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Oxytocin modulates the pulsatile secretion of prostaglandin F2αin initiated luteolysis in cattle
Research in Veterinary Science 1998, 66, 1–5 Article No. rvsc.1998.0230, available online at http://idealibrary.com on IDE
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Oxytocin modulates the pulsatile secretion of prostaglandin F2α in initiated luteolysis in cattle J. KOTWICA*, D. SKARZYNSKI, G. MISZKIEL, Institute of Animal Reproduction and Food Research, Polish Academy of Sciences, 10–718 Olsztyn-Kortowo, Poland, P. MELIN, Ferring Research Institute AB, S-200 61 Malmo, Sweden, K. OKUDA, Division of Animal Science, Okayama University, Okayama 700, Japan
SUMMARY Subluteolytic doses of prostaglandin F2α analogue (oestrophan) given i.m. and oxytocin (OT) antagonist (CAP) and noradrenaline (NA) infused into the abdominal aorta were used to test the importance of luteal OT in pulsatile secretion of prostaglandin F2α (PGF) during luteolysis in heifers (n = 17). In experiment 1, heifers were pre-infused for 30 minutes with saline on either day 17 of the oestrous cycle (group 1; n = 4) or on day 18 of the oestrous cycle (group 2; n = 3), and with CAP (8 mg per animal) on day 17 of the oestrous cycle (group 3; n = 4). Next, heifers were injected with oestrophan (30 µg per animal). Injection of oestrophan in Group 3 increased OT concentrations (P < 0.001) to values similar to those observed during spontaneous luteolysis (50 to 70 pg ml–1). PGFM concentrations in this group also increased (P < 0.001), but were lower (P < 0.05) than the values in groups 1 and 2. CAP given prior to oestrophan decreased both PGFM elevation (P < 0.06) and its area under the curve (P < 0.01), compared to the saline pretreated heifers. In experiment 2 NA (4 mg) was infused twice for 30 minutes at five hour intervals to release OT on day 17 of the oestrous cycle (n = 6). However, during hormone analysis it appeared that three of six heifers had elevated PGFM concentrations (group 1) and three others did not (group 2). NA caused the correlated increase of progesterone and OT secretion (r = 0.68; P < 0.05) in both groups but it only influenced PGF secretion in group 1 only (P < 0.05). We postulate that OT can amplify and modulate the course of induced luteolysis as a regulator of the amplitude of pulsatile PGF secretion. PGF analogue stimulates secretion of endogenous PGF from the uterus in cattle and this may be an important component of the luteolytic response to exogenous PGF.
AFTER formation of oxytocin (OT) receptors in the uterus, OT stimulates the release of uterine prostaglandin F2α (PGF) in bovines in vivo (Homanics and Silvia 1988, Silvia and Taylor 1989) and in vitro (Silvia and Homanics 1988) and hence it has been suggested to be a trigger for luteolysis in ruminants (Flint and Sheldrick 1983; McCracken et al 1984). However, some data suggest that OT is not necessary for initiation of luteolysis. Moore et al (1986) showed that pulsatile OT release during luteolysis in ewes is preceded by an output of endometrial PGF. In hysterectomized cows, OT pulses were not observed on the expected days of physiological luteolysis (Schams et al 1985), probably indicating the importance of uterine secretion of PGF in OT release. In ovariectomized ewes treated with progesterone and with oestrogens, a pulsatile release of endogenous, uterine PGF was observed without the influence of OT (Lye et al 1983). Moreover, treatment of heifers every four hours with 8 mg of highly specific oxytocin receptor antagonist, (CAP) (which completely blocked OT receptors) from day 15 until evident signs of oestrus affects neither luteolysis nor oestrous cycle duration (Kotwica et al 1997). Finally, the depletion of OT stored in the corpus luteum (CL) by means of noradrenaline (NA), by 50 per cent during the early luteal phase (Jaroszewski and Kotwica 1994) or by 75 to 82 per cent at the mid- and late luteal phases (Kotwica and Skarzynski 1993), did not change the duration of the oestrous cycle in cattle. In our recent study (Skarzynski et al, 1997), the injection of 30 µg of prostaglandin F2α analogue (oestrophan) in cattle evoked an *Corresponding author 0034-5288/99/010001 + 05 $18.00/0
increase of up to 70 pg ml–1, a level similar to that observed during spontaneous luteolysis (Schams et al 1985, Kotwica et al 1997). Pulses of OT in this study were accompanied by synchronous peaks of PGF. Therefore it was assumed that the use of low, subluteolytic doses of oestrophan and NA, both of which release ovarian OT, together with an OT antagonist preventing its physiological effect, would test the importance of luteal OT in pulsatile secretion of PGF and the resulting luteolysis in cattle. OT
MATERIALS AND METHODS Animals and surgery The experiments were carried out in accordance with the principles for the care and use of research animals (Instruction by Director of Institute No. 29, from 10 June 1991). Mature heifers weighing 380 to 450 kg with oestrous cycles of normal length (18 to 22 days) were used in this study. Those with developed CL (7 to 14 days of oestrous cycle) confirmed by rectal palpation were synchronized with 500 µg oestrophan (Spofa, Czech Republic). The day of the first signs of oestrus was designated day 0 of the oestrous cycle. The day before experiments a silastic catheter (OD 1.7 mm, ID 1.1 mm) was inserted into the abdominal aorta through the coccygeal artery (Kotwica et al 1990) to facilitate the infusion of either saline, OT antagonist (1-deamino-2- D-Tyr(0Et)-4-Thr-8-Orn-oxytocin; CAP527, Ferring AB, Malmo, Sweden) or NA (Polfa, Poland). The tip of the catheter was located cranially to the origin of © 1999 W. B. Saunders Company Ltd
J. Kotwica, D. Skarzynski, G. Miszkiel, P. Melin, K. Okuda
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TABLE 1: PGFM and oxytocin (OT) concentrations (mean±SEM) after two injections of prostaglandin F2α analogue (30 µg) at a five hour interval in each heifer on day 17 of the oestrous cycle as measured by height of peak and area under the curve. Group (pg ml–1)
Hormone (relative units)
Saline pretreatment (n = 4)
PGFM
pretreatment (n = 4)
PGFM
CAP
OT
OT
Height of the peak
Area under the curve
134 ± 11 68 ± 12
14790 ± 414 3920 ± 478
109 ± 5 55 ± 8
12790 ± 266* 3790 ± 255
*Values are lower compared to saline pretreatment group (P < 0.01).
ovarian artery and caudally to the both the renal vein and artery (Kotwica et al 1990). The second catheter was inserted into the jugular vein for blood sample collection. Blood samples were collected every five to 10 minutes during experiments and were prevented from clotting by the addition of 30 µM EDTA and 1 per cent aspirin. After centrifugation at 1000 g for 10 minutes, plasma was collected and stored at – 20°C until hormone analysis. Experiment 1 Group 1. On day 17 of the oestrous cycle heifers (n = 4) were injected twice with 30 µg of oestrophan, a dose sufficient to stimulate OT secretion (Skarzynski et al 1997) and similar to that observed during spontaneous luteolysis. This treatment was preceded with an infusion of saline (30 minutes). Group 2. Oestrophan (30 µg) was injected on day 18 of the oestrous cycle in three heifers pretreated with saline as in the previous group.
