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We identified three types of monosynaptic cholinergic inputs spatially arranged onto medial substantia nigra dopaminergic neurons in male and female mice: cotransmitted acetylcholine (ACh)/GABA, GABA-only, and ACh only. There was a predominant GABA-only conductance along lateral dendrites and soma-centered ACh/GABA cotransmission. In response to repeated stimulation, the GABA conductance found on lateral dendrites decremented less than the proximally located GABA conductance, and was more effective at inhibiting action potentials. While soma-localized ACh/GABA cotransmission showed depression of the GABA component with repeated stimulation, ACh-mediated nicotinic responses were largely maintained. We investigated whether this differential change in inhibitory/excitatory inputs leads to altered neuronal excitability. We found that a depolarizing current or glutamate preceded by cotransmitted ACh/GABA was more effective in eliciting an action potential compared with current, glutamate, or ACh/GABA alone. This enhanced excitability was abolished with nicotinic receptor inhibitors, and modulated by T- and L-type calcium channels, thus establishing that activity of multiple classes of ion channels integrates to shape neuronal excitability.SIGNIFICANCE STATEMENTOur laboratory has previously discovered a population of substantia nigra dopaminegic neurons (DA) that receive cotransmitted ACh and GABA. This study used subcellular optogenetic stimulation of cholinergic presynaptic terminals to map the functional ACh and GABA synaptic inputs across the somatodendritic extent of substantia nigra DA neurons. We determined spatially clustered GABA-only inputs on the lateral dendrites while cotransmitted ACh and GABA clustered close to the soma. We have shown that the action of GABA and ACh in cotransmission spatially clustered near the soma play a critical role in enhancing glutamate-mediated neuronal excitability through the activation of T- and L-type voltage-gated calcium channels.

The alteration of the endocannabinoid tone usually associates with changes in the expression and/or function of the cannabinoid CB

Activation of cannabinoid 1 receptors (CB1 R) modulates multiple behaviours, including exploration, motor coordination and response to psychostimulants. It is known that CB1 R expressed by either excitatory or inhibitory neurons mediates different behavioural responses to CB1 R activation, yet the involvement of CB1 R expressed by medium spiny neurons (MSNs), the neuronal subpopulation that expresses the highest level of CB1 R in the CNS, remains unknown. We report a new genetically modified mouse line that expresses functional CB1 R in MSN on a CB1 R knockout (KO) background (CB1 R(MSN) mice). The absence of cannabimimetic responses measured in CB1 R KO mice was not rescued in CB1 R(MSN) mice, nor was decreased spontaneous locomotion, impaired instrumental behaviour or reduced amphetamine-triggered hyperlocomotion measured in CB1 R KO mice. Significantly, reduced novel environment exploration of an open field and absence of amphetamine sensitization (AS) measured in CB1 R KO mice were fully rescued in CB1 R(MSN) mice. Impaired motor coordination in CB1 R KO mice measured on the Rotarod was partially rescued in CB1 R(MSN) mice. Thus, CB1 R expressed by MSN control exploration, motor coordination, and AS. Our study demonstrates a new functional roles for cell specific CB1 R expression and their causal link in the control of specific behaviors.

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.).

The use and abuse of cannabis can be associated with significant pathophysiology, however, it remains unclear whether (1) acute administration of Δ-9-tetrahydrocannabinol (THC) during early adulthood alters the cannabinoid type 1 (CB1 ) receptor localization and expression in cells of the brain, and (2) THC produces structural brain changes. Here we use electron microscopy and a highly sensitive pre-embedding immunogold method to examine CB1 receptors in the hippocampus cornu ammonis subfield 1 (CA1) 30 min after male mice were exposed to a single THC injection (5 mg/kg). The findings show that acute exposure to THC can significantly decrease the percentage of CB1 receptor immunopositive terminals making symmetric synapses, mitochondria, and astrocytes. The percentage of CB1 receptor-labeled terminals forming asymmetric synapses was unaffected. Lastly, CB1 receptor expression was significantly lower at terminals of symmetric and asymmetric synapses as well as in mitochondria. Structurally, CA1 dendrites were significantly larger, and contained more spines and mitochondria following acute THC administration. The area of the dendritic spines, synaptic terminals, mitochondria, and astrocytes decreased significantly following acute THC exposure. Altogether, these results indicate that even a single THC exposure can have a significant impact on CB1 receptor expression, and can alter CA1 ultrastructure, within 30 min of drug exposure. These changes may contribute to the behavioral alterations experienced by young individuals shortly after cannabis intoxication.

