title: Prolonged exposure to WIN55,212-2 causes downregulation of the CB1 receptor and the development of tolerance to its anticonvulsant effects in the hippocampal neuronal culture model of acquired epilepsy.
abstract: Cannabinoids have been shown to cause CB1-receptor-dependent anticonvulsant activity in both in vivo and in vitro models of status epilepticus (SE) and acquired epilepsy (AE). It has been further demonstrated in these models that the endocannabinoid system functions in a tonic manner to suppress seizure discharges through a CB1-receptor-dependent pathway. Although acute cannabinoid treatment has anticonvulsant activity, little is known concerning the effects of prolonged exposure to CB1 agonists and development of tolerance on the epileptic phenotype. This study was carried out to evaluate the effects of prolonged exposure to the CB1 agonist WIN55,212-2 on seizure activity in a hippocampal neuronal culture model of low-Mg(2+) induced spontaneous recurrent epileptiform discharges (SREDs). Following low-Mg(2+) induced SREDs, cultures were returned to maintenance media containing 10, 100 or 1000 nM WIN55,212-2 from 4 to 24 h. Whole-cell current-clamp analysis of WIN55,212-2 treated cultures revealed a concentration-dependent increase in SRED frequency. Immunocytochemical staining revealed that WIN55,212-2 treatment induced a concentration-dependent downregulation of the CB1 receptor in neuronal processes and at both glutamatergic and GABAergic presynaptic terminals. Prolonged exposure to the inactive enantiomer WIN55,212-3 in low-Mg(2+) treated cultures had no effect on the frequency of SREDs or CB1 receptor staining. The results from this study further substantiate a role for a tonic CB1-receptor-dependent endocannabinoid regulation of seizure discharge and suggest that prolonged exposure to cannabinoids results in the development of tolerance to the anticonvulsant effects of cannabinoids and an exacerbation of seizure activity in the epileptic phenotype.
authors.affiliation: Department of Neurology, Virginia Commonwealth University School of Medicine, Richmond, VA 23298, USA. rblair@vcu.edu
authors.forename: Robert E
authors.initials: RE
authors.lastname: Blair
authors.forename: Laxmikant S
authors.initials: LS
authors.lastname: Deshpande
authors.forename: Sompong
authors.initials: S
authors.lastname: Sombati
authors.forename: Maurice R
authors.initials: MR
authors.lastname: Elphick
authors.forename: Billy R
authors.initials: BR
authors.lastname: Martin
authors.forename: Robert J
authors.initials: RJ
authors.lastname: DeLorenzo
mesh_terms: D000818:Animals; D000927:Anticonvulsants / Q000008:administration & dosage / Q000494:pharmacology*; D048588:Benzoxazines / Q000008:administration & dosage / Q000494:pharmacology*; D002478:Cells, Cultured; D004305:Dose-Response Relationship, Drug; D004827:Epilepsy / Q000188:drug therapy* / Q000503:physiopathology; D018698:Glutamic Acid / Q000378:metabolism; D006624:Hippocampus / Q000187:drug effects* / Q000503:physiopathology; D007150:Immunohistochemistry; D008274:Magnesium / Q000378:metabolism; D009025:Morpholines / Q000008:administration & dosage / Q000494:pharmacology*; D009281:Naphthalenes / Q000008:administration & dosage / Q000494:pharmacology*; D009474:Neurons / Q000187:drug effects* / Q000502:physiology; D018408:Patch-Clamp Techniques; D017729:Presynaptic Terminals / Q000187:drug effects / Q000502:physiology; D017966:Pyramidal Cells / Q000187:drug effects / Q000502:physiology; D051381:Rats; D043884:Receptor, Cannabinoid, CB1 / Q000819:agonists / Q000378:metabolism*; D013997:Time Factors; D005680:gamma-Aminobutyric Acid / Q000378:metabolism
chemical_list: D000927:Anticonvulsants; D048588:Benzoxazines; D009025:Morpholines; D009281:Naphthalenes; D043884:Receptor, Cannabinoid, CB1; D018698:Glutamic Acid; D005680:gamma-Aminobutyric Acid; C070417:(3R)-((2,3-dihydro-5-methyl-3-((4-morpholinyl)methyl)pyrrolo-(1,2,3-de)-1,4-benzoxazin-6-yl)(1-naphthalenyl))methanone; D008274:Magnesium