RESUMEN
Cannabinoids, the bioactive constituents of cannabis, exert a wide array of effects on the brain by engaging Type 1 cannabinoid receptor (CB1R). Accruing evidence supports that cannabinoid action relies on context-dependent factors, such as the biological characteristics of the target cell, suggesting that cell population-intrinsic molecular cues modulate CB1R-dependent signaling. Here, by using a yeast two-hybrid-based high-throughput screening, we identified BiP as a potential CB1R-interacting protein. We next found that CB1R and BiP interact specifically in vitro, and mapped the interaction site within the CB1R C-terminal (intracellular) domain and the BiP C-terminal (substrate-binding) domain-α. BiP selectively shaped agonist-evoked CB1R signaling by blocking an "alternative" Gq/11 protein-dependent signaling module while leaving the "classical" Gi/o protein-dependent inhibition of the cAMP pathway unaffected. In situ proximity ligation assays conducted on brain samples from various genetic mouse models of conditional loss or gain of CB1R expression allowed to map CB1R-BiP complexes selectively on terminals of GABAergic neurons. Behavioral studies using cannabinoid-treated male BiP+/- mice supported that CB1R-BiP complexes modulate cannabinoid-evoked anxiety, one of the most frequent undesired effects of cannabis. Together, by identifying BiP as a CB1R-interacting protein that controls receptor function in a signaling pathway- and neuron population-selective manner, our findings may help to understand the striking context-dependent actions of cannabis in the brain.SIGNIFICANCE STATEMENT Cannabis use is increasing worldwide, so innovative studies aimed to understand its complex mechanism of neurobiological action are warranted. Here, we found that cannabinoid CB1 receptor (CB1R), the primary molecular target of the bioactive constituents of cannabis, interacts specifically with an intracellular protein called BiP. The interaction between CB1R and BiP occurs selectively on terminals of GABAergic (inhibitory) neurons, and induces a remarkable shift in the CB1R-associated signaling profile. Behavioral studies conducted in mice support that CB1R-BiP complexes act as fine-tuners of anxiety, one of the most frequent undesired effects of cannabis use. Our findings open a new conceptual framework to understand the striking context-dependent pharmacological actions of cannabis in the brain.
Asunto(s)
Encéfalo/metabolismo , Cannabinoides/metabolismo , Neuronas GABAérgicas/metabolismo , Proteínas de Choque Térmico/metabolismo , Receptor Cannabinoide CB1/metabolismo , Transducción de Señal/fisiología , Animales , Chaperón BiP del Retículo Endoplásmico , Células HEK293 , Proteínas de Choque Térmico/genética , Humanos , Ratones , Ratones Noqueados , Receptor Cannabinoide CB1/genéticaRESUMEN
Cannabis use disorder is frequent in schizophrenia patients, and it is associated with an earlier age of onset and poor schizophrenia prognosis. Serotonin 2A receptors (5-HT2AR) have been involved in psychosis and, like Akt kinase, are known to be modulated by THC. Likewise, endocannabinoid system dysregulation has been suggested in schizophrenia. The presence of these molecules in blood makes them interesting targets, as they can be evaluated in patients by a minimally invasive technique. The aim of the present study was to evaluate 5-HT2AR protein expression and the Akt functional status in platelet homogenates of subjects diagnosed with schizophrenia, cannabis use disorder, or both conditions, compared with age- and sex-matched control subjects. Additionally, endocannabinoids and pro-inflammatory interleukin-6 (IL-6) levels were also measured in the plasma of these subjects. Results showed that both platelet 5-HT2AR and the active phospho (Ser473)Akt protein expression were significantly increased in schizophrenia subjects, whereas patients with a dual diagnosis of schizophrenia and cannabis use disorder did not show significant changes. Similarly, plasma concentrations of anandamide and other lipid mediators such as PEA and DEA, as well as the pro-inflammatory IL-6, were significantly increased in schizophrenia, but not in dual subjects. Results demonstrate that schizophrenia subjects show different circulating markers pattern depending on the associated diagnosis of cannabis use disorder, supporting the hypothesis that there could be different underlying mechanisms that may explain clinical differences among these groups. Moreover, they provide the first preliminary evidence of peripherally measurable molecules of interest for bigger prospective studies in these subpopulations.
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Cannabis , Abuso de Marihuana , Esquizofrenia , Humanos , Esquizofrenia/metabolismo , Interleucina-6 , Estudios Prospectivos , Proteínas Proto-Oncogénicas c-akt , Agonistas de Receptores de Cannabinoides , BiomarcadoresRESUMEN
Although human epidermal growth factor receptor 2 (HER2)-targeted therapies have dramatically improved the clinical outcome of HER2-positive breast cancer patients, innate and acquired resistance remains an important clinical challenge. New therapeutic approaches and diagnostic tools for identification, stratification, and treatment of patients at higher risk of resistance and recurrence are therefore warranted. Here, we unveil a mechanism controlling the oncogenic activity of HER2: heteromerization with the cannabinoid receptor CB2R. We show that HER2 physically interacts with CB2R in breast cancer cells, and that the expression of these heteromers correlates with poor patient prognosis. The cannabinoid Δ9-tetrahydrocannabinol (THC) disrupts HER2-CB2R complexes by selectively binding to CB2R, which leads to (i) the inactivation of HER2 through disruption of HER2-HER2 homodimers, and (ii) the subsequent degradation of HER2 by the proteasome via the E3 ligase c-CBL. This in turn triggers antitumor responses in vitro and in vivo. Selective targeting of CB2R transmembrane region 5 mimicked THC effects. Together, these findings define HER2-CB2R heteromers as new potential targets for antitumor therapies and biomarkers with prognostic value in HER2-positive breast cancer.
