RESUMEN
Alzheimer's disease (AD) is the most common cause of dementia, which affects more than 5 million individuals in the USA. Unfortunately, no effective therapies are currently available to prevent development of AD or to halt progression of the disease. It has been proposed that monoacylglycerol lipase (MAGL), the key enzyme degrading the endocannabinoid 2-arachidonoylglycerol (2-AG) in the brain, is a therapeutic target for AD based on the studies using the APP transgenic models of AD. While inhibition of 2-AG metabolism mitigates ß-amyloid (Aß) neuropathology, it is still not clear whether inactivation of MAGL alleviates tauopathies as accumulation and deposition of intracellular hyperphosphorylated tau protein are the neuropathological hallmark of AD. Here we show that JZL184, a potent MAGL inhibitor, significantly reduced proinflammatory cytokines, astrogliosis, phosphorylated GSK3ß and tau, cleaved caspase-3, and phosphorylated NF-kB while it elevated PPARγ in P301S/PS19 mice, a tau mouse model of AD. Importantly, tau transgenic mice treated with JZL184 displayed improvements in spatial learning and memory retention. In addition, inactivation of MAGL ameliorates deteriorations in expression of synaptic proteins in P301S/PS19 mice. Our results provide further evidence that MAGL is a promising therapeutic target for AD.
Asunto(s)
Enfermedad de Alzheimer/metabolismo , Ácidos Araquidónicos/antagonistas & inhibidores , Ácidos Araquidónicos/metabolismo , Benzodioxoles/uso terapéutico , Cognición/efectos de los fármacos , Endocannabinoides/antagonistas & inhibidores , Endocannabinoides/metabolismo , Glicéridos/antagonistas & inhibidores , Glicéridos/metabolismo , Piperidinas/uso terapéutico , Proteínas tau/metabolismo , Enfermedad de Alzheimer/tratamiento farmacológico , Enfermedad de Alzheimer/genética , Enfermedad de Alzheimer/patología , Animales , Benzodioxoles/farmacología , Cognición/fisiología , Femenino , Mediadores de Inflamación/antagonistas & inhibidores , Mediadores de Inflamación/metabolismo , Masculino , Aprendizaje por Laberinto/efectos de los fármacos , Aprendizaje por Laberinto/fisiología , Ratones , Ratones Transgénicos , Monoacilglicerol Lipasas/antagonistas & inhibidores , Monoacilglicerol Lipasas/metabolismo , Piperidinas/farmacología , Tauopatías/tratamiento farmacológico , Tauopatías/genética , Tauopatías/metabolismo , Tauopatías/patología , Proteínas tau/genéticaRESUMEN
A wide body of evidence supports an integral role for mesolimbic dopamine (DA) in motivated behavior. In brief, drugs that increase DA in mesolimbic terminal regions, like cocaine, enhance motivation, while drugs that decrease DA concentration reduce motivation. Data from our laboratory and others shows that phasic activation of mesolimbic DA requires signaling at cannabinoid type-1 (CB1) receptors in the ventral tegmental area (VTA), and systemic delivery of CB1 receptor antagonists reduces DA cell activity and attenuates motivated behaviors. Recent findings demonstrate that cocaine mobilizes the endocannabinoid 2-arachidonoylglycerol (2-AG) in the VTA to cause phasic activation of DA neurons and terminal DA release. It remains unclear, however, if cocaine-induced midbrain 2-AG signaling contributes to the motivation-enhancing effects of cocaine. To examine this, we trained male and female rats on a progressive ratio (PR) task for a food reinforcer. Each rat underwent a series of tests in which they were pretreated with cocaine alone or in combination with systemic or intra-VTA administration of the CB1 receptor antagonist rimonabant or the 2-AG synthesis inhibitor tetrahydrolipstatin (THL). Cocaine increased motivation, measured by augmented PR breakpoints, while rimonabant dose-dependently decreased motivation. Importantly, intra-VTA administration of rimonabant or THL, at doses that did not decrease breakpoints on their own, blocked systemic cocaine administration from increasing breakpoints in male and female rats. These data suggest that cocaine-induced increases in motivation require 2-AG signaling at CB1 receptors in the VTA and may provide critical insight into cannabinoid-based pharmacotherapeutic targets for the successful treatment of substance abuse.
Asunto(s)
Ácidos Araquidónicos/antagonistas & inhibidores , Cocaína/farmacología , Endocannabinoides/antagonistas & inhibidores , Glicéridos/antagonistas & inhibidores , Motivación/efectos de los fármacos , Receptor Cannabinoide CB1/antagonistas & inhibidores , Área Tegmental Ventral/efectos de los fármacos , Animales , Condicionamiento Operante/efectos de los fármacos , Femenino , Masculino , Ratas , Ratas Long-Evans , Recompensa , Rimonabant/farmacología , AutoadministraciónRESUMEN
BACKGROUND: Cannabinoids induce biphasic effects on memory depending on stress levels. We previously demonstrated that different stress intensities, experienced soon after encoding, impaired rat short-term recognition memory in a time-of-day-dependent manner, and that boosting endocannabinoid anandamide (AEA) levels restored memory performance. Here, we examined if two different stress intensities and time-of-day alter hippocampal endocannabinoid tone, and whether these changes modulate short-term memory. METHODS: Male Sprague-Dawley rats were subjected to an object recognition task and exposed, at two different times of the day (i.e., morning or afternoon), to low or high stress conditions, immediately after encoding. Memory retention was assessed 1 hr later. Hippocampal AEA and 2-arachidonoyl glycerol (2-AG) content and the activity of their primary degrading enzymes, fatty acid amide hydrolase (FAAH) and monoacylglycerol lipase (MAGL), were measured soon after testing. RESULTS: Consistent with our previous findings, low stress impaired 1-hr memory performance only in the morning, whereas exposure to high stress impaired memory independently of testing time. Stress exposure decreased AEA levels independently of memory alterations. Interestingly, exposure to high stress decreased 2-AG content and, accordingly, increased MAGL activity, selectively in the afternoon. Thus, to further evaluate 2-AG's role in the modulation of short-term recognition memory, rats were given bilateral intra-hippocampal injections of the 2-AG hydrolysis inhibitor KML29 immediately after training, then subjected to low or high stress conditions and tested 1 hr later. CONCLUSIONS: KML29 abolished the time-of-day-dependent impairing effects of stress on short-term memory, ameliorating short-term recognition memory performance.
