Omega-3 Recovers Cannabinoid 1 Receptor Expression in the Adult Mouse Brain after Adolescent Binge Drinking.

Adolescent binge drinking is a social problem with a long-lasting impact on cognitive functions. The cannabinoid type-1 (CB1) receptor of the endocannabinoid system (ECS) is involved in brain synaptic plasticity, cognition and behavior via receptor localization at specific subcellular compartments of the cortical, limbic and motor regions. Alcohol (EtOH) intake affects the ECS, CB1 and their functions. Evidence indicates that binge drinking during adolescence impairs memory via the abrogation of CB1-dependent synaptic plasticity in the hippocampus. However, the impact of EtOH consumption on global CB1 receptor expression in the adult brain is unknown. We studied this using optical density analysis throughout brain regions processed for light microscopy (LM) immunohistotochemistry. CB1 staining decreased significantly in the secondary motor cortex, cerebellum, cingulate cortex, amygdala and nucleus accumbens. Next, as omega-3 (n-3) polyunsaturated fatty acids (PUFAs) rescue synaptic plasticity and improve EtOH-impaired cognition, we investigated whether docosahexaenoic acid (DHA) and eicosapentaenoic acid (EPA) had any effect on CB1 receptors. N-3 intake during EtOH abstinence restored CB1 immunostaining in the secondary motor cortex, cerebellum and amygdala, and ameliorated receptor density in the cingulate cortex. These results show that n-3 supplementation recovers CB1 receptor expression disrupted by EtOH in distinct brain regions involved in motor functions and cognition.

The effect of omega-3 fatty acids on alcohol-induced damage.

Alcohol is the most widely consumed psychoactive substance in the world that has a severe impact on many organs and bodily systems, particularly the liver and nervous system. Alcohol use during pregnancy roots long-lasting changes in the newborns and during adolescence has long-term detrimental effects especially on the brain. The brain contains docosahexaenoic acid (DHA), a major omega-3 (n-3) fatty acid (FA) that makes up cell membranes and influences membrane-associated protein function, cell signaling, gene expression and lipid production. N-3 is beneficial in several brain conditions like neurodegenerative diseases, ameliorating cognitive impairment, oxidative stress, neuronal death and inflammation. Because alcohol decreases the levels of n-3, it is timely to know whether n-3 supplementation positively modifies alcohol-induced injuries. The aim of this review is to summarize the state-of-the-art of the n-3 effects on certain conditions caused by alcohol intake, focusing primarily on brain damage and alcoholic liver disease.

Environmental Enrichment Rescues Endocannabinoid-Dependent Synaptic Plasticity Lost in Young Adult Male Mice after Ethanol Exposure during Adolescence.

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.

Intermittent ethanol exposure during adolescence impairs cannabinoid type 1 receptor-dependent long-term depression and recognition memory in adult mice.

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 (CB1) 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, CB1 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 CB1 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 CB1 receptor distribution and proportion of immunopositive excitatory synaptic terminals in the medial DGML. Furthermore, there was decreased binding of [35S]guanosine-5*-O-(3-thiotriphosphate) ([35S] 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 CB1 receptor expression, eCB-eLTD, and reduced recognition memory, but that these functional deficits can be restored by treatments that increase endogenous 2-arachidonoylglycerol.

Deletion of the cannabinoid CB1 receptor impacts on the ultrastructure of the cerebellar parallel fiber-Purkinje cell synapses.

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.

Endocannabinoid long-term depression revealed at medial perforant path excitatory synapses in the dentate gyrus.

The endocannabinoid system modulates synaptic plasticity in the hippocampus, but a link between long-term synaptic plasticity and the type 1 cannabinoid (CB1) receptor at medial perforant path (MPP) synapses remains elusive. Here, immuno-electron microscopy in adult mice showed that ∼26% of the excitatory synaptic terminals in the middle 1/3 of the dentate molecular layer (DML) contained CB1 receptors, and field excitatory postsynaptic potentials evoked by MPP stimulation were inhibited by CB1 receptor activation. In addition, MPP stimulation at 10 Hz for 10 min triggered CB1 receptor-dependent excitatory long-term depression (eCB-eLTD) at MPP synapses of wild-type mice but not on CB1-knockout mice. This eCB-eLTD was group I mGluR-dependent, required intracellular calcium influx and 2-arachydonoyl-glycerol (2-AG) synthesis but did not depend on N-methyl-d-aspartate (NMDA) receptors. Overall, these results point to a functional role for CB1 receptors with eCB-eLTD at DML MPP synapses and further involve these receptors in memory processing within the adult brain.