Omega-3 fatty acids (FA) are essential long-chain polyunsaturated FA, amongst others, α-linolenic acid (ALA), eicosapentaenoic acid (EPA) and docosahexaenoic acid (DHA). The main food source of omega-3 is the oily fish which is found in salmon, anchovy or tuna. A diet enriched with omega-3 is known to favour healthy states by promoting molecular and functional changes during brain damage recovery, membranes fluidity, energy metabolism regulation, release of signalling molecules or gene expression. Likewise, the activation of signalling pathways by omega-3 improves neural transmission and plasticity and decreases oxidative stress and inflammation in cells, particularly in neurons. Therefore, omega-3 supplements have been used to prevent or treat many human disorders. This review is intended to provide the stateof- the art of omega-3 as a natural component with beneficial therapeutic properties in cardiovascular and neurodegenerative diseases (Alzheimer and Parkinson), cancer, alcoholism and overweight. Lastly, some insights into the potential benefits of omega-3 supplementation to dodge or treat some other diseases in the future are also considered.; Kate luzeko omega-3 mantenugaiak, azido α-linolenikoa (ALA), azido eikosapentaenoikoa (EPA) eta azido dokosahexaenoikoa (DHA), dietaren bitartez bereganatzen diren gantz-azido (GA) poliasegabeak dira. Propietate antioxidatzaileak barne hartzen dituzten hiru osagai horien elikagai-iturri nagusia arrain koipetsua (izokina, antxoa, hegalaburra…) eta horretatik eratorritako arrain-olioa dira batez ere. Omega-3 GA osagarriaz aberastutako dietak aldaketa molekular zein funtzional mesedegarriak eragiten ditu garunaren garapen prozesuan, zenbait garun lesioren berreskurapenean parte hartzen. Gehigarri horrek mintz zelularraren fluidotasuna areagotzen du, eta metabolismoaren erregulazioan parte hartzen du, seinaleztapen molekulen askapena sustatuz eta gene espresioan eraginez. Bi ekintza horien bidez seinaleztapen bideak aktibatzen dira, eta ondorioz garun plastikotasuna eta transmisio sinaptikoa suspertu. Areago, omega-3 GAk zeluletan oro har, eta neuronetan bereziki, oxidazio-estresak eta hanturak eragindako kalteak murriztu ditzake. Horregatik guztiagatik, omega-3 osagarria hainbat patologietan prebentzioan edo tratamenduan erabili da. Berrikuspen honek laburbiltzen ditu kate luzeko omega-3 GAetan aberastutako tratamenduak bihotz hodietako gaixotasunetan, minbizian, neuroendekapenezko gaixotasunetan (Alzheimer eta Parkinson), alkoholismoan eta gainpisuan, oinarrizko ikerkuntzan eta ikerketa klinikoan frogatu eta egiaztatu diren aurrerapen terapeutiko berriak; eta etorkizunera begira beste hainbat gaixotasuni aurrea hartzeko edo haiek tratatzeko potentzialtasun handiko eta albo ondoriorik gabeko osagarri ez-inbaditzaile aproposa izan daitekeela iradokitzen du.