Asunto(s)
Ácidos Araquidónicos/metabolismo , Endocannabinoides/metabolismo , Glicéridos/metabolismo , Hipocampo/metabolismo , Memoria a Corto Plazo/fisiología , Amidohidrolasas/genética , Animales , Ácidos Araquidónicos/antagonistas & inhibidores , Ácidos Araquidónicos/genética , Benzodioxoles/farmacología , Emociones/fisiología , Endocannabinoides/antagonistas & inhibidores , Endocannabinoides/genética , Glicéridos/antagonistas & inhibidores , Glicéridos/genética , Hipocampo/efectos de los fármacos , Hipocampo/fisiología , Humanos , Masculino , Monoacilglicerol Lipasas/genética , Piperidinas/farmacología , Alcamidas Poliinsaturadas/metabolismo , Ratas , Ratas Sprague-Dawley , Transducción de Señal/genéticaRESUMEN
BACKGROUND: Ethanol (EtOH) self-administration is particularly sensitive to the modulation of CB1 signaling in the nucleus accumbens (NAc) shell, and EtOH consumption increases extracellular levels of the endogenous cannabinoid CB1 receptor agonist 2-arachidonoyl glycerol (2-AG) in this brain region. Stimulation of CB1 receptor with agonists increases EtOH consumption, suggesting that EtOH-induced increases in 2-AG might sustain motivation for EtOH intake. METHODS: In order to further explore this hypothesis, we analyzed the alterations in operant EtOH self-administration induced by intra-NAc shell infusions of 2-AG itself, the CB1 inverse agonist SR141716A, the 2-AG clearance inhibitor URB602, anandamide, and the cyclooxygenase-2 (COX-2) inhibitor nimesulide. RESULTS: Surprisingly, self-administration of 10% EtOH was dose-dependently reduced by either intra-NAc shell SR141716A or 2-AG infusions. Similar effects were found by intra-NAc shell infusions of URB602, suggesting again a role for accumbal 2-AG on the modulation of EtOH intake. Intra-NAc shell anandamide did not alter EtOH self-administration, pointing to a specific role for 2-AG in the modulation of EtOH self-administration. Finally, the inhibitory effect of intra-NAc shell 2-AG on EtOH intake was significantly reversed by pretreatment with nimesulide, suggesting that oxidative metabolites of 2-AG might mediate these inhibitory effects on operant self-administration. CONCLUSIONS: We propose that 2-AG signaling in the NAc exerts an inhibitory influence on EtOH consumption through a non-CB1 receptor mechanism involving the COX-2 pathway.
Asunto(s)
Consumo de Bebidas Alcohólicas/tratamiento farmacológico , Ácidos Araquidónicos/farmacología , Inhibidores de la Ciclooxigenasa 2/farmacología , Endocannabinoides/farmacología , Glicéridos/farmacología , Núcleo Accumbens/efectos de los fármacos , Animales , Ácidos Araquidónicos/antagonistas & inhibidores , Compuestos de Bifenilo/farmacología , Relación Dosis-Respuesta a Droga , Endocannabinoides/antagonistas & inhibidores , Glicéridos/antagonistas & inhibidores , Masculino , Núcleo Accumbens/fisiología , Alcamidas Poliinsaturadas/farmacología , Ratas , Ratas Wistar , Receptor Cannabinoide CB1/efectos de los fármacos , Receptor Cannabinoide CB1/metabolismo , Rimonabant/farmacología , Autoadministración , Sulfonamidas/farmacologíaRESUMEN
While the endocannabinoid 2-arachidonoylglycerol (2-AG) is thought to enhance the proliferation and differentiation of oligodendrocyte progenitor cells (OPCs) in vitro, less is known about how endogenous 2-AG may influence the migration of these cells. When we assessed this in Agarose drop and Boyden chemotaxis chamber assays, inhibiting the sn-1-diacylglycerol lipases α and ß (DAGLs) that are responsible for 2-AG synthesis significantly reduced the migration of OPCs stimulated by platelet-derived growth factor-AA (PDGF) and basic fibroblast growth factor (FGF). Likewise, antagonists of the CB1 and CB2 cannabinoid receptors (AM281 and AM630, respectively) produced a similar inhibition of OPC migration. By contrast, increasing the levels of endogenous 2-AG by blocking its degradation (impairing monoacylglycerol lipase activity with JZL-184) significantly increased OPC migration, as did agonists of the CB1, CB2 or CB1/CB2 cannabinoid receptors. This latter effect was abolished by selective CB1 or CB2 antagonists, strongly suggesting that cannabinoid receptor activation specifically potentiates OPC chemotaxis and chemokinesis in response to PDGF/FGF. Furthermore, the chemoattractive activity of these cannabinoid receptor agonists on OPCs was even evident in the absence of PDGF/FGF. In cultured brain slices prepared from the corpus callosum of postnatal rat brains, DAGL or cannabinoid receptor inhibition substantially diminished the in situ migration of Sox10+ OPCs. Overall, these results reveal a novel function of endogenous 2-AG in PDGF and FGF induced OPC migration, highlighting the importance of the endocannabinoid system in regulating essential steps in oligodendrocyte development.
