, and the organum vasculosum of the lamina terminalis (OVLT) constitute the primary structures sensing thirst signals and modulating water intake [6-10]. These subcortical regions are connected with the neocortex [11]. In particular, insular and anterior cingulate cortices (IC and ACC, respectively) have been shown to receive indirect innervations from the SFO and OVLT in rats [11] and to be involved in the control of water intake [12-15]. Type-1 cannabinoid receptors (CB1) modulate consummatory behaviors, such as feeding [16-26]. However, the role of CB1 receptors in the control of water intake is still a matter of debate [27-31]. Here, we show that endogenous activation of CB1 in cortical glutamatergic neurons of the ACC promotes water intake. Notably, presynaptic CB1 receptors of ACC glutamatergic neurons are abundantly located in the basolateral amygdala (BLA), a key area in the regulation of water intake. The selective expression of CB1 receptors in the ACC-to-BLA-projecting neurons is sufficient to stimulate drinking behavior. Moreover, chemogenetic stimulation of these projecting neurons suppresses drinking behavior, further supporting the role of this neuronal population in the control of water intake. Altogether, these data reveal a novel cortico-amygdalar mechanism involved in the regulation of drinking behavior.

SUMMARYA complex array of different inhibitory interneurons tightly controls hippocampal activity, but how such diversity specifically impacts on memory processes is scantly known. We found that a small subclass of type-1 cannabinoid receptor (CB1)-expressing hippocampal interneurons determines episodic-like memory consolidation by linking dopamine D1 receptor signaling to GABAergic transmission.Mice lacking CB1 in D1-positive cells (D1-CB1-KO) displayed impaired long-term, but not short-term, object recognition memory. Re-expression of CB1 in hippocampal, but not striatal, D1-positive cells rescued this memory impairment. Learning induced a facilitation of in vivo hippocampal long-term potentiation (LTP), which was abolished in mutant mice. Chemogenetic and pharmacological experiments revealed that both CB1-mediated memory and associated LTP facilitation involves the local control of GABAergic inhibition in a D1-dependent manner.This study reveals that CB1-/D1-expressing interneurons shape hippocampal circuits to sustain recognition memory, thereby identifying a mechanism linking the diversity of hippocampal interneurons to specific behavioral and cognitive outcomes.

Water intake is regulated by neocortical top-down circuits, but their identity and the cellular mechanisms involved are scantly known. Here, we show that endogenous activation of type-1 cannabinoid receptors (CB1) promotes water intake and that endocannabinoid modulation of excitatory projections from the anterior cingulate cortex to the basolateral amygdala is sufficient to guarantee physiological drinking. These data reveal a new circuit involved in the homeostatic control of water intake.

International audience; The lack of intrinsic motivation to engage in, and adhere to, physical exercise has major health consequences. However, the neurobiological bases of exercise motivation are still unknown. This study aimed at examining whether the endocannabinoid system (ECS) is involved in this process. To do so, we developed an operant conditioning paradigm wherein mice unlocked a running wheel with nose pokes. Using pharmacological tools and conditional mutants for cannabinoid type-1 (CB1) receptors, we provide evidence that CB1 receptors located on GABAergic neurons are both necessary and sufficient to positively control running motivation. Conversely, this receptor population proved dispensable for the modulation of running duration per rewarded sequence. Although the ECS mediated the motivation for another reward, namely palatable food, such a regulation was independent from CB1 receptors on GABAergic neurons. In addition, we report that the lack of CB1 receptors on GABAergic neurons decreases the preference for running over palatable food when mice were proposed an exclusive choice between the two rewards. Beyond providing a paradigm that enables motivation processes for exercise to be dissected either singly or in concurrence, this study is the first to our knowledge to identify a neurobiological mechanism that might contribute to sedentary behavior

International audience; The retrieval of odor-related memories shapes animal behavior. The anterior piriform cortex (aPC) is the largest part of the olfactory cortex, and it plays important roles in olfactory processing and memory. However, it is still unclear whether specific cellular mechanisms in the aPC control olfactory memory, depending on the appetitive or aversive nature of the stimuli involved. Cannabinoid-type 1 (CB1) receptors are present in the aPC (aPC-CB1), but their potential impact on olfactory memory was never explored. Here, we used a combination of behavioral, genetic, anatomical, and electrophysiological approaches to characterize the functions of aPC-CB1 receptors in the regulation of appetitive and aversive olfactory memory. Pharmacological blockade or genetic deletion of aPC-CB1 receptors specifically impaired the retrieval of conditioned odor preference (COP). Interestingly, expression of conditioned odor aversion (COA) was unaffected by local CB1 receptor blockade, indicating that the role of aPC endocannabinoid signaling is selective for retrieval of appetitive memory. Anatomical investigations revealed that CB1 receptors are highly expressed on aPC GABAergic interneurons, and ex vivo electrophysiological recordings showed that their pharmacological activation reduces miniature inhibitory post-synaptic currents (mIPSCs) onto aPC semilunar (SL), but not pyramidal principal neurons. COP retrieval, but not COA, was associated with a specific CB1-receptor-dependent decrease of mIPSCs in SL cells. Altogether, these data indicate that aPC-CB1 receptor-dependent mechanisms physiologically control the retrieval of olfactory memory, depending on odor valence and engaging modulation of local inhibitory transmission.