Cannabinoid receptors are widely expressed throughout the hippocampal formation, but are particularly dense in the dentate gyrus (DG) subregion. We, and others, have shown in mice that cannabinoid type 1 receptors (CB1Rs) are involved in a long-term depression (LTD) that can be induced by prolonged 10 Hz stimulation of the medial perforant path (MPP)-granule cell synaptic input to the DG. Here, we extend this work to examine the involvement of CB1Rs in other common forms of LTD in the hippocampus of juvenile male and female Sprague–Dawley rats (Rattus norvegicus). We found, as in mice, that prolonged 10 Hz stimulation (6000 pulses) could reliably induce a form of LTD that was dependent upon CB1R activation. In addition, we also discovered a role for both CB1R and mGluR proteins in LTD induced with 1 Hz low-frequency stimulation (1 Hz-LTD; 900 pulses) and in LTD induced by bath application of the group I mGluR agonist (RS)-3,5-Dihydroxyphenylglycine (DHPG; DHPG-LTD). This study elucidates an essential role for endocannabinoid receptors in a number of forms of LTD in the rat DG, and identifies a novel role for CB1Rs as potential therapeutic targets for conditions that involve impaired LTD in the DG.

Astrocytes take up glucose from the bloodstream to provide energy to the brain, thereby allowing neuronal activity and behavioural responses1,2,3,4,5. By contrast, astrocytes are under neuronal control through specific neurotransmitter receptors5,6,7. However, whether the activation of astroglial receptors can directly regulate cellular glucose metabolism to eventually modulate behavioural responses is unclear. Here we show that activation of mouse astroglial type-1 cannabinoid receptors associated with mitochondrial membranes (mtCB1) hampers the metabolism of glucose and the production of lactate in the brain, resulting in altered neuronal functions and, in turn, impaired behavioural responses in social interaction assays. Specifically, activation of astroglial mtCB1 receptors reduces the phosphorylation of the mitochondrial complex I subunit NDUFS4, which decreases the stability and activity of complex I. This leads to a reduction in the generation of reactive oxygen species by astrocytes and affects the glycolytic production of lactate through the hypoxia-inducible factor 1 pathway, eventually resulting in neuronal redox stress and impairment of behavioural responses in social interaction assays. Genetic and pharmacological correction of each of these effects abolishes the effect of cannabinoid treatment on the observed behaviour. These findings suggest that mtCB1 receptor signalling can directly regulate astroglial glucose metabolism to fine-tune neuronal activity and behaviour in mice., This work was funded by: INSERM, the European Research Council (Endofood, ERC–2010–StG–260515 and CannaPreg, ERC-2014-PoC-640923, MiCaBra, ERC-2017-AdG-786467), Fondation pour la Recherche Medicale (FRM, DRM20101220445), the Human Frontiers Science Program, Region Nouvelle Aquitaine and Agence Nationale de la Recherche (ANR; NeuroNutriSens ANR-13-BSV4-0006, CaCoVi ANR 18-CE16-0001-02, MitObesity ANR 18-CE14-0029-01, ORUPS ANR-16-CE37-0010-01 and BRAIN ANR-10-LABX-0043) (to G.M.); NIH/NIDA (1R21DA037678-01), Spanish Ministry of Science, Innovation and Universities (MCINU/FEDER; grants SAF2016-78114-R and RED2018-102576-T), Instituto de Salud Carlos III (CB16/10/00282), an EU BATCure grant (666918), Junta de Castilla y León (Escalera de Excelencia CLU-2017-03), Ayudas Equipos Investigación Biomedicina 2017 Fundación BBVA and Fundación Ramón Areces (to J.P.B.); Instituto de Salud Carlos III (PI18/00285; RD16/0019/0018), the European Regional Development Fund, the European Union’s Horizon 2020 Research and Innovation Programme (grant agreement 686009), Junta de Castilla y León (IES007P17) and Fundación Ramón Areces to (A.A.); French State/ANR/IdEx (ANR-10-IDEX-03-02), Eu-Fp7 (FP7-PEOPLE-2013-IEF-623638) and Ramon y Cajal Investigator Program (RYC-2017-21776) (to A.B.-G.); FRM (SPF20121226369) (to R.S.); FRM (ARF20140129235) (to L.B.); Spanish Ministry of Science, Innovation and Universities (MCINU/FEDER; grants SAF2015-64945-R and RTI2018-095311-B-I00) (to M.G.); Canada Research Chair, Alzheimer Society of Canada—Brain Canada (17-09), Natural Sciences and Engineering Research Council (RGPIN-2015-05880), Canadian Breast Cancer Foundation (2015-317342), Canadian Health Research Institute (CIHR, 388808), New Brunswick Innovation Foundation, New Brunswick Health Research Foundation and Université de Moncton (to E.H.-C.); Basque Government (IT1230-19), Red de Trastornos Adictivos, Instituto de Salud Carlos III (ISC-III), European Regional Development Funds-European Union (ERDF-EU; RD16/0017/0012) and MINECO/FEDER, UE (SAF2015-65034-R) (to P.G.); University of the Basque Country PhD contract (PIF 16/251) (to S.A.); POP contract (BES-2016-076766, BES-C-2016-0051) (to I.B.-D.R.); French State/ANR (ANR-10-IDEX and TRAIL ANR-10-LABX-57); French-Swiss ANR-FNS (ANR-15- CE37-0012) (to A.-K.B.-S.); SFB/TRR 58 ‘Fear, anxiety, anxiety disorders’ (subproject A04); and CRC 1193 ‘Neurobiology of resilience’ (subproject B04) (to B.L.).