Asunto(s)
Ácidos Araquidónicos/fisiología , Movimiento Celular , Endocannabinoides/fisiología , Glicéridos/fisiología , Oligodendroglía/fisiología , Células Madre/fisiología , Animales , Ácidos Araquidónicos/antagonistas & inhibidores , Ácidos Araquidónicos/biosíntesis , Ácidos Araquidónicos/metabolismo , Agonistas de Receptores de Cannabinoides/farmacología , Antagonistas de Receptores de Cannabinoides/farmacología , Movimiento Celular/efectos de los fármacos , Células Cultivadas , Cuerpo Calloso/citología , Endocannabinoides/antagonistas & inhibidores , Endocannabinoides/biosíntesis , Endocannabinoides/metabolismo , Glicéridos/antagonistas & inhibidores , Glicéridos/biosíntesis , Glicéridos/metabolismo , Ratas WistarRESUMEN
BACKGROUND: Acute neurological insults caused by infection, systemic inflammation, ischemia, or traumatic injury are often associated with breakdown of the blood-brain barrier (BBB) followed by infiltration of peripheral immune cells, cytotoxic proteins, and water. BBB breakdown and extravasation of these peripheral components into the brain parenchyma result in inflammation, oxidative stress, edema, excitotoxicity, and neurodegeneration. These downstream consequences of BBB dysfunction can drive pathophysiological processes and play a substantial role in the morbidity and mortality of acute and chronic neurological insults, and contribute to long-term sequelae. Preserving or rescuing BBB integrity and homeostasis therefore represents a translational research area of high therapeutic potential. METHODS: Induction of general and localized BBB disruption in mice was carried out using systemic administration of LPS and focal photothrombotic ischemic insult, respectively, in the presence and absence of the monoacylglycerol lipase (MAGL) inhibitor, CPD-4645. The effects of CPD-4645 treatment were assessed by gene expression analysis performed on neurovascular-enriched brain fractions, cytokine and inflammatory mediator measurement, and functional assessment of BBB permeability. The mechanism of action of CPD-4645 was studied pharmacologically using inverse agonists/antagonists of the cannabinoid receptors CB1 and CB2. RESULTS: Here, we demonstrate that the neurovasculature exhibits a unique transcriptional signature following inflammatory insults, and pharmacological inhibition of MAGL using a newly characterized inhibitor rescues the transcriptional profile of brain vasculature and restores its functional homeostasis. This pronounced effect of MAGL inhibition on blood-brain barrier permeability is evident following both systemic inflammatory and localized ischemic insults. Mechanistically, the protective effects of the MAGL inhibitor are partially mediated by cannabinoid receptor signaling in the ischemic brain insult. CONCLUSIONS: Our results support considering MAGL inhibitors as potential therapeutics for BBB dysfunction and cerebral edema associated with inflammatory brain insults.
Asunto(s)
Ácidos Araquidónicos/antagonistas & inhibidores , Ácidos Araquidónicos/metabolismo , Barrera Hematoencefálica/metabolismo , Lesiones Encefálicas/tratamiento farmacológico , Lesiones Encefálicas/metabolismo , Permeabilidad Capilar/fisiología , Endocannabinoides/antagonistas & inhibidores , Endocannabinoides/metabolismo , Glicéridos/antagonistas & inhibidores , Glicéridos/metabolismo , Animales , Barrera Hematoencefálica/efectos de los fármacos , Lesiones Encefálicas/inducido químicamente , Permeabilidad Capilar/efectos de los fármacos , Inhibidores Enzimáticos/farmacología , Inhibidores Enzimáticos/uso terapéutico , Hidrólisis/efectos de los fármacos , Lipopolisacáridos , Masculino , Ratones , Ratones Endogámicos C57BL , Monoacilglicerol Lipasas/antagonistas & inhibidores , Monoacilglicerol Lipasas/metabolismoRESUMEN
Current therapeutics are not effective for orofacial neuropathic pain, and better options are needed. The present study used inferior orbital nerve (ION)-injured mice to investigate the effect of inhibiting monoacylglycerol lipase (MAGL), an enzyme that degrades the major endocannabinoid 2-arachydonoylgycerol (2-AG) in orofacial neuropathic pain. The head-withdrawal threshold to mechanical stimulation of the whisker pad was reduced on days 3, 5, and 7 after ION injury. Injection of JZL184, a selective inhibitor of MAGL, on day 7 after ION injury attenuated the reduction in head-withdrawal threshold at 2 h after administration. Moreover, the numbers of MAGL-immunoreactive neurons in the trigeminal subnucleus caudalis (Vc) and upper cervical spinal cord (C1-C2) were significantly greater in ION-injured mice than in sham-operated mice but were reduced after administration of JZL184. The increase in MAGL immunoreactivity suggests that increased 2-AG production is followed by rapid enzymatic degradation of 2-AG. JZL184 inhibited this degradation and thus increased 2-AG concentration in the brain, particularly in the Vc and C1-C2 regions, thus attenuating pain. Our findings suggest that inhibition of 2-AG degradation by MAGL inhibitors is a promising therapeutic option for treatment of orofacial neuropathic pain.
Asunto(s)
Ácidos Araquidónicos/antagonistas & inhibidores , Endocannabinoides/antagonistas & inhibidores , Dolor Facial/prevención & control , Glicéridos/antagonistas & inhibidores , Neuralgia/prevención & control , Traumatismos del Nervio Trigémino/complicaciones , Animales , Ácidos Araquidónicos/metabolismo , Conducta Animal , Benzodioxoles/farmacología , Endocannabinoides/metabolismo , Inhibidores Enzimáticos/farmacología , Dolor Facial/etiología , Glicéridos/metabolismo , Masculino , Ratones Endogámicos C57BL , Monoacilglicerol Lipasas/antagonistas & inhibidores , Monoacilglicerol Lipasas/metabolismo , Neuralgia/etiología , Piperidinas/farmacologíaRESUMEN
Cell-cell communication plays a central role in the guidance of migrating neuronal precursor cells during the development of the cerebral cortex. Endocannabinoids (eCBs) have previously been shown to be one of the central factors regulating neuronal migration. In this study the effects of eCBs on different parameters, expected to affect embryonic cortical neuronal motility have been analyzed in neurosphere-derived neuroblasts using time-lapse microscopy. Increased endogenous production of the endocannabinoid 2-arachidonyl glycerol (2-AG) causes bursts of neuroblast motility. The neuroblasts move longer distances and show a low frequency of turning, and the number of neuron-neuron contacts are reduced. Similar changes occur interfering with the function of the metabotropic glutamate receptor 5 (mGluR5) or its transducer canonical transient receptor potential channel 3 (TRPC3) or the neuregulin receptor ErbB4. Blocking of 2-AG production reverses these effects. The data suggest that eCB-regulated neuronal motility is controlled by mGluR5/TRPC3 activity possibly via NRG/ErbB4 signaling.