A complex array of different inhibitory interneurons tightly controls hippocampal activity, but how such diversity specifically impacts on memory processes is scantly known. We found that a small subclass of type-1 cannabinoid receptor (CB1)-expressing hippocampal interneurons determines episodic-like memory consolidation by linking dopamine D1 receptor signaling to GABAergic transmission. Mice lacking CB1 in D1-positive cells (D1-CB1-KO) displayed impaired long-term, but not short-term, object recognition memory. Re-expression of CB1 in hippocampal, but not striatal, D1-positive cells rescued this memory impairment. Learning induced a facilitation of in vivo hippocampal long-term potentiation (LTP), which was abolished in mutant mice. Chemogenetic and pharmacological experiments revealed that both CB1-mediated memory and associated LTP facilitation involves the local control of GABAergic inhibition in a D1-dependent manner. This study reveals that CB1-/D1-expressing interneurons shape hippocampal circuits to sustain recognition memory, thereby identifying a mechanism linking the diversity of hippocampal interneurons to specific behavioral and cognitive outcomes.

International audience; Recent advances in neuroscience have positioned brain circuits as key units in controlling behavior, implying that their positive or negative modulation necessarily leads to specific behavioral outcomes. However, emerging evidence suggests that the activation or inhibition of specific brain circuits can actually produce multimodal behavioral outcomes. This study shows that activation of a receptor at different subcellular locations in the same neuronal circuit can determine distinct behaviors. Pharmacological activation of type 1 cannabinoid (CB1) receptors in the striatonigral circuit elicits both antinociception and catalepsy in mice. The decrease in nociception depends on the activation of plasma membrane-residing CB1 receptors (pmCB1), leading to the inhibition of cytosolic PKA activity and substance P release. By contrast, mitochondrial-associated CB1 receptors (mtCB1) located at the same terminals mediate cannabinoid-induced catalepsy through the decrease in intra-mitochondrial PKA-dependent cellular respiration and synaptic transmission. Thus, subcellular-specific CB1 receptor signaling within striatonigral circuits determines multimodal control of behavior.

Astroglial type‐1 cannabinoid (CB1) receptors are involved in synaptic transmission, plasticity and behavior by interfering with the so‐called tripartite synapse formed by pre‐ and post‐synaptic neuronal elements and surrounding astrocyte processes. However, little is known concerning the subcellular distribution of astroglial CB1 receptors. In particular, brain CB1 receptors are mostly localized at cells' plasmalemma, but recent evidence indicates their functional presence in mitochondrial membranes. Whether CB1 receptors are present in astroglial mitochondria has remained unknown. To investigate this issue, we included conditional knock‐out mice lacking astroglial CB1 receptor expression specifically in glial fibrillary acidic protein (GFAP)‐containing astrocytes (GFAP‐CB1‐KO mice) and also generated genetic rescue mice to re‐express CB1 receptors exclusively in astrocytes (GFAP‐CB1‐RS). To better identify astroglial structures by immunoelectron microscopy, global CB1 knock‐out (CB1‐KO) mice and wild‐type (CB1‐WT) littermates were intra‐hippocampally injected with an adeno‐associated virus expressing humanized renilla green fluorescent protein (hrGFP) under the control of human GFAP promoter to generate GFAPhrGFP‐CB1‐KO and ‐WT mice, respectively. Furthermore, double immunogold (for CB1) and immunoperoxidase (for GFAP or hrGFP) revealed that CB1 receptors are present in astroglial mitochondria from different hippocampal regions of CB1‐WT, GFAP‐CB1‐RS and GFAPhrGFP‐CB1‐WT mice. Only non‐specific gold particles were detected in mouse hippocampi lacking CB1 receptors. Altogether, we demonstrated the existence of a precise molecular architecture of the CB1 receptor in astrocytes that will have to be taken into account in evaluating the functional activity of cannabinergic signaling at the tripartite synapse.

