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BACKGROUND AND PURPOSE: Transient hypofunction of the NMDA receptor represents a convergence point for the onset and further development of psychiatric disorders, including schizophrenia. Although the cumulative evidence indicates dysregulation of the hippocampal formation in schizophrenia, the integrity of the synaptic transmission and plasticity conveyed by the somatosensorial inputs to the dentate gyrus, the perforant pathway synapses, have barely been explored in this pathological condition. EXPERIMENTAL APPROACH: We identified a series of synaptic alterations of the lateral and medial perforant paths in animals postnatally treated with the NMDA antagonist MK-801. This dysregulation suggests decreased cognitive performance, for which the dentate gyrus is critical. KEY RESULTS: We identified alterations in the synaptic properties of the lateral and medial perforant paths to the dentate gyrus synapses in slices from MK-801-treated animals. Altered glutamate release and decreased synaptic strength precede an impairment in the induction and expression of long-term potentiation (LTP) and CB1 receptor-mediated long-term depression (LTD). Remarkably, by inhibiting the degradation of 2-arachidonoylglycerol (2-AG), an endogenous ligand of the CB1 receptor, we restored the LTD in animals treated with MK-801. Additionally, we showed for the first time, that spatial discrimination, a cognitive task that requires dentate gyrus integrity, is impaired in animals exposed to transient hypofunction of NMDA receptors. CONCLUSION AND IMPLICATIONS: Dysregulation of glutamatergic transmission and synaptic plasticity from the entorhinal cortex to the dentate gyrus has been demonstrated, which may explain the cellular dysregulations underlying the altered cognitive processing in the dentate gyrus associated with schizophrenia.
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Giro Denteado , Maleato de Dizocilpina , Plasticidade Neuronal , Via Perfurante , Receptores de N-Metil-D-Aspartato , Animais , Receptores de N-Metil-D-Aspartato/antagonistas & inibidores , Receptores de N-Metil-D-Aspartato/metabolismo , Giro Denteado/efeitos dos fármacos , Giro Denteado/metabolismo , Maleato de Dizocilpina/farmacologia , Via Perfurante/efeitos dos fármacos , Via Perfurante/fisiologia , Plasticidade Neuronal/efeitos dos fármacos , Masculino , Ratos , Endocanabinoides/metabolismo , Receptor CB1 de Canabinoide/metabolismo , Receptor CB1 de Canabinoide/antagonistas & inibidores , Ratos Wistar , Sinapses/efeitos dos fármacos , Sinapses/metabolismo , Antagonistas de Aminoácidos Excitatórios/farmacologia , Potenciação de Longa Duração/efeitos dos fármacosRESUMO
Mitochondrial dysfunction plays a key role in the development of neurodegenerative disorders. In contrast, the regulation of the endocannabinoid system has been shown to promote neuroprotection in different neurotoxic paradigms. The existence of an active form of the cannabinoid receptor 1 (CB1R) in mitochondrial membranes (mitCB1R), which might exert its effects through the same signaling mechanisms as the cell membrane CB1R, has been shown to regulate mitochondrial activity. Although there is evidence suggesting that some cannabinoids may induce protective effects on isolated mitochondria, substantial evidence on the role of cannabinoids in mitochondria remains to be explored. In this work, we developed a toxic model of mitochondrial dysfunction induced by exposure of brain mitochondria to the succinate dehydrogenase inhibitor 3-nitropropionic acid (3-NP). Mitochondria were also pre-incubated with the endogenous agonist anandamide (AEA) and the synthetic CB1R agonist WIN 55212-2 to evaluate their protective effects. Mitochondrial reduction capacity, reactive oxygen species (ROS) formation, and mitochondrial swelling were assessed as toxic markers. While 3-NP decreased the mitochondrial reduction capacity and augmented mitochondrial ROS formation and swelling, both AEA and WIN 55212-2 ameliorated these toxic effects. To explore the possible involvement of mitCB1R activation on the protective effects of AEA and WIN 55212-2, mitochondria were also pre-incubated in the presence of the selective CB1R antagonist AM281, which completely reverted the protective effects of the cannabinoids to levels similar to those evoked by 3-NP. These results show partial protective effects of cannabinoids, suggesting that mitCB1R activation may be involved in the recovery of compromised mitochondrial activity, related to reduction of ROS formation and further prevention of mitochondrial swelling.
