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The effect of treatment with a GnRH analogue on postpartum reproductive performance in Friesian cows
Animal Reproduction Science, 12 (1986) 99--107 Elsevier Science Publishers B.V., Amsterdam - - Printed in The Netherlands
THE EFFECT OF TREATMENT WITH A GnRH ANALOGUE ON POSTPARTUM REPRODUCTIVE PERFORMANCE IN FRIESIAN
99
COWS
M.B. ABOUL-ELA 1 and F.E. EL-KERABY 2
1Faculty of Agriculture, University of Mansoura, El Mansoura (Egypt) 2Animal Production Research Institute, Ministry of Agriculture, Cairo (Egypt) (Accepted 18 March 1986)
ABSTRACT Aboul-Ela, M.B. and E1-Keraby, F.E., 1986. The effect o f treatment with a GnRH analogue on p o s t p a r t u m reproductive performance in Friesian cows. Anita. Reprod. Sci., 12; 99--107. Three trials were conducted to examine the effect of GnRH injection on the reproductive performance of Friesian cows. In the first trial 100 ~g gonadotrophin releasing hormone synthetic analogue (GnRH) was given at the time o f A.I. to 32 cows while a control group received a saline placebo injection. Conception rate to first insemination was significantly higher in treated than control cows (81.3 v. 54.8%, P < 0.05). In the second trial, two groups of 19 cows each received either 100 #g GnRH or saline at 15 days postpartum. A total of 60 cows was used in the third trial in which GnRH was given at either 50 or 100 ug dose level on either days 7 or 15 postpartum while control cows were untreated. In b o t h trials GnRH treatment reduced the intervals from calving to complete uterine involution, first ovulation and first detected oestrus. The effect was most pronounced when 100 ug GnRH was given on day 7 postpartum. Number of days open and number of services per conception were appreciably reduced when 100 #g GnRH was given at either 7 or 15 days postpartum with maximal effect when given 15 days after calving.
INTRODUCTION I t is g e n e r a l l y b e l i e v e d t h a t a c a l v i n g i n t e r v a l o f 1 2 m o n t h s is a n i d e a l goal for economic dairy cattle production. Longer calving intervals have been reported for Friesian cows in Egypt (E1-Keraby and Aboul-Ela, 1982). C a l v i n g t o c o n c e p t i o n i n t e r v a l is d e p e n d e n t o n t h e r e - e s t a b l i s h m e n t o f n o r mal ovarian cycles, the occurrence of oestrous behaviour and its accurate detection, and the pregnancy rate following service (Peters, 1984). There have been many attempts to enhance the resumption of ovarian activity and oestrus early in the postpartum period by a single injection of 100--500 g g G n R H g i v e n a t d i f f e r e n t t i m e s a f t e r c a l v i n g (e.g. S c h a m s e t al., 1 9 7 3 ; B r i t t e t al., 1 9 7 4 ; B o i t i e t al., 1 9 8 2 ; B a l l a n d L a m m i n g , 1 9 8 3 ) a n d t h e s e have given variable results. Administration of GnRH at the time of A.I. has
0378-4320/86[$03.50
© 1986 Elsevier Science Publishers BoV.
