Signaling Pathways from Cannabinoid Receptor-1 Activation to Inhibition of N-Methyl-D-Aspartic Acid Mediated Calcium Influx and Neurotoxicity in Dorsal Root Ganglion Neurons (original) (raw)
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Journal of Neurochemistry, 2002
The ability of cannabinoid CB 1 receptors to influence glutamatergic excitatory neurotransmission has fueled interest in how these receptors and their endogenous ligands may interact in conditions of excitotoxic insults. The present study characterized the impact of stimulated and inhibited CB 1 receptor function on NMDA-induced excitotoxicity. Neonatal (6-day-old) rat pups received a systemic injection of a mixed CB 1 /CB 2 receptor agonist (WIN55,212-2) or their respective antagonists (SR141716A for CB 1 and SR144528 for CB 2 ) prior to an unilateral intrastriatal microinjection of NMDA. The NMDA-induced excitotoxic damage in the ipsilateral forebrain was not influenced by agonist-stimulated CB 1 receptor function. In contrast, blockade of CB 1 , but not CB 2 , receptor activity evoked a robust neuroprotective response by reducing the infarct area and the number of cortical degenerating neurons. These results suggest a critical involvement of CB 1 receptor tonus on neuronal survival following NMDA receptorinduced excitotoxicity in vivo. Abbreviations used: CB 1 receptor, cannabinoid receptor subtype 1; CB 2 receptor, cannabinoid receptor subtype 2; GFAP, glial fibrilary acidic protein; KPBS, potassium phosphate-buffered saline; PBS, phosphate-buffered saline; D 9 -THC, D 9 -tetrahydrocannabinol; TdT, terminal deoxynucleotidyltransferase; TUNEL, terminal deoxynucleotidyltransferase-mediated dUTP nick-end labeling.
Molecular Mechanisms of Cannabinoid Protection from Neuronal Excitotoxicity
Molecular Pharmacology, 2005
Nonstandard abbreviations: CB1 cannabinoid receptor (CB1R); protein kinase A (PKA); dibutyryl-cyclic adenosine monophosphate (dbcAMP); R(+)-[2, 3-dihydro-5methyl-3-[(morpholinyl)methyl]pyrrolo[1,2,3-de]-1,4-benzoxazin-yl]-(1naphthalenyl)methanone mesylate (R(+)-WIN 55212); N-(piperidin-1-yl)-5-(4chlorophenyl)-1-(2,4-cichlorophenyl)-4-methyl-1H-pyrazole-3-carboxamide hydrochloride (SR141716A); 7-nitroindazole (7-NI); N-ω-nitro-L-arginine methyl ester (L-NAME); N[2-((p-bromocinnamyl)amino)ethyl]-5-isoquinolinesulfonamide (H-89); pertussis toxin (PTX); nitric oxide synthase (NOS); minimal essential medium (MEM); lactate dehydrogenase (LDH); Hepes-buffered control salt solution (HCSS); fluorescein isothiocyanate (FITC)
Cannabinoids enhance NMDA-elicited Ca2+ signals in cerebellar granule neurons in culture
The Journal of neuroscience : the official journal of the Society for Neuroscience, 1999
A physiological role for cannabinoids in the CNS is indicated by the presence of endogenous cannabinoids and cannabinoid receptors. However, the cellular mechanisms of cannabinoid actions in the CNS have yet to be fully defined. In the current study, we identified a novel action of cannabinoids to enhance intracellular Ca2+ responses in CNS neurons. Acute application of the cannabinoid receptor agonists R(+)-methanandamide, R(+)-WIN, and HU-210 (1-50 nM) dose-dependently enhanced the peak amplitude of the Ca2+ response elicited by stimulation of the NMDA subtype of glutamate receptors (NMDARs) in cerebellar granule neurons. The cannabinoid effect was blocked by the cannabinoid receptor antagonist SR141716A and the Gi/Go protein inhibitor pertussis toxin but was not mimicked by the inactive cannabinoid analog S(-)-WIN, indicating the involvement of cannabinoid receptors. In current-clamp studies neither R(+)-WIN nor R(+)-methanandamide altered the membrane response to NMDA or passive...