(Fig 1a). It was correlated with an increase in progesterone (r = 0.74, P < 0.001) and PGFM (r = 0.65, P < 0.01) concentrations. The increase of PGFM clearly followed the release of OT. In group 2, the concentration of progesterone in plasma before oestrophan injection was in the range of 0.5 to 1.5 ng ml–1. In these heifers, luteolysis was already underway. Injection of oestrophan only increased PGFM (Fig 1b) in blood (P < 0.01). There were no significant changes in OT concentration. Furthermore oestrophan given after CAP infusion (group 3) increased (P < 0.001) OT concentrations (Fig 1c) to values similar to those observed during spontaneous luteolysis (50 to 70 pg ml–1). Concentrations of PGFM were also increased (P < 0.001), but were lower (P < 0.05) when compared to the saline pretreatment heifers (groups 1 and 2). Moreover, both PGFM elevation and the AUC were reduced by 19.5 per cent (P < 0.06) and by 14.5 per cent (P < 0.01), respectively, compared to the saline pretreatment (Table 1). Experiment 2 During hormones analysis it appeared that in three heifers concentrations before treatment were already above 100 pg ml–1 whereas in three others they were in the range of 50 to 65 pg ml–1 (P < 0.05). Thus, heifers were divided into two groups; those with elevated (group 1; Fig 2a) and those with low (group 2; Fig 2b) PGFM plasma concentrations before NA infusion. Concomitantly, the concentration of progesterone in heifers in group 2 was over 5 ng ml–1, whereas in group 1, progesterone concentrations were lower (P < 0.06) as measured by the AUC. Infusion of NA into these heifers caused an increase in progesterone and OT secretion, which were correlated (r = 0.68: P < 0.05) in both groups. However, NA increased PGF secretion (P < 0.05) compared to the pre-treatment period in group 1 only (Fig 2a). PGFM
Group 3. Eight mg of CAP is sufficient to completely block OT receptors in heifers within 4 hours (Kotwica et al 1997). Therefore, on day 17 of the oestrous cycle the heifers (n = 4) were infused twice for 30 minutes with 8 mg of CAP-527 every five hours and subsequently injected (i.m.) with a subluteolytic dose of oestrophan (30 µg). We assumed that the oestrophan would release ovarian OT, while its receptors were occupied by CAP. Thus, endogenous PGF release could not be modified by OT. Experiment 2 NA (4 mg) was infused twice for 30 minutes at a five hour internal in heifers (n = 6) on day 17 of the oestrous cycle to release OT and to determine whether it would induce PGF secretion if secreted in physiological pulses (i.e. 50 to 70 pg ml–1).
RESULTS Experiment 1 The concentration of progesterone in the control period of heifers in group 1 ranged from 7 to 12 ng ml–1. In this group a sharp increase in OT concentration (P < 0.001) was observed within five minutes of oestrophan injection
Hormone analysis Progesterone was determined (Kotwica et al 1990) using rabbit progesterone antiserum (IFP4) characterised previously (Kotwica et al. 1994). The sensitivity of the procedure was 15 pg tube–1. The intra- and interassay coefficients of variation were 8.1 and 15.4 per cent, respectively. The precision, of the procedure is expressed by the linear regression equation (y = 1.034x–0.13). Concentrations of OT were determined (Schams et al 1979) using rabbit OT antiserum (R-1), as characterised previously (Kotwica and Skarzynski 1993). The efficiency of extraction was 85 per cent. The sensitivity of the method was 3 pg ml–1. Intra- and interassay coefficients of variation
Effect of oxytocin on PGF2α secretion in cattle 180
200
12
(a) Saline
3
15
(a)
NA
NA
Saline 150
135
10
4
0
PGFM
180 (b)
12
Saline
Saline
135 8 90 4 45
Progesterone (ng ml–1)
(pg ml–1); Oxytocin (pg ml–1)
0
0
0
100 5 50
0
0 200 (b)
10 100 5 50
0
0 0
12 Cap
NA
150
–60
180 (c)
15 NA
Progesterone (ng ml–1)
45
PGFM
90
(pg ml–1); Oxytocin (pg ml–1)
8
60
120 180 240 300 360 420 Time (minutes)
Cap
135 8
Fig. 2 Peripheral concentration of oxytocin (●), PGFM (● ● ) and progesterone (bars) (mean ± SEM) in heifers infused twice five hours apart with noradrena–1 –1 line (NA; 0.3 µg kg min ) for 30 minutes (horizontal lines) on day 17 of the oestrous cycle. Heifers had elevated (a; n = 3) and low (b; n = 3) concentrations of PGFM before treatment.
90 4 45
0
–60
0
60
120 180 240 300 360 420
0
Time (minutes) Fig. 1 Peripheral concentration of oxytocin (●), PGFM (● ● ) and progesterone (bars) (mean ± SEM) in heifers injected twice i.m. (arrows) five hours apart with an analogue of prostaglandin F2α (30 µg per animal) and preinfused into abdominal aorta for 30 minutes with: saline on day 17 of the oestrous cycle (a; n = 4), saline on day 18 of the oestrous cycle (b; n = 3), and oxytocin antagonist, CAP (8 mg per animal) on day 17 of the oestrous cycle (c; n = 4).
were 7.5 and 14.6 per cent, respectively. The relationship between real (x) and determined (y) amounts of four different concentrations of OT added to the plasma samples and included in each assay is expressed by the linear regression equation (y = 0.99x + 0.14). Plasma 13,14-dihydro-15-keto-prostaglandin F2α (PGFM) was measured via the procedure described by Homanics and Silvia (1988) using antiserum for PGFM (WS-44685). The sensitivity of the assay was 65 pg ml–1 and intra- and interassay coefficients of variation were 10.9 and 14.8 per cent, respectively. The precision of the procedure is expressed by the linear regression equation (y = 0.96x–4.4).
In order to avoid the criticism that the PGFM antibody might cross-react with PGF2α analogue or with its metabolite, oestrophan was incubated with blood (10 ng ml–1) at 37°C for 30, 60 and 120 minutes and, plasma was the subjected to RIA for PGFM. The values obtained (n = 5 each time) were not different from the same plasma samples incubated without oestrophan. Statistical analysis Baseline concentrations of OT and PGFM were defined as the average concentration from time – 60 to 0 min, the period prior to oestrophan injection. The amounts of PGF and OT released after oestrophan and NA treatment were measured by calculating the area under the curve (AUC) within two hours of treatment, using the GraphPad PRISM program (San Diego, CA, U.S.A.). The AUC for progesterone was calculated over the whole experiment. The correlation between OT and PGFM after treatment was calculated. The differences between mean (± SEM) values were determined by a one-way analysis of variance.