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Neoplasias de la Mama/líquido cefalorraquídeo , Terapia Molecular Dirigida , Receptor Cannabinoide CB2/genética , Receptor ErbB-2/genética , Neoplasias de la Mama/tratamiento farmacológico , Neoplasias de la Mama/genética , Neoplasias de la Mama/patología , Línea Celular Tumoral , Dronabinol/farmacología , Resistencia a Antineoplásicos/genética , Femenino , Regulación Neoplásica de la Expresión Génica/efectos de los fármacos , Humanos , Recurrencia Local de Neoplasia/tratamiento farmacológico , Recurrencia Local de Neoplasia/genética , Recurrencia Local de Neoplasia/patología , Multimerización de Proteína/efectos de los fármacos , Proteínas Proto-Oncogénicas c-cbl/genética , Receptor Cannabinoide CB2/química , Receptor ErbB-2/química , Transducción de SeñalRESUMEN
Cereblon/CRBN is a substrate-recognition component of the Cullin4A-DDB1-Roc1 E3 ubiquitin ligase complex. Destabilizing mutations in the human CRBN gene cause a form of autosomal recessive non-syndromic intellectual disability (ARNSID) that is modelled by knocking-out the mouse Crbn gene. A reduction in excitatory neurotransmission has been proposed as an underlying mechanism of the disease. However, the precise factors eliciting this impairment remain mostly unknown. Here we report that CRBN molecules selectively located on glutamatergic neurons are necessary for proper memory function. Combining various in vivo approaches, we show that the cannabinoid CB1 receptor (CB1R), a key suppressor of synaptic transmission, is overactivated in CRBN deficiency-linked ARNSID mouse models, and that the memory deficits observed in these animals can be rescued by acute CB1R-selective pharmacological antagonism. Molecular studies demonstrated that CRBN interacts physically with CB1R and impairs the CB1R-Gi/o-cAMP-PKA pathway in a ubiquitin ligase-independent manner. Taken together, these findings unveil that CB1R overactivation is a driving mechanism of CRBN deficiency-linked ARNSID and anticipate that the antagonism of CB1R could constitute a new therapy for this orphan disease.
Asunto(s)
Proteínas Adaptadoras Transductoras de Señales , Trastornos de la Memoria , Ubiquitina-Proteína Ligasas , Animales , Ratones , Proteínas Adaptadoras Transductoras de Señales/genética , Proteínas Adaptadoras Transductoras de Señales/metabolismo , Mutación , Ubiquitina/metabolismo , Ubiquitina-Proteína Ligasas/genética , Ubiquitina-Proteína Ligasas/metabolismo , Receptor Cannabinoide CB1/genética , Receptor Cannabinoide CB1/metabolismo , Trastornos de la Memoria/genética , Trastornos de la Memoria/metabolismoRESUMEN
Prepulse inhibition of the startle response enables measuring animal behavior and helps us understand core aspects of neuropsychiatric diseases. Prepulse inhibition is considered a translational indicator of sensorimotor gating deficits present in schizophrenia patients and is crucial in the characterization of animal models of schizophrenia-like behaviors. Hallucinogenic drugs acting through 5-HT2A receptors, such as psilocybin, lysergic acid diethylamide (LSD), and dimethoxyiodoamphetamine (DOI), produce symptoms in healthy subjects comparable to those seen in schizophrenia and can be used in rodent models for mimicking some of these behaviors. Here we describe a protocol for the evaluation of prepulse inhibition of the startle response in CD1-Swiss male mice after a single dose of the hallucinogenic drug DOI.
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Inhibición Prepulso , Esquizofrenia , Ratones , Masculino , Animales , Reflejo de Sobresalto/fisiología , Filtrado SensorialRESUMEN
Cannabinoids exert pleiotropic effects on the brain by engaging the cannabinoid CB1 receptor (CB1R), a presynaptic metabotropic receptor that regulates key neuronal functions in a highly context-dependent manner. We have previously shown that CB1R interacts with growth-associated protein of 43 kDa (GAP43) and that this interaction inhibits CB1R function on hippocampal excitatory synaptic transmission, thereby impairing the therapeutic effect of cannabinoids on epileptic seizures in vivo. However, the underlying molecular features of this interaction remain unexplored. Here, we conducted mechanistic experiments on HEK293T cells co-expressing CB1R and GAP43 and show that GAP43 modulates CB1R signalling in a strikingly selective manner. Specifically, GAP43 did not affect the archetypical agonist-evoked (i) CB1R/Gi/o protein-coupled signalling pathways, such as cAMP/PKA and ERK, or (ii) CB1R internalization and intracellular trafficking. In contrast, GAP43 blocked an alternative agonist-evoked CB1R-mediated activation of the cytoskeleton-associated ROCK signalling pathway, which relied on the GAP43-mediated impairment of CB1R/Gq/11 protein coupling. GAP43 also abrogated CB1R-mediated ROCK activation in mouse hippocampal neurons, and this process led in turn to a blockade of cannabinoid-evoked neurite collapse. An NMR-based characterization of the CB1R-GAP43 interaction supported that GAP43 binds directly and specifically through multiple amino acid stretches to the C-terminal domain of the receptor. Taken together, our findings unveil a CB1R-Gq/11-ROCK signalling axis that is selectively impaired by GAP43 and may ultimately control neurite outgrowth.
