title: Synthetic and endogenous cannabinoids protect retinal neurons from AMPA excitotoxicity in vivo, via activation of CB1 receptors: Involvement of PI3K/Akt and MEK/ERK signaling pathways.
abstract: Cannabinoids have been suggested to protect retinal ganglion cells in different models of toxicity, but their effects on other retinal neurons are poorly known. We investigated the neuroprotective actions of the endocannabinoid N-arachidonoyl ethanolamine (Anandamide/AEA) and the synthetic cannabinoids R1-Methanandamide (MethAEA) and HU-210, in an in vivo retinal model of AMPA excitotoxicity, and the mechanisms involved in the neuroprotection. Sprague-Dawley rats were intravitreally injected with PBS or AMPA in the absence or presence of the cannabinoid agonists. Brain nitric oxide synthase (bNOS) and choline acetyltransferase (ChAT) immunoreactivity (IR), as well as TUNEL staining, assessed the AMPA-induced retinal amacrine cell loss and the dose-dependent neuroprotection afforded by cannabinoids. The CB1 receptor selective antagonist AM251 and the PI3K/Akt inhibitor wortmannin reversed the cannabinoid-induced neuroprotection, suggesting the involvement of CB1 receptors and the PI3K/Akt pathway in cannabinoids' actions. Experiments with the CB2 agonist JWH015 and [(3)H]CP55940 radioligand binding suggested that the CB2 receptor is not involved in the neuroprotection. AEA and HU-210 induced phosphorylation of Akt but only AEA induced phosphorylation of ERK1/2 kinases, as revealed by western blot analysis. To investigate the role of caspase-3 in the AMPA-induced cell death, the caspase-3 inhibitor Z-DEVD-FMK was co-injected with AMPA. Z-DEVD-FMK had no effect on AMPA excitotoxicity. Moreover, no difference was observed in the phosphorylation of SAPK/JNK kinases between PBS- and AMPA-treated retinas. These results suggest that endogenous and synthetic cannabinoids protect retinal amacrine neurons from AMPA excitotoxicity in vivo via a mechanism involving the CB1 receptors, and the PI3K/Akt and/or MEK/ERK1/2 signaling pathways.
authors.affiliation: Department of Pharmacology, School of Medicine, University of Crete, Heraklion, Crete 71003 GR, Greece. Electronic address: despina.kokona@insel.ch.
authors.forename: Despina
authors.initials: D
authors.lastname: Kokona
authors.affiliation: Department of Pharmacology, School of Medicine, University of Crete, Heraklion, Crete 71003 GR, Greece. Electronic address: thermos@med.uoc.gr.
authors.forename: Kyriaki
authors.initials: K
authors.lastname: Thermos
mesh_terms: D025042:Amacrine Cells / Q000187:drug effects* / Q000378:metabolism / Q000473:pathology; D000818:Animals; D017209:Apoptosis; D001095:Arachidonic Acids / Q000494:pharmacology; D063386:Cannabinoid Receptor Agonists / Q000494:pharmacology; D002795:Choline O-Acetyltransferase / Q000378:metabolism; D004305:Dose-Response Relationship, Drug; D013759:Dronabinol / Q000031:analogs & derivatives / Q000494:pharmacology; D063388:Endocannabinoids / Q000494:pharmacology; D004791:Enzyme Inhibitors / Q000494:pharmacology; D018690:Excitatory Amino Acid Agonists / Q000633:toxicity; D019084:Fluorescent Antibody Technique, Indirect; D020287:In Situ Nick-End Labeling; D058449:Intravitreal Injections; D020935:MAP Kinase Signaling System / Q000502:physiology*; D008297:Male; D018696:Neuroprotective Agents / Q000494:pharmacology*; D052248:Nitric Oxide Synthase Type I / Q000378:metabolism; D019869:Phosphatidylinositol 3-Kinases / Q000378:metabolism*; D000081082:Phosphoinositide-3 Kinase Inhibitors; D053284:Polyunsaturated Alkamides / Q000494:pharmacology; D051057:Proto-Oncogene Proteins c-akt / Q000037:antagonists & inhibitors / Q000378:metabolism*; D051381:Rats; D017207:Rats, Sprague-Dawley; D043884:Receptor, Cannabinoid, CB1 / Q000037:antagonists & inhibitors / Q000378:metabolism*; D018350:alpha-Amino-3-hydroxy-5-methyl-4-isoxazolepropionic Acid / Q000633:toxicity*
chemical_list: D001095:Arachidonic Acids; D063386:Cannabinoid Receptor Agonists; D002795:Choline O-Acetyltransferase; D013759:Dronabinol; D063388:Endocannabinoids; D004791:Enzyme Inhibitors; D018690:Excitatory Amino Acid Agonists; D018696:Neuroprotective Agents; D052248:Nitric Oxide Synthase Type I; D019869:Phosphatidylinositol 3-Kinases; D000081082:Phosphoinositide-3 Kinase Inhibitors; D053284:Polyunsaturated Alkamides; D051057:Proto-Oncogene Proteins c-akt; D043884:Receptor, Cannabinoid, CB1; D018350:alpha-Amino-3-hydroxy-5-methyl-4-isoxazolepropionic Acid; C088155:methanandamide; C496322:Nos1 protein, rat; C062018:HU 211; C078814:anandamide
keywords: Amacrine cells; CB1 receptors; Cannabinoids; Excitotoxicity; MEK/ERK1/2 signaling pathway; Neuroprotection; PI3K/Akt signaling pathway; Retina