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been reported to improve conception rate (Nakao et al., 1983). The present study aims at investigating (a) the effect of GnRH treatment in the early postpartum period on the reproductive performance of Friesian dairy cows; (b) the effect of GnRH injection at the time of insemination on conception rate. M A T E R I A L S AND METHODS
In this study, three trials were conducted at Sakha Experimental S t a t i o n , Ministry of Agriculture, located almost in the middle of the Nile Delta. Pure Friesian cows were used and were subjected to the general management system applied on the farm. They were housed in half-shedded open yards and were fed hay, rice straw, concentrate feed mixture and green fodder, whenever available, according to their requirements for maintenance and milk production. Average daily milk yield during the first 90 days postpartum was 15.4 + 1.36 kg. After parturition, all cows were left to suckle their calves for 3 days. Thereafter, t h e y were machine milked twice daily. Cows were checked for standing oestrus twice daily at 08.00 and 15.00 h. Starting at 40 days postpartum, cows were artificially inseminated with frozen semen when seen on heat. Pregnancy diagnosis was carried out by rectal palpation at about 60 days after the last insemination. The GnRH synthetic analogue used in this study (Gonadorelin, Hoechst U.K. Ltd.) was given intramuscularly (i.m.). Trial 1. This trial involved 63 cows (second--fifth lactations) which were subjected to oestrous d e t e c t i o n and put for insemination during October and November 1983. They were r a n d o m l y chosen to receive i.m. injection of either 100 ug GnRH analogue (n = 32) or 2.5 ml saline (n = 31) as a placebo (control) at the time of insemination. Observation for standing oestrus continued for 30 days after A.I. Conception rate from first insemination was calculated from the results of pregnancy diagnosis. Differences between the two groups were statistically tested using the Fisher Exact test according to Zar (1974). Trial 2. A total of 38 cows (second--fourth lactations) which calved in July--August 1983 was used. Cows of the same parity were allocated at r a n d o m to either of two groups, 19 cows in each. Cows of group 1 received i.m. injection of 100 pg GnRH analogue each on day 15 postpartum while each cow in group 2 (control) received 2.5 ml saline as a placebo. The ovaries and uterus were examined per rectum twice weekly starting 7 days after calving to estimate dates of first ovulation and complete uterine involution. The latter was diagnosed using the procedure outlined by Buch et al. (1955). Days from parturition to first ovulation (POV), first oestrus (POE), com-
I01
plete uterine involution (UI) and conception (days open, DO) were recorded a l o n g w i t h t h e n u m b e r o f services p e r c o n c e p t i o n (S/C). D a t a w e r e statistically a n a l y s e d using g r o u p e d t-test (Steel a n d T o r r i e , 1 9 6 0 ) .
Trial 3. S i x t y c o w s ( s e c o n d - - f i f t h l a c t a t i o n s ) w h i c h calved d u r i n g N o v e m b e r 1 9 8 3 - - J a n u a r y 1 9 8 4 w e r e used. C o w s o f t h e s a m e p a r i t y w e r e r a n d o m l y divided i n t o five g r o u p s , 12 in each: g r o u p 1 was u n t r e a t e d as c o n t r o l ; g r o u p s 2 a n d 3 r e c e i v e d e i t h e r 50 o r 1 0 0 p g G n R H a n a l o g u e p e r c o w , respectively, o n d a y 7 p o s t p a r t u m ; while g r o u p s 4 a n d 5 r e c e i v e d e i t h e r 50 or 100 p g G n R H a n a l o g u e p e r c o w , r e s p e c t i v e l y , o n d a y 15 p o s t p a r t u m . R e c o r d s o f P O V , P O E , UI, D O a n d S/C w e r e r e c o r d e d as in trial 2. D a t a o b t a i n e d w e r e s u b j e c t e d t o s t a n d a r d analysis o f v a r i a n c e (Steel a n d T o r r i e , 1960). RESULTS
Trial 1. T r e a t m e n t w i t h G n R H at t h e t i m e o f A.I. r e s u l t e d in a significant ( P < 0.05) increase in c o n c e p t i o n r a t e f r o m first i n s e m i n a t i o n o v e r t h a t o f t h e c o n t r o l g r o u p ( 8 1 . 3 v. 54.8%). Trial 2. U t e r i n e i n v o l u t i o n was c o m p l e t e d earlier in t h e t r e a t e d t h a n t h e c o n t r o l c o w s ( T a b l e 1). T h e d i f f e r e n c e h o w e v e r ( 2 5 . 4 v. 2 8 . 2 d a y s ) , was n o t significant. First o v u l a t i o n ( P O V ) o c c u r r e d a b o u t 5 d a y s earlier in t h e G n R H t r e a t e d c o w s t h a n t h e c o n t r o l s , t h e d i f f e r e n c e b e i n g highly significant ( P < 0.01). A significant ( P < 0.01) d i f f e r e n c e o f 9.0 d a y s in P O E was f o u n d b e t w e e n t h e t w o groups. This d i f f e r e n c e was c o n s i s t e n t w i t h t h a t o f P O V . G n R H - t r e a t e d c o w s c o n c e i v e d , o n average, a b o u t 2 1 . 4 d a y s earlier t h a n t h e c o n t r o l c o w s ; a d i f f e r e n c e w h i c h was statistically significant ( P < 0.05). TABLE 1 Means and standard errors (~-+ SE) of postpartum reproductive traits as affected by GnRH treatment (100 ug on day 15 postpartum) Trait
Control
Treated
Number of cows Days to :
19
19
Complete uterine involution First ovulation First oestrus Conception Number of services per conception
28.2 32.5 36.7 86.4
-+ 2.71 +- 1.57 + 2.33 • 8.74
2.2 -+ 0.37
25.4 27.4 27.7 65.0
+ 1.72 + 1.76 + 1.82 + 4.92
1.2 + 0.15
Significance of differences
NS ** ** * *
NS = Not significant, * = Significant (P < 0.05), ** = Significant (P < 0.01).