RESUMEN
The study of psychiatric and neurological diseases requires the substrate in which the disorders occur, that is, the nervous tissue. Currently, several types of human bio-specimens are being used for research, including postmortem brains, cerebrospinal fluid, induced pluripotent stem (iPS) cells, and induced neuronal (iN) cells. However, these samples are far from providing a useful predictive, diagnostic, or prognostic biomarker. The olfactory epithelium is a region close to the brain that has received increased interest as a research tool for the study of brain mechanisms in complex neuropsychiatric and neurological diseases. The olfactory sensory neurons are replaced by neurogenesis throughout adult life from stem cells on the basement membrane. These stem cells are multipotent and can be propagated in neurospheres, proliferated in vitro and differentiated into multiple cell types including neurons and glia. For all these reasons, olfactory epithelium provides a unique resource for investigating neuronal molecular markers of neuropsychiatric and neurological diseases. Here, we describe the isolation and culture of human differentiated neurons and glial cells from olfactory epithelium of living subjects by an easy and non-invasive exfoliation method that may serve as a useful tool for the research in brain diseases.
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Técnicas de Cultivo de Célula , Diferenciación Celular , Separación Celular , Neurogénesis , Neuroglía , Neuronas , Mucosa Olfatoria , Humanos , Membrana Basal/citología , Biomarcadores/análisis , Adhesión Celular , Técnicas de Cultivo de Célula/métodos , Proliferación Celular , Separación Celular/métodos , Células Cultivadas , Medios de Cultivo/química , Citometría de Flujo , Inmunohistoquímica , Magnetismo , Células-Madre Neurales/citología , Neuroglía/citología , Neuronas/citología , Mucosa Olfatoria/citología , Especificidad de ÓrganosRESUMEN
Munc18-1 and syntaxin-1 are crucial interacting molecules for synaptic membrane fusion and neurotransmitter release. Contrasting abnormalities of several proteins of the exocytotic machinery, including the formation of SNARE (synaptobrevin, SNAP-25 and syntaxin-1) complexes, have been reported in schizophrenia. This study quantified in the dorsolateral prefrontal cortex (PFC, Brodmann area 9) the immunocontent of munc18-1a/b isoforms, syntaxin-1A, other presynaptic proteins (synaptotagmin, synaptophysin), and SNARE complexes, as well as the effects of psychoactive drug exposure, in schizophrenia (SZ, n=24), non-schizophrenia suicide (SD, n=13) and major depression (MD, n=15) subjects compared to matched controls (n=39). SZ was associated with normal expression of munc18-1a/b and increased syntaxin-1A (+44%). The presence of antipsychotic drugs reduced the basal content of munc18-1a isoform (-23%) and synaptobrevin (-32%), and modestly reduced that of up-regulated syntaxin-1A (-16%). Munc18-1a and syntaxin-1A protein expression correlated positively in controls but showed a markedly opposite pattern in SZ, regardless of antipsychotic treatment. Thus, the ratio of syntaxin-1A to munc18-1a showed a net increase in SZ (+53/114%). The SNARE complex (75 kDa) was found unaltered in antipsychotic-free and reduced (-28%) in antipsychotic-treated SZ subjects. None of these abnormalities were observed in SD and MD subjects, unexposed or exposed to psychoactive drugs. The results reveal some exocytotic dysfunctions in SZ that are probably related to an imbalance of the interaction between munc18-1a and SNARE (mainly syntaxin-1A) complex. Moreover, antipsychotic drug treatment is associated with lower content of key proteins of the exocytotic machinery, which could result in a destabilization/impairment of neurosecretion.