DISCUSSION The aim of this study was to determine whether endogenous OT released just before luteolysis by a subluteolytic
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J. Kotwica, D. Skarzynski, G. Miszkiel, P. Melin, K. Okuda
dose of oestrophan or by NA was able to release PGF from the uterus. A lack of PGFM response after NA infusion was observed in spite of pulsatile OT release (Fig 2b), compared to the response seen during spontaneous luteolysis. Moreover, blocking OT receptors did not prevent increases of PGFM in blood (Fig 1c). However, if the progesterone level was below 1.5 ng ml–1 (Fig 1b), oestrophan did not affect OT secretion but increased PGF release secretion. These results indicate that OT is not necessary for initiating pulsatile PGF release during luteolysis in cattle, but that exogenous PGF2α stimulates utero-ovarian release of PGF as found in cyclic (Wade and Lewis 1996) and pregnant (Weems et al 1993) ewes. The stimulus seems to be a via direct effect on the utero-ovarian unit, rather than a result of PGF-induced changes in progesterone or the effects of ovarian OT on the uterus (Wade and Lewis 1996). Although the increase in PGF secretion appeared after the OT peak, these increases were independent of each other, as clearly illustrated if oestrophan injection preceded the blockage of OT receptors by CAP (Fig 1c). During the late luteal phase (15 to 19 days of cycle) exogenous OT (100 IU) caused a release of PGF (Howard and Britt 1987, Silvia and Taylor 1989, Parkinson et al 1992, Lafrance and Goff 1993). However, after i.v. injection in cattle, this dose increased the concentration of OT in peripheral blood above 2000 pg ml–1 (Howard and Britt 1987). This is 40 times higher than the physiological level of OT pulses seen during luteolysis (Schams et al 1985, Kotwica et al 1993, Kotwica et al 1997). Thus, it seems that only pharmacological doses of OT are able to release PGF from the uterus in experiments in vivo. It has been shown that within a few minutes of an intra-arterial injection of 10 or 20 IU of OT in heifers on days 16 to 17 of the cycle, peripheral concentrations of OT increased to 200 and 350 to 500 pg ml–1, respectively (Kotwica et al 1993). However, this did not cause changes in PGF secretion. Treatment with 50 IU of OT increased OT in blood to 1200 pg ml–1 which increased PGFM concentration in blood plasma from 50 to 70 pg ml–1 to 100 to 200 pg ml–1. During spontaneous luteolysis, the levels of pulsatile PGF secretion reached 300 to 500 pg ml–1, but OT pulses were approximately 70 pg ml–1 (Schams et al 1985, Kotwica et al 1993, 1997. Moreover, treatment of heifers with an OT receptor blocker from day 15 to evident signs of oestrus caused no changes in progesterone or PGF secretion and did not affect cycle duration compared to the control heifers (Kotwica et al 1997). The highest increase of OTinduced PGF secretion has been observed just before and during CL regression in cattle (Mirando et al 1993). However OT receptor changes in endometrium and inositol hydrolysis after OT treatment clearly occurred later and were negatively correlated with progesterone on day 18 and positively related to those of circulating concentrations of oestradiol on day 17 of the cycle (Parkinson et al 1990). This conclusion is similar those of Meyer et al (1988) and Soloff and Fields (1989). It also suggests that the time of initiation of luteolysis appears to be independent of changes in OT receptor synthesis, but depends on the duration of the high plasma progesterone concentration characteristic of the luteal phase (Kotwica 1988, Geisert et al 1992, Silvia and Raw 1993). On the basis of the data presented, OT may play a supportive and modulatory role in luteolysis. In experiment 1, when OT receptors were blocked, the magnitude of PGF release
decreased (Table 1). In ovariectomized ewes treated with progesterone for 15 days, PGF secretion was reduced without a concomitant release of OT (Silvia and Raw 1993). These authors suggest that ovarian OT is not required to initiate pulsatile secretion of PGF, but may be required to achieve full pulse amplitude. When luteolysis was in progress, NA released OT and later increased PGF secretion (Fig 2b). This may be a direct influence of NA on the uterus. NA was found to affect PGF release from the human uterus before and during ovulation (Quass et al 1987, Tanaka et al 1993) and from the human decidua (Tada et al 1991). In rats, catecholamines increase PGF secretion via β-adrenergic receptors (Ishikawa and Fuchs 1978). Moreover, blocking both α- and β-adrenergic receptors in the human uterus decreases catecholamine-induced PGF release (Quass and Zahradnik 1985). However, indirect actions of NA on PGF secretion by OT release are also possible. We postulated that OT is not the main factor in the initiation of luteolysis in cattle and this supports data from Blair et al (1997). However, it may play a supportive and modulatory role in induced luteolysis as a regulator of the amplitude of pulsatile PGF secretion. PGF analogue stimulates the secretion of endogenous PGF from the uterus in cattle and this may be the essential component of the luteolytic action of exogenous PGF, as suggested in ewes (Wade and Lewis 1996).
ACKNOWLEDGEMENTS We thank Dr S. Okrasa and Dr G. Kotwica (University of Agriculture and Technology, Olsztyn, Poland) and Dr W.J. Silvia (University of Kentucky, Lexington, U.S.A.) for progesterone, oxytocin and PGFM antiserum. CAP was kindly donated by Ferring AB, Sweden. This study was supported by The Polish Academy of Sciences.