British Journal of Pharmacology, 2004
1 The prototypical aminoalkylindole cannabinoid WIN 55,212-2 (WIN-2) has been shown to produce antihyperalgesia through a peripheral mechanism of action. However, it is not known whether WIN-2 exerts this action directly via cannabinoid receptors located on primary afferents or if other, perhaps indirect or noncannabinoid, mechanisms are involved. To address this question, we have examined the specific actions of WIN-2 on trigeminal ganglion (TG) neurons in vitro by quantifying its ability to modulate the evoked secretion of the proinflammatory neuropeptide CGRP as well as the inflammatory mediator-induced generation of cAMP. 2 WIN-2 evoked CGRP release from TG neurons in vitro (EC 50 ¼ 26 mM) in a concentration-and calcium-dependent manner, which was mimicked by the cannabinoid receptor-inactive enantiomer WIN 55,212-3 (WIN-3). Moreover, WIN-2-evoked CGRP release was attenuated by the nonselective cation channel blocker ruthenium red but not by the vanilloid receptor type 1 (TRPV1) antagonist capsazepine, suggesting that, unlike certain endogenous and synthetic cannabinoids, WIN-2 is not a TRPV1 agonist but rather acts at an as yet unidentified cation channel.
…, 2001
Cannabinoids modulate nociceptive processing in models of acute, inflammatory and neuropathic pain. We have investigated the location and function of cannabinoid receptors on cultured neonatal dorsal root ganglion (DRG) neurones and F-11 cells, a dorsal root ganglion×neuroblastoma hybridoma which displays several of the features of authentic DRG neurones. CB 1 receptor immunolabelling was observed on the cell bodies and as fine puncta on processes of both cultured DRG neurones and F-11 cells. Additionally, fluorescence-activated cell sorting (FACS) analysis provided evidence that both CB 1 and CB 2 receptors are expressed on populations of cells within the cultured DRG and F-11 cells. The cannabinoid receptor agonist (+)-WIN55212 (10 and 100 nM) inhibited the mean voltage-activated Ca 2+ current in DRG neurones by 21% and 30%, respectively. The isomer, (Ϫ)-WIN55212 (10 and 100 nM) produced significantly less inhibition of 6% and 10% respectively. The CB 1 selective receptor antagonist SR141716A (100 nM) enhanced the peak high voltage-activated Ca 2+ current by 24% and simultaneous application of SR141716A (100 nM) and (+)-WIN55212 (100 nM) resulted in a significant attenuation of the inhibition obtained with (+)-WIN55212 alone. These data give functional evidence for the hypothesis that the analgesic actions of cannabinoids may be mediated by presynaptic inhibition of transmitter release in sensory neurones.
Journal of neurochemistry, 2002
The ability of cannabinoid CB(1) receptors to influence glutamatergic excitatory neurotransmission has fueled interest in how these receptors and their endogenous ligands may interact in conditions of excitotoxic insults. The present study characterized the impact of stimulated and inhibited CB(1) receptor function on NMDA-induced excitotoxicity. Neonatal (6-day-old) rat pups received a systemic injection of a mixed CB(1) /CB(2) receptor agonist (WIN55,212-2) or their respective antagonists (SR141716A for CB(1) and SR144528 for CB(2) ) prior to an unilateral intrastriatal microinjection of NMDA. The NMDA-induced excitotoxic damage in the ipsilateral forebrain was not influenced by agonist-stimulated CB(1) receptor function. In contrast, blockade of CB(1), but not CB(2), receptor activity evoked a robust neuroprotective response by reducing the infarct area and the number of cortical degenerating neurons. These results suggest a critical involvement of CB(1) receptor tonus on neurona...