Asunto(s)
Comunicación Celular/fisiología , Movimiento Celular/fisiología , Endocannabinoides/metabolismo , Neuronas/metabolismo , Receptor del Glutamato Metabotropico 5/metabolismo , Canales Catiónicos TRPC/metabolismo , Animales , Ácidos Araquidónicos/antagonistas & inhibidores , Ácidos Araquidónicos/metabolismo , Comunicación Celular/efectos de los fármacos , Movimiento Celular/efectos de los fármacos , Células Cultivadas , Endocannabinoides/antagonistas & inhibidores , Receptores ErbB/metabolismo , Glicéridos/antagonistas & inhibidores , Glicéridos/metabolismo , Ratones Endogámicos C57BL , Proteínas Asociadas a Microtúbulos/metabolismo , Células-Madre Neurales/citología , Células-Madre Neurales/efectos de los fármacos , Células-Madre Neurales/metabolismo , Neurregulina-1/metabolismo , Neuronas/citología , Neuronas/efectos de los fármacos , Receptor Cannabinoide CB1/metabolismo , Receptor ErbB-4/metabolismoRESUMEN
An improved understanding of the endocannabinoid system has provided new avenues of drug discovery and development toward the management of pain and other behavioral maladies. Exogenous cannabinoid type 1 (CB1) receptor agonists such as Δ9-tetrahydrocannabinol are increasingly used for their medicinal actions; however, their utility is constrained by concern regarding abuse-related subjective effects. This has led to growing interest in the clinical benefit of indirectly enhancing the activity of the highly labile endocannabinoids N-arachidonoylethanolamine [AEA (or anandamide)] and/or 2-arachidonoylglycerol (2-AG) via catabolic enzyme inhibition. The present studies were conducted to determine whether such actions can lead to CB1 agonist-like subjective effects, as reflected in CB1-related discriminative stimulus effects in laboratory subjects. Squirrel monkeys (n = 8) that discriminated the CB1 full agonist AM4054 (0.01 mg/kg) from vehicle were used to study, first, the inhibitors of fatty acid amide hydrolase (FAAH) or monoacylglycerol lipase (MGL) alone or in combination [FAAH (URB597, AM4303); MGL (AM4301); FAAH/MGL (JZL195, AM4302)] and, second, the ability of the endocannabinoids AEA and 2-AG to produce CB1 agonist-like effects when administered alone or after enzyme inhibition. Results indicate that CB1-related discriminative stimulus effects were produced by combined, but not selective, inhibition of FAAH and MGL, and that these effects were nonsurmountably antagonized by low doses of rimonabant. Additionally, FAAH or MGL inhibition revealed CB1-like subjective effects produced by AEA but not by 2-AG. Taken together, the present data suggest that therapeutic effects of combined, but not selective, enhancement of AEA or 2-AG activity via enzyme inhibition may be accompanied by CB1 receptor-mediated subjective effects.
Asunto(s)
Amidohidrolasas/antagonistas & inhibidores , Aprendizaje Discriminativo/efectos de los fármacos , Drogas en Investigación/farmacología , Endocannabinoides/farmacología , Inhibidores Enzimáticos/farmacología , Monoacilglicerol Lipasas/antagonistas & inhibidores , Receptor Cannabinoide CB1/agonistas , Adamantano/administración & dosificación , Adamantano/efectos adversos , Adamantano/análogos & derivados , Adamantano/farmacología , Amidohidrolasas/metabolismo , Animales , Ácidos Araquidónicos/administración & dosificación , Ácidos Araquidónicos/agonistas , Ácidos Araquidónicos/antagonistas & inhibidores , Ácidos Araquidónicos/farmacología , Conducta Animal/efectos de los fármacos , Antagonistas de Receptores de Cannabinoides/administración & dosificación , Antagonistas de Receptores de Cannabinoides/efectos adversos , Antagonistas de Receptores de Cannabinoides/farmacología , Cannabinol/administración & dosificación , Cannabinol/efectos adversos , Cannabinol/análogos & derivados , Cannabinol/farmacología , Relación Dosis-Respuesta a Droga , Agonismo de Drogas , Antagonismo de Drogas , Drogas en Investigación/administración & dosificación , Drogas en Investigación/efectos adversos , Endocannabinoides/administración & dosificación , Endocannabinoides/agonistas , Endocannabinoides/antagonistas & inhibidores , Inhibidores Enzimáticos/administración & dosificación , Inhibidores Enzimáticos/efectos adversos , Glicéridos/administración & dosificación , Glicéridos/agonistas , Glicéridos/antagonistas & inhibidores , Glicéridos/farmacología , Inyecciones Intramusculares , Inyecciones Intravenosas , Ligandos , Masculino , Monoacilglicerol Lipasas/metabolismo , Proteínas del Tejido Nervioso/agonistas , Proteínas del Tejido Nervioso/antagonistas & inhibidores , Proteínas del Tejido Nervioso/metabolismo , Alcamidas Poliinsaturadas , Receptor Cannabinoide CB1/antagonistas & inhibidores , Receptor Cannabinoide CB1/metabolismo , SaimiriRESUMEN
We investigated the effects of S-777469 (1-[[6-Ethyl-1-[4-fluorobenzyl]-5-methyl-2-oxo-1, 2-dihydropyridine-3-carbonyl]amino]-cyclohexanecarboxylic acid), a novel cannabinoid type 2 receptor (CB2) agonist, on 1-fluoro-2,4-dinitrobenzene (DNFB)-induced ear inflammation and mite antigen-induced dermatitis in mice. The oral administration of S-777469 significantly suppressed DNFB-induced ear swelling in a dose-dependent manner. In addition, S-777469 significantly alleviated mite antigen-induced atopic dermatitis-like skin lesions in NC/Nga mice. A histological analysis revealed that S-777469 significantly reduced the epidermal thickness and the number of mast cells infiltrating skin lesions. We demonstrated that S-777469 inhibited mite antigen-induced eosinophil accumulation in skin lesions and an endogenous CB2 ligand, 2-arachidonoylglycerol (2-AG)-induced eosinophil migration in vitro. Moreover, we confirmed that 2-AG levels significantly increased in skin lesions of mite antigen-induced dermatitis model. Together, these results suggest that S-777469 inhibits skin inflammation in mice by blocking the activities of 2-AG.