REFERENCES BLAIR, R.M., SAATMAN, R., LIOU, S.S., FORTUNE, J.E. & HANSEL, W. (1997) Roles of leukotrienes in bovine corpus luteum regression: an in vivo microdialysis study. Proceedings of the Society for Experimental Biology and Medicine 216, 72–80 FLINT, A.P.F. & SHELDRICK, E.L. (1983) Secretion of oxytocin by the corpus luteum in sheep. Progress in Brain Research 60, 521–530 GEISERT, R.D., SHORT, E.C. & ZAVY, M.T. (1992) Maternal recognition of pregnancy. Animal Reproduction Science 28, 287–298 HOMANICS, G.E. & SILVIA, W.J. (1988) Effects of progesterone and estradiol on uterine secretion of prostaglandin F2α in response to oxytocin in ovariectomized ewes. Biology of Reproduction 38, 804–811 HOWARD, H.J. & BRITT, J.H. (1987) Prostaglandin F metabolite after exogenous oxytocin in cows given hCG during diestrus. Biology of Reproduction 36(Suppl. 1), 274 ISHIKAWA, M. & FUCHS, A.R. (1978) Effects of epinephrine and oxytocin on the release of prostaglandin F from the rat uterus in vitro. Prostaglandins 15, 89–101 JAROSZEWSKI, J. & KOTWICA, J. (1994) Reduction of oxytocin content in corpus luteum does not affect the duration of the estrous cycle in cattle. Reproductive Nutrition and Development 34, 175–182 KOTWICA J. (1988) Ovarian oxytocin in bovine oestrous cycle regulation. Acta Academiae Agriculture et Technicae Olstenensis Zootechnica 31 (Suppl. C), 1–45 (in Polish) KOTWICA, J., SKARZYNSKI, D. & JAROSZEWSKI, J. (1990) Coccygeal artery as a route for administration of drugs into the reproductive tract of cattle. The Veterinary Record 127, 38–40 KOTWICA, J. & SKARZYNSKI, D. (1993a) Influence of oxytocin removal from the corpus luteum on secretory function and duration of the oestrous cycle in cattle. Journal of Reproduction and Fertility 97, 411–417 KOTWICA, J., SKARZYNSKI, D. & JAROSZEWSKI, J. (1993b) Secretion of PGF stimulated by different doses of oxytocin and released spontaneously during luteolysis in cattle. Journal of Reproduction and Fertility Abstract Series 12, 60
Effect of oxytocin on PGF2α secretion in cattle KOTWICA, J., SKARZYNSKI, D., JAROSZEWSKI, J. & BOGACKI, M. (1994) Noradrenaline affects secretory function of corpus luteum independently on prostaglandins in conscious cattle. Prostaglandins 48, 1–10 KOTWICA, J., SKARZYNSKI, D., BOGACKI, M., MELIN, P. & STAROSTKA B. (1997) The use of an oxytocin antagonist to study the function of ovarian oxytocin during luteolysis in cattle. Theriogenology 48, 1287–1299 LAFRANCE, M. & GOFF, A.K. (1993) Effects of progesterone and estradiol-17β on oxytocin-induced release of prostaglandin in heifers. Biology of Reproduction 34 (Suppl. 1), 161 LYE, S. J., SPRAGUE, C. L. & CHALLIS, H. H. D. (1983) Modulation of ovine myometrial activity by estradiol-17β. The possible involvement of prostaglandins. Canadian Journal of Physiology and Pharmacology 61, 729–735 MCCRACKEN, J. A., SCHRAMM, W. & OKULICZ, W. C. (1984) Hormone receptor control of pulsatile secretion of PGF2α from the uterus luteolysis and abrogation in early pregnancy. Animal Reproduction Science 7, 31–55 MEYER, H.H.D., MITTERMEIER, T. & SCHAMS, D. (1988) Dynamics of oxytocin, estrogen and progestin receptors in the bovine endometrium during the estrous cycle. Acta Endocrinologica 118, 96–104 MIRANDO, M. A., BECKER, W. C. & WHITEAKER, S.S. (1993) Relationship among endometrial oxytocin receptors, oxytocin-stimulated phosphoinositide hydrolysis and prostaglandin F2α secretion in vitro, and plasma concentrations of ovarian steroids before and during corpus luteum regression in cyclic heifers. Biology of Reproduction 48, 874–882 MOORE, L. G., CHOY, V. J. & ELLIOT, R. L. WATKINS, W. B. (1986) Evidence for the release of prostaglandin F2α inducing the release of ovarian oxytocin during luteolysis in the ewe. Journal Reproduction and Fertility 75, 159–166 PARKINSON, T.J., JENNER, L.J. & LAMMING, G.E. (1990) Comparison of oxytocin/prostaglandin F-2α interrelationship in cyclic and pregnant cows. Journal of Reproduction and Fertility 90, 337–345 PARKINSON, T. J., WATHES, D. C., JENNER, L. J. & LAMMING, G. E. (1992) Plasma and luteal concentrations of oxytocin in cyclic and early-pregnant cattle. Journal of Reproduction and Fertility 94, 161–167 QUAAS, L. & ZAHRADNIK, H. P. (1985) The effects of α- and β-adrenergic stimulation on contractility and prostaglandin (prostaglandins E2 and F2α and 6-ketoprostaglandin 1α) strips. American Journal of Obstetrics and Gynecology 157, 856–865 QUASS, L., GOPPINGER, A. & ZAHRADNIK, H. P. (1987) The effect of acetylosalicylic acid and indomethacin on catecholamine- and oxytocin-induced contractility and prostaglandin (6-keto-PGF1α, PGF2α) – production of human pregnant myometrial strips. Prostaglandins 34, 257–269
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SCHAMS, D., SCHMIDT-POLEX, B. & KRUSE, V. (1979) Oxytocin determination by radioimmunoassay in cattle. Acta Endocrinologica 92, 258–270 SCHAMS, D., SCHALLENBERGER, E., MEYER, H. H. D., BULLERMANN, B., BREITINGER, H.J., KOLL, R., ENZENHOFER, G., KRUIP, T.A.M., WALTERS, D.L. & KARG, H. (1985) Ovarian oxytocin during estrous cycle in cattle. In Oxytocin, Clinical and Laboratory Studies. Eds. J. A. Amico and A.G. Robinson. Amsterdam, Elsevier Biomedical, pp 317–334 SILVIA, W.J. & HOMANICS, G.E. (1988) Role of phospholipase C in mediating oxytocin-induced release of prostaglandin F2α from ovine endometrial tissue. Prostaglandins 35, 535–548 SILVIA, W. J. & TAYLOR, M. L. (1989) Relationship between uterine secretion of prostaglandin F2α induced by oxytocin and endogenous concentration of estradiol and progesterone at three stages of the bovine oestrous cycle. Journal of Animal Science 67, 2347–2353 SILVIA, W.J. & RAW, R.E. (1993) Regulation of pulsatile secretion of prostaglandin F2α from the ovine uterus by ovarian steroids. Journal of Reproduction and Fertility 98, 341–347 SKARZYNSKI, D., BOGACKI, M. & KOTWICA, J. (1997) Changes in ovarian oxytocin secretion as an indicator of corpus luteum response to prostaglandin F2α treatment in cattle. Theriogenology 48, 733–742 SOLOFF, M.S. & FIELDS, M.J. (1989) Changes in uterine oxytocin receptor concentrations throughout the estrous cycle of the cow. Biology of Reproduction 40, 283–287 TANAKA, N., MIYAZAKI, K., TASHIRO, H., MIZUTANI, H. & OKAMURA, H. (1993) Changes in adenylyl cyclase activity in human endometrium during the menstrual cycle and in human decidua during pregnancy. Journal of Reproduction and Fertility 98, 33–39 TADA, K., KUDO, T. & KISHIMOTO, Y. (1991) Effect of L-dopa or dopamine on human decidual prostaglandin synthesis. Acta Medicine Okayama 45, 333–336 WADE, D.E. & LEWIS, G. S. (1996) Exogenous prostaglandin F2α stimulates uteroovarian release of prostaglandin F2α in sheep: possible component of luteolytic mechanism of action of exogenous prostaglandin F2α. Domestic Animal Endocrinology 13, 383–398 WEEMS, Y. S., VINCENT, D. L., NUSSER, K. D., TANAKA, Y., MILLERPATRICK, K., LEDGERWOOD, K. S. & WEEMS, C. W. (1993) Effect of prostaglandin F2α on uterine or ovarian secretion of prostaglandins E and F2α in vivo in 90–100 day hysterectomised, intact or ovariectomized pregnant ewes. Prostaglandins 46, 277–296 Accepted July 29, 1998