The distribution and roles of the cannabinoid CB

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).

The cannabinoid CB1 receptor localizes to the glutamatergic parallel fiber (PF) terminals of the cerebellar granule cells and participates in synaptic plasticity, motor control and learning that are impaired in CB1 receptor knockout (CB 1 -KO) mice. However, whether ultrastructural changes at the PF-Purkinje cell (PC) synapses occur in CB 1 -KO remains unknown. We studied this in the vermis of the spinocerebellar lobule V and the vestibulocerebellar lobule X of CB 1 -KO and wild-type (CB 1 -WT) mice by electron microscopy. Lobule V, but not lobule X, of CB 1 -KO had significantly less and longer synapses than in CB 1 -WT. PF terminals were significantly larger in both lobules of CB 1 -KO with no changes in PC dendritic spines. The PF terminals in lobule V of CB 1 -KO contained less synaptic vesicles and lower vesicle density; by contrast, vesicle density in lobule X of CB 1 -KO remained unchangeable relative to CB 1 -WT. There were as many vesicles in lobule V of CB 1 -KO as in CB 1 -WT, but their distribution decreased drastically at 300 nm of the active zone. In lobule X of CB 1 -KO, less vesicles were found within 150 nm from the presynaptic membrane; however, no vesicles were at 450-600 nm of the active zone. A significant higher amount of synaptic vesicles close to the active zone in lobule V and X of CB 1 -KO was observed. In conclusion, the absence of CB1 receptors strikingly and distinctively impacts on the ultrastructural architecture of the PF-PC synapses located in cerebellar lobules that differ in vulnerability to damage and motor functions.

INTRODUCTION Despite a steady decline in the incidence of gastric cancer (GC) over recent decades, the prognosis remains poor with 5-year survival rates of 21.9% in Europe. The AJCC Cancer Staging Manual (8th Edition) establishes the necessity to evaluate personalized survival models. These models would help in the decision-making process. Proteomic and genomic molecular analyses have provided a more in-depth knowledge of GC. MATERIAL AND METHODS 19 patients affected by 21 tumors operated between 2012-2014 were prospectively included. Healthy tissue and tumor samples were taken from each case and deposited in a tissue biobank for subsequent proteomic analysis by Proteomics Department. RESULTS The following histopathological types were identified: (73.69%) intestinal, (10.52%) diffuse, and (15.78%) undetermined, poorly differentiated, and mixed neuroendocrine of the classification WHO. According to the size and extent of tumor invasion, the percentages of the sample were T1 or T2 (26.31%) and T3 or T4 (73.68%). Furthermore, 57.89% of the patients were found to have lymph-node metastases (N 1, 2, 3 +). After the differences in protein intensities in healthy tissue and tumor tissue were analyzed, it was found that in the healthy tissue, EGFR1, MAPK1 and MAPK3 were predictive variables of overall survival when the tumor stage was T3 or T4 CONCLUSIONS It would be convenient to redefine the classification and treatment plan in gastric cancer by integrating the study at a molecular level.