Asunto(s)
Antipsicóticos/metabolismo , Trastorno Depresivo Mayor/metabolismo , Proteínas Munc18/metabolismo , Corteza Prefrontal/metabolismo , Proteínas SNARE/metabolismo , Esquizofrenia , Suicidio , Sintaxina 1/metabolismo , Animales , Regulación hacia Abajo , Exocitosis/fisiología , Humanos , Masculino , Isoformas de Proteínas/metabolismo , Proteínas R-SNARE/metabolismo , Ratas , Ratas Sprague-Dawley , Esquizofrenia/tratamiento farmacológico , Esquizofrenia/metabolismo , Sinaptotagminas/metabolismoRESUMEN
Accumulating evidence has proven that both exogenous cannabinoids as well as imbalances in the endocannabinoid system are involved in the onset and development of mental disorders such as anxiety, depression, or schizophrenia. Extensive recent research in this topic has mainly focused on the molecular mechanisms by which cannabinoid agonists may contribute to the pathophysiology of these disorders. Initially, serotonin neurotransmitter garnered most attention due to its relationship to mood disorders and mental diseases, with little attention to specific receptors. To date, the focus has redirected toward the understanding of different serotonin receptors, through a demonstration of its versatile pharmacology and synergy with different modulators. Serotonin 2A receptors are a good example of this phenomenon, and the complex signaling that they trigger appears of high relevance in the context of mental disorders, especially in schizophrenia. This chapter will analyze most relevant attributes of serotonin 2A receptors and the endocannabinoid system, and will highlight the evidence toward the functional bidirectional interaction between these elements in the brain as well as the impact of the endocannabinoid system dysregulation on serotonin 2A receptors functionality.
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Cannabinoides , Esquizofrenia , Moduladores de Receptores de Cannabinoides , Cannabinoides/farmacología , Endocannabinoides , Humanos , Receptor de Serotonina 5-HT2A , Esquizofrenia/tratamiento farmacológicoRESUMEN
The mechanistic target of rapamycin (also known as mammalian target of rapamycin) (mTOR)-dependent signaling pathway plays an important role in protein synthesis, cell growth, and proliferation, and has been linked to the development of the central nervous system. Recent studies suggest that mTOR signaling pathway dysfunction could be involved in the etiopathogenesis of schizophrenia. The main goal of this study was to evaluate the status of mTOR signaling pathway in postmortem prefrontal cortex (PFC) samples of subjects with schizophrenia. For this purpose, we quantified the protein expression and phosphorylation status of the mTOR downstream effector ribosomal protein S6 as well as other pathway interactors such as Akt and GSK3ß. Furthermore, we quantified the status of these proteins in the brain cortex of rats chronically treated with the antipsychotics haloperidol, clozapine, or risperidone. We found a striking decrease in the expression of total S6 and in its active phosphorylated form phospho-S6 (Ser235/236) in the brain of subjects with schizophrenia compared to matched controls. The chronic treatment with the antipsychotics haloperidol and clozapine affected both the expression of GSK3ß and the activation of Akt [phospho-Akt (Ser473)] in rat brain cortex, while no changes were observed in S6 and phospho-S6 (Ser235/236) protein expression with any antipsychotic treatment. These findings provide further evidence for the involvement of the mTOR-dependent signaling pathway in schizophrenia and suggest that a hypofunctional S6 may have a role in the etiopathogenesis of this disorder.
RESUMEN
Disruption of the hypothalamic-pituitary-adrenal axis is an established finding in patients with anxiety and/or depression. Chronic corticosterone administration in animals has been proposed as a model for the study of these stress-related disorders and the antidepressant action. Alterations of the central noradrenergic system and specifically of inhibitory α2-adrenoceptors seem to be part of the pathophysiology of depression and contribute to the antidepressant activity. The present study evaluates in male rats the effect of chronic corticosterone treatment during 35 days (16-20â¯mgâ¯kg-1 day-1) on the sensitivity of α2-adrenoceptors expressed in the somatodendritic and terminal noradrenergic areas locus coeruleus (LC) and prefrontal cortex (PFC), respectively. Further, the effect of chronic fluoxetine treatment (5â¯mgâ¯kg-1, i.p., since the 15th day) on the sensitivity of α2-adrenoceptors was examined under control conditions and in corticosterone-treated rats. The α2-adrenoceptor functionality was analysed in vitro by agonist-mediated [35S]GTPγS binding stimulation and in vivo through the modulation of noradrenaline (NA) release evaluated by dual-probe microdialysis. The concentration-effect curves of the [35S]GTPγS binding stimulation by the agonist UK14304 (5-bromo-N-(4,5-dihydro-1H-imidazol-2-yl)-6-quinoxalinamine) demonstrated a desensitization of cortical α2-adrenoceptors induced by corticosterone (-logEC50â¯=â¯6.7⯱â¯0.2 vs 8.2⯱â¯0.3 in controls) that was reverted by fluoxetine treatment (-logEC50â¯=â¯7.5⯱â¯0.3). Local administration of the α2-adrenoceptor antagonist RS79948 ((8aR,12aS,13aS)-5,8,8a,9,10,11,12,12a,13,13a-decahydro-3-methoxy-12-(ethylsulfonyl)-6H-isoquino[2,1-g][1,6]naphthyridine) (0.1-100⯵molâ¯L-1) into the LC induced a concentration-dependent NA increase in the PFC of the control group (Emaxâ¯=â¯191⯱â¯30%) but non-significant effect was observed in corticosterone-treated rats (Emaxâ¯=â¯133⯱â¯46%), reflecting a desensitization of α2-adrenoceptors that control the firing of noradrenergic neurons. Fluoxetine treatment did not alter the corticosterone-induced desensitization in this area (Emaxâ¯=â¯136⯱â¯19%). No effect of fluoxetine on α2-adrenoceptor functionality was observed in control animals (Emaxâ¯=â¯223⯱â¯30%). In PFC, the local administration of RS79948 increased NA in controls (Emaxâ¯=â¯226⯱â¯27%) without effect in the corticosterone group (Emaxâ¯=â¯115⯱â¯26%), suggesting a corticosterone-induced desensitization of terminal α2-adrenoceptors. Fluoxetine administration prevented the desensitization induced by corticosterone in the PFC (Emaxâ¯=â¯233⯱â¯33%) whereas desensitized α2-adrenoceptors in control animals (Emaxâ¯=â¯-24⯱â¯10%). These data indicate that chronic corticosterone increases noradrenergic activity by acting at different α2-adrenoceptor subpopulations. Treatment with the antidepressant fluoxetine seems to counteract these changes by acting mainly on presynaptic α2-adrenoceptors expressed in terminal areas.