Summary By priming brain circuits, associations between low-salience stimuli often guide future behavioral choices through a process known as mediated or inferred learning. However, the precise neurobiological mechanisms of these incidental associations are largely unknown. Using sensory preconditioning procedures, we show that type 1 cannabinoid receptors (CB1R) in hippocampal GABAergic neurons are necessary and sufficient for mediated but not direct learning. Deletion and re-expression of CB1R in hippocampal GABAergic neurons abolishes and rescues mediated learning, respectively. Interestingly, paired presentations of low-salience sensory cues induce a specific protein synthesis-dependent enhancement of hippocampal CB1R expression and facilitate long-term synaptic plasticity at inhibitory synapses. CB1R blockade or chemogenetic manipulations of hippocampal GABAergic neurons upon preconditioning affect incidental associations, as revealed by impaired mediated learning. Thus, CB1R-dependent control of inhibitory hippocampal neurotransmission mediates incidental associations, allowing future associative inference, a fundamental process for everyday life, which is altered in major neuropsychiatric diseases. Video Abstract Download : Download video (49MB)

Cannabis-induced acute psychotic-like states (CIAPS) represent a growing health issue, but their underlying neurobiological mechanisms are poorly understood. The use of antipsychotics and benzodiazepines against CIAPS is limited by side-effects and/or by their ability to tackle only certain aspects of psychosis. Thus, safer wide-spectrum treatments are currently needed. Although the blockade of cannabinoid type-1 receptor (CB1) had been suggested as a therapeutical means against CIAPS, the use of orthosteric CB1 receptor full antagonists is strongly limited by undesired side effects and low efficacy. The neurosteroid pregnenolone has been recently shown to act as a potent endogenous allosteric signal-specific inhibitor of CB1 receptors. Thus, we tested in mice the potential therapeutic use of pregnenolone against acute psychotic-like effects of Δ9-tetrahydrocannabinol (THC), the main psychoactive component of cannabis. We found that pregnenolone blocks a wide spectrum of THC-induced endophenotypes typically associated with psychotic-like states, including impairments in cognitive functions, somatosensory gating and social interaction. In order to capture THC-induced positive psychotic-like symptoms (e.g. perceptual delusions), we adapted a behavioral paradigm based on associations between different sensory modalities and selective devaluation, allowing the measurement of mental sensory representations in mice. Acting at hippocampal CB1 receptors, THC impaired the correct processing of mental sensory representations (reality testing) in an antipsychotic- and pregnenolone-sensitive manner. Overall, this work reveals that signal-specific inhibitors mimicking pregnenolone effects can be considered as promising new therapeutic tools to treat CIAPS.

Bidirectional communication between neurons and astrocytes shapes synaptic plasticity and behavior. D-serine is a necessary co-agonist of synaptic N-methyl-D-aspartate receptors (NMDARs), but the physiological factors regulating its impact on memory processes are scantly known. We show that astroglial CB

The internal state of the organism is an important modulator of perception and behavior. The link between hunger, olfaction and feeding behavior is one of the clearest examples of these connections. At the neurobiological level, olfactory circuits are the targets of several signals (i.e. hormones and nutrients) involved in energy balance. This indicates that olfactory areas are potential sensors of the internal state of the organism. Thus, the aim of this manuscript is to review the literature showing the interplay between metabolic signals in olfactory circuits and its impact on food intake.

Cannabinoid receptor type 1 (CB1)-dependent signaling in the brain is known to modulate food intake. Recent evidence has actually shown that CB1 can both inhibit and stimulate food intake in fasting/refeeding conditions, depending on the specific neuronal circuits involved. However, the exact brain sites where this bimodal control is exerted and the underlying neurobiological mechanisms are not fully understood yet. Using pharmacological and electrophysiological approaches, we show that local CB1 blockade in the paraventricular nucleus of the hypothalamus (PVN) increases fasting-induced hyperphagia in rats. Furthermore, local CB1 blockade in the PVN also increases the orexigenic effect of the gut hormone ghrelin in animals fed ad libitum. At the electrophysiological level, CB1 blockade in slices containing the PVN potentiates the decrease of the activity of PVN neurons induced by long-term application of ghrelin. Hence, the PVN is (one of) the site(s) where signals associated with the body's energy status determine the direction of the effects of endocannabinoid signaling on food intake.

Comment in Sensory systems: the hungry sense. [Nat Rev Neurosci. 2014] Inhaling: endocannabinoids and food intake. [Nat Neurosci. 2014]; International audience; Hunger arouses sensory perception, eventually leading to an increase in food intake, but the underlying mechanisms remain poorly understood. We found that cannabinoid type-1 (CB1) receptors promote food intake in fasted mice by increasing odor detection. CB1 receptors were abundantly expressed on axon terminals of centrifugal cortical glutamatergic neurons that project to inhibitory granule cells of the main olfactory bulb (MOB). Local pharmacological and genetic manipulations revealed that endocannabinoids and exogenous cannabinoids increased odor detection and food intake in fasted mice by decreasing excitatory drive from olfactory cortex areas to the MOB. Consistently, cannabinoid agonists dampened in vivo optogenetically stimulated excitatory transmission in the same circuit. Our data indicate that cortical feedback projections to the MOB crucially regulate food intake via CB1 receptor signaling, linking the feeling of hunger to stronger odor processing. Thus, CB1 receptor-dependent control of cortical feedback projections in olfactory circuits couples internal states to perception and behavior.