The use and abuse of alcohol (EtOH) is one of the world’s main health issues that strikingly impacts on our society, as heavy episodic drinking is becoming more and more common in the adolescence when the brain is particularly vulnerable to EtOH. However, molecular, anatomical, functional and behavioral alterations improve inyoung adult mice brains by an enriched environment (EE) exposure after adolescence EtOH consumption [21]. It remains unknown whether these beneficial effects are maintained over a long period of time after cessation of EtOH consumption. The aim of this study was to measure the long-term behavioral consequences of EtOH consumption and to explore the effects of EE in adulthood. For this goal, we treated C57BL/6J male mice with 20% EtOH or water during the 4 weeks of adolescence (p32-p56) followed by an abstinence period (p56-p90). Finally, they were exposed to EE for two weeks (p90-p104) and behavioral tests were conducted at their full adulthood: thigmotaxis for anxiety-like behaviour; novel object recognition test (NORT) for object recognition memory; novel object location test (NOLT) for location memory and beam walking balance test (BWBT) for motor coordination and balance. Object and spatial recognition memory were significantly lower in EtOH-treated mice. Also, motor coordination and balance were impaired after EtOH intake. Noticeably, memory and motor deficits reversed to control values after EE. In conclusion, we show that EE recovers the long-term behavioral and motor deficits after abusive EtOH consumption during adolescence. These results point to the beneficial effects EE have in EtOH addiction.; Alkohola (EtOH) munduan gehien kontsumitzen den substantzia psikoaktiboa da eta nerabezaroko alkoholaren kontsumo intentsiboa geroz eta ohikoagoa da. Adin tarte horretan burmuina garatzen ari da eta hainbat garun-atal zaurgarriagoak dira neurotoxikoen kalteen aurrean; hipokanpoa eta garuntxoa, esaterako. Ingurune aberastuak (IAk), aldaketa molekular, anatomiko zein funtzionalak eragiten ditu garunaren garapen prozesuan eta alkoholaren ondorioz helduaro goiztiarreko saguek galdutako portaera gaitasunen berreskurapena sustatzen du. Hala ere, IAk eragindako efektu mesedegarri horiek epe luzerago batean mantentzen diren aztertzeke dago. Ikerketa honen helburuak hurrengoak dira: nerabezaroko gehiegizko alkohol kontsumoak helduaroan eragiten dituen portaera aldaketak ikertzea eta parametro hauetan IAk izan ditzakeen onurak aztertzea. Horretarako, C57BL/6J sagu arrei nerabezaroko 4 astetan zehar (p32-p56) alkohol edo ur tratamendua eman zaie. Ondoren, helduaro goiztiarrean (p56-p90) animaliak abstinentzia egoeran mantendu dira eta helduaroan (p90-p104) saguen kumaldi erdia IAko baldintzetan jarri da 2 astez. Abstinentzia tarte horren azken egunetan portaera probak burutu dira: eremu irekiaren proba, antsietate maila neurtzeko; objektu berrien ezagutze proba, ezagutze oroimenerako; objektuen kokaleku berriaren ezagutze proba, oroimen espazialerako eta oreka proba, oreka eta koordinazio motorrerako. Alkohol taldeko saguek bereizketa indize baxuagoak erakutsi dituzte bai ezagutze oroimen proban baita oroimen espazialean ere, alkohol kontsumoaren ondoriozko narriadura kognitibo adierazgarria iradokiz. Antzeko emaitzak behatu dira oreka proban ere, non alkohol taldeko saguek (EtOH) oreka eta koordinazio motorra kaltetuta erakutsi duten. Interesgarriki, animaliak IAko baldintzapean jartzean objektuak eta kokalekuak bereizteko gaitasuna berreskuratzen dute eta oreka eta koordinazio maila hobetzen dute helduaroan, kontrol taldekoen (H2O) antzeko balioetaraino. IAk alkoholaren ondoriozko helduaroko efektu kaltegarriak leheneratzeko gaitasuna duela erakutsi du.