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Ácidos Araquidônicos , Benzoxazinas , Encéfalo , Endocanabinoides , Mitocôndrias , Morfolinas , Naftalenos , Fármacos Neuroprotetores , Nitrocompostos , Alcamidas Poli-Insaturadas , Propionatos , Ratos Wistar , Espécies Reativas de Oxigênio , Animais , Nitrocompostos/toxicidade , Propionatos/farmacologia , Propionatos/toxicidade , Mitocôndrias/metabolismo , Mitocôndrias/efeitos dos fármacos , Endocanabinoides/metabolismo , Endocanabinoides/farmacologia , Benzoxazinas/farmacologia , Ácidos Araquidônicos/farmacologia , Morfolinas/farmacologia , Espécies Reativas de Oxigênio/metabolismo , Alcamidas Poli-Insaturadas/farmacologia , Encéfalo/efeitos dos fármacos , Encéfalo/metabolismo , Encéfalo/patologia , Masculino , Fármacos Neuroprotetores/farmacologia , Naftalenos/farmacologia , Dilatação Mitocondrial/efeitos dos fármacos , Ratos , Receptor CB1 de Canabinoide/metabolismoRESUMO
Patterns of drug ingestion may have a dissimilar impact on the brain, and therefore also the development of drug addiction. One pattern is binge intoxication that refers to the ingestion of a high amount of drug on a single occasion followed by an abstinence period of variable duration. In this study, our goal was to contrast the effect of continuous low amounts with intermittent higher amounts of Arachidonyl-chloro-ethylamide (ACEA), a CB1R agonist, on amphetamine seeking and ingestion, and describe the effects on the expression of CB1R and CRFR1 in the central nucleus of the amygdala (CeA) and in the nucleus accumbens shell (NAcS). Adult male Wistar rats were treated with a daily administration of vehicle or 20 µg of ACEA, or four days of vehicle followed by 100 µg of ACEA on the fifth day, for a total of 30 days. Upon completion of this treatment, the CB1R and CRFR1 expression in the CeA and NAcS was evaluated by immunofluorescence. Additional groups of rats were evaluated for their anxiety levels (elevated plus maze, EPM), amphetamine (AMPH) self-administration (ASA) and breakpoint (A-BP), as well as AMPH-induced conditioned place preference (A-CPP). Results indicated that ACEA induced changes in the CB1R and CRFR1 expression in both the NAcS and CeA. An increase in anxiety-like behavior, ASA, A-BP and A-CPP was also observed. Since the intermittent administration of 100 µg of ACEA induced the most evident changes in most of the parameters studied, we concluded that binge-like ingestion of drugs induces changes in the brain that may make the subject more vulnerable to developing drug addiction.
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Anfetamina , Núcleo Accumbens , Ratos , Masculino , Animais , Núcleo Accumbens/metabolismo , Anfetamina/farmacologia , Ratos Wistar , Tonsila do Cerebelo , Condicionamento ClássicoRESUMO
The endocannabinoid system (ECS) refers to a complex cell-signaling system highly conserved among species formed by numerous receptors, lipid mediators (endocannabinoids) and synthetic and degradative enzymes. It is widely distributed throughout the body including the CNS, where it participates in synaptic signaling, plasticity and neurodevelopment. Besides, the olfactory ensheathing glia (OEG) present in the olfactory system is also known to play an important role in the promotion of axonal growth and/or myelination. Therefore, both OEG and the ECS promote neurogenesis and oligodendrogenesis in the CNS. Here, we investigated if the ECS is expressed in cultured OEG, by assessing the main markers of the ECS through immunofluorescence, western blotting and qRT-PCR and quantifying the content of endocannabinoids in the conditioned medium of these cells. After that, we investigated whether the production and release of endocannabinoids regulate the differentiation of oligodendrocytes co-cultured with hippocampal neurons, through Sholl analysis in oligodendrocytes expressing O4 and MBP markers. Additionally, we evaluated through western blotting the modulation of downstream pathways such as PI3K/Akt/mTOR and ERK/MAPK, being known to be involved in the proliferation and differentiation of oligodendrocytes and activated by CB1, which is the major endocannabinoid responsive receptor in the brain. Our data show that OEG expresses key genes of the ECS, including the CB1 receptor, FAAH and MAGL. Besides, we were able to identify AEA, 2-AG and AEA related mediators palmitoylethanolamide (PEA) and oleoylethanolamide (OEA), in the conditioned medium of OEG cultures. These cultures were also treated with URB597 10-9 M, a FAAH selective inhibitor, or JZL184 10-9 M, a MAGL selective inhibitor, which led to the increase in the concentrations of OEA and 2-AG in the conditioned medium. Moreover, we found that the addition of OEG conditioned medium (OEGCM) enhanced the complexity of oligodendrocyte process branching in hippocampal mixed cell cultures and that this effect was inhibited by AM251 10-6 M, a CB1 receptor antagonist. However, treatment with the conditioned medium enriched with OEA or 2-AG did not alter the process branching complexity of premyelinating oligodendrocytes, while decreased the branching complexity in mature oligodendrocytes. We also observed no change in the phosphorylation of Akt and ERK 44/42 in any of the conditions used. In conclusion, our data show that the ECS modulates the number and maturation of oligodendrocytes in hippocampal mixed cell cultures.