Type-1 cannabinoid (CB1) and leptin (ObR) receptors regulate metabolic and astroglial functions, but the potential links between the two systems in astrocytes were not investigated so far. Genetic and pharmacological manipulations of CB1 receptor expression and activity in cultured cortical and hypothalamic astrocytes demonstrated that cannabinoid signaling controls the levels of ObR expression. Lack of CB1 receptors also markedly impaired leptin-mediated activation of signal transducers and activators of transcription 3 and 5 (STAT3 and STAT5) in astrocytes. In particular, CB1 deletion determined a basal overactivation of STAT5, thereby leading to the downregulation of ObR expression, and leptin failed to regulate STAT5-dependent glycogen storage in the absence of CB1 receptors. These results show that CB1 receptors directly interfere with leptin signaling and its ability to regulate glycogen storage, thereby representing a novel mechanism linking endocannabinoid and leptin signaling in the regulation of brain energy storage and neuronal functions.

Background We have shown that the endogenous stimulation of cannabinoid type-1 (CB 1 ) receptors is a prerequisite for voluntary running in mice, but the precise mechanisms through which the endocannabinoid system exerts a tonic control on running performance remain unknown. Methods We analyzed the respective impacts of constitutive/conditional CB 1 receptor mutations and of CB 1 receptor blockade on wheel-running performance. We then assessed the consequences of ventral tegmental area (VTA) CB 1 receptor blockade on the wheel-running performances of wildtype (gamma-aminobutyric acid [GABA]- CB 1 +/+ ) and mutant (GABA- CB 1 –/– ) mice for CB 1 receptors in brain GABA neurons. Using in vivo electrophysiology, the consequences of wheel running on VTA dopamine (DA) neuronal activity were examined in GABA- CB 1 +/+ and GABA- CB 1 –/– mice. Results Conditional deletion of CB 1 receptors from brain GABA neurons, but not from several other neuronal populations or from astrocytes, decreased wheel-running performance in mice. The inhibitory consequences of either the systemic or the intra-VTA administration of CB 1 receptor antagonists on running behavior were abolished in GABA- CB 1 –/– mice. The absence of CB 1 receptors from GABAergic neurons led to a depression of VTA DA neuronal activity after acute/repeated wheel running. Conclusions This study provides evidence that CB 1 receptors on VTA GABAergic terminals exert a permissive control on rodent voluntary running performance. Furthermore, it is shown that CB 1 receptors located on GABAergic neurons impede negative consequences of voluntary exercise on VTA DA neuronal activity. These results position the endocannabinoid control of inhibitory transmission as a prerequisite for wheel-running performance in mice.

International audience; Activation of type 1 cannabinoid receptors (CB1R) decreases GABA and glutamate release in cortical and subcortical regions, with complex outcomes on cortical network activity. To date there have been few attempts to disentangle the region-and cell-specific mechanisms underlying the effects of cannabinoids on cortical network activity in vivo. Here we addressed this issue by combining in vivo electrophysiological recordings with local and systemic pharmacological manipulations in conditional mutant mice lacking CB1R expression in different neuronal populations. First we report that cannabinoids induce hypersynchronous thalamocortical oscillations while decreasing the amplitude of faster cortical oscillations. Then we demonstrate that CB1R at striatonigral synapses (basal ganglia direct pathway) mediate the thalamocortical hypersynchrony, whereas activation of CB1R expressed in cortical glutamatergic neurons decreases cortical synchrony. Finally we show that activation of CB1 expressed in cortical glutamatergic neurons limits the cannabinoid-induced thalamocortical hypersynchrony. By reporting that CB1R activations in cortical and subcortical regions have contrasting effects on cortical synchrony, our study bridges the gap between cellular and in vivo network effects of cannabinoids. Incidentally, the thalamocortical hypersynchrony we report suggests a potential mechanism to explain the sensory "high" experienced during recreational consumption of marijuana.