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 use and abuse of cannabis can be associated with significant pathophysiology, however, it remains unclear whether (1) acute administration of Δ-9-tetrahydrocannabinol (THC) during early adulthood alters the cannabinoid type 1 (CB

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.

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.

Binge drinking (BD) is a common pattern of ethanol (EtOH) consumption by adolescents. The brain effects of the acute EtOH exposure are well-studied; however, the long-lasting cognitive and neurobehavioral consequences of BD during adolescence are only beginning to be elucidated. Environmental enrichment (EE) has long been known for its benefits on the brain and may serve as a potential supportive therapy following EtOH exposure. In this study, we hypothesized that EE may have potential benefits on the cognitive deficits associated with BD EtOH consumption. Four-week-old C57BL/6J male mice were exposed to EtOH following an intermittent 4-day drinking-in-the-dark procedure for 4 weeks. Then they were exposed to EE during EtOH withdrawal for 2 weeks followed by a behavioral battery of tests including novel object recognition, novel location, object-in-place, rotarod, beam walking balance, tail suspension, light-dark box and open field that were run during early adulthood. Young adult mice exposed to EE significantly recovered recognition, spatial and associative memory as well as motor coordination skills and balance that were significantly impaired after adolescent EtOH drinking with respect to controls. No significant permanent anxiety or depressive-like behaviors were observed. Taken together, an EE exerts positive effects on the long-term negative cognitive deficits as a result of EtOH consumption during adolescence.

Cannabinoid type-1 (CB1 ) receptors are widely distributed in the brain and play important roles in astrocyte function and the modulation of neuronal synaptic transmission and plasticity. However, it is currently unknown how CB1 receptor expression in astrocytes is affected by long-term exposure to stressors. Here we examined CB1 receptors in astrocytes of ethanol (EtOH)-exposed adolescent mice to determine its effect on CB1 receptor localization and density in adult brain. 4-8-week-old male mice were exposed to 20 percent EtOH over a period of 4 weeks, and receptor localization was examined after 4 weeks in the hippocampal CA1 stratum radiatum by pre-embedding immunoelectron microscopy. Our results revealed a significant reduction in CB1 receptor immunoparticles in astrocytic processes of EtOH-exposed mice when compared with controls (positive astrocyte elements: 21.50 ± 2.80 percent versus 37.22 ± 3.12 percent, respectively), as well as a reduction in particle density (0.24 ± 0.02 versus 0.35 ± 0.02 particles/μm). The majority of CB1 receptor metal particles were in the range of 400-1200 nm from synaptic terminals in both control and EtOH. Altogether, the decrease in the CB1 receptor expression in hippocampal astrocytes of adult mice exposed to EtOH during adolescence reveals a long lasting effect of EtOH on astrocytic CB1 receptors. This deficiency may also have negative consequences for synaptic function.

