Nitric oxide synthase activity and progesterone release by isolated corpora lutea of rabbits in the early and mid-luteal phases of pseudopregnancy are modulated differently by prostaglandin E-2 and prostaglandin F-2alpha via adenylate cyclase and phospholipase C

Abstract

By examining in vitro the effects of prostaglandin E-2 (PGE-2) and prostaglandin F-2 (PGF-2 ) induced in the corpora lutea (CL) of pseudopregnant rabbits, we have demonstrated that these prostaglandins modulate luteal nitric oxide synthase (NOS) activity and progesterone production differently, depending on the age of the CL. On CL obtained on day 4 of pseudopregnancy (day-4), PGE-2 was found to depress NOS total activity to 13% of control and to significantly increase basal progesterone secretion by 61%, while PGF-2 had no effect. On day-9 CL, PGE-2 was ineffective, but PGF-2 caused a 2·5-fold increase of NOS activity and a marked decrease in progesterone production. Using specific inhibitors, we found that the regulatory actions of PGE-2 in vitro are mediated via the adenyl cyclase/protein kinase A (PKA) second messenger system, while the PGF-2 -induced luteolytic effects on day-9 CL depend upon activation of the phospholipase C/protein kinase C (PKC) system. The different responsiveness of day-4 and day-9 CL to PGE-2 and PGF-2 could depend on receptor availability for these two prostaglandins, even if other cellular mechanisms cannot be excluded. The present study supports a functional role for NOS in regulating the steroidogenic capacity of rabbit CL, and reveals a novel interaction between a stimulatory G-protein-coupled receptor and PKC/PKA-mediated signal transduction modulating NOS activity.

Novel insights into the mechanisms of pregnancy establishment: regulation of prostaglandin synthesis and signaling in the pig

“…PTGER2 and PTGER4 are coupled to adenylate cyclase and generate cAMP that in turn activates the protein kinase A signaling pathway. PGE 2 stimulates luteal progesterone secretion through a cAMP-mediated pathway in the CL in humans (Hahlin et al 1988, Harris et al 2001, rabbits (Boiti et al 2000), and ruminants (Fitz et al 1984, Weems et al 2006. Moreover, the luteoprotective action of PGE 2 may be mediated by elevated content of vascular endothelial growth factor (VEGFA) in the luteal cells on days 10-12 of pregnancy (Kowalczyk et al 2008).…”

Direct actions of ACTH on ovarian function of pseudopregnant rabbits

“…Each incubation set of wells was divided into 6 experimental groups of 5 181 wells as follows: (I) medium alone as control; (II) ACTH (ACTH 1-24, 1 M); (III) ACTH plus adenylyl cyclase 182 (AC) inhibitor (2-O-methyladenosine, 2 M); (IV) ACTH plus PKA inhibitor (H-9, 3 M); and (V) ACTH plus 183 calcium channel blocker (verapamil). The culture plates were incubated at 37° C in air with 5% CO 2 as reported 184 elsewhere (Boiti et al, 2000). The medium of each well was collected after 4h of incubation and stored 185 immediately at -20° C for later determination of progesterone, PGF 2 , and PGE 2 .…”

“…citrulline using a commercial NOS assay kit according to the procedure previously described (Boiti et al, 2000). 225 226…”

Expression of type I GNRH receptor and in vivo and in vitro GNRH-I effects in corpora lutea of pseudopregnant rabbits

“…Therefore, the main objectives of the present study were to examine the in vivo and in vitro modulatory effects of GNRH on rabbit CL during early-, mid-, and late-luteal stages of pseudopregnancy. With this end in view, experiments were devised to clarify the presence of GNRHR-I and GNRH-I and their possible post-receptorial mechanisms, as well as the effects of GNRH-I on the production of progesterone, PGE 2 , and PGF 2a and on the activities of COX-1 and -2 and nitric oxide synthase (NOS), the enzymes involved in the regulation of rabbit CL life span (Boiti et al 2000, 2005, Zerani et al 2007). …”