Cellular activity in the brain depends on the high energetic support provided by mitochondria, the cell organelles which use energy sources to generate ATP. Acute cannabinoid intoxication induces amnesia in humans and animals, and the activation of type-1 cannabinoid receptors present at brain mitochondria membranes (mtCB1) can directly alter mitochondrial energetic activity. Although the pathological impact of chronic mitochondrial dysfunctions in the brain is well established, the involvement of acute modulation of mitochondrial activity in high brain functions, including learning and memory, is unknown. Here, we show that acute cannabinoid-induced memory impairment in mice requires activation of hippocampal mtCB1 receptors. Genetic exclusion of CB1 receptors from hippocampal mitochondria prevents cannabinoid-induced reduction of mitochondrial mobility, synaptic transmission and memory formation. mtCB1 receptors signal through intra-mitochondrial Gαi protein activation and consequent inhibition of soluble-adenylyl cyclase (sAC). The resulting inhibition of protein kinase A (PKA)-dependent phosphorylation of specific subunits of the mitochondrial electron transport system eventually leads to decreased cellular respiration. Hippocampal inhibition of sAC activity or manipulation of intra-mitochondrial PKA signalling or phosphorylation of the Complex I subunit NDUFS2 inhibit bioenergetic and amnesic effects of cannabinoids. Thus, the G protein-coupled mtCB1 receptors regulate memory processes via modulation of mitochondrial energy metabolism. By directly linking mitochondrial activity to memory formation, these data reveal that bioenergetic processes are primary acute regulators of cognitive functions.

This study has investigated the role of hypothalamic and amygdalar type-1 cannabinoid (CB1) receptors in the emotional and neuroendocrine responses to stress. To do so, we used the Cre/loxP system to generate conditional mutant mice lacking the CB1 gene in neurons expressing the transcription factor single-minded 1 (Sim1). This choice was dictated by former evidence for Sim1-Cre transgenic mice bearing Cre activity in all areas expressing Sim1, which chiefly includes the hypothalamus (especially the paraventricular nucleus, the supraoptic nucleus, and the posterior hypothalamus) and the mediobasal amygdala. Genomic DNA analyses in Sim1-CB1(-/-) mice indicated that the CB1 allele was excised from the hypothalamus and the amygdala, but not from the cortex, the striatum, the thalamus, the nucleus accumbens, the brainstem, the hippocampus, the pituitary gland, and the spinal cord. Double-fluorescent in situ hybridization experiments further indicated that Sim1-CB1(-/-) mice displayed a weaker CB1 receptor mRNA expression in the paraventricular nucleus of the hypothalamus and the mediobasal part of the amygdala, compared to wild-type animals. Individually housed Sim1-CB1(-/-) mice and their Sim1-CB1(+/+) littermates were exposed to anxiety and fear memory tests under basal conditions as well as after acute/repeated social stress. A principal component analysis of the behaviors of Sim1-CB1(-/-) and Sim1-CB1(+/+) mice in anxiety tests (open field, elevated plus-maze, and light/dark box) revealed that CB1 receptors from Sim1-expressing neurons exert tonic, albeit opposite, controls of locomotor and anxiety reactivity to novel environments. No difference between genotypes was observed during the recall of contextual fear conditioning or during active avoidance learning. Sim1-CB1(-/-), but not Sim1-CB1(+/+), mice proved sensitive to an acute social stress as this procedure reverted the increased ambulation in the center of the open field. The stimulatory influence of repeated social stress on body and adrenal weights, water intake, and sucrose preference was similar in the two genotypes. On the other hand, repeated social stress abolished the decrease in cued-fear conditioned expression that was observed in Sim1-CB1(-/-) mice, compared to Sim1-CB1(+/+) mice. This study suggests that CB1 receptors located on Sim1-expressing neurons exert a tonic control on locomotor reactivity, unconditioned anxiety, and cued-fear expression under basal conditions as well as after acute or repeated stress.

The central nervous system control of food intake has been extensively studied, hence, several neurotransmitter systems regulating this function are now clearly identified, for example, the endocannabinoid and serotoninergic systems. The former stimulates feeding while the latter inhibits it. Oleamide (Ole) is a cannabimimetic molecule affecting both systems. In this work, we tested the orexigenic and anorectic potential of Ole when administered into the nucleus accumbens shell (NAcS), a brain region that has been related to the orexigenic effects of cannabinoids. Additionally, we tested if Ole administered into this nucleus affects the activity of the hypothalamic nuclei involved in feeding behaviour, just as other cannabinoids do. We found a hyperphagic effect of Ole that is mediated through CB1 activation. The combination of Ole and the CB1 antagonist, AM251, produced a hypophagia that was fully blocked by SB212084, a 5-HT2C receptor antagonist. We also show that blockade of 5-HT2C and 5-HT2A receptors in the NAcS stimulates food intake. Finally, the combination of Ole and AM251 activates hypothalamic nuclei, an effect also blocked by SB242084. In conclusion, we show, for the first time, that Ole administered into the NAcS has a dual effect on feeding behaviour, acting through cannabinoid and serotonin receptors. These effects probably result from a downstream interaction with the hypothalamus.

