Cannabinoid receptors type 1 and 2 are involved in the embryonic resorption induce by LPS

SALAZAR ANA INES; DOMINGUEZ RUBIO ANA PAULA; DAVIO CARLOS; FRANCHI, ANA MARIA

Boston

Congreso; 12th Congress of the International Society for Immunology Reproduction (ISIR); 2013

Endocannabinoids are a family of endogenous messengers that modulate diverse physiological responses, including different events in reproduction. In our laboratory, we have demonstrated that the pregnant mice uterus expresses the cannabinoid receptors (CB) and the enzymes that synthesized and degrade anandamide, one of the most abundant cannabinoid in this reproductive organ. In addition, we have seen that the endocannabinoids mediate the deleterious effect of LPS on the uterus, causing an increase in the levels of nitric oxide and prostaglandins. In this study we describe the apoptotic effect of LPS (1ug/ml) and the cannabinoid meta-anandamida (m-AEA), a non hydrolyzable analogue of anandamida (AEA), in murine uterine fragments in the 7th day of pregnancy from Balb/C mice. We analyzed their nucleus structure making a Hoechst stain. We also measure caspase activity by a luminescence kit. Both LPS as m-AEA, increased the amount of apoptotic bodies and the levels of caspase activity. The apoptotic effect was reverted with the co-incubation of cannabinoid receptors type 1 and 2 antagonists. Throughout gestation it is vital that the uterus remains relatively quiescent permitting the fetus to grow and mature in preparation for extrauterine life. Cyclic AMP promotes the relaxation of myometrial, and other, smooth muscle cells via activation of cyclic AMP-dependent protein kinase (PKA). Based on this, we decided to measure the cyclic AMP levels in our model. Cyclic AMP content was determined by competition of [3H]cAMP for protein kinase A. We measured the cyclic AMP levels in presence of LPS and different concentrations of m- AEA. Both molecules produced a decreased in cyclic AMP synthesis, indicating that receptors are coupled to Gi protein. This effect was reverted with the co-incubation of cannabinoid receptor type 2 antagonist. These results show that both LPS and AEA produce tissue damage and apoptosis, and a diminished of cyclic AMP levels. The endocannabinoid receptors could participate in the LPS mechanism of action. We postulate that apoptosis and the regulation of cyclic AMP levels are key events in the embryonic resorption induce by LPS, and that this effect could be mediated by endocannabinoids.