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Objective: The aim of this study was to identify and validate the reference genes in cultured human odontoblasts to quantify their cannabinoid receptor transcripts. Methods: The most stably transcribed genes in cultured human odontoblast cells were identified using the RefGenes tool and were selected for real-time polymerase chain reaction (PCR) amplification. Human odontoblast cells were differentiated from mesenchymal stem cells using a transforming growth factor-ß-supplemented differentiation medium, and total RNA was purified. Reverse transcription-quantitative PCR and relative quantification analyses were performed using the Schefe's method. The relative expression dataset was analyzed to select the most stable genes. Results: The analysis showed that the transcripts of cholinergic receptor nicotinic beta 2 subunit, LIM homeobox transcription factor 1 beta, and family with sequence similarity 223 member B presented the lowest standard deviation (SD) in expression (SD: 0.2, 0.17, and 0.16, respectively). These genes showed similar expression levels as the target genes (cannabinoid receptors). Significant differences were found in the relative expression levels of cannabinoid receptors using the selected genes compared to those calculated using beta actin transcripts as references (p < 0.05). Conclusions: The strategy reported here for searching and verifying new reference genes will aid in the accurate and reliable expression of cannabinoid receptors in human odontoblast cells.
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The sleep-wake cycle is a complex process that includes wake (W), non-rapid-eye-movement (NREM) and rapid-eye-movement (REM) sleep. Each phase is regulated by specialized brain structures that, by means of different neurotransmitters, maintain the constant expression of the sleep-wake cycle. Molecules like orexin, serotonin, noradrenaline, histamine, for waking; GABA, adenosine, prostaglandins, for NREM sleep and acetylcholine and glutamate for REM sleep, among other molecules are responsible for the expression and maintenance of each phase. When the endocannabinoid system was being described for the first time, almost three decades ago, oleamide's sleep promoting properties were highlighted. Nowadays, enough evidence has been cumulated to support the endocannabinoid system role in the sleep-wake cycle regulation. The endocannabinoids oleamide anandamide, and 2-arachidonylglycerol promote NREM and/or REM sleep via the CB1R, thereby making this system a target to treat sleep disorders, such as insomnia.
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Canabinoides , Encéfalo , Eletroencefalografia , Neurotransmissores , Sono , Sono REM , VigíliaRESUMO
The aim of this work was to determine whether Capsaicin may exert a vascular regulation through the activation of CB1 and/or CB2 receptors causing vasorelaxation in the rat aorta. Our results show the location of TRPV1 mainly in the endothelial and smooth muscle cells membrane. Nevertheless, Capsaicin caused vasorelaxation of this artery through a mechanism independent of TRPV1, since the specific antagonists Capsazepine and SB-366791 did not block the effect of Capsaicin. Because the significant expression of CB1 and CB2 receptors has been previously reported in the rat aorta, we used antagonists for these two receptors prior to the addition of Capsaicin. In these experiments, we found that the inhibition of CB1 using AM281, decreases the vasorelaxant effect caused by Capsaicin. On the other hand, the vasorelaxant effect is not altered in the presence of the CB2 receptor antagonist AM630. Furthermore, a partial decrease of the effect of Capsaicin was also seen when L-type calcium channels are blocked. A complete block of Capsaicin-induced vasorelaxation was achieved using a combination of Verapamil and AM281. In accordance to our results, Capsaicin-induced vasorelaxation of the rat aorta is neither dependent of TRPV1 or CB2 receptors, but rather it is strongly suggested that a tandem mechanism between inactivation of L-type calcium channels and the direct activation of CB1 receptors is involved. These findings are supported by CB1 docking simulation which predicted a binding site on CB1 receptors for Capsaicin.
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Aorta/efeitos dos fármacos , Canais de Cálcio Tipo L/metabolismo , Capsaicina/farmacologia , Endotélio Vascular/efeitos dos fármacos , Receptor CB1 de Canabinoide/metabolismo , Vasodilatação/efeitos dos fármacos , Animais , Bloqueadores dos Canais de Cálcio/farmacologia , Masculino , Ratos , Ratos Wistar , Receptor CB2 de Canabinoide/metabolismo , Canais de Cátion TRPV/metabolismoRESUMO
BACKGROUND: Cannabinoid receptor 1 has its crystallographic structure available in complex with agonists and inverse agonists, which paved the way to establish an understanding of the structural basis of interactions with ligands. Dipyrone is a prodrug with analgesic capabilities and is widely used in some countries. Recently some evidence of a dipyrone metabolite acting over the Cannabinoid Receptor 1has been shown. OBJECTIVE: Our goal here is to explore the dipyrone metabolite 4-aminoantipyrine as a Cannabinoid Receptor 1 agonist, reviewing dipyrone characteristics, and investigating the structural basis for its interaction with the Cannabinoid Receptor 1. METHOD: We reviewed here recent functional studies related to the dipyrone metabolite focusing on its action as a Cannabinoid Receptor 1 agonist. We also analyzed protein-ligand interactions for this complex obtained through docking simulations against the crystallographic structure of the Cannabinoid Receptor 1. RESULTS: Analysis of the crystallographic structure and docking simulations revealed that most of the interactions present in the docked pose were also present in the crystallographic structure of Cannabinoid Receptor 1 and agonist. CONCLUSION: Analysis of the complex of 4-aminoantipyrine and Cannabinoid Receptor 1 revealed the pivotal role played by residues Phe 170, Phe 174, Phe 177, Phe 189, Leu 193, Val 196, and Phe 379, besides the conserved hydrogen bond at Ser 383. The mechanistic analysis and the present computational study suggest that the dipyrone metabolite 4-aminoantipyrine interacts with the Cannabinoid Receptor 1.