International audience; Bidirectional communication between neurons and astrocytes shapes synaptic plasticity and behavior. D-serine is a necessary co-agonist of synaptic N-methyl-D-aspartate receptors (NMDARs), but the physiological factors regulating its impact on memory processes are scantly known. We show that astroglial CB1 receptors are key determinants of object recognition memory by determining the availability of D-serine at hippocampal synapses. Mutant mice lacking CB1 receptors from astroglial cells (GFAP-CB1-KO) displayed impaired object recognition memory and decreased in vivo and in vitro long-term potentiation (LTP) at CA3-CA1 hippocampal synapses. Activation of CB1 receptors increased intracellular astroglial Ca2+ levels and extracellular levels of D-serine in hippocampal slices. Accordingly, GFAP-CB1-KO displayed lower occupancy of the co-agonist binding site of synaptic hippocampal NMDARs. Finally, elevation of D-serine levels fully rescued LTP and memory impairments of GFAP-CB1-KO mice. These data reveal a novel mechanism of in vivo astroglial control of memory and synaptic plasticity via the D-serine-dependent control of NMDARs.

The dorsolateral striatum (DLS) has been implicated in the learning of habits and procedural memories. Extinction of this kind of memories has been poorly studied. The DLS expresses high levels of the cannabinergic receptor one (CB1), and, lately, it has been suggested that the activation of CB1 in this structure is indispensable for long-term depression (LTD) development. We performed experiments in a T-maze and evaluated the effects of intrastriatal and intrahipocampal administration of the CB1 antagonist AM251 on extinction and on c-Fos expression. We also administered anandamide to evaluate if an artificial increase of endocannabinoids facilitates extinction. Our results indicate clearly a dose-response blockade of extinction induced by AM251 injected into the striatum but a facilitation of extinction when administered into the hippocampus. Anandamide did not induce any observable changes. AM251 effects were accompanied by an increase in c-Fos immunoreactivity in the DLS and its decrease in the hippocampal region, suggesting that the activation of CB1 in the striatum is necessary for the extinction of procedural memories. These findings could be important in some neurological conditions, such as obsessive-compulsive disorder in which striatal activity seems to be abnormal.

One of the adaptive abilities of the brain is the generation of a strategy to optimize acquisition of information, i.e., learning. In this study, we explored the role of environmental conditions (the light–dark cycle) and of the endocannabinoid anandamide in rats to select a strategy to solve the Barnes maze (BM). To determine the effects of manipulating the cannabinergic system on a spatial task in relation to the light–dark cycle. Rats received an intrahippocampal or intrastriatal administration of anandamide, AM251, or their combination at two different points of the light–dark cycle (1300 and 0100 hours), and their performance in the BM was evaluated. In addition, we determined the expression of the cannabinoid 1 receptor (CB1R) in the hippocampus and striatum throughout the light–dark cycle. Results indicate that rats solved the BM by using a spatial strategy during the light phase and a procedural (serial) strategy during the dark phase of the cycle. CB1R expression varied in the hippocampus, being higher at 1300 hours and lower at 0100 hours, whereas its expression remained unchanged in the striatum. Changes in the brain, which include changes in the endocannabinoid system, prompt it to use different strategies (spatial and procedural, or others not evaluated in this study) to cope with the environmental demands. These cerebral changes are adaptive responses to the light–dark cycle.

Background and purpose: Evidence indicates that the endocannabinoid, 2-arachidonoylglycerol (2-AG), increases food intake when injected into the nucleus accumbens shell (NAcS), thereby potentially activating hypothalamic nuclei involved in food intake regulation. We aimed to evaluate potential orexigenic effects of the endocannabinoid anandamide and of AA5HT, a fatty acid amide hydrolase (FAAH) inhibitor, and OMDM-1, an inhibitor of anandamide uptake, injected in the NAcS, as well as the effect of these treatments on activation of hypothalamic nuclei.

The CB1 cannabinoid receptor, the main molecular target of endocannabinoids and cannabis active components, is the most abundant G protein-coupled receptor in the mammalian brain. Of note, CB1 receptors are expressed at the synapses of two opposing (i.e., GABAergic/inhibitory and glutamatergic/excitatory) neuronal populations, so the activation of one and/or another receptor population may conceivably evoke different effects. Despite the widely reported neuroprotective activity of the CB1 receptor in animal models, the precise pathophysiological relevance of those two CB1 receptor pools in neurodegenerative processes is unknown. Here, we first induced excitotoxic damage in the mouse brain by (i) administering quinolinic acid to conditional mutant animals lacking CB1 receptors selectively in GABAergic or glutamatergic neurons, and (ii) manipulating corticostriatal glutamatergic projections remotely with a designer receptor exclusively activated by designer drug pharmacogenetic approach. We next examined the alterations that occur in the R6/2 mouse, a well-established model of Huntington disease, upon (i) fully knocking out CB1 receptors, and (ii) deleting CB1 receptors selectively in corticostriatal glutamatergic or striatal GABAergic neurons. The data unequivocally identify the restricted population of CB1 receptors located on glutamatergic terminals as an indispensable player in the neuroprotective activity of (endo)cannabinoids, therefore suggesting that this precise receptor pool constitutes a promising target for neuroprotective therapeutic strategies.