Asunto(s)
Neuronas Adrenérgicas/efectos de los fármacos , Antidepresivos de Segunda Generación/farmacología , Corticosterona/farmacología , Fluoxetina/farmacología , Locus Coeruleus/efectos de los fármacos , Corteza Prefrontal/efectos de los fármacos , Receptores Adrenérgicos alfa 2/efectos de los fármacos , Neuronas Adrenérgicas/metabolismo , Agonistas de Receptores Adrenérgicos alfa 2/farmacología , Antagonistas de Receptores Adrenérgicos alfa 2/farmacología , Animales , Tartrato de Brimonidina/farmacología , Cuerpo Celular/efectos de los fármacos , Cuerpo Celular/metabolismo , Dendritas/efectos de los fármacos , Dendritas/metabolismo , Modelos Animales de Enfermedad , Guanosina 5'-O-(3-Tiotrifosfato) , Sistema Hipotálamo-Hipofisario/metabolismo , Técnicas In Vitro , Isoquinolinas/farmacología , Locus Coeruleus/metabolismo , Masculino , Microdiálisis , Naftiridinas/farmacología , Norepinefrina/metabolismo , Sistema Hipófiso-Suprarrenal/metabolismo , Corteza Prefrontal/metabolismo , Terminales Presinápticos/efectos de los fármacos , Terminales Presinápticos/metabolismo , Ratas , Receptores Adrenérgicos alfa 2/metabolismo , Estrés Psicológico/metabolismo , Radioisótopos de AzufreRESUMEN
Long-term use of potent cannabis during adolescence increases the risk of developing schizophrenia later in life, but to date, the mechanisms involved remain unknown. Several findings suggest that the functional selectivity of serotonin 2A receptor (5-HT2AR) through inhibitory G-proteins is involved in the molecular mechanisms responsible for psychotic symptoms. Moreover, this receptor is dysregulated in the frontal cortex of schizophrenia patients. In this context, studies involving cannabis exposure and 5-HT2AR are scarce. Here, we tested in mice the effect of an early chronic Δ9-tetrahydrocannabinol (THC) exposure on cortical 5-HT2AR expression, as well as on its in vivo and in vitro functionality. Long-term exposure to THC induced a pro-hallucinogenic molecular conformation of the 5-HT2AR and exacerbated schizophrenia-like responses, such as prepulse inhibition disruption. Supersensitive coupling of 5-HT2AR toward inhibitory Gαi1-, Gαi3-, Gαo-, and Gαz-proteins after chronic THC exposure was observed, without changes in the canonical Gαq/11-protein pathway. In addition, we found that inhibition of Akt/mTOR pathway by rapamycin blocks the changes in 5-HT2AR signaling pattern and the supersensitivity to schizophrenia-like effects induced by chronic THC. The present study provides the first evidence of a mechanistic explanation for the relationship between chronic cannabis exposure in early life and increased risk of developing psychosis-like behaviors in adulthood.
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Dronabinol/farmacología , Alucinógenos/farmacología , Proteína Oncogénica v-akt/efectos de los fármacos , Receptor de Serotonina 5-HT2A/efectos de los fármacos , Transducción de Señal/efectos de los fármacos , Serina-Treonina Quinasas TOR/farmacología , Anfetaminas/farmacología , Animales , Química Encefálica/efectos de los fármacos , Masculino , Ratones , Proteína Oncogénica v-akt/metabolismo , Receptor de Serotonina 5-HT2A/metabolismo , Receptores Acoplados a Proteínas G/efectos de los fármacos , Reflejo de Sobresalto/efectos de los fármacos , Psicología del Esquizofrénico , Agonistas de Receptores de Serotonina/farmacología , Sirolimus/farmacología , Serina-Treonina Quinasas TOR/metabolismoRESUMEN
Mental disorders have a high prevalence compared with many other health conditions and are the leading cause of disability worldwide. Several studies performed in the last years support the involvement of the endocannabinoid system in the etiopathogenesis of different mental disorders. The present review will summarize the latest information on the role of the endocannabinoid system in psychiatric disorders, specifically depression, anxiety, and schizophrenia. We will focus on the findings from human brain studies regarding alterations in endocannabinoid levels, cannabinoid receptors and endocannabinoid metabolizing enzymes in patients suffering mental disorders. Studies carried out in humans have consistently demonstrated that the endocannabinoid system is fundamental for emotional homeostasis and cognitive function. Thus, deregulation of the different elements that are part of the endocannabinoid system may contribute to the pathophysiology of several mental disorders. However, the results reported are controversial. In this sense, different alterations in gene and/or protein expression of CB1 receptors have been shown depending on the technical approach used or the brain region studied. Despite the current discrepancies regarding cannabinoid receptors changes in depression and schizophrenia, present findings point to the endocannabinoid system as a pivotal neuromodulatory pathway relevant in the pathophysiology of mental disorders.