Cognitive and neurobehavioral benefits of an enriched environment on young adult mice after chronic ethanol consumption during adolescence.

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.

Environmental Enrichment Reverses Tyrosine Kinase Inhibitor-Mediated Impairment Through BDNF-TrkB Pathway.

Exposure to an enriched environment (EE) has neuroprotective benefits and improves recovery from brain injury due to, among other, increased neurotrophic factor expression. Through these neurotrophins, important cortical and hippocampal changes occur. Vandetanib acts as a tyrosine kinase inhibitor of cell receptors, among others, the vascular endothelial growth factor receptor (VEGFR). Our aim was to investigate the effectiveness of EE counteracting cognitive and cellular effects after tyrosine kinase receptor blockade. Animals were reared under standard laboratory condition or EE; both groups received vandetanib or vehicle. Visuospatial learning was tested with Morris water maze. Neuronal, interneuronal, and vascular densities were measured by inmunohistochemistry and histochemistry techniques. Quantifications were performed in the hippocampus and in the visual cortex. Brain-derived neurotrophic factor (BDNF), tyrosine kinase B receptor (TrkB), Akt, and Erk were measured by Western blot technique. Vandetanib produces a significant decrease in vascular and neuronal densities and reduction in the expression of molecules involved in survival and proliferation processes such as phospho-Akt/Akt and phospho-Erk/Erk. These results correlated to a cognitive impairment in visuospatial test. On the other hand, animals reared in an EE are able to reverse the negative effects, activating PI3K-AKT and MAP kinase pathways mediated by BDNF-TrkB binding. Present results provide novel and consistent evidences about the usefulness of living in EE as a strategy to improve deleterious effects of blocking neurotrophic pathways by vandetanib and the notable role of the BDNF-TrkB pathway to balance the neurovascular unit and cognitive effects.

Targeting the endocannabinoid system: future therapeutic strategies.

The endocannabinoid system (ECS) is involved in many physiological regulation pathways in the human body, which makes this system the target of many drugs and therapies. In this review, we highlight the latest studies regarding the role of the ECS and the drugs that target it, with a particular focus on the basis for the discovery of new cannabinoid-based drugs. In addition, we propose some key steps, such as the creation of a cannabinoid-receptor interaction matrix (CRIM) and the use of metabolomics, toward the development of improved and more specific drugs for each relevant disease.

Increased physical activity is not enough to recover astrocytic population from dark-rearing. Synergy with multisensory enrichment is required.

Elimination of sensory inputs (deprivation) modifies the properties of the sensory cortex and serves as a model for studying plasticity during postnatal development. Many studies on the effects of deprivation have been performed in the visual cortex using dark-rearing as a visual deprivation model. It induces changes in all cellular and molecular components, including astrocytes, which play an important role in the development, maintenance, and plasticity of the cortex, mediated by cytokines which have been termed angioglioneurins. When one sense is deprived, a compensatory mechanism called cross-modal plasticity increases performance in the remaining senses. Environmental enrichment is so far the best-known method to compensate sensorial deprivation. The aim of this work is to study the effects of exercise alone, and of an enriched environment combined with exercise, on astroglial population in order to observe the effects of exercise by itself, or the potential synergistic effect during the rat visual system development. Pregnant Sprague-Dawley rats were raised in one of the following rearing conditions: in total darkness and enriched environment conditions with physical exercise, and in total darkness with voluntary physical exercise. Astrocytic density was estimated by immunohistochemistry for S-100ß protein and quantifications were performed in layer IV. The somatosensorial cortex barrel field was also studied as control. Our main result shows that an enriched environment combined with voluntary physical exercise manages to reverse the negative effects induced by darkness over the astroglial population of both the visual and the somatosensory cortices. On the other hand, exercise alone only produces effects upon the astroglial population of the somatosensory cortex, and less so when combined with an enriched environment.

Angiogenic signalling pathways altered in gliomas: selection mechanisms for more aggressive neoplastic subpopulations with invasive phenotype.