Complex interactions between periphery and the brain regulate food intake in mammals. Cannabinoid type-1 (CB 1 ) receptor antagonists are potent hypophagic agents, but the sites where this acute action is exerted and the underlying mechanisms are not fully elucidated. To dissect the mechanisms underlying the hypophagic effect of CB 1 receptor blockade, we combined the acute injection of the CB 1 receptor antagonist rimonabant with the use of conditional CB 1 -knockout mice, as well as with pharmacological modulation of different central and peripheral circuits. Fasting/refeeding experiments revealed that CB 1 receptor signaling in many specific brain neurons is dispensable for the acute hypophagic effects of rimonabant. CB 1 receptor antagonist-induced hypophagia was fully abolished by peripheral blockade of β-adrenergic transmission, suggesting that this effect is mediated by increased activity of the sympathetic nervous system. Consistently, we found that rimonabant increases gastrointestinal metabolism via increased peripheral β-adrenergic receptor signaling in peripheral organs, including the gastrointestinal tract. Blockade of both visceral afferents and glutamatergic transmission in the nucleus tractus solitarii abolished rimonabant-induced hypophagia. Importantly, these mechanisms were specifically triggered by lipid-deprivation, revealing a nutrient-specific component acutely regulated by CB 1 receptor blockade. Finally, peripheral blockade of sympathetic neurotransmission also blunted central effects of CB 1 receptor blockade, such as fear responses and anxiety-like behaviors. These data demonstrate that, independently of their site of origin, important effects of CB 1 receptor blockade are expressed via activation of peripheral sympathetic activity. Thus, CB 1 receptors modulate bidirectional circuits between the periphery and the brain to regulate feeding and other behaviors.

To maximize their chances of survival, animals need to rapidly and efficiently respond to aversive situations. These responses can be classified as active or passive and depend on the specific nature of threats, but also on individual fear coping styles. In this study, we show that the control of excitatory and inhibitory brain neurons by type-1 cannabinoid (CB1) receptors is a key determinant of fear coping strategies in mice. In classical fear conditioning, a switch between initially predominant passive fear responses (freezing) and active behaviors (escape attempts and risk assessment) develops over time. Constitutive genetic deletion of CB1receptors inCB1−/−mice disrupted this pattern by favoring passive responses. This phenotype can be ascribed to endocannabinoid control of excitatory neurons, because it was reproduced in conditional mutant mice lacking CB1receptors from cortical glutamatergic neurons. CB1receptor deletion from GABAergic brain neurons led to the opposite phenotype, characterized by the predominance of active coping. The CB1receptor agonist Δ9-tetrahydrocannabinol exerted a biphasic control of fear coping strategies, with lower and higher doses favoring active and passive responses, respectively. Finally, viral re-expression of CB1receptors in the amygdala ofCB1−/−mice restored the normal switch between the two coping strategies. These data strongly suggest that CB1receptor signaling bimodally controls the spontaneous adoption of active or passive coping strategies in individuals. This primary function of the endocannabinoid system in shaping individual behavioral traits should be considered when studying the mechanisms of physiological and pathological fear.

Oleoylethanolamide (OEA) is an endogenous molecule related to endocannabinoids (eCBs) that induces satiety. It binds to the peroxisome-proliferator-activated receptor alpha (PPAR alpha). PPAR alpha is involved in feeding regulation and it has been proposed to play a role in sleep modulation. The objective of the present work is to show if this molecule modifies the sleep-waking cycle through central mechanisms. We have found that the peripheral administration of OEA reduces food intake and increases waking with a concomitant reduction of rapid eye movement sleep. Additionally, this treatment produces deactivation of the lateral hypothalamus, as inferred from the c-Fos expression evaluation. Finally, intra-lateral hypothalamus injection of OEA has mirrored the effects induced by this molecule when it is peripherally administered. In conclusion, we show for the very first time that OEA can modify the sleep-waking cycle and food intake, apparently mediated by the lateral hypothalamus.