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Agonistas de Receptores de Canabinoides/farmacologia , Ampirona , Analgésicos , Canabinoides , DipironaRESUMO
BACKGROUND Formation of schistosomal granulomata surrounding the ova can result in schistosomiasis-associated liver fibrosis (SSLF). The current standard of treatment is praziquantel (PZQ), which cannot effectively reverse SSLF. The role of the cannabinoid (CB) receptor family in liver fibrosis has recently been highlighted. OBJECTIVES This study aimed to assess the therapeutic effect of CB1 receptor antagonism in reversing SSLF in a murine model of Schistosoma mansoni infection. METHODS One hundred male Swiss albino mice were divided equally into five groups: healthy uninfected control (group I), infected control (group II), PZQ treated (group III), rimonabant (RIM) (SR141716, a CB1 receptor antagonist)-treated (group IV) and group V was treated with combined PZQ and RIM. Liver sections were obtained for histopathological examination, alpha-1 smooth muscle actin (α-SMA) immunostaining and assessment of CB1 receptor expression using real-time polymerase chain reaction (RT-PCR). FINDINGS The most effective reduction in fibrotic marker levels and granuloma load was achieved by combined treatment with PZQ+RIM (group V): CB1 receptor expression (H = 26.612, p < 0.001), number of α-SMA-positive cells (F = 57.086, p < 0.001), % hepatic portal fibrosis (F = 42.849, p < 0.001) and number of granulomata (F = 69.088, p < 0.001). MAIN CONCLUSIONS Combining PZQ with CB1 receptor antagonists yielded the best results in reversing SSLF. To our knowledge, this is the first study to test this regimen in S. mansoni infection.
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Humanos , Fibrose/diagnóstico , Tifo Endêmico Transmitido por Pulgas/transmissão , Fígado/fisiopatologia , Receptores de CanabinoidesRESUMO
Activation of the hypothalamic-pituitary-adrenal axis (HPA) is critical for survival when the organism is exposed to a stressful stimulus. The endocannabinoid system (ECS) is currently considered an important neuromodulator involved in numerous pathophysiological processes and whose primary function is to maintain homeostasis. In the tissues constituting the HPA axis, all the components of the ECS are present and the activation of this system acts in parallel with changes in the activity of numerous neurotransmitters, including nitric oxide (NO). NO is widely distributed in the brain and adrenal glands and recent studies have shown that free radicals, and in particular NO, may play a crucial role in the regulation of stress response. Our objective was to determine the participation of the endocannabinoid and NOergic systems as probable mediators of the neuroendocrine HPA axis response to a psychophysical acute stress model in the adult male rat. Animals were pre-treated with cannabinoid receptors agonists and antagonists at central and systemic level prior to acute restraint exposure. We also performed in vitro studies incubating adrenal glands in the presence of ACTH and pharmacological compounds that modifies ECS components. Our results showed that the increase in corticosterone observed after acute restraint stress is blocked by anandamide administered at both central and peripheral level. At hypothalamic level both cannabinoid receptors (CB1 and CB2) are involved, while in the adrenal gland, anandamide has a very potent effect in suppressing ACTH-induced corticosterone release that is mainly mediated by vanilloid TRPV1 receptors. We also observed that stress significantly increased hypothalamic mRNA levels of CB1 as well as adrenal mRNA levels of TRPV1 receptor. In addition, anandamide reduced the activity of the nitric oxide synthase enzyme during stress, indicating that the anti-stress action of endocannabinoids may involve a reduction in NO production at hypothalamic and adrenal levels. In conclusion, an endogenous cannabinoid tone maintains the HPA axis in a stable basal state, which is lost with a noxious stimulus. In this case, the ECS dampens the response to stress allowing the recovery of homeostasis. Moreover, our work further contributes to in vitro evidence for a participation of the endocannabinoid system by inhibiting corticosterone release directly at the adrenal gland level.