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Oxytocin modulates the pulsatile secretion of prostaglandin F2α in initiated luteolysis in cattle J. KOTWICA*, D. SKARZYNSKI, G. MISZKIEL, Institute of Animal Reproduction and Food Research, Polish Academy of Sciences, 10–718 Olsztyn-Kortowo, Poland, P. MELIN, Ferring Research Institute AB, S-200 61 Malmo, Sweden, K. OKUDA, Division of Animal Science, Okayama University, Okayama 700, Japan
SUMMARY Subluteolytic doses of prostaglandin F2α analogue (oestrophan) given i.m. and oxytocin (OT) antagonist (CAP) and noradrenaline (NA) infused into the abdominal aorta were used to test the importance of luteal OT in pulsatile secretion of prostaglandin F2α (PGF) during luteolysis in heifers (n = 17). In experiment 1, heifers were pre-infused for 30 minutes with saline on either day 17 of the oestrous cycle (group 1; n = 4) or on day 18 of the oestrous cycle (group 2; n = 3), and with CAP (8 mg per animal) on day 17 of the oestrous cycle (group 3; n = 4). Next, heifers were injected with oestrophan (30 µg per animal). Injection of oestrophan in Group 3 increased OT concentrations (P < 0.001) to values similar to those observed during spontaneous luteolysis (50 to 70 pg ml–1). PGFM concentrations in this group also increased (P < 0.001), but were lower (P < 0.05) than the values in groups 1 and 2. CAP given prior to oestrophan decreased both PGFM elevation (P < 0.06) and its area under the curve (P < 0.01), compared to the saline pretreated heifers. In experiment 2 NA (4 mg) was infused twice for 30 minutes at five hour intervals to release OT on day 17 of the oestrous cycle (n = 6). However, during hormone analysis it appeared that three of six heifers had elevated PGFM concentrations (group 1) and three others did not (group 2). NA caused the correlated increase of progesterone and OT secretion (r = 0.68; P < 0.05) in both groups but it only influenced PGF secretion in group 1 only (P < 0.05). We postulate that OT can amplify and modulate the course of induced luteolysis as a regulator of the amplitude of pulsatile PGF secretion. PGF analogue stimulates secretion of endogenous PGF from the uterus in cattle and this may be an important component of the luteolytic response to exogenous PGF.
AFTER formation of oxytocin (OT) receptors in the uterus, OT stimulates the release of uterine prostaglandin F2α (PGF) in bovines in vivo (Homanics and Silvia 1988, Silvia and Taylor 1989) and in vitro (Silvia and Homanics 1988) and hence it has been suggested to be a trigger for luteolysis in ruminants (Flint and Sheldrick 1983; McCracken et al 1984). However, some data suggest that OT is not necessary for initiation of luteolysis. Moore et al (1986) showed that pulsatile OT release during luteolysis in ewes is preceded by an output of endometrial PGF. In hysterectomized cows, OT pulses were not observed on the expected days of physiological luteolysis (Schams et al 1985), probably indicating the importance of uterine secretion of PGF in OT release. In ovariectomized ewes treated with progesterone and with oestrogens, a pulsatile release of endogenous, uterine PGF was observed without the influence of OT (Lye et al 1983). Moreover, treatment of heifers every four hours with 8 mg of highly specific oxytocin receptor antagonist, (CAP) (which completely blocked OT receptors) from day 15 until evident signs of oestrus affects neither luteolysis nor oestrous cycle duration (Kotwica et al 1997). Finally, the depletion of OT stored in the corpus luteum (CL) by means of noradrenaline (NA), by 50 per cent during the early luteal phase (Jaroszewski and Kotwica 1994) or by 75 to 82 per cent at the mid- and late luteal phases (Kotwica and Skarzynski 1993), did not change the duration of the oestrous cycle in cattle. In our recent study (Skarzynski et al, 1997), the injection of 30 µg of prostaglandin F2α analogue (oestrophan) in cattle evoked an *Corresponding author 0034-5288/99/010001 + 05 $18.00/0
increase of up to 70 pg ml–1, a level similar to that observed during spontaneous luteolysis (Schams et al 1985, Kotwica et al 1997). Pulses of OT in this study were accompanied by synchronous peaks of PGF. Therefore it was assumed that the use of low, subluteolytic doses of oestrophan and NA, both of which release ovarian OT, together with an OT antagonist preventing its physiological effect, would test the importance of luteal OT in pulsatile secretion of PGF and the resulting luteolysis in cattle. OT
MATERIALS AND METHODS Animals and surgery The experiments were carried out in accordance with the principles for the care and use of research animals (Instruction by Director of Institute No. 29, from 10 June 1991). Mature heifers weighing 380 to 450 kg with oestrous cycles of normal length (18 to 22 days) were used in this study. Those with developed CL (7 to 14 days of oestrous cycle) confirmed by rectal palpation were synchronized with 500 µg oestrophan (Spofa, Czech Republic). The day of the first signs of oestrus was designated day 0 of the oestrous cycle. The day before experiments a silastic catheter (OD 1.7 mm, ID 1.1 mm) was inserted into the abdominal aorta through the coccygeal artery (Kotwica et al 1990) to facilitate the infusion of either saline, OT antagonist (1-deamino-2- D-Tyr(0Et)-4-Thr-8-Orn-oxytocin; CAP527, Ferring AB, Malmo, Sweden) or NA (Polfa, Poland). The tip of the catheter was located cranially to the origin of © 1999 W. B. Saunders Company Ltd
J. Kotwica, D. Skarzynski, G. Miszkiel, P. Melin, K. Okuda
2
TABLE 1: PGFM and oxytocin (OT) concentrations (mean±SEM) after two injections of prostaglandin F2α analogue (30 µg) at a five hour interval in each heifer on day 17 of the oestrous cycle as measured by height of peak and area under the curve. Group (pg ml–1)
Hormone (relative units)
Saline pretreatment (n = 4)
PGFM
pretreatment (n = 4)
PGFM
CAP
OT
OT
Height of the peak
Area under the curve
134 ± 11 68 ± 12
14790 ± 414 3920 ± 478
109 ± 5 55 ± 8
12790 ± 266* 3790 ± 255
*Values are lower compared to saline pretreatment group (P < 0.01).
ovarian artery and caudally to the both the renal vein and artery (Kotwica et al 1990). The second catheter was inserted into the jugular vein for blood sample collection. Blood samples were collected every five to 10 minutes during experiments and were prevented from clotting by the addition of 30 µM EDTA and 1 per cent aspirin. After centrifugation at 1000 g for 10 minutes, plasma was collected and stored at – 20°C until hormone analysis. Experiment 1 Group 1. On day 17 of the oestrous cycle heifers (n = 4) were injected twice with 30 µg of oestrophan, a dose sufficient to stimulate OT secretion (Skarzynski et al 1997) and similar to that observed during spontaneous luteolysis. This treatment was preceded with an infusion of saline (30 minutes). Group 2. Oestrophan (30 µg) was injected on day 18 of the oestrous cycle in three heifers pretreated with saline as in the previous group.