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Encéfalo/metabolismo , Endocannabinoides/metabolismo , Trastornos Mentales/metabolismo , Trastornos de Ansiedad/genética , Trastornos de Ansiedad/metabolismo , Depresión/genética , Depresión/metabolismo , Emociones , Humanos , Receptores de Cannabinoides/metabolismo , Esquizofrenia/genética , Esquizofrenia/metabolismoRESUMEN
Cognitive deficits are considered a key feature of schizophrenia, and they usually precede the onset of the illness and continue after psychotic symptoms appear. Current antipsychotic drugs have little or no effect on the cognitive deficits of this disorder. Prolyl oligopeptidase (POP) is an 81-kDa monomeric serine protease that is expressed in brain and other tissues. POP inhibitors have shown neuroprotective, anti-amnesic and cognition-enhancing properties. Here we studied the potential of IPR19, a new POP inhibitor, for the treatment of the cognitive symptoms related to schizophrenia. The efficacy of the inhibitor was evaluated in mouse models based on subchronic phencyclidine and acute dizocilpine administration, and in adult offspring from mothers with immune reaction induced by polyinosinic:polycytidylic acid administration during pregnancy. Acute IPR19 administration (5mg/kg, i.p.) reversed the cognitive performance deficits of the three mouse models in the novel object recognition test, T-maze, and eight-arm radial maze. The compound also ameliorates deficits of the prepulse inhibition response. The in vitro inhibitory efficacy and selectivity, brain penetration and exposure time after injection of IPR19 were also addressed. Our results indicate that the inhibition of POP using IPR19 may offer a promising strategy to develop drugs to ameliorate the cognitive deficits of schizophrenia.
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Trastornos del Conocimiento/tratamiento farmacológico , Prolina/análogos & derivados , Psicotrópicos/farmacología , Esquizofrenia/tratamiento farmacológico , Psicología del Esquizofrénico , Animales , Línea Celular Tumoral , Cognición/efectos de los fármacos , Cognición/fisiología , Trastornos del Conocimiento/enzimología , Trastornos del Conocimiento/etiología , Modelos Animales de Enfermedad , Relación Dosis-Respuesta a Droga , Humanos , Masculino , Aprendizaje por Laberinto/efectos de los fármacos , Aprendizaje por Laberinto/fisiología , Ratones Endogámicos C57BL , Actividad Motora/efectos de los fármacos , Actividad Motora/fisiología , Poli I-C , Inhibición Prepulso/efectos de los fármacos , Inhibición Prepulso/fisiología , Prolina/química , Prolina/farmacocinética , Prolina/farmacología , Prolina/toxicidad , Prolil Oligopeptidasas , Psicotrópicos/química , Psicotrópicos/farmacocinética , Psicotrópicos/toxicidad , Reconocimiento en Psicología/efectos de los fármacos , Reconocimiento en Psicología/fisiología , Esquizofrenia/complicaciones , Esquizofrenia/enzimología , Serina Endopeptidasas/metabolismo , Inhibidores de Serina Proteinasa/química , Inhibidores de Serina Proteinasa/farmacocinética , Inhibidores de Serina Proteinasa/farmacología , Inhibidores de Serina Proteinasa/toxicidadRESUMEN
RATIONALE: Noradrenergic system plays a critical role in the hypothalamic-pituitary-adrenal (HPA) axis regulation and the stress response. A dysregulated HPA axis may be indicative of an increased biological vulnerability for depression. In addition, a variety of studies have focused on specific alterations of α2-adrenoceptors as a mechanism involved in the pathogenesis of mood disorders and antidepressant response. OBJECTIVES: This study aimed to evaluate the effect of subchronic corticosterone administration on rat brain α2-adrenoceptor functionality by in vitro [35S]GTPγS binding stimulation assays and in vivo dual-probe microdialysis determination of extracellular noradrenaline concentrations. RESULTS: Implantation of a time release corticosterone pellet during 14 days induced sustained changes in endocrine function. However, there were no differences in α2-adrenoceptor agonist UK14304-induced stimulation of [35S]GTPγS binding in prefrontal cortex (PFC) between corticosterone-treated and control rats. In the same way, the in vivo evaluation of α2-adrenoceptor-mediated noradrenaline release responses to the α2-adrenoceptor agonist clonidine local administration into the locus coeruleus (LC), and the PFC did not show differences between the groups. CONCLUSIONS: The present results show that subchronic corticosterone administration does not induce changes on functionality of α2-adrenoceptors neither in the LC nor in noradrenergic cortical terminal areas.