The impact of pannexin-1 (Panx1) channels on synaptic transmission is poorly understood. Here, we show that selective block of Panx1 in single postsynaptic hippocampal CA1 neurons from male rat or mouse brain slices causes intermittent, seconds long increases in the frequency of sEPSC following Schaffer collateral stimulation. The increase in sEPSC frequency occurred without an effect on evoked neurotransmission. Consistent with a presynaptic origin of the augmented glutamate release, the increased sEPSC frequency was prevented by bath-applied EGTA-AM or TTX. Manipulation of a previously described metabotropic NMDAR pathway (i.e., by preventing ligand binding to NMDARs with competitive antagonists or blocking downstream Src kinase) also increased sEPSC frequency similar to that seen when Panx1 was blocked. This facilitated glutamate release was absent in transient receptor potential vanilloid 1 (TRPV1) KO mice and prevented by the TRPV1 antagonist, capsazepine, suggesting it required presynaptic TRPV1. We show presynaptic expression of TRPV1 by immunoelectron microscopy and link TRPV1 to Panx1 because Panx1 block increases tissue levels of the endovanilloid, anandamide. Together, these findings demonstrate an unexpected role for metabotropic NMDARs and postsynaptic Panx1 in suppression of facilitated glutamate neurotransmission.SIGNIFICANCE STATEMENTThe postsynaptic ion and metabolite channel, pannexin-1, is regulated by metabotropic NMDAR signaling through Src kinase. This pathway suppresses facilitated release of presynaptic glutamate during synaptic activity by regulating tissue levels of the transient receptor potential vanilloid 1 agonist anandamide.

Binge drinking is a significant problem in adolescent populations, and because of the reciprocal interactions between ethanol (EtOH) consumption and the endocannabinoid (eCB) system, we sought to determine if adolescent EtOH intake altered the localization and function of the cannabinoid 1 (CB(1)) receptors in the adult brain. Adolescent mice were exposed to a 4-day-per week drinking in the dark (DID) procedure for a total of 4 weeks and then tested after a 2-week withdrawal period. Field excitatory postsynaptic potentials (fEPSPs), evoked by medial perforant path (MPP) stimulation in the dentate gyrus molecular layer (DGML), were significantly smaller. Furthermore, unlike control animals, CB(1) receptor activation did not depress fEPSPs in the EtOH-exposed animals. We also examined a form of excitatory long-term depression that is dependent on CB(1) receptors (eCB-eLTD) and found that it was completely lacking in the animals that consumed EtOH during adolescence. Histological analyses indicated that adolescent EtOH intake significantly reduced the CB(1) receptor distribution and proportion of immunopositive excitatory synaptic terminals in the medial DGML. Furthermore, there was decreased binding of [(35)S]guanosine-5*-O-(3-thiotriphosphate) ([(35)S] GTPγS) and the guanine nucleotide-binding (G) protein Gαi2 subunit in the EtOH-exposed animals. Associated with this, there was a significant increase in monoacylglycerol lipase (MAGL) mRNA and protein in the hippocampus of EtOH-exposed animals. Conversely, deficits in eCB-eLTD and recognition memory could be rescued by inhibiting MAGL with JZL184. These findings indicate that repeated exposure to EtOH during adolescence leads to long-term deficits in CB(1) receptor expression, eCB-eLTD, and reduced recognition memory, but that these functional deficits can be restored by treatments that increase endogenous 2-arachidonoylglycerol.

Hippocampal slices are widely used for in vitro electrophysiological experiments to study underlying mechanisms for synaptic transmission and plasticity, and there is a growing appreciation for sex differences in synaptic plasticity. To date, several studies have shown that the process of making slices from male animals can induce synaptogenesis in cornu ammonis area 1 (CA1) pyramidal cells, but there is a paucity of data for females and other brain regions. In the current study we use microcrystals of the lipophilic carbocyanine dye DiI (1,1′-dioctadecyl-3,3,3′,3′-tetramethylindocarbocyanine perchlorate) to stain individual neurons in the CA1 and dentate gyrus (DG) hippocampal subfields of postnatal day 21 male and female rats. We show that the preparation of sections for electrophysiology produces significant increases in spines in sections obtained from females, similar to that observed in males. We also show that the procedures used for in vitro electrophysiology also result in significant spine increases in the DG and CA1 subfields. These results demonstrate the utility of this refined DiI procedure for staining neuronal dendrites and spines. They also show, for the first time, that in vitro electrophysiology slice preparations enhance spine numbers on hippocampal cells equivalently in both juvenile females and males.NEW & NOTEWORTHY This study introduces a new DiI technique that elucidates differences in spine numbers in juvenile female and male hippocampus, and shows that slice preparations for hippocampal electrophysiology in vitro may mask these differences.

The cannabinoid CB