2.0a + 0.17
1.67 ab + 0:19
+- 2 . 1 9 ab ± 2.87 ab ± 2.97 a + 5.99
12
12
29.3 24.7 33.8 70.8
50 #g on day 7
Untreated
31.5 + 2.96 35.1 + 2.18 48.3 ± 4.68 80-8a ± 5.55
2
1
reproductive
± 1.73 a ± 1.05 a -+ 2 . 0 5 b + 2.94
1.33 b ± 0.14
23.5 19.3 23.9 54.2
12
100 ~g on day 7
3
Groups
+ 1.65 b + 2.16 b ± 5.88 a ± 4.39 1.5fl ab ± 0.19
26.5 25.9 39.6 73.4
12
50 ~g on day 15
4
+ 2.09 ab + 2.41 ab + 2.64 b + 2.96 1.25 b ± 0.13
25.5 24.3 29.0 56.1
12
100 ~g on day 15
5
traits as affected by dose level and time of GnRH treatment
N 8 = N o t s i g n i f i c a n t , * S i g n i f i c a n t ( P < 0 . 0 5 ) , * * = S i g n i f i c a n t ( P < 0 , 0 1 ). 1 M e a n s h a v i n g t h e s a m e s u p e r s c r i p t a r e n o t s i g n i f i c a n t l y (P < 0 . 0 5 ) d i f f e r e n t ,
Number of services per conception
Complete uterine involution First ovulation First oestrus Conception
Number of cows Days to :
Trait
M e a n s 1 a n d s t a n d a r d el-~ors (~ ± S E ) o f p o s t p a r t u m
TABLE 2
*
NS ** ** **
Significance of variation among groups
O
103
The within-group individual variation was greater in DO compared with those of other intervals studied, with the GnRH-treated cows showing smaller individual variation than the control cows ( C V - - 4 5 . 2 v. 33.8%). In the control group, this was partially due to animals (n = 5) that had to be inseminated three times before t h e y conceived, b u t it was mainly due to three animals that did n o t show oestrus for periods of 45 days or more after t h e y manifested their first oestrus. Rectal examination of these animals revealed that they did n o t have a palpable corpus luteum. On the other hand, only one of the treated cows had a period of anoestrus, with no palpable corpus luteum detected, for a b o u t 40 days. The GnRH-treated cows required, on average, 1.2 inseminations per conception, whereas the control animals required, on average, 2.2 inseminations to conceive (Table 1), The difference between the two groups was statistically significant ( P < 0.05). Trial 3. Longer UI was observed in untreated cows than in all treated groups (Table 2). The overall differences among groups were, however, n o t statistically significant. Among the treated groups, complete uterine involution occurred earliest (23.5 days) in cows receiving 100 pg G n R H at 7 days postpartum. Significant ( P < 0.01) reduction in POV occurred due to G n R H injection, as c o m p a r e d with untreated cows. First postpartum ovulation occurred markedly earlier (at 19.3 days) in group 3 (Receiving 100/~g G n R H on day 7 postpartum) than in the other treated groups, in which the averages of POV were similar (Table 2). Similar to the first ovulation, first detected oestrus was recorded significantly ( P < 0.01) earlier in all treated groups compared with the untreated control. Slightly longer POE was observed in cows treated on day 15 p o s t p a r t u m (groups 4 and 5) than in the corresponding groups receiving the same dose of G n R H on day 7 postpartum (groups 2 and 3, respectively). It is worth noting that incidence of the first ovulation not being associated with signs of oestrus (silent heat) was 33.3% in cows receiving 100 pg G n R H while it was 58.3% in both untreated cows and those receiving 50 #g GnRH. Overall differences in the number of days open (DO) were statistically significant ( P < 0.01). The latest animals to conceive were those of the control group (80.8 days) b u t t h e y did n o t significantly differ in DO from the two groups receiving 50 pg G n R H (groups 2 and 4), the latter having closely similar values (70.8 and 73.4 days). On the other hand, conception occurred significantly earlier in the cows receiving 100 pg G n R H (groups 3 and 5) than in other groups. Time of G n R H treatment after parturition, at high or low dose, did not have a significant effect on DO. The control animals required the greatest n u m b e r of services per conception (2.0). G n R H treatment at a dose of 50 pg either on day 7 or 15 p o s t p a r t u m insignificantly reduced S/C. An appreciable and significant ( P < 0 . 