Asunto(s)
Inflamación/tratamiento farmacológico , Piridonas/farmacología , Piridonas/uso terapéutico , Receptor Cannabinoide CB2/agonistas , Piel/efectos de los fármacos , Piel/patología , Animales , Ácidos Araquidónicos/antagonistas & inhibidores , Ácidos Araquidónicos/metabolismo , Ensayos de Migración de Leucocitos , Dermatitis Atópica/tratamiento farmacológico , Dermatitis Atópica/metabolismo , Dinitrofluorobenceno , Relación Dosis-Respuesta a Droga , Endocannabinoides/antagonistas & inhibidores , Endocannabinoides/metabolismo , Glicéridos/antagonistas & inhibidores , Glicéridos/metabolismo , Inflamación/inducido químicamente , Masculino , Ratones , Infestaciones por Ácaros/tratamiento farmacológico , Infestaciones por Ácaros/metabolismoRESUMEN
PURPOSE: Cannabinoids, such as Δ9-THC, act through an endogenous signaling system in the vertebrate eye that reduces IOP via CB1 receptors. Endogenous cannabinoid (eCB) ligand, 2-arachidonoyl glycerol (2-AG), likewise activates CB1 and is metabolized by monoacylglycerol lipase (MAGL). We investigated ocular 2-AG and its regulation by MAGL and the therapeutic potential of harnessing eCBs to lower IOP. METHODS: We tested the effect of topical application of 2-AG and MAGL blockers in normotensive mice and examined changes in eCB-related lipid species in the eyes and spinal cord of MAGL knockout (MAGL-/-) mice using high performance liquid chromatography/tandem mass spectrometry (HPLC/MS/MS). We also examined the protein distribution of MAGL in the mouse anterior chamber. RESULTS: 2-Arachidonoyl glycerol reliably lowered IOP in a CB1- and concentration-dependent manner. Monoacylglycerol lipase is expressed prominently in nonpigmented ciliary epithelium. The MAGL blocker KML29, but not JZL184, lowered IOP. The ability of CB1 to lower IOP is not desensitized in MAGL-/- mice. Ocular monoacylglycerols, including 2-AG, are elevated in MAGL-/- mice but, in contrast to the spinal cord, arachidonic acid and prostaglandins are not changed. CONCLUSIONS: Our data confirm a central role for MAGL in metabolism of ocular 2-AG and related lipid species, and that endogenous 2-AG can be harnessed to reduce IOP. The MAGL blocker KML29 has promise as a therapeutic agent, while JZL184 may have difficulty crossing the cornea. These data, combined with the relative specificity of MAGL for ocular monoacylglycerols and the lack of desensitization in MAGL-/- mice, suggest that the development of an optimized MAGL blocker offers therapeutic potential for treatment of elevated IOP.
Asunto(s)
Ácidos Araquidónicos/fisiología , Endocannabinoides/fisiología , Glicéridos/fisiología , Presión Intraocular/fisiología , Monoacilglicerol Lipasas/fisiología , Administración Tópica , Animales , Cámara Anterior/metabolismo , Ácidos Araquidónicos/antagonistas & inhibidores , Ácidos Araquidónicos/metabolismo , Ácidos Araquidónicos/farmacología , Benzodioxoles , Cuerpo Ciliar/metabolismo , Córnea/metabolismo , Endocannabinoides/antagonistas & inhibidores , Endocannabinoides/metabolismo , Endocannabinoides/farmacología , Glicéridos/antagonistas & inhibidores , Glicéridos/metabolismo , Glicéridos/farmacología , Inmunohistoquímica , Presión Intraocular/efectos de los fármacos , Ratones , Ratones Endogámicos C57BL , Ratones Noqueados , Monoacilglicerol Lipasas/antagonistas & inhibidores , Monoacilglicerol Lipasas/metabolismo , Monoglicéridos/metabolismo , Piperidinas , Conejos , Espectrometría de Masas en TándemRESUMEN
Chronic nicotine exposure (CNE) alters synaptic transmission in the ventral tegmental area (VTA) in a manner that enhances dopaminergic signaling and promotes nicotine use. The present experiments identify a correlation between enhanced production of the endogenous cannabinoid 2-arachidonoylglycerol (2-AG) and diminished release of the inhibitory neurotransmitter GABA in the VTA following CNE. To study the functional role of on-demand 2-AG signaling in GABAergic synapses, we used 1,2,3-triazole urea compounds to selectively inhibit 2-AG biosynthesis by diacylglycerol lipase (DAGL). The potency and selectivity of these inhibitors were established in rats in vitro (rat brain proteome), ex vivo (brain slices), and in vivo (intracerebroventricular administration) using activity-based protein profiling and targeted metabolomics analyses. Inhibition of DAGL (2-AG biosynthesis) rescues nicotine-induced VTA GABA signaling following CNE. Conversely, enhancement of 2-AG signaling in naïve rats by inhibiting 2-AG degradation recapitulates the loss of nicotine-induced GABA signaling evident following CNE. DAGL inhibition reduces nicotine self-administration without disrupting operant responding for a nondrug reinforcer or motor activity. Collectively, these findings provide a detailed characterization of selective inhibitors of rat brain DAGL and demonstrate that excessive 2-AG signaling contributes to a loss of inhibitory GABAergic constraint of VTA excitability following CNE.
Asunto(s)
Lipoproteína Lipasa/antagonistas & inhibidores , Nicotina/farmacología , Área Tegmental Ventral/efectos de los fármacos , Animales , Ácidos Araquidónicos/análisis , Ácidos Araquidónicos/antagonistas & inhibidores , Ácidos Araquidónicos/fisiología , Endocannabinoides/análisis , Endocannabinoides/antagonistas & inhibidores , Endocannabinoides/fisiología , Glicéridos/análisis , Glicéridos/antagonistas & inhibidores , Glicéridos/fisiología , Masculino , Ratas , Ratas Wistar , Autoadministración , Área Tegmental Ventral/fisiología , Ácido gamma-Aminobutírico/metabolismoRESUMEN
In eukaryotic cells, phosphatidic acid (PA) and diacylglycerol (DAG), are at the origin of all membrane glycerolipids. Their interconversion is achieved by dephosphorylation of PA and phosphorylation of DAG: they form therefore a metabolic hub. PA and DAG are also known to be versatile signaling molecules. Two independent pharmacological screenings conducted on plant and human targets, led to the discovery of a new family of compounds acting on enzymes binding to either PA or DAG, in biological contexts that seemed initially independent. On the one hand, in plants, monogalactosyldiacylglycerol synthases (MGDG synthases or MGD) are responsible for the synthesis of MGDG, which is the most profuse lipid of photosynthetic membranes, and thus essential for metabolism and development. MGD use DAG as substrate. On the other hand, in mammals, phospholipases D (PLD), that produce PA, are involved in a variety of signaling cascades that control a broad spectrum of cellular functions, and play a role in the development of cancers. The two independent pharmacological screenings described in this review aimed to identify inhibitory molecules of either MGD of the plant model Arabidopsis, or human PLD. In both cases, the obtained molecules are piperidinyl-benzimidazolone derivatives, thereby allowing to propose this family of molecules as a novel source of inspiration for the search of compounds interfering with glycerolipid metabolism, that could be useful for other biological and therapeutics contexts.