(Fig 1a). It was correlated with an increase in progesterone (r = 0.74, P < 0.001) and PGFM (r = 0.65, P < 0.01) concentrations. The increase of PGFM clearly followed the release of OT. In group 2, the concentration of progesterone in plasma before oestrophan injection was in the range of 0.5 to 1.5 ng ml–1. In these heifers, luteolysis was already underway. Injection of oestrophan only increased PGFM (Fig 1b) in blood (P < 0.01). There were no significant changes in OT concentration. Furthermore oestrophan given after CAP infusion (group 3) increased (P < 0.001) OT concentrations (Fig 1c) to values similar to those observed during spontaneous luteolysis (50 to 70 pg ml–1). Concentrations of PGFM were also increased (P < 0.001), but were lower (P < 0.05) when compared to the saline pretreatment heifers (groups 1 and 2). Moreover, both PGFM elevation and the AUC were reduced by 19.5 per cent (P < 0.06) and by 14.5 per cent (P < 0.01), respectively, compared to the saline pretreatment (Table 1). Experiment 2 During hormones analysis it appeared that in three heifers concentrations before treatment were already above 100 pg ml–1 whereas in three others they were in the range of 50 to 65 pg ml–1 (P < 0.05). Thus, heifers were divided into two groups; those with elevated (group 1; Fig 2a) and those with low (group 2; Fig 2b) PGFM plasma concentrations before NA infusion. Concomitantly, the concentration of progesterone in heifers in group 2 was over 5 ng ml–1, whereas in group 1, progesterone concentrations were lower (P < 0.06) as measured by the AUC. Infusion of NA into these heifers caused an increase in progesterone and OT secretion, which were correlated (r = 0.68: P < 0.05) in both groups. However, NA increased PGF secretion (P < 0.05) compared to the pre-treatment period in group 1 only (Fig 2a). PGFM
Group 3. Eight mg of CAP is sufficient to completely block OT receptors in heifers within 4 hours (Kotwica et al 1997). Therefore, on day 17 of the oestrous cycle the heifers (n = 4) were infused twice for 30 minutes with 8 mg of CAP-527 every five hours and subsequently injected (i.m.) with a subluteolytic dose of oestrophan (30 µg). We assumed that the oestrophan would release ovarian OT, while its receptors were occupied by CAP. Thus, endogenous PGF release could not be modified by OT. Experiment 2 NA (4 mg) was infused twice for 30 minutes at a five hour internal in heifers (n = 6) on day 17 of the oestrous cycle to release OT and to determine whether it would induce PGF secretion if secreted in physiological pulses (i.e. 50 to 70 pg ml–1).
RESULTS Experiment 1 The concentration of progesterone in the control period of heifers in group 1 ranged from 7 to 12 ng ml–1. In this group a sharp increase in OT concentration (P < 0.001) was observed within five minutes of oestrophan injection
Hormone analysis Progesterone was determined (Kotwica et al 1990) using rabbit progesterone antiserum (IFP4) characterised previously (Kotwica et al. 1994). The sensitivity of the procedure was 15 pg tube–1. The intra- and interassay coefficients of variation were 8.1 and 15.4 per cent, respectively. The precision, of the procedure is expressed by the linear regression equation (y = 1.034x–0.13). Concentrations of OT were determined (Schams et al 1979) using rabbit OT antiserum (R-1), as characterised previously (Kotwica and Skarzynski 1993). The efficiency of extraction was 85 per cent. The sensitivity of the method was 3 pg ml–1. Intra- and interassay coefficients of variation
Effect of oxytocin on PGF2α secretion in cattle 180
200
12
(a) Saline
3
15
(a)
NA
NA
Saline 150
135
10
4
0
PGFM
180 (b)
12
Saline
Saline
135 8 90 4 45
Progesterone (ng ml–1)
(pg ml–1); Oxytocin (pg ml–1)
0
0
0
100 5 50
0
0 200 (b)
10 100 5 50
0
0 0
12 Cap
NA
150
–60
180 (c)
15 NA
Progesterone (ng ml–1)
45
PGFM
90
(pg ml–1); Oxytocin (pg ml–1)
8
60
120 180 240 300 360 420 Time (minutes)
Cap
135 8
Fig. 2 Peripheral concentration of oxytocin (●), PGFM (● ● ) and progesterone (bars) (mean ± SEM) in heifers infused twice five hours apart with noradrena–1 –1 line (NA; 0.3 µg kg min ) for 30 minutes (horizontal lines) on day 17 of the oestrous cycle. Heifers had elevated (a; n = 3) and low (b; n = 3) concentrations of PGFM before treatment.
90 4 45
0
–60
0
60
120 180 240 300 360 420
0
Time (minutes) Fig. 1 Peripheral concentration of oxytocin (●), PGFM (● ● ) and progesterone (bars) (mean ± SEM) in heifers injected twice i.m. (arrows) five hours apart with an analogue of prostaglandin F2α (30 µg per animal) and preinfused into abdominal aorta for 30 minutes with: saline on day 17 of the oestrous cycle (a; n = 4), saline on day 18 of the oestrous cycle (b; n = 3), and oxytocin antagonist, CAP (8 mg per animal) on day 17 of the oestrous cycle (c; n = 4).
were 7.5 and 14.6 per cent, respectively. The relationship between real (x) and determined (y) amounts of four different concentrations of OT added to the plasma samples and included in each assay is expressed by the linear regression equation (y = 0.99x + 0.14). Plasma 13,14-dihydro-15-keto-prostaglandin F2α (PGFM) was measured via the procedure described by Homanics and Silvia (1988) using antiserum for PGFM (WS-44685). The sensitivity of the assay was 65 pg ml–1 and intra- and interassay coefficients of variation were 10.9 and 14.8 per cent, respectively. The precision of the procedure is expressed by the linear regression equation (y = 0.96x–4.4).
In order to avoid the criticism that the PGFM antibody might cross-react with PGF2α analogue or with its metabolite, oestrophan was incubated with blood (10 ng ml–1) at 37°C for 30, 60 and 120 minutes and, plasma was the subjected to RIA for PGFM. The values obtained (n = 5 each time) were not different from the same plasma samples incubated without oestrophan. Statistical analysis Baseline concentrations of OT and PGFM were defined as the average concentration from time – 60 to 0 min, the period prior to oestrophan injection. The amounts of PGF and OT released after oestrophan and NA treatment were measured by calculating the area under the curve (AUC) within two hours of treatment, using the GraphPad PRISM program (San Diego, CA, U.S.A.). The AUC for progesterone was calculated over the whole experiment. The correlation between OT and PGFM after treatment was calculated. The differences between mean (± SEM) values were determined by a one-way analysis of variance.