0 5 ) reduction in S/C occurred in
104 c6ws treated with 100 #g GnRH, as compared to the untreated group. The lowest S/C was t h a t of cows receiving 100 #g on day 15 postpartum. The effect of time of GnRH injection after calving on S/C was very small and insignificant.
Relation between various reproductive traits The data obtained in trial 3 were used to calculate the within-groups correlation coefficients between different traits studied. Out of these, the highest correlation was obtained between POV and POE (r= 0.697) which is, obviously, n o t unexpected. UI was positively correlated with each of POV and S/C ( r = 0 . 3 3 0 and 0.355, respectively), but its strongest correlation (r= 0.464) was t h a t with DO. On the other hand, poor and insignificant correlation was f o u n d between UI and POE (r = 0.166). The correlation between DO and POE (r = 0.544) was stronger than that between DO and POV (r = 0.41). Furthermore, both POV and POE were insignificantly correlated with S/C (r= 0.219 and 0.221, respectively). DISCUSSION The improved conception rate resulting from GnRH injection at the time of A.I. in the first trial supports the findings of Mori and Takahashi (1978), Schels and Mostafawi (1978) and Nakao et al. (1983). GnRH injection causes a luteinizing h o r m o n e (LH) surge (Zolman et al., 1973), and enhances ovulation (Leidl et al., 1979) and thus eliminates the occurrence of delayed ovulation, relative to the time of A.I. It has been reported t h a t in 10% or more of cows,: ovulation might be delayed in relation to oestrus, resulting in a mistiming of A.I. and low conception rate (Bostedt, 1976; Karg et al., 1980). The effect of postpartum t r e a t m e n t with GnRH on uterine involution was consistent in both trials 2 and 3. The effect was more pronounced when a 100 #g dose of GnRH was given 7 days postpartum. GnRH enhances uterine involution in cows with retained placenta (Bostedt and Maurer, 1982) and also in cows after normal parturition without retained placenta (Britt et al., 1974). The role of G n R H in uterine involution, however, remains unelucidated. It has been reported that GnRH injection in the early postpartum period enhances the resumption of ovarian function and the occurrence of first ovulation (Schams et al., 1973; Britt et al. 1974). The results of b o t h trials 2 and 3 support this view. Although GnRH given at either high or low dose and either at 7 or 15 days postpartum significantly enhanced the occurrence of first ovulation, the effect was most pronounced when a dose of 100 #g was given on day 7 postpartum. The responsiveness of the anterior pituitary to GnRH stimulation has been reported to be regained by day 7 or 8 postpartum in lactating dairy cows (Kesler et al.,
105 1977). The results of b o t h trials 2 and 3 (Tables 1 and 2) indicate that there was a time lapse which ranged from 9.3 to 17.7 days on average, between GnRH treatment and the occurrence of first ovulation. Peters and Lamming (1984) suggested that the induced LH release resulting from G n