Role of CB< sub> 2 receptors in neuroprotective effects of cannabinoids
Molecular and cellular …, 2008
CB 2 receptors, the so-called peripheral cannabinoid receptor type, were first described in the immune system, but they have been recently identified in the brain in healthy conditions and, in particular, after several types of cytotoxic stimuli. Specifically, CB 2 receptors were identified in microglial cells, astrocytes and, to a lesser extent, in certain subpopulations of neurons. Given the lack of psychoactivity demonstrated by selective CB 2 receptor agonists, this receptor becomes an interesting target for the treatment of neurological diseases, in particular, the case of certain neurodegenerative disorders in which induction/up-regulation of CB 2 receptors has been already demonstrated. These disorders include Alzheimer's disease, Huntington's chorea, amyotrophic lateral sclerosis and others. Interestingly, in experimental models of these disorders, the activation of CB 2 receptors has been related to a delayed progression of neurodegenerative events, in particular, those related to the toxic influence of microglial cells on neuronal homeostasis. The present article will review the evidence supporting that CB 2 receptors might represent a key element in the endogenous response against different types of cytotoxic events, and that this receptor type may be a clinicallypromising target for the control of brain damage in neurodegenerative disorders.
Brain Research, 2006
Its presynaptic location suggests a role for cannabinoids in modulating the release of neurotransmitters from axon terminals by retrograde signaling. The neuroprotective effects of cannabinoid agonists in animal models of ischemia, seizures, hypoxia, Multiple Sclerosis, Huntington and Parkinson disease have been demonstrated in several reports. The proposed mechanism for the neuroprotection ranges from antioxidant effects, reduction of microglial activation and anti-inflammatory reaction to receptor-mediated reduction of glutamate release. In the present work, we analyzed the morphological changes induced by a chronic treatment with the synthetic cannabinoid receptor agonist, WIN 55,212-2, in four brain regions where the CB1 cannabinoid receptor is present in high density: the CA1 hippocampal area, corpus striatum, cerebellum and frontal cortex. After a twice-daily treatment for 14 days with the cannabinoid receptor agonist (3 mg/kg sc, each dose) to male Wistar rats (150-170 g), the expression of neurofilaments (Nf-160 and Nf-200), microtubuleassociated protein-2 (MAP-2), synaptophysin (Syn) and glial fibrillary acidic protein (GFAP) was studied by immunohistochemistry and digital image analysis. Ultrastructural study of the synapses was done using electron microscopy. After the treatment, a significant increase in the expression of neuronal cytoskeletal proteins (Nf-160, Nf-200, MAP-2) was observed, but we did not find changes in the expression of GFAP, the main astroglial cytoskeletal protein. In cerebellum, there was an increase in Syn expression and in the number of synaptic vesicles, while, in the hippocampus, an increase in the Syn expression and in the thickness of the postsynaptic densities was observed. The results obtained from these studies provide evidences on the absence of astroglial reaction and a sprouting phenomena induced by the WIN treatment that might be a key contributor to the long-term neuroprotective effects observed after cannabinoid treatments in different models of central nervous system (CNS) injury reported in the literature.
Δ9-Tetrahydrocannabinol protects hippocampal neurons from excitotoxicity
2007
Excitotoxic neuronal death underlies many neurodegenerative disorders. Because cannabinoid receptor agonists act presynaptically to inhibit glutamate release, we examined the effects of Win 55212-2, a full agonist at CB 1 receptors, and Δ 9-tetrahydrocannabinol (THC), a partial agonist, on the survival of neurons exposed to an excitotoxic pattern of synaptic activity. Reducing the extracellular Mg 2+ concentration ([Mg 2+ ] o) to 0.1 mM evoked an aberrant pattern of glutamatergic activity that produced synaptically mediated death of rat hippocampal neurons in culture. Neuronal viability was quantified with a multiwell fluorescence plate scanner equipped to detect propidium iodide fluorescence. Win 55212-2 (100 nM) and THC (100 nM) significantly reduced 0.1 mM [Mg 2+ ] o-induced cell death by 77 ± 11% and 84 ± 8%, respectively. Interestingly, the protection afforded by THC was not significantly different from that produced by Win 55212-2, suggesting that attenuation without a complete block of excitatory activity is sufficient for neuroprotection. The effect of prolonged drug exposure on the neuroprotection afforded by cannabinoid receptor agonists was also studied. When cultures were pretreated for 24 h with Win 55212-2 (100 nM) or THC (100 nM), inhibition of 0.1 mM [Mg 2+ ] o-induced toxicity was significantly reduced to 39 ± 19% and 45 ± 13%, respectively. Thus, desensitization of CB 1 receptors diminishes the neuroprotective effects of cannabinoids. This study demonstrates the importance of agonist efficacy and the duration of treatment on the neuroprotective effects of cannabinoids. It will be important to consider these effects on neuronal survival when evaluating pharmacologic treatments that modulate the endocannabinoid system.
