title: High levels of anandamide, an endogenous cannabinoid, block the growth of sheep preimplantation embryos by inducing apoptosis and reversible arrest of cell proliferation.
abstract: BACKGROUND
  The process of implantation is mediated by various molecules, one of which is anandamide (AEA), a lipid signalling ligand belonging to the family of endocannabinoids. AEA exerts its effects on implantation by binding to the Type 1 Cannabinoid Receptor (CB1-R), expressed in both blastocysts and uterus. We wanted to know whether the endocannabinoid signalling system was present also in the sheep reproductive tract and which kind of effect(s) AEA had on the development of sheep blastocysts in vitro.
  
  
  METHODS
  We analysed the expression and activity of the endocannabinoid system in sheep reproductive tracts and blastocysts. Hatched sheep blastocysts were then exposed to AEA and its effect(s) were determined by TUNEL assay and by measuring the rate of necrosis and 5-bromo-deoxyuridine incorporation.
  
  
  RESULTS
  We show that the AEA signalling system is present in sheep and that high concentrations of AEA induce apoptosis and inhibit cell proliferation via a CB1-R-dependent mechanism. Indeed, AEA effects were blocked when sheep blastocysts were cultured in the presence of the CB1-R antagonist SR161417A. Moreover, AEA inhibition of cell proliferation was reversible, as arrested embryos resumed a normal growth rate upon AEA removal from the medium.
  
  
  CONCLUSIONS
  Our results suggest that disturbed regulation of AEA signalling via CB1-R may be associated with pregnancy failure. AEA could lower the quality of blastocysts by inducing apoptosis and inhibiting cell proliferation, thus making them incompetent for implantation.
authors.affiliation: Department of Comparative Biomedical Sciences, University of Teramo 64100, Teramo, Italy.
authors.forename: M Y
authors.initials: MY
authors.lastname: Turco
authors.forename: K
authors.initials: K
authors.lastname: Matsukawa
authors.forename: M
authors.initials: M
authors.lastname: Czernik
authors.forename: V
authors.initials: V
authors.lastname: Gasperi
authors.forename: N
authors.initials: N
authors.lastname: Battista
authors.forename: L
authors.initials: L
authors.lastname: Della Salda
authors.forename: P A
authors.initials: PA
authors.lastname: Scapolo
authors.forename: P
authors.initials: P
authors.lastname: Loi
authors.forename: M
authors.initials: M
authors.lastname: Maccarrone
authors.forename: G
authors.initials: G
authors.lastname: Ptak
mesh_terms: D000818:Animals; D017209:Apoptosis / Q000187:drug effects*; D001095:Arachidonic Acids / Q000378:metabolism / Q000494:pharmacology*; D001755:Blastocyst / Q000187:drug effects*; D063385:Cannabinoid Receptor Modulators / Q000378:metabolism; D049109:Cell Proliferation / Q000187:drug effects*; D047108:Embryonic Development / Q000187:drug effects*; D063388:Endocannabinoids; D005260:Female; D020287:In Situ Nick-End Labeling; D009336:Necrosis; D053284:Polyunsaturated Alkamides / Q000378:metabolism / Q000494:pharmacology*; D043884:Receptor, Cannabinoid, CB1 / Q000378:metabolism; D012756:Sheep; D015398:Signal Transduction; D014599:Uterus / Q000378:metabolism
chemical_list: D001095:Arachidonic Acids; D063385:Cannabinoid Receptor Modulators; D063388:Endocannabinoids; D053284:Polyunsaturated Alkamides; D043884:Receptor, Cannabinoid, CB1; C078814:anandamide