DISCUSSION The aim of this study was to determine whether endogenous OT released just before luteolysis by a subluteolytic
4
J. Kotwica, D. Skarzynski, G. Miszkiel, P. Melin, K. Okuda
dose of oestrophan or by NA was able to release PGF from the uterus. A lack of PGFM response after NA infusion was observed in spite of pulsatile OT release (Fig 2b), compared to the response seen during spontaneous luteolysis. Moreover, blocking OT receptors did not prevent increases of PGFM in blood (Fig 1c). However, if the progesterone level was below 1.5 ng ml–1 (Fig 1b), oestrophan did not affect OT secretion but increased PGF release secretion. These results indicate that OT is not necessary for initiating pulsatile PGF release during luteolysis in cattle, but that exogenous PGF2α stimulates utero-ovarian release of PGF as found in cyclic (Wade and Lewis 1996) and pregnant (Weems et al 1993) ewes. The stimulus seems to be a via direct effect on the utero-ovarian unit, rather than a result of PGF-induced changes in progesterone or the effects of ovarian OT on the uterus (Wade and Lewis 1996). Although the increase in PGF secretion appeared after the OT peak, these increases were independent of each other, as clearly illustrated if oestrophan injection preceded the blockage of OT receptors by CAP (Fig 1c). During the late luteal phase (15 to 19 days of cycle) exogenous OT (100 IU) caused a release of PGF (Howard and Britt 1987, Silvia and Taylor 1989, Parkinson et al 1992, Lafrance and Goff 1993). However, after i.v. injection in cattle, this dose increased the concentration of OT in peripheral blood above 2000 pg ml–1 (Howard and Britt 1987). This is 40 times higher than the physiological level of OT pulses seen during luteolysis (Schams et al 1985, Kotwica et al 1993, Kotwica et al 1997). Thus, it seems that only pharmacological doses of OT are able to release PGF from the uterus in experiments in vivo. It has been shown that within a few minutes of an intra-arterial injection of 10 or 20 IU of OT in heifers on days 16 to 17 of the cycle, peripheral concentrations of OT increased to 200 and 350 to 500 pg ml–1, respectively (Kotwica et al 1993). However, this did not cause changes in PGF secretion. Treatment with 50 IU of OT increased OT in blood to 1200 pg ml–1 which increased PGFM concentration in blood plasma from 50 to 70 pg ml–1 to 100 to 200 pg ml–1. During spontaneous luteolysis, the levels of pulsatile PGF secretion reached 300 to 500 pg ml–1, but OT pulses were approximately 70 pg ml–1 (Schams et al 1985, Kotwica et al 1993, 1997. Moreover, treatment of heifers with an OT receptor blocker from day 15 to evident signs of oestrus caused no changes in progesterone or PGF secretion and did not affect cycle duration compared to the control heifers (Kotwica et al 1997). The highest increase of OTinduced PGF secretion has been observed just before and during CL regression in cattle (Mirando et al 1993). However OT receptor changes in endometrium and inositol hydrolysis after OT treatment clearly occurred later and were negatively correlated with progesterone on day 18 and positively related to those of circulating concentrations of oestradiol on day 17 of the cycle (Parkinson et al 1990). This conclusion is similar those of Meyer et al (1988) and Soloff and Fields (1989). It also suggests that the time of initiation of luteolysis appears to be independent of changes in OT receptor synthesis, but depends on the duration of the high plasma progesterone concentration characteristic of the luteal phase (Kotwica 1988, Geisert et al 1992, Silvia and Raw 1993). On the basis of the data presented, OT may play a supportive and modulatory role in luteolysis. In experiment 1, when OT receptors were blocked, the magnitude of PGF release
decreased (Table 1). In ovariectomized ewes treated with progesterone for 15 days, PGF secretion was reduced without a concomitant release of OT (Silvia and Raw 1993). These authors suggest that ovarian OT is not required to initiate pulsatile secretion of PGF, but may be required to achieve full pulse amplitude. When luteolysis was in progress, NA released OT and later increased PGF secretion (Fig 2b). This may be a direct influence of NA on the uterus. NA was found to affect PGF release from the human uterus before and during ovulation (Quass et al 1987, Tanaka et al 1993) and from the human decidua (Tada et al 1991). In rats, catecholamines increase PGF secretion via β-adrenergic receptors (Ishikawa and Fuchs 1978). Moreover, blocking both α- and β-adrenergic receptors in the human uterus decreases catecholamine-induced PGF release (Quass and Zahradnik 1985). However, indirect actions of NA on PGF secretion by OT release are also possible. We postulated that OT is not the main factor in the initiation of luteolysis in cattle and this supports data from Blair et al (1997). However, it may play a supportive and modulatory role in induced luteolysis as a regulator of the amplitude of pulsatile PGF secretion. PGF analogue stimulates the secretion of endogenous PGF from the uterus in cattle and this may be the essential component of the luteolytic action of exogenous PGF, as suggested in ewes (Wade and Lewis 1996).
ACKNOWLEDGEMENTS We thank Dr S. Okrasa and Dr G. Kotwica (University of Agriculture and Technology, Olsztyn, Poland) and Dr W.J. Silvia (University of Kentucky, Lexington, U.S.A.) for progesterone, oxytocin and PGFM antiserum. CAP was kindly donated by Ferring AB, Sweden. This study was supported by The Polish Academy of Sciences.