The angiogenesis process is a key event for glioma survival, malignancy and growth. The start of angiogenesis is mediated by a cascade of intratumoural events: alteration of the microvasculature network; a hypoxic microenvironment; adaptation of neoplastic cells and synthesis of pro-angiogenic factors. Due to a chaotic blood flow, a consequence of an aberrant microvasculature, tissue hypoxia phenomena are induced. Hypoxia inducible factor 1 is a major regulator in glioma invasiveness and angiogenesis. Clones of neoplastic cells with stem cell characteristics are selected by HIF-1. These cells, called "glioma stem cells" induce the synthesis of vascular endothelial growth factor. This factor is a pivotal mediator of angiogenesis. To elucidate the role of these angiogenic mediators during glioma growth, we have used a rat endogenous glioma model. Gliomas induced by prenatal ENU administration allowed us to study angiogenic events from early to advanced tumour stages. Events such as microvascular aberrations, hypoxia, GSC selection and VEGF synthesis may be studied in depth. Our data showed that for the treatment of gliomas, developing anti-angiogenic therapies could be aimed at GSCs, HIF-1 or VEGF. The ENU-glioma model can be considered to be a useful option to check novel designs of these treatment strategies.

Enriched and deprived sensory experience induces structural changes and rewires connectivity during the postnatal development of the brain.

During postnatal development, sensory experience modulates cortical development, inducing numerous changes in all of the components of the cortex. Most of the cortical changes thus induced occur during the critical period, when the functional and structural properties of cortical neurons are particularly susceptible to alterations. Although the time course for experience-mediated sensory development is specific for each system, postnatal development acts as a whole, and if one cortical area is deprived of its normal sensory inputs during early stages, it will be reorganized by the nondeprived senses in a process of cross-modal plasticity that not only increases performance in the remaining senses when one is deprived, but also rewires the brain allowing the deprived cortex to process inputs from other senses and cortices, maintaining the modular configuration. This paper summarizes our current understanding of sensory systems development, focused specially in the visual system. It delineates sensory enhancement and sensory deprivation effects at both physiological and anatomical levels and describes the use of enriched environment as a tool to rewire loss of brain areas to enhance other active senses. Finally, strategies to apply restorative features in human-deprived senses are studied, discussing the beneficial and detrimental effects of cross-modal plasticity in prostheses and sensory substitution devices implantation.

Effect of intracortical vascular endothelial growth factor infusion and blockade during the critical period in the rat visual cortex.

VEGF is the major angiogenic and vascular permeability factor in health and disease. Vascular development depends on function, and in sensory areas is experience-dependent. Our aim was to investigate, qualitatively and quantitatively, the effects of intracortical infusion and neutralisation of VEGF during the first days of the critical visual period, when peak levels of endogenous VEGF secretion are reached. VEGF was intracortically delivered into middle cortical layers of P18 Long-Evans rats. Another cohort received anti-VEGF. Vehicle (PBS)-infused and non-operated animals were used as controls. Various immunopathological analyses were performed: Endothelial Barrier Antigen (EBA) for the BBB integrity and GFAP for astroglial response. Vascular density was measured by Butyryl Cholinesterase Histochemistry, neuronal density by NeuN immunohistochemistry and apoptosis by TUNEL staining. VEGF levels were measured by Western Blot. Decreased vascular permeability was evoked in VEGF-infused rats whilst EBA expression remained constant, suggesting a preserved BBB function. When VEGF was blocked, tissue showed a higher degree of extravasation and a decreased number of EBA-positive vessels surrounding the injury. Lesion induced by cannula implantation annulled the normal increase in vascular density and the decrease in neuronal density during this time. VEGF rescued in part the vascular increase, and also prevented physiological and pathological neuronal death. VEGF blockade induced a higher amount of neural loss and lower astrocytic reaction. Our results support the role of VEGF as extending beyond vascularization, preventing physiological and pathological neuronal death, not only in the injured hemisphere but also in the intact one suggesting a process of transhemispheric diaschisis.

Vascular endothelial growth factor: adaptive changes in the neuroglialvascular unit.

Brain postnatal development is modulated by adaptation and experience. Experience-mediated changes increase neuronal activity leading to increased metabolic demands that involve adaptive changes including ones at the microvascular network. Therefore, vascular environment plays a key role in central nervous system (CNS) development and function in health and disease. Trophic factors are crucial in CNS development and cell survival in adults. They participate in protection and proliferation of neuronal, glial and endothelial cells. Among the most important molecules are: the proangiogenic vascular endothelial growth factor (VEGF), the neurotrophin brain derived neurotrophic factor (BDNF), insulin growth factor (IGF-I) and the glycoprotein erythropoietin (EPO). We propose the term angioglioneurins to define molecules acting on the three components of the neurogliovascular unit. We have previously reported the effects of environmental modifications on the three components of the neurogliovascular unit during the postnatal development. We have also described the main role played by VEGF in the experience-induced postnatal changes. Angioglioneurin administration, alone or in combination with other neuroprotective strategies such as environmental enrichment, has been proposed as a non-invasive therapeutic strategy against several CNS diseases.