Asunto(s)
Descubrimiento de Drogas/métodos , Evaluación Preclínica de Medicamentos , Inhibidores Enzimáticos/aislamiento & purificación , Glicéridos/antagonistas & inhibidores , Glicéridos/metabolismo , Metabolismo de los Lípidos/efectos de los fármacos , Galactosiltransferasas/antagonistas & inhibidores , Humanos , Invenciones , Fosfolipasa D/antagonistas & inhibidores , PlantasRESUMEN
The endocannabinoid ligand 2-arachidonoylglycerol (2-AG) is inactivated primarily by monoacylglycerol lipase (MAGL). We have shown recently that chronic treatments with MAGL inhibitor JZL184 produce antidepressant- and anxiolytic-like effects in a chronic unpredictable stress (CUS) model of depression in mice. However, the underlying mechanisms remain poorly understood. Adult hippocampal neurogenesis has been implicated in animal models of anxiety and depression and behavioral effects of antidepressants. We tested whether CUS and chronic JZL184 treatments affected adult neurogenesis and synaptic plasticity in the dentate gyrus (DG) of mouse hippocampus. We report that CUS induced depressive-like behaviors and decreased the number of bromodeoxyuridine-labeled neural progenitor cells and doublecortin-positive immature neurons in the DG, while chronic JZL184 treatments prevented these behavioral and cellular deficits. We also investigated the effects of CUS and chronic JZL184 on a form long-term potentiation (LTP) in the DG known to be neurogenesis-dependent. CUS impaired LTP induction, whereas chronic JZL184 treatments restored LTP in CUS-exposed mice. These results suggest that enhanced adult neurogenesis and long-term synaptic plasticity in the DG of the hippocampus might contribute to antidepressant- and anxiolytic-like behavioral effects of JZL184.
Asunto(s)
Antidepresivos/farmacología , Ácidos Araquidónicos/antagonistas & inhibidores , Conducta Animal/fisiología , Benzodioxoles/farmacología , Agonistas de Receptores de Cannabinoides/metabolismo , Giro Dentado/fisiopatología , Depresión/fisiopatología , Endocannabinoides/antagonistas & inhibidores , Glicéridos/antagonistas & inhibidores , Monoacilglicerol Lipasas/antagonistas & inhibidores , Neurogénesis/fisiología , Plasticidad Neuronal/fisiología , Piperidinas/farmacología , Estrés Psicológico/fisiopatología , Animales , Ansiolíticos/farmacología , Conducta Animal/efectos de los fármacos , Giro Dentado/efectos de los fármacos , Depresión/tratamiento farmacológico , Hidrólisis/efectos de los fármacos , Masculino , Ratones , Ratones Endogámicos C57BL , Neurogénesis/efectos de los fármacos , Plasticidad Neuronal/efectos de los fármacosRESUMEN
Increased vascular sensitivity to angiotensin II (Ang II) is a marker of a hypertensive human pregnancy. Recent evidence of interactions between the renin-angiotensin system and the endocannabinoid system suggests that anandamide and 2-arachidonoylglycerol may modulate Ang II contraction. We hypothesized that these interactions may contribute to the enhanced vascular responses in hypertensive pregnancy. We studied Ang II contraction in isolated uterine artery (UA) at early gestation in a rat model that mimics many features of preeclampsia, the transgenic human angiotensinogen×human renin (TgA), and control Sprague-Dawley rats. We determined the role of the cannabinoid receptor 1 by blockade with SR171416A, and the contribution of anandamide and 2-arachidonoylglycerol degradation to Ang II contraction by inhibiting their hydrolyzing enzyme fatty acid amide hydrolase (with URB597) or monoacylglycerol lipase (with JZL184), respectively. TgA UA showed increased maximal contraction and sensitivity to Ang II that was inhibited by indomethacin. Fatty acid amide hydrolase blockade decreased Ang IIMAX in Sprague-Dawley UA, and decreased both Ang IIMAX and sensitivity in TgA UA. Monoacylglycerol lipase blockade had no effect on Sprague-Dawley UA and decreased Ang IIMAX and sensitivity in TgA UA. Blockade of the cannabinoid receptor 1 in TgA UA had no effect. Immunolocalization of fatty acid amide hydrolase and monoacylglycerol lipase showed a similar pattern between groups; fatty acid amide hydrolase predominantly localized in endothelium and monoacylglycerol lipase in smooth muscle cells. We demonstrated an increased Ang II contraction in TgA UA before initiation of the hypertensive phenotype. Anandamide and 2-arachidonoylglycerol reduced Ang II contraction in a cannabinoid receptor 1-independent manner. These renin-angiotensin system-endocannabinoid system interactions may contribute to the enhanced vascular reactivity in early stages of hypertensive pregnancy.
