CELLULAR MECHANISMS MEDIATING THE STIMULATION OF OVINE ENDOMETRIAL SECRETION OF PROSTAGLANDIN-F2-ALPHA IN RESPONSE TO OXYTOCIN - ROLE OF PHOSPHOLIPASE-C AND DIACYLGLYCEROL

The first objective was to describe and evaluate the relationship betw een the ability of oxytocin to stimulate the activity of phospholipase (PL) C and its ability to stimulate the release of prostaglandin (PG) F-2 alpha in ovine endometrial tissue. Caruncular endometrial tissue was collected from ovariectomized ewes after completion of an 11-day s teroid replacement protocol. In experiment 1, explants were incubated either in the presence (10(-6) M) or absence of oxytocin for 0, 1, 3, 10, 30 or 100 min to examine the time-course for activation of PLC and release of PGF(2 alpha) in response to oxytocin. An increase in the a ctivity of PLC was detected at 3 min while an increase in the release of PGF(2 alpha) was not detected until 10 min (P<0.05). In experiment 2, explants were incubated in the presence of various oxytocin analogu es (10(-6) M) to compare their abilities to activate PLC and release P GF(2 alpha). Oxytocin and three receptor angonists stimulated the acti vity of PLC and the release of PGF(2 alpha) (P<0.05) while two oxytoci n receptor antagonists had no effect on either response. In experiment 3, explants were incubated inthe presence of oxytocin or arginine vas opressin at 10(-9) to 10(-6) M to establish dose-response curves for t he activation of PLC and release of PGF(2 alpha). For both hormones, s ignificant increases (P<0.05) in the release of PGF(2 alpha) were obse rved at 10(-8) M while increases in PLC activity were not detected unt il 10(-7) M was used. In experiment 4, explants were pretreated with e ither U-73122 (an inhibitor of PLC activity) or U-73343 (an inactive a nalogue of U-73122). Explants were then treated with control medium, o xytocin or AlF4-. Both oxytocin and AlF4- stimulated the activity of P LC and the release of PGF(2 alpha) (P<0.05). U-73122 blocked the abili ty of oxytocin to stimulate the release of PGF(2 alpha), (P<0.05) but had no effect on its ability to stimulate the activity of PLC (P>0.1). Based on the results from these experiments, the role of PLC in media ting the stimulatory effect of oxytocin on the release of PGF(2 alpha) remains unclear. The second objective was to evaluate the role of dia cylglycerol (DAG) in mediating the stimulatory effect of oxytocin on e ndometrial secretion of PGF(2 alpha). In experiment 5, explants were i ncubated in vitro with varying doses of two DAG analogues. Both analog ues stimulated the release of PGF(2 alpha) at 10(-6) M (P<0.05), the h ighest dose tested. Corresponding inactive control compounds had no st imulatory effect. In experiment 6, explants were incubated with two sy nthetic DAGs and two indole-derived analogues of DAG. The indole deriv atives stimulated the release of PGF(2 alpha). The synthetic DAGs were less effective in stimulating the release of PGF(2 alpha) at the dose s tested. In experiment 7, explants were preincubated with R59022 or L iCl. R59022 enhanced both the basal and oxytocin-stimulated released o f PGF(2 alpha) (P=0.07). LiCl promoted an increase in the accumulation of inositol trisphosphate (P<0.05) but had no effect on the release o f PGF(2 alpha) (P>0.5). These data indicate that DAG stimulates releas e of PGF(2 alpha) from ovine endometrial tissue and may mediate the st imulatory effect of oxytocin on release of PGF(2 alpha).