A nascent literature has postulated endocannabinoids (eCBs) as strong sleep-inducing lipids, particularly rapid-eye-movement sleep (REMs), nevertheless the exact mechanisms behind this effect remain to be determined. Anandamide and 2-arachidonyl glycerol, two of the most important eCBS, are synthesized in the hippocampus. This structure also expresses a high concentration of cannabinoid receptor 1 (CB1). Recent extensive literature supports eCBs as important regulators of hippocampal activity. It has also been shown that these molecules vary their expression on the hippocampus depending on the light-dark cycle. In this context we decided to analyze the effect of intrahippocampal administration of the eCB anandamide (ANA) on the sleep-waking cycle at two points of the light-dark cycle. Our data indicate that the administration of ANA directly into the hippocampus increases REMs in a dose dependent manner during the dark but not during the light phase of the cycle. The increase of REMs was blocked by the CB1 antagonist AM251. This effect was specific for the hippocampus since ANA administrations in the surrounding cortex did not elicit any change in REMs. These results support the idea of a direct relationship between hippocampal activity and sleep mechanisms by means of eCBs. The data presented here show, for the first time that eCBs administered into the hippocampus trigger REMs and support previous studies where chemical stimulation of limbic areas triggered sleep.

It has been suggested that sleep has a restorative function; however, experimental support is limited. Hence, we investigated whether changes in the level of antiapoptotic BCL-2 protein and proapoptotic BAX protein occur during sleep deprivation (SD) and sleep rebound, and evaluated the spontaneous changes in these proteins, along the light–dark cycle, in the adult male Wistar rat. Estimations were made in the prefrontal cortex, hippocampus, striatum, and pons. We observed that BCL-2 exhibited diurnal variations in the prefrontal cortex and striatum, whereas BAX varied in the striatum and showed only small variations in the pons as measured by immunoblotting. The BCL-2/BAX ratio exhibited diurnal variations in the prefrontal cortex and striatum. BCL-2 and BAX levels were affected by 24 hr of total SD and 24 hr of sleep rebound. SD decreased the BCL-2/BAX ratio in the prefrontal cortex and pons. Sleep rebound increased the BCL-2/BAX ratio in the hippocampus. In conclusion, the BCL-2/BAX ratio is high during the dark phase as compared with the light phase in the prefrontal cortex and during the light phase as compared with the dark phase in the striatum. SD decreased the BCL-2/BAX ratio in the prefrontal cortex and pons, whereas sleep rebound increased it in the hippocampus. These changes point out structures in the brain that express these proteins as a response to the light–dark cycle. Similarly, SD and sleep rebound seem to change these proteins expression in some other brain structures, suggesting that cellular vulnerability might be altered by these changes. © 2009 Wiley-Liss, Inc.

International audience; Expression of aversive memories is key for survival, but the underlying brain mechanisms are not fully understood. Medial habenular (MHb) axons corelease glutamate and acetylcholine onto target postsynaptic interpeduncular (IPN) neurons, but their role in aversive memories has not been addressed so far. We found that cannabinoid type 1 receptors (CB1R), key regulators of aversive responses, are present at presynaptic terminals of MHb neurons in the IPN. Conditional deletion of CB1R from MHb neurons reduces fear-conditioned freezing and abolishes conditioned odor aversion in mice, without affecting neutral or appetitively motivated memories. Interestingly, local inhibition of nicotinic, but not glutamatergic receptors in the target region IPN before retrieval, rescues these phenotypes. Finally, optogenetic electrophysiological recordings of MHb-to-IPN circuitry revealed that blockade of CB1R specifically enhances cholinergic, but not glutamatergic, neurotransmission. Thus, presynaptic CB1R control expression of aversive memories by selectively modulating cholinergic transmission at MHb synapses in the IPN.

Olfaction has a direct influence on behavior and cognitive processes. There are different neuromodulatory systems in olfactory circuits that control the sensory information flowing through the rest of the brain. The presence of the cannabinoid type-1 (CB1) receptor, (the main cannabinoid receptor in the brain), has been shown for more than 20 years in different brain olfactory areas. However, only over the last decade have we started to know the specific cellular mechanisms that link cannabinoid signaling to olfactory processing and the control of behavior. In this review, we aim to summarize and discuss our current knowledge about the presence of CB1 receptors, and the function of the endocannabinoid system in the regulation of different olfactory brain circuits and related behaviors. This research was funded by Fondation pour la Recherche Médicale (FRM, FDT20170436845) (to G.T.); The Basque Government (ITI230-19), Red de Trastornos Adictivos, Instituto de Salud Carlos III (ISC-III) and European Regional Development Funds-European Union (ERDF-EU; grant RD16/0017/0012), MINECO/FEDER, UE (SAF2015-65034-R) (to P.G.); EU–FP7 (PAINCAGE, HEALTH-603191), European Research Council (Endofood, ERC–2010–StG–260515; CannaPreg, ERC-2014-PoC-640923, Micabra) (to G.M.); Ikerbasque (The Basque Foundation for Science) and MINECO (Ministerio de Economía y Competitividad) PGC2018-093990-A-I00 (to E.S.-G.).