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Endocanabinoides/farmacologia , Receptor CB1 de Canabinoide/metabolismo , Estresse Psicológico/tratamento farmacológico , Animais , Ácidos Araquidônicos/metabolismo , Ácidos Araquidônicos/farmacologia , Encéfalo/metabolismo , Corticosterona/farmacologia , Endocanabinoides/metabolismo , Sistema Hipotálamo-Hipofisário/metabolismo , Masculino , Sistemas Neurossecretores/efeitos dos fármacos , Sistemas Neurossecretores/metabolismo , Óxido Nítrico/metabolismo , Sistema Hipófise-Suprarrenal/metabolismo , Alcamidas Poli-Insaturadas/metabolismo , Alcamidas Poli-Insaturadas/farmacologia , Ratos , Ratos Sprague-Dawley , Transdução de Sinais/fisiologia , Guanilil Ciclase Solúvel/efeitos dos fármacos , Estresse Psicológico/metabolismo , Canais de Cátion TRPV/efeitos dos fármacos , Canais de Cátion TRPV/metabolismoRESUMO
Drug dependence seems to involve a learning and memory process. Since learning and memory depend on protein synthesis, drug dependence may depend on protein synthesis, too. Drug-induced reward is a crucial effect for the development of drug-dependence. We used chloramphenicol (CAP, a protein synthesis inhibitor), to evaluate its effects on amphetamine (amph)-seeking behavior, on CB1R expression and on protein synthesis in general, in specific areas of the brain. Two groups of Wistar adult male rats were subjected to amph-induced conditioned place preference (CPP). Rats in group 1 received amph and were kept in the chamber for 30min. Once this period elapsed, they received a subcutaneous injection of saline (veh) and were returned to their home-cage. Rats in group 2 were also treated with amph but received CAP (150mg/kgsc) instead of saline. Once CPP was evaluated rats were sacrificed and the prefrontal cortex (PFC), the nucleus accumbens (NAcc) and the hippocampus (Hipp) were isolated and prepared for CB1R Western blot analysis. A vivarium reared group of rats was added as a non-experimentally manipulated control group. Results indicate that group 1 developed CPP while increasing CB1R expression in the NAcc. Group 2 did not develop CPP, had lower CB1R expression in the PFC and lacked the CB1R increase in the NAcc observed in the amph+veh group. These results support the notion that among the underlying mechanisms for amph-seeking reward is an increase in CB1R, further supporting an interaction between dopamine/endocannabinoids in CPP learning.
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Anfetamina/antagonistas & inibidores , Anfetamina/farmacologia , Estimulantes do Sistema Nervoso Central/antagonistas & inibidores , Estimulantes do Sistema Nervoso Central/farmacologia , Cloranfenicol/farmacologia , Condicionamento Operante/efeitos dos fármacos , Núcleo Accumbens/metabolismo , Córtex Pré-Frontal/metabolismo , Inibidores da Síntese de Proteínas/farmacologia , Receptor CB1 de Canabinoide/biossíntese , Receptor CB1 de Canabinoide/efeitos dos fármacos , Animais , Masculino , Memória/efeitos dos fármacos , Núcleo Accumbens/efeitos dos fármacos , Córtex Pré-Frontal/efeitos dos fármacos , Desempenho Psicomotor/efeitos dos fármacos , Ratos , Ratos WistarRESUMO
Abstract Background The endocannabinoid system (eCBs) is one of the modulatory systems widely expressed in the brain. It consists of receptors expressed in the cytoplasmic (CB1 and CB2), the mitochondrial membrane (CB1), and the endogenous ligands known as endocannabinoids, such as anandamide, 2AG and oleamide. CB1 has been found in excitatory and inhibitory neurons in the pre- and post-synaptic membranes. It is expressed in several brain areas such as the hippocampus, dorsal, and ventral striatum, amygdala and prefrontal cortex. The eCBs has been involved in the regulation of learning and memory, mood, energy balance, sleep, and drug addiction. Objective Integrate existing information about the eCBs and its role in brain function and mental health. Method Review of the information of basic and clinical relevance obtained from indexed scientific journals (PubMed/Medline, Scopus). Results Basic and clinical research on eCBs related to central nervous system function is described. Discussion and conclusion At present, the study of eCBs is of importance. The development of drugs that affect this system may be clinically useful to control different debilitating diseases. This is an area of interest to the scientific community and health care providers.
Resumen Antecedentes El sistema de endocannabinoides (eCBs) es uno de los sistemas moduladores más ampliamente expresados en el cerebro. Se compone de receptores expresados en la membrana citoplasmática (CB1 y CB2) y en la membrana mitocondrial (CB1) y ligandos endógenos conocidos como endocannabinoides, como anandamida, 2AG y oleamida. El CB1 se ha encontrado en neuronas excitadoras e inhibidoras, en las membranas pre- y pos-sináptica, en varias áreas cerebrales como el hipocampo, el estriado dorsal y ventral, y en la amígdala y la corteza prefrontal. El eCBs se ha relacionado con la regulación del aprendizaje y la memoria, del estado afectivo, del equilibrio energético, del sueño y del proceso de la adicción a las drogas. Objetivo Integrar la información existente sobre el eCBs y su función sobre los procesos cerebrales y la salud mental. Método Revisión de la información de relevancia básica y clínica obtenida de revistas científicas indexadas (PubMed/Medline, Scopus). Resultados Se describe de manera concisa información de interés básico y clínico de la investigación sobre el eCBs relacionada con la función del sistema nervioso central. Discusión y conclusión En la actualidad, el estudio del eCBs es indispensable debido a su potencial terapéutico. El desarrollo de fármacos que afecten este sistema puede ser clínicamente útil para controlar diferentes enfermedades debilitantes. Ésta es un área de interés para la comunidad científica y los proveedores de salud.
