title: Binding affinity and agonist activity of putative endogenous cannabinoids at the human neocortical CB1 receptor.
abstract: We investigated the affinity of putative endocannabinoids (2-arachidonylglycerol, 2-AG; noladin ether, virodhamine) for the human neocortical CB1 receptor. Functional activity of these compounds (including anandamide, AEA) was determined by examining basal and forskolin-stimulated cAMP formation. Assays were performed with synaptosomes, prepared from fresh human neocortical tissue. Receptor affinity was assessed from competition binding experiments with the CB1/2 agonist [3H]-CP55.940 in absence or presence of a protease inhibitor to assess enzymatic stability. Noladin ether and virodhamine inhibited [3H]-CP55.940 binding (Ki: 98, 1740 nM, respectively). Protease inhibition decreased the Ki value of virodhamine (Ki: 912 nM), but left that of noladin ether unchanged. 2-AG almost lacked affinity (Ki lymphoblasic )10 microM). Basal cAMP formation was unaffected by AEA and noladin ether, but strongly enhanced by 2-AG and virodhamine. Forskolin-stimulated cAMP formation was inhibited by AEA and noladin ether (IC50: 69, 427 nM, respectively) to the same extent as by CP55.940 (Imax each approximately 30%). Inhibitions by AEA or noladin ether were blocked by the CB1 receptor antagonist AM251. Virodhamine increased forskolin-stimulated cAMP formation, also in presence of AM251, by approximately 20%. 2-AG had no effect; in presence of AM251, however, 10 microM 2-AG stimulated cAMP formation by approximately 15%. Our results suggest, that AEA and noladin ether are full CB1 receptor agonists in human neocortex, whereas virodhamine may act as a CB1 receptor antagonist/inverse agonist. Particularly the (patho)physiological role of 2-AG should be further investigated, since its CB1 receptor affinity and agonist activity especially in humans might be lower than generally assumed.
authors.affiliation: Sektion Klinische Neuropharmakologie, Neurozentrum, Breisacherstrasse 64, D-79106 Freiburg, Germany.
authors.forename: Marc
authors.initials: M
authors.lastname: Steffens
authors.forename: Josef
authors.initials: J
authors.lastname: Zentner
authors.forename: Jürgen
authors.initials: J
authors.lastname: Honegger
authors.forename: Thomas J
authors.initials: TJ
authors.lastname: Feuerstein
mesh_terms: D000293:Adolescent; D000328:Adult; D000818:Animals; D001665:Binding Sites / Q000187:drug effects; D063385:Cannabinoid Receptor Modulators / Q000378:metabolism* / Q000494:pharmacology; D002648:Child; D002675:Child, Preschool; D000242:Cyclic AMP / Q000378:metabolism; D004305:Dose-Response Relationship, Drug; D005260:Female; D006801:Humans; D008297:Male; D008875:Middle Aged; D019579:Neocortex / Q000187:drug effects / Q000378:metabolism*; D011485:Protein Binding / Q000187:drug effects / Q000502:physiology; D051381:Rats; D017208:Rats, Wistar; D043884:Receptor, Cannabinoid, CB1 / Q000819:agonists* / Q000378:metabolism*
chemical_list: D063385:Cannabinoid Receptor Modulators; D043884:Receptor, Cannabinoid, CB1; D000242:Cyclic AMP