Cellular mechanisms by which oxytocin mediates ovine endometrial prostaglandin F-2 alpha synthesis: Role of G(i) proteins and mitogen-activated protein kinases

Oxytocin stimulates a rapid increase in ovine endometrial prostaglandin (PG ) F-2 alpha synthesis. The overall objective of these experiments was to in vestigate the cellular mechanisms by which oxytocin induces endometrial PGF (2 alpha). synthesis. The objective of experiment I was to determine whethe r G(i) proteins mediate oxytocin-induced PGF(2 alpha) synthesis. Uteri were collected from four ovary-intact ewes on Day 14 postestrus. Caruncular end ometrial explants were dissected and subjected to in vitro incubation. Pert ussis toxin, an inhibitor of Gi proteins, had no effect on the ability of o xytocin to induce PGF(2 alpha). synthesis (P > 0.10). The objective of expe riment 2 was to determine whether any of the three mitogen-activated protei n kinases (MAPKs), extracellular signal regulated protein kinase (ERK1/2), c-Jun N-terminal/stress-activated protein kinase (JNK/SAPK), or p38 MAPK, m ediate oxytocin-induced PGF(2 alpha) synthesis. Eleven ovary-intact ewes we re given an injection of oxytocin (10 IU; i.v.; n = 5) or physiological sal ine (i.v.; n = 6) on Day 15 postestrus. Uteri were collected 15 min after i njection and caruncular endometrium was dissected. Endometrial homogenates were prepared and subjected to Western blotting. Membranes were probed for both total and phosphorylated forms of all three classes of MAPK. All class es of MAPK were detected in ovine endometrium, but oxytocin treatment had n o effect on the expression of these proteins (P > 0.10). ERK1/2 was the onl y phosphorylated MAPK detected and its concentrations were higher in oxytoc in-treated ewes (P < 0.01). The objective of experiment 3 was to further in vestigate the role of ERK1/2 during oxytocin-induced PGF(2 alpha) synthesis . Uteri were collected from four ovary-intact ewes on Day 14 postestrus. Ca runcular endometrial explants were dissected and subjected to in vitro incu bation. PD98059, a specific inhibitor of ERK1/2 activity, blocked the abili ty of oxytocin to stimulate PGF(2 alpha) synthesis in a dose-dependent mann er (P < 0.05). These results indicate that the ovine oxytocin receptor is n ot coupled to G(i) proteins. These results indicate that oxytocin induces p hosphorylation of ERK1/2 and that this MAPK appears to mediate oxytocin-ind uced PGF(2 alpha) synthesis in ovine endometrium.