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In view of the fact that human marijuana users often show dry mouth symptom, the present study was attempted to examine the localization of CB1, which was originally identified in brain, in the submandibular and sublingual salivary glands of postnatal developing male mice by immunohistochemistry. In submandibular gland, CB1-immunoreactivity was positive in a majority of acinar cells in forms of granular appearance in their apical cytoplasm, while it was negative in the ducts at newborn stage. The immunoreactivity decreased in the acinar cells at P1W and no immunoreactivity was detected in the acinar cells at P3W and thereafter. The immunoreactivity was positive in ductal cells at P3W and it remained positive thereafter until P8W stage. The immunoreaction was distinct on the apical plasmalemma of the intercalated ductal cells, while it was distinct on the basal plasmalemma of the granular convoluted ductal cells. The enhanced immunostaining on the lateral plasmalemma of the granular ductal cells was discerned only on P6W. In sublingual gland, CB1-immunoreactivity was detected in the demilune acinar cells and ductal cells only on P4W. Furthermore, CB1-immunoreactivity was shown to occur in the salivary ganglionic neurons, suggesting the CB1-inhibitory action in the saliva secretion through the parasympathetic nervous transmission.
En vista de que los usuarios humanos de la marihuana a menudo presentan síntomas de sequedad oral, en el presente estudio se intentó examinar la localización de CB1, que se identificó originalmente en el cerebro, en las glándulas salivales submandibulares y sublinguales durante el desarrollo postnatal en ratones machos. En la glándula submandibular, la inmunoreactividad CB1 fue positiva en la mayoría de las células acinares de apariencia granular en su citoplasma apical, mientras que fue negativa en los conductos en la etapa de recién nacidos. La inmunorreactividad disminuyó en las células acinares en P1W y no se detectó inmunoreactividad en las células acinares en P3W. La inmunoreactividad fue positiva en las células ductales en P3W y se mantuvo positiva hasta la etapa P8W. La inmunorreacción se observó en el plasmalema apical de las células ductales intercaladas, mientras que fue distinta en el plasmalema basal de las células ductales contorneadas granulares. La inmunotinción mejorada en el plasmalema lateral de las células ductales granulares fue distingible sólo en P6W. En la glándula sublingual, se detectó inmunoreactividad CB1 en las células acinares y se observaron células ductales solamente en P4W. Además, se demostró que la inmunoreactividad CB1 se produce en las neuronas ganglionares salivales, lo que sugiere la acción CB1 inhibitoria en la secreción de saliva a través de la transmisión parasimpática nerviosa.
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Animais , Camundongos , Glândulas Salivares/metabolismo , Receptor CB1 de Canabinoide/metabolismo , Imuno-Histoquímica , Animais Recém-NascidosRESUMO
Diabetes and obesity are two universal health problems that constitute a research objective of several groups due to the lack of efficient and safe drug treatment. In this sense, cannabinoid receptor 1 (CB1) has attracted interest because of its role in food intake and metabolic balance, two targets in the control of metabolic syndrome. In this work, novel 1,5-diaryl pyrazole derivatives were synthesized in accordance with the pKi prediction of a previously reported CoMFA model by our group. To further investigate the biological activity of these compounds in metabolic disorders, their hypoglycemic activity in an in vivo model was tested. Interestingly, a high degree of correlation was observed between the predicted pKi and hypoglycemic effect 7 h after administration. Compounds 4, 9 and 13 showed the most significant plasma glucose reduction with decreases of 60%, 64% and 60% respectively. This result not only surpasses the activity of the lead rimonabant, but also that of the reference drug glibenclamide. Moreover, PASS prediction and molecular docking in an excellent validated homology model of CB1 suggest that these compounds would probably act as CB1 antagonists/inverse agonists and therefore, anti-obesity agents. The ligand-receptor complexes demonstrate that 1,5-diaryl pyrazole derivatives bind to the proposed binding site where a hydrophobic moiety interacts with the phenyl rings in the pyrazole nucleus and Lys192 forms a hydrogen bond with the oxygen of the carbonyl group. Dynamics simulations were also carried out to study the stability of the ligand-receptor complexes where the most active compounds showed smaller ΔG values and more hydrogen bonds throughout the simulation. These compounds are considered useful leads for further optimization in the treatment of such metabolic illnesses.