Asunto(s)
Antiinflamatorios/farmacología , Corticosterona/farmacología , Receptores Adrenérgicos/efectos de los fármacos , Animales , Clonidina/farmacología , Corticosterona/sangre , Modelos Animales de Enfermedad , Sistema Hipotálamo-Hipofisario/metabolismo , Locus Coeruleus/efectos de los fármacos , Masculino , Microdiálisis , Norepinefrina/metabolismo , Sistema Hipófiso-Suprarrenal/metabolismo , Corteza Prefrontal/efectos de los fármacos , Corteza Prefrontal/metabolismo , Ratas , Ratas Sprague-DawleyRESUMEN
Cannabinoid receptors are able to couple to different families of G proteins when activated by an agonist drug. It has been suggested that different intracellular responses may be activated depending on the ligand. The goal of the present study was to characterize the pattern of G protein subunit stimulation triggered by three different cannabinoid ligands, Δ9-THC, WIN55212-2, and ACEA in mouse brain cortex. Stimulation of the [35S]GTPγS binding coupled to specific immunoprecipitation with antibodies against different subtypes of G proteins (Gαi1, Gαi2, Gαi3, Gαo, Gαz, Gαs, Gαq/11, and Gα12/13), in the presence of Δ9-THC, WIN55212-2 and ACEA (submaximal concentration 10 µM) was determined by scintillation proximity assay (SPA) technique in mouse cortex of wild type, CB1 knock-out, CB2 knock-out and CB1/CB2 double knock-out mice. Results show that, in mouse brain cortex, cannabinoid agonists are able to significantly stimulate not only the classical inhibitory Gαi/o subunits but also other G subunits like Gαz, Gαq/11, and Gα12/13. Moreover, the specific pattern of G protein subunit activation is different depending on the ligand. In conclusion, our results demonstrate that, in mice brain native tissue, different exogenous cannabinoid ligands are able to selectively activate different inhibitory and non-inhibitory Gα protein subtypes, through the activation of CB1 and/or CB2 receptors. Results of the present study may help to understand the specific molecular pathways involved in the pharmacological effects of cannabinoid-derived drugs.
RESUMEN
The orphan G protein-coupled receptor GPR55 has been directly or indirectly related to basic alterations that drive malignant growth: uncontrolled cancer cell proliferation, sustained angiogenesis, and cancer cell adhesion and migration. However, little is known about the involvement of this receptor in metastasis. Here, we show that elevated GPR55 expression in human tumors is associated with the aggressive basal/triple-negative breast cancer population, higher probability to develop metastases, and therefore poor patient prognosis. Activation of GPR55 by its proposed endogenous ligand lysophosphatidylinositol confers pro-invasive features on breast cancer cells both in vitro and in vivo. Specifically, this effect is elicited by coupling to Gq/11 heterotrimeric proteins and the subsequent activation, through ERK, of the transcription factor ETV4/PEA3. Together, these data show that GPR55 promotes breast cancer metastasis, and supports the notion that this orphan receptor may constitute a new therapeutic target and potential biomarker in the highly aggressive triple-negative subtype.
Asunto(s)
Lisofosfolípidos/farmacología , Receptores Acoplados a Proteínas G/fisiología , Neoplasias de la Mama Triple Negativas/patología , Proteínas E1A de Adenovirus/fisiología , Línea Celular Tumoral , Quinasas MAP Reguladas por Señal Extracelular/fisiología , Femenino , Subunidades alfa de la Proteína de Unión al GTP Gq-G11/fisiología , Humanos , Metástasis de la Neoplasia , Proteínas Proto-Oncogénicas/fisiología , Proteínas Proto-Oncogénicas c-ets , Receptores de Cannabinoides , Proteína de Unión al GTP rhoA/fisiologíaRESUMEN
This study examined the time course effects (8, 16 and 31 days) of fluoxetine administration (1 mg/kg, p.o./day) on serotonin transporter (5-HTT), opioid, tyrosine hydroxylase (TH) and cannabinoid CB1 receptor gene expressions in selected regions of the rat brain. Treatment with fluoxetine progressively decreased (35-55%) 5-HTT gene expression in dorsal raphe nucleus at 8, 16 and 31 days. The results revealed that fluoxetine administration decreased (30%) proenkephalin gene expression in nucleus accumbens shell (AcbS) and caudate-putamen (CPu) (31 days) but was without effect in nucleus accumbens core AcbC. A pronounced and time related decrease (25-65%) in prodynorphin gene expression was detected in AcbC, AcbS, CPu, hypothalamic supraoptic and paraventricular nuclei at all time points as well as in proopiomelanocortin gene expression (20-30%) in the arcuate nucleus (ARC) of the hypothalamus. On days 16 and 31, tyrosine hydroxylase gene expression in ventral tegmental area and substantia nigra and cannabinoid CB1 receptor gene expression in the CPu decreased (approximately 45-50% from vehicle). In conclusion, fluoxetine by inhibiting the reuptake of serotonin produced pronounced and time related alterations in genes involved in the regulation of emotional behaviour, suggesting that these neuroplastic changes may be involved, at least in part, in the clinical efficacy of this drug in neuropsychiatric disorders.