Cannabinoids and Dopamine Receptors' Action on Calcium Current in Rat Neurons
The Canadian Journal of Neurological Sciences, 2005
The brain CB1 cannabinoid receptor is a member of the Gprotein-coupled receptor superfamily. 1 Among the wide variety of effects induced by activation of CB1 cannabinoid receptors are: inhibition of glutamatergic and GABA A synaptic transmission, 2-4 inhibition of dopamine release in rat brain, 5 and inhibition of serotonin release in mouse brain. 6 Thus, the CB1 cannabinoid receptor modulates neuronal excitability and neurotransmitter release, and thereby regulates Ca 2+-and K +currents. 7 Activation of CB1 receptors has been shown to inhibit N-and P-/Q-type Ca 2+ channels in cultured hippocampal neurons and in heterologous expression systems. 8-9 On the other hand, CB1 receptors activate inwardly rectifying K+ channels, 10 and it was recently shown that cannabinoids decrease the K + M-current in hippocampal CA1 neurons. 11 Additionally, there is evidence for associations and signaling interactions between cannabinoids ABSTRACT: Objective: To study the effects of cannabinoid, glutamate, and dopamine agonists and antagonists on the calcium current in rat sympathetic neurons. Methods: Calcium current was recorded using the whole-cell variant of the patch-clamp technique. After expression in neuronal membranes of the cannabinoid CB1, glutamate mGluR2, or dopamine D1 receptor (by microinjection of the relevant receptor's cDNA into the neuron's nucleus) agonists' and antagonists' effects were observed. Results: Applications of agonists of the expressed receptor (0.1-10 µM) decreased the calcium current. The calcium current was increased after application of cannabinoid antagonists (AM251 and AM630); these compounds thus act as inverse agonists in this preparation. Glutamate and dopamine antagonists had no effects on the calcium current by themselves. Combined application of cannabinoids and dopamine, but not glutamate, agonists produced a decrement in the calcium current that was bigger than either of the effects seen when one agonist was applied alone. Conclusions: These results suggest that cannabinoid with dopamine receptors have an interactive inhibitory effect on the calcium current in this preparation, indicating that within the nervous system, receptor interactions may be important in the regulation of ion-channel functions. RÉSUMÉ: Effet des cannabinoïdes et des récepteurs dopaminergiques sur les courants calciques dans des neurones de rat. Objectif: Étudier les effets d'agonistes et d'antagonistes de cannabinoïdes, du glutamate et de la dopamine sur le courant calcique dans des neurones sympathiques de rat. Méthodes: Le courant calcique a été enregistré au moyen de la technique patch-clamp sur cellules intactes. Des effets agonistes et antagonistes ont été observés après expression dans les membranes neuronales du récepteur cannabinoïde de type CB1, du récepteur métabotropique du glutamate mGluR2 ou du récepteurs D1 de la dopamine par micro injection de l'ADNc correspondant au récepteur dans le noyau du neurone. Résultats: L'application d'agonistes du récepteur exprimé (0,1 à 10 mmol) diminuait le courant calcique. Le courant calcique était augmenté suite à l'application d'antagonistes cannabinoïdes tels l'AM251 et l'AM630. Ces substances agissent donc comme des agonistes inverses dans cette préparation. Les antagonistes du glutamate et de la dopamine n'avaient pas d'effet sur le courant calcique par eux-mêmes. Une application combinée d'agonistes de cannabinoïdes et de dopamine diminuait davantage le courant calcique que l'application de chacun d'eux seul, ce qui n'était pas le cas des agonistes du glutamate. Conclusions: Ces résultats suggèrent un effet inhibiteur interactif des récepteurs cannabinoïdes et des récepteurs dopaminergiques sur le courant calcique dans cette préparation. Ceci indique que, dans le système nerveux, l'interaction de récepteurs pourrait être importante dans la régulation des fonctions des canaux ioniques.