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Diabetes Mellitus Experimental/tratamento farmacológico , Desenho de Fármacos , Hidrazinas/síntese química , Hidrazinas/farmacologia , Hipoglicemiantes/síntese química , Hipoglicemiantes/farmacologia , Pirazóis/síntese química , Pirazóis/farmacologia , Relação Quantitativa Estrutura-Atividade , Animais , Humanos , Hidrazinas/química , Hipoglicemiantes/química , Masculino , Modelos Moleculares , Simulação de Dinâmica Molecular , Pirazóis/química , Ratos , Ratos Wistar , Receptor CB1 de Canabinoide/agonistas , Receptor CB1 de Canabinoide/antagonistas & inibidores , TermodinâmicaRESUMO
Sleep is a universal experience and a necessary ingredient to life. Young adult humans benefit from spending 8 h a day, every day, sleeping. While the function(s) of sleep is not completely understood, it is known that sleep is critical to the survival of the species. In humans, it restores alertness, helps consolidate memory and «recharge¼ cognitive abilities which are impaired at the end of the activity-phase of the cycle. Humans who do not pay their toll to sleep, for one night, for example, experience difficulties maintaining wakefulness the next day. This condition may put in danger their lives, particularly if they work in the transportation industry (i. e. taxi cabs, truck or trailers-drivers, pilots and/or operating heavy machinery among many other activities). In the past, when humans were more exposed to predators, to be sleepy in the savannah was synonymous with dying. Interestingly enough, the maladaptive strategies exhibited by a sleepy subject (which put at risk his life), are reversed by sleep. It is widely believed that sleep has a restorative function. However, what precisely is being restored during sleep remains a topic of speculation and on-going research. Sleep deprivation in humans results in cognitive deterioration and increased sleepiness, which might compromise survival as aforementioned. It is known that in rats prolonged sleep deprivation leads to death. In humans, it results in sleepiness, decreased attention, compromises memory and learning skills, and may affect motor control. The negative effects of sleep loss are reversed by recovery sleep, which may show features of delta and/or REM-sleep rebound. In this context, insomnia is a condition with known negative consequences to the health of the affected individual and frequently conveys negative effects to the family nucleus and to society in general. It has been estimated that 9% to 15% of the adult population suffers from chronic insomnia. Psychophysiologic insomnia (or primary insomnia) is likely the most prevalent type of insomnia. The patient with insomnia frequently develops an aversive response to sleep and to all aspects related to this activity. The manifestations of insomnia may include difficulty falling (and staying) asleep, frequent awakenings, early morning awakenings and/or un-refreshing quality of sleep. As a result, affected individuals frequently complain of daytime consequences such as decreased concentration, negative effects on memory (and learning), and daytime fatigue. They may also complain of headaches, fuzziness (or grogginess) and might experience manifestations of excessive sleepiness (which might represent a hazard when driving and/or operating machinery). Stress and anxiety frequently represent precipitating and/or perpetuating factors in the development of insomnia. In regards to stress, the role of the hypothalamus-pituitary-adrenal (HPA) axis in preserving homeostasis has been amply studied. The HPA axis involves the participation of peptides such as corticotropin-releasing hormone (CRH), corticotropin itself and cortisol. The autonomic nervous system activates the amygdaloid complex further enhancing the stress response. When patients are unable to control their stress response, the magnified response may be manifested as an anxiety disorder. According to the DSM-IV, the diagnosis of generalized anxiety disorder (GAD) is based on persistent symptoms of excessive anxiety and worry. Patients with GAD as well as those suffering from other anxiety disorders such as PTSD and panic attacks may manifest symptoms of insomnia. Several models of stress have been proposed to better understand these conditions. For example, prenatal stress has been suggested to increase vulnerability to life events and some reports have suggested impaired sleep among some of the animal models that have been studied. Specifically, it has been reported that rats who are deprived of maternal care suffer from dysregulation of the orexinergic system. Consequently, affected rats may have manifestations of sleep-wake dysregulation. There is more. Rats born to a low care maternal provider (which induces an early stress response) have been found to have methylated the gene that encodes the glucocorticoid receptors, which is reflected in a low expression of receptors. As a result, these animals release more corticosterone in response to stressful situations and are less efficient in managing stress. Furthermore, they have a lower expression of the CB1 receptor in several areas of the brain, thus suggesting that the systems responsible for reducing excitability of the brain (and consequently reducing the subjective sensation of fear and anxiety) are shattered. Models of insomnia evaluating the possible role of an inadequate stress-response have not been thoroughly studied. Potential pharmacologic interventions using such a theoretical framework have not been systematically studied and thus offer a venue for novel pharmacological interventions. The addition of new therapies would be particularly useful as the clinical management of patients with chronic insomnia remains a challenging area in medical practice. This despite the availability of multiple approved hypnotic medications in the physician's armamentarium. To date there is no hypnotic medication which can be considered ideal for the treatment of chronic insomnia. Issues of tolerance and dependence remain relevant concerns for those hypnotic medications, which are considered most effective in the treatment of this condition. Research identifying new compounds based on molecules whose physiologic action is to induce sleep may render safer, more efficient pharmacological interventions to treat insomnia. Following this line of thinking, we have tested the effects of endocannabinoids (eCBs). The eCBs represent a family of molecules, lipids in nature, which bind to the same receptors to which marijuana is known to bind. The active metabolite of marijuana (delta-9-tetrahydrocannabinol [THC]) is known to bind to the CB1 receptors and produce a series of effects including relaxation and sleep. Following the discovery of several eCBs by the research groups of Mechoulam and Lerner, we have tested anandamide and oleamide as sleep inducers. Results have indicated that both molecules and a third one, 2-arachidonyl glycerol, induce sleep (mainly REM sleep). In the present review insomnia is speculated to be a consequence of chronic stress. Several animal models of early stress are also discussed to better understand the role of stress in the causation of insomnia. The current limitations in the availability of ideal hypnotic medications prompt us to argue in favor of continued efforts to find additional, novel pharmacologic interventions to treat this condition. In this context, the potential use of endocannabinoid compounds is proposed as a possible new line of hypnotic medications. While eCBs have been used so far only in animal models, they have been amply successful in promoting the expression of non-REM and REM sleep. The endocannabinoid system has the potential to induce sleep and thus suggest that endocannabinoid agonists offer a new research venue for the exploration of novel pharmacologic interventions in the treatment of insomnia.