Asunto(s)
Angiotensina II/farmacología , Endocannabinoides/antagonistas & inhibidores , Hipertensión Inducida en el Embarazo/fisiopatología , Preñez/fisiología , Arteria Uterina/fisiología , Vasoconstricción/efectos de los fármacos , Amidohidrolasas/antagonistas & inhibidores , Amidohidrolasas/metabolismo , Animales , Ácidos Araquidónicos/antagonistas & inhibidores , Ácidos Araquidónicos/metabolismo , Benzamidas/farmacología , Benzodioxoles/farmacología , Presión Sanguínea/fisiología , Carbamatos/farmacología , Modelos Animales de Enfermedad , Endocannabinoides/metabolismo , Femenino , Glicéridos/antagonistas & inhibidores , Glicéridos/metabolismo , Humanos , Hidrólisis , Masculino , Monoglicéridos/antagonistas & inhibidores , Monoglicéridos/metabolismo , Piperidinas/farmacología , Alcamidas Poliinsaturadas/antagonistas & inhibidores , Alcamidas Poliinsaturadas/metabolismo , Embarazo , Ratas , Ratas Sprague-Dawley , Ratas Transgénicas , Arteria Uterina/efectos de los fármacos , Vasoconstricción/fisiologíaRESUMEN
2-arachidonyl glycerol (2-AG) allosterically potentiates GABA(A) receptors via a binding site located in transmembrane segment M4 of the ß2 subunit. Two amino acid residues have been described that are essential for this effect. With the aim to further describe this potential drug target, we performed a cysteine scanning of the entire M4 and part of M3. All four residues in M4 affecting the potentiation here and the two already identified residues locate to the same side of the α-helix. This side is exposed to M3, where further residues were identified. From the fact that the important residues span > 18 Å, we conclude that the hydrophobic tail of the bound 2-AG molecule must be near linear and that the site mainly locates to the inner leaflet but stretches far into the membrane. The influence of the structure of the head group of the ligand molecule on the activity of the molecule was also investigated. We present a model of 2-AG docked to the GABA(A) receptor.
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Ácidos Araquidónicos/metabolismo , Endocannabinoides/metabolismo , Glicéridos/metabolismo , Receptores de GABA-A/metabolismo , Animales , Ácidos Araquidónicos/antagonistas & inhibidores , Sitios de Unión , Cisteína/metabolismo , Relación Dosis-Respuesta a Droga , Endocannabinoides/antagonistas & inhibidores , Etanolaminas/farmacología , Ácidos Grasos Insaturados/farmacología , Femenino , Glicéridos/antagonistas & inhibidores , Humanos , Cinética , Modelos Moleculares , Mutación/genética , Mutación/fisiología , Oocitos/metabolismo , Mutación Puntual/genética , Mutación Puntual/fisiología , Alcamidas Poliinsaturadas/farmacología , Receptor Cannabinoide CB1/agonistas , Xenopus laevisRESUMEN
The endocannabinoid 2-arachidonoylglycerol (2-AG) mediates activity-dependent depression of excitatory neurotransmission at central synapses, but the molecular regulation of 2-AG synthesis is not well understood. Here we identify a functional interaction between the 2-AG synthetic enzyme diacylglycerol lipase-α (DGLα) and calcium/calmodulin dependent protein kinase II (CaMKII). Activated CaMKII interacted with the C-terminal domain of DGLα, phosphorylated two serine residues and inhibited DGLα activity. Consistent with an inhibitory role for CaMKII in 2-AG synthesis, in vivo genetic inhibition of CaMKII increased striatal DGL activity and basal levels of 2-AG, and CaMKII inhibition augmented short-term retrograde endocannabinoid signaling at striatal glutamatergic synapses. Lastly, blockade of 2-AG breakdown using concentrations of JZL-184 that have no effect in wild-type mice produced a hypolocomotor response in mice with reduced CaMKII activity. These findings provide mechanistic insights into the molecular regulation of striatal endocannabinoid signaling with implications for physiological control of motor function.
Asunto(s)
Ácidos Araquidónicos/metabolismo , Proteína Quinasa Tipo 2 Dependiente de Calcio Calmodulina/fisiología , Cuerpo Estriado/metabolismo , Endocannabinoides/metabolismo , Glicéridos/metabolismo , Lipoproteína Lipasa/metabolismo , Transducción de Señal/fisiología , Animales , Ácidos Araquidónicos/antagonistas & inhibidores , Benzodioxoles/farmacología , Proteína Quinasa Tipo 2 Dependiente de Calcio Calmodulina/antagonistas & inhibidores , Cuerpo Estriado/efectos de los fármacos , Endocannabinoides/antagonistas & inhibidores , Técnicas de Silenciamiento del Gen/métodos , Glicéridos/antagonistas & inhibidores , Células HEK293 , Humanos , Lipoproteína Lipasa/antagonistas & inhibidores , Masculino , Ratones , Ratones Endogámicos C57BL , Ratones Transgénicos , Piperidinas/farmacología , Transducción de Señal/efectos de los fármacosRESUMEN
2-Arachidonoylglycerol (2-AG) is recognized as a potent endocannabinoid, which reduces synaptic transmission through cannabinoid CB(1) receptors, and is hydrolyzed by monoacylglycerol lipase (MGL) to arachidonic acid (AA), a cyclooxygenase substrate. We already reported that centrally administered MGL and cyclooxygenase inhibitors each reduced the intracerebroventricularly (i.c.v.) administered bombesin-induced secretion of adrenal catecholamines, while a centrally administered CB(1)-antagonist potentiated the response, indirectly suggesting bidirectional roles of brain 2-AG (stimulatory and inhibitory roles) in the bombesin-induced response. In the present study, we separately examined these bidirectional roles using 2-AG and 2-AG ether (2-AG-E) (stable 2-AG analog for MGL) in rats. 2-AG (0.5 µmol/animal, i.c.v.), but not 2-AG-E (0.5 µmol/animal, i.c.v.), elevated basal plasma catecholamines with JZL184 (MGL inhibitor)- and indomethacin (cyclooxygenase inhibitor)-sensitive brain mechanisms. 2-AG-E (0.1 µmol/animal, i.c.v.) effectively reduced the bombesin (1 nmol/animal, i.c.v.)-induced elevation of plasma catecholamines with rimonabant (CB(1) antagonist)-sensitive brain mechanisms. Immunohistochemical studies demonstrated the bombesin-induced activation of diacylglycerol lipase α (2-AG-producing enzyme)-positive spinally projecting neurons in the hypothalamic paraventricular nucleus, a control center of central adrenomedullary outflow. These results directly indicate bidirectional roles of brain 2-AG, a stimulatory role as an AA precursor and an inhibitory role as an endocannabinoid, in the bombesin-induced central adrenomedullary outflow in rats.