REFERENCES BLAIR, R.M., SAATMAN, R., LIOU, S.S., FORTUNE, J.E. & HANSEL, W. (1997) Roles of leukotrienes in bovine corpus luteum regression: an in vivo microdialysis study. Proceedings of the Society for Experimental Biology and Medicine 216, 72–80 FLINT, A.P.F. & SHELDRICK, E.L. (1983) Secretion of oxytocin by the corpus luteum in sheep. Progress in Brain Research 60, 521–530 GEISERT, R.D., SHORT, E.C. & ZAVY, M.T. (1992) Maternal recognition of pregnancy. Animal Reproduction Science 28, 287–298 HOMANICS, G.E. & SILVIA, W.J. (1988) Effects of progesterone and estradiol on uterine secretion of prostaglandin F2α in response to oxytocin in ovariectomized ewes. Biology of Reproduction 38, 804–811 HOWARD, H.J. & BRITT, J.H. (1987) Prostaglandin F metabolite after exogenous oxytocin in cows given hCG during diestrus. Biology of Reproduction 36(Suppl. 1), 274 ISHIKAWA, M. & FUCHS, A.R. (1978) Effects of epinephrine and oxytocin on the release of prostaglandin F from the rat uterus in vitro. Prostaglandins 15, 89–101 JAROSZEWSKI, J. & KOTWICA, J. (1994) Reduction of oxytocin content in corpus luteum does not affect the duration of the estrous cycle in cattle. Reproductive Nutrition and Development 34, 175–182 KOTWICA J. (1988) Ovarian oxytocin in bovine oestrous cycle regulation. Acta Academiae Agriculture et Technicae Olstenensis Zootechnica 31 (Suppl. C), 1–45 (in Polish) KOTWICA, J., SKARZYNSKI, D. & JAROSZEWSKI, J. (1990) Coccygeal artery as a route for administration of drugs into the reproductive tract of cattle. The Veterinary Record 127, 38–40 KOTWICA, J. & SKARZYNSKI, D. (1993a) Influence of oxytocin removal from the corpus luteum on secretory function and duration of the oestrous cycle in cattle. Journal of Reproduction and Fertility 97, 411–417 KOTWICA, J., SKARZYNSKI, D. & JAROSZEWSKI, J. (1993b) Secretion of PGF stimulated by different doses of oxytocin and released spontaneously during luteolysis in cattle. Journal of Reproduction and Fertility Abstract Series 12, 60
Effect of oxytocin on PGF2α secretion in cattle KOTWICA, J., SKARZYNSKI, D., JAROSZEWSKI, J. & BOGACKI, M. (1994) Noradrenaline affects secretory function of corpus luteum independently on prostaglandins in conscious cattle. Prostaglandins 48, 1–10 KOTWICA, J., SKARZYNSKI, D., BOGACKI, M., MELIN, P. & STAROSTKA B. (1997) The use of an oxytocin antagonist to study the function of ovarian oxytocin during luteolysis in cattle. Theriogenology 48, 1287–1299 LAFRANCE, M. & GOFF, A.K. (1993) Effects of progesterone and estradiol-17β on oxytocin-induced release of prostaglandin in heifers. Biology of Reproduction 34 (Suppl. 1), 161 LYE, S. J., SPRAGUE, C. L. & CHALLIS, H. H. D. (1983) Modulation of ovine myometrial activity by estradiol-17β. The possible involvement of prostaglandins. Canadian Journal of Physiology and Pharmacology 61, 729–735 MCCRACKEN, J. A., SCHRAMM, W. & OKULICZ, W. C. (1984) Hormone receptor control of pulsatile secretion of PGF2α from the uterus luteolysis and abrogation in early pregnancy. Animal Reproduction Science 7, 31–55 MEYER, H.H.D., MITTERMEIER, T. & SCHAMS, D. (1988) Dynamics of oxytocin, estrogen and progestin receptors in the bovine endometrium during the estrous cycle. Acta Endocrinologica 118, 96–104 MIRANDO, M. A., BECKER, W. C. & WHITEAKER, S.S. (1993) Relationship among endometrial oxytocin receptors, oxytocin-stimulated phosphoinositide hydrolysis and prostaglandin F2α secretion in vitro, and plasma concentrations of ovarian steroids before and during corpus luteum regression in cyclic heifers. Biology of Reproduction 48, 874–882 MOORE, L. G., CHOY, V. J. & ELLIOT, R. L. WATKINS, W. B. (1986) Evidence for the release of prostaglandin F2α inducing the release of ovarian oxytocin during luteolysis in the ewe. Journal Reproduction and Fertility 75, 159–166 PARKINSON, T.J., JENNER, L.J. & LAMMING, G.E. (1990) Comparison of oxytocin/prostaglandin F-2α interrelationship in cyclic and pregnant cows. Journal of Reproduction and Fertility 90, 337–345 PARKINSON, T. J., WATHES, D. C., JENNER, L. J. & LAMMING, G. E. (1992) Plasma and luteal concentrations of oxytocin in cyclic and early-pregnant cattle. Journal of Reproduction and Fertility 94, 161–167 QUAAS, L. & ZAHRADNIK, H. P. (1985) The effects of α- and β-adrenergic stimulation on contractility and prostaglandin (prostaglandins E2 and F2α and 6-ketoprostaglandin 1α) strips. American Journal of Obstetrics and Gynecology 157, 856–865 QUASS, L., GOPPINGER, A. & ZAHRADNIK, H. P. (1987) The effect of acetylosalicylic acid and indomethacin on catecholamine- and oxytocin-induced contractility and prostaglandin (6-keto-PGF1α, PGF2α) – production of human pregnant myometrial strips. Prostaglandins 34, 257–269
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SCHAMS, D., SCHMIDT-POLEX, B. & KRUSE, V. (1979) Oxytocin determination by radioimmunoassay in cattle. Acta Endocrinologica 92, 258–270 SCHAMS, D., SCHALLENBERGER, E., MEYER, H. H. D., BULLERMANN, B., BREITINGER, H.J., KOLL, R., ENZENHOFER, G., KRUIP, T.A.M., WALTERS, D.L. & KARG, H. (1985) Ovarian oxytocin during estrous cycle in cattle. In Oxytocin, Clinical and Laboratory Studies. Eds. J. A. Amico and A.G. Robinson. Amsterdam, Elsevier Biomedical, pp 317–334 SILVIA, W.J. & HOMANICS, G.E. (1988) Role of phospholipase C in mediating oxytocin-induced release of prostaglandin F2α from ovine endometrial tissue. Prostaglandins 35, 535–548 SILVIA, W. J. & TAYLOR, M. L. (1989) Relationship between uterine secretion of prostaglandin F2α induced by oxytocin and endogenous concentration of estradiol and progesterone at three stages of the bovine oestrous cycle. Journal of Animal Science 67, 2347–2353 SILVIA, W.J. & RAW, R.E. (1993) Regulation of pulsatile secretion of prostaglandin F2α from the ovine uterus by ovarian steroids. Journal of Reproduction and Fertility 98, 341–347 SKARZYNSKI, D., BOGACKI, M. & KOTWICA, J. (1997) Changes in ovarian oxytocin secretion as an indicator of corpus luteum response to prostaglandin F2α treatment in cattle. Theriogenology 48, 733–742 SOLOFF, M.S. & FIELDS, M.J. (1989) Changes in uterine oxytocin receptor concentrations throughout the estrous cycle of the cow. Biology of Reproduction 40, 283–287 TANAKA, N., MIYAZAKI, K., TASHIRO, H., MIZUTANI, H. & OKAMURA, H. (1993) Changes in adenylyl cyclase activity in human endometrium during the menstrual cycle and in human decidua during pregnancy. Journal of Reproduction and Fertility 98, 33–39 TADA, K., KUDO, T. & KISHIMOTO, Y. (1991) Effect of L-dopa or dopamine on human decidual prostaglandin synthesis. Acta Medicine Okayama 45, 333–336 WADE, D.E. & LEWIS, G. S. (1996) Exogenous prostaglandin F2α stimulates uteroovarian release of prostaglandin F2α in sheep: possible component of luteolytic mechanism of action of exogenous prostaglandin F2α. Domestic Animal Endocrinology 13, 383–398 WEEMS, Y. S., VINCENT, D. L., NUSSER, K. D., TANAKA, Y., MILLERPATRICK, K., LEDGERWOOD, K. S. & WEEMS, C. W. (1993) Effect of prostaglandin F2α on uterine or ovarian secretion of prostaglandins E and F2α in vivo in 90–100 day hysterectomised, intact or ovariectomized pregnant ewes. Prostaglandins 46, 277–296 Accepted July 29, 1998