El sueño es una actividad fundamental para el bienestar y la preservación del la salud. El no dormir resulta en consecuencias potencialmente letales. Por ejemplo, ratas experimentalmente privadas de sueño total (o de sueño MOR) mueren al cabo de algunas semanas de experimentación. Asimismo, en humanos, la privación de tan solo una noche de sueño conlleva consecuencias importantes. Hipersomnolencia y disminución de las habilidades cognoscitivas son consecuencias de la privación de sueño. El individuo privado de sueño corre el riesgo de cometer errores que potencialmente pueden poner en riesgo su vida o integridad física, así como la de otros. Se ha sugerido que el sueño cumple con la muy importante tarea de ofrecer las condiciones para que se lleven a cabo diversos procesos de restauración y de reorganización neuronal así como el procesamiento de información y consolidación de la memoria. La ausencia de sueño interfiere con estos procesos con el consecuente deterioro de la conducta adaptativa del sujeto. El insomnio es un trastorno que deteriora de manera importante la calidad de vida de las personas que lo padecen. Afecta aproximadamente al 10% de la población. El insomnio se presenta en diversas formas. La clasificación internacional de los trastornos del dormir considera 11 tipos de insomnio. Entre ellos, el insomnio psicofisiológico representa el tipo de insomnio que más frecuentemente se manifiesta en la población. Se trata de un padecimiento donde el paciente desarrolla una aversión a dormir y a todo lo que se relacione con ello. Este tipo de insomnio tiene un componente de estrés que precipita la aparición del insomnio y puede contribuir a los elementos que ayudan a perpetuarlo (insomnio crónico). Se han desarrollado diversos modelos animales para el estudio del estrés y sus consecuencias. Por ejemplo, el estrés temprano inducido por privación del cuidado maternal. Asimismo, por inducción de estrés en la madre (rata) gestante. Sin embargo, se han explotado poco para evaluar el insomnio y mejor aún, para ensayar fármacos que puedan beneficiar al paciente insomne. A pesar de contar con una gran variedad de medicamentos hipnóticos, en la actualidad no existe un hipnótico ideal. Los tratamientos más efectivos con los que contamos conllevan riesgos importantes de tolerancia, adicción y potencialmente efectos colaterales. Por ello, la búsqueda de nuevos fármacos con propiedades inductoras de sueño es inaplazable. Especialmente de fármacos que sean capaces de inducir las fases de sueño delta y sueño MOR sin causar sonambulismo, somnolencia residual y/o efectos negativos en la memoria. En este contexto se discute el potencial uso terapéutico de los endocanabinoides (eCBs), ya que son ansiolíticos e inductores de las fases profundas de sueño (delta y sueño MOR). Los eCBs son moléculas endógenas que tienen una actividad semejante a la de la mariguana. Esto es debido a que tanto la mariguana como los eCBs afectan a un mismo receptor, que es el receptor canabinoide 1 (CB1). Los eCBs tienen un potencial terapéutico que hasta ahora no se ha explotado en beneficio de los pacientes que sufren de insomnio y/o ansiedad. Por ello, en esta revisión se analiza el insomnio desde el punto de vista clínico, se detallan sus características para que el médico clínico no experto pueda reconocerlo y potencialmente tratarlo; también se persigue subrayar la influencia potencial del estrés en la fisiopatogénesis de este trastorno. A pesar de que hay cada vez más información acerca de la síntesis y degradación de los eCBs (lo que es muy importante porque estos mecanismos pueden ser afectados por fármacos que inhiban la degradación o la faciliten, dependiendo de las necesidades terapéuticas) no discutiremos estos temas que se vuelven más especializados. Sin embargo, es importante que se conozca y se discuta su posible uso para beneficio del paciente. Esta revisión se centra en discutir los potenciales beneficios causados por la activación del receptor CB1 en el paciente insomne para reducir su dolencia de mal dormir.