Asunto(s)
Química Encefálica/efectos de los fármacos , Endorfinas/biosíntesis , Endorfinas/genética , Fluoxetina/farmacología , Receptor Cannabinoide CB1/biosíntesis , Receptor Cannabinoide CB1/genética , Inhibidores Selectivos de la Recaptación de Serotonina/farmacología , Animales , Autorradiografía , Encefalinas/biosíntesis , Encefalinas/genética , Expresión Génica/efectos de los fármacos , Procesamiento de Imagen Asistido por Computador , Hibridación in Situ , Cinética , Masculino , Glicoproteínas de Membrana/efectos de los fármacos , Glicoproteínas de Membrana/metabolismo , Proteínas de Transporte de Membrana/efectos de los fármacos , Proteínas de Transporte de Membrana/metabolismo , Proteínas del Tejido Nervioso/efectos de los fármacos , Proteínas del Tejido Nervioso/metabolismo , Proopiomelanocortina/biosíntesis , Proopiomelanocortina/genética , Precursores de Proteínas/biosíntesis , Precursores de Proteínas/genética , ARN Mensajero/biosíntesis , ARN Mensajero/genética , Ratas , Ratas Wistar , Proteínas de Transporte de Serotonina en la Membrana Plasmática , Tirosina 3-Monooxigenasa/metabolismoRESUMEN
Despite advances in neonatal care, hypoxic-ischemic brain injury is still a serious clinical problem, which is responsible for many cases of perinatal mortality, cerebral palsy, motor impairment and cognitive deficits. Resveratrol, a natural polyphenol with important anti-oxidant and anti-inflammatory properties, is present in grapevines, peanuts and pomegranates. The aim of the present work was to evaluate the possible neuroprotective effect of resveratrol when administered before or immediately after a hypoxic-ischemic brain event in neonatal rats by analyzing brain damage, the mitochondrial status and long-term cognitive impairment. Our results indicate that pretreatment with resveratrol protects against brain damage, reducing infarct volume, preserving myelination and minimizing the astroglial reactive response. Moreover its neuroprotective effect was found to be long lasting, as behavioral outcomes were significantly improved at adulthood. We speculate that one of the mechanisms for this neuroprotection may be related to the maintenance of the mitochondrial inner membrane integrity and potential, and to the reduction of reactive oxygen species. Curiously, none of these protective features was observed when resveratrol was administered immediately after hypoxia-ischemia.
Asunto(s)
Lesiones Encefálicas/prevención & control , Trastornos del Conocimiento/prevención & control , Hipoxia-Isquemia Encefálica/tratamiento farmacológico , Fármacos Neuroprotectores/farmacología , Estilbenos/farmacología , Animales , Animales Recién Nacidos , Astrocitos/efectos de los fármacos , Astrocitos/metabolismo , Astrocitos/patología , Conducta Animal/efectos de los fármacos , Encéfalo/efectos de los fármacos , Encéfalo/metabolismo , Encéfalo/patología , Lesiones Encefálicas/patología , Lesiones Encefálicas/psicología , Trastornos del Conocimiento/psicología , Modelos Animales de Enfermedad , Femenino , Hipoxia-Isquemia Encefálica/patología , Hipoxia-Isquemia Encefálica/psicología , Masculino , Potencial de la Membrana Mitocondrial/efectos de los fármacos , Mitocondrias/efectos de los fármacos , Mitocondrias/metabolismo , Proteína Básica de Mielina/metabolismo , Fármacos Neuroprotectores/administración & dosificación , Ratas , Ratas Sprague-Dawley , Especies Reactivas de Oxígeno/metabolismo , Resveratrol , Estilbenos/administración & dosificaciónRESUMEN
OBJECTIVE: To describe the expression of cannabinoid receptors CB1 and CB2 and cannabinoid-degrading enzymes fatty acid amide hydrolase (FAAH) and monoglyceride lipase (MGLL) in human granulosa cells and to investigate their differential distribution with respect to CB1 at various stages during the nuclear maturation of the oocyte. DESIGN: Analysis of granulosa cells from germinal vesicle (GV), metaphase I (MI), and MII oocytes by quantitative reverse transcriptase-polymerase chain reaction, Western blot, and indirect immunofluorescence assays. SETTING: Academic research laboratory. PATIENT(S): Patients from the Human Reproduction Unit of Cruces University Hospital undergoing intracytoplasmic sperm injection. INTERVENTION(S): We analyzed the granulosa cells of 300 oocytes from 53 patients. The oocyte maturation stages were 75 at GV stage, 51 at MI, and 174 at MII. MAIN OUTCOME MEASURE(S): The mRNA and protein expression of CB1, CB2, FAAH, and MGLL and localization in granulosa cells at each oocyte maturation stage. RESULT(S): CB1, FAAH, and MGLL are present in human granulosa cells during oocyte maturation, but the presence of CB2 receptor is not entirely clear in those cells. CB1 and FAAH were detected in the periphery of the granulosa cells from the GV to the MII oocytes, and they colocalized in some portions of the cell membrane. On the other hand, MGLL immunostaining was more homogeneous across the cell and overlapped with CB1 only weakly. CONCLUSION(S): The presence of the cannabinoid system in granulosa cells suggests a possible role of this system in the nuclear maturation of the oocyte.