Asunto(s)
Médula Suprarrenal/efectos de los fármacos , Ácidos Araquidónicos/farmacología , Bombesina/farmacología , Agonistas de Receptores de Cannabinoides/farmacología , Catecolaminas/metabolismo , Endocannabinoides/farmacología , Glicéridos/farmacología , Neurotransmisores/agonistas , Médula Suprarrenal/metabolismo , Animales , Ácidos Araquidónicos/administración & dosificación , Benzodioxoles/administración & dosificación , Benzodioxoles/farmacología , Bombesina/administración & dosificación , Agonistas de Receptores de Cannabinoides/administración & dosificación , Antagonistas de Receptores de Cannabinoides/farmacología , Catecolaminas/sangre , Inhibidores de la Ciclooxigenasa/farmacología , Interacciones Farmacológicas , Endocannabinoides/administración & dosificación , Glicéridos/administración & dosificación , Glicéridos/antagonistas & inhibidores , Indometacina/administración & dosificación , Indometacina/farmacología , Inyecciones Intraventriculares , Lipoproteína Lipasa/metabolismo , Masculino , Monoacilglicerol Lipasas/antagonistas & inhibidores , Neurotransmisores/administración & dosificación , Neurotransmisores/antagonistas & inhibidores , Núcleo Hipotalámico Paraventricular/metabolismo , Piperidinas/administración & dosificación , Piperidinas/farmacología , Pirazoles/administración & dosificación , Pirazoles/farmacología , Ratas , RimonabantRESUMEN
BACKGROUND AND PURPOSE: The development of potent and selective inhibitors of the biosynthesis of the endocannabinoid 2-arachidonoylglycerol (2-AG) via DAG lipases (DAGL) α and ß is just starting to be considered as a novel and promising source of pharmaceuticals for the treatment of disorders that might benefit from a reduction in endocannabinoid tone, such as hyperphagia in obese subjects. EXPERIMENTAL APPROACH: Three new fluorophosphonate compounds O-7458, O-7459 and O-7460 were synthesized and characterized in various enzymatic assays. The effects of O-7460 on high-fat diet intake were tested in mice. KEY RESULTS: Of the new compounds, O-7460 exhibited the highest potency (IC50 = 690 nM) against the human recombinant DAGLα, and selectivity (IC50 > 10 µM) towards COS-7 cell and human monoacylglycerol lipase (MAGL), and rat brain fatty acid amide hydrolase. Competitive activity-based protein profiling confirmed that O-7460 inhibits mouse brain MAGL only at concentrations ≥ 10 µM, and showed that this compound has only one major 'off-target', that is, the serine hydrolase KIAA1363. O-7460 did not exhibit measurable affinity for human recombinant CB1 or CB2 cannabinoid receptors (Ki > 10 µM). In mouse neuroblastoma N18TG2 cells stimulated with ionomycin, O-7460 (10 µM) reduced 2-AG levels. When administered to mice, O-7460 dose-dependently (0-12 mg·kg⻹, i.p.) inhibited the intake of a high-fat diet over a 14 h observation period, and, subsequently, slightly but significantly reduced body weight. CONCLUSIONS AND IMPLICATIONS: O-7460 might be considered a useful pharmacological tool to investigate further the role played by 2-AG both in vitro and in vivo under physiological as well as pathological conditions.
Asunto(s)
Fármacos Antiobesidad/uso terapéutico , Ácidos Araquidónicos/antagonistas & inhibidores , Endocannabinoides/antagonistas & inhibidores , Inhibidores Enzimáticos/uso terapéutico , Glicéridos/antagonistas & inhibidores , Glicerofosfolípidos/uso terapéutico , Lipoproteína Lipasa/antagonistas & inhibidores , Obesidad/tratamiento farmacológico , Ácidos Oléicos/uso terapéutico , Organofosfonatos/uso terapéutico , Animales , Fármacos Antiobesidad/administración & dosificación , Fármacos Antiobesidad/farmacología , Ácidos Araquidónicos/metabolismo , Conducta Animal/efectos de los fármacos , Línea Celular , Chlorocebus aethiops , Relación Dosis-Respuesta a Droga , Endocannabinoides/metabolismo , Ingestión de Energía/efectos de los fármacos , Inhibidores Enzimáticos/farmacología , Glicéridos/metabolismo , Glicerofosfolípidos/administración & dosificación , Glicerofosfolípidos/farmacología , Humanos , Hipotálamo/efectos de los fármacos , Hipotálamo/enzimología , Hipotálamo/metabolismo , Lipoproteína Lipasa/genética , Lipoproteína Lipasa/metabolismo , Hígado/efectos de los fármacos , Hígado/enzimología , Hígado/metabolismo , Masculino , Ratones , Ratones Endogámicos C57BL , Proteínas del Tejido Nervioso/antagonistas & inhibidores , Proteínas del Tejido Nervioso/genética , Proteínas del Tejido Nervioso/metabolismo , Neuronas/efectos de los fármacos , Neuronas/enzimología , Neuronas/metabolismo , Obesidad/enzimología , Obesidad/metabolismo , Ácidos Oléicos/administración & dosificación , Ácidos Oléicos/farmacología , Organofosfonatos/administración & dosificación , Organofosfonatos/farmacología , Ratas , Proteínas Recombinantes/química , Proteínas Recombinantes/metabolismo , Esterol Esterasa/antagonistas & inhibidores , Esterol Esterasa/metabolismoRESUMEN
The GPR119 receptor plays an important role in the secretion of incretin hormones in response to nutrient consumption. We have studied the ability of an array of naturally occurring endocannabinoid-like lipids to activate GPR119 and have identified several lipid receptor agonists. The most potent receptor agonists identified were three N-acylethanolamines: oleoylethanolamine (OEA), palmitoleoylethanolamine, and linoleylethanolamine (LEA), all of which displayed similar potency in activating GPR119. Another lipid, 2-oleoylglycerol (2-OG), also activated GPR119 receptor but with significantly lower potency. Endogenous levels of endocannabinoid-like lipids were measured in intestine in fasted and refed mice. Of the lipid GPR119 agonists studied, the intestinal levels of only OEA, LEA, and 2-OG increased significantly upon refeeding. Intestinal levels of OEA and LEA in the fasted mice were low. In the fed state, OEA levels only moderately increased, whereas LEA levels rose drastically. 2-OG was the most abundant of the three GPR119 agonists in intestine, and its levels were radically elevated in fed mice. Our data suggest that, in lean mice, 2-OG and LEA may serve as physiologically relevant endogenous GPR119 agonists that mediate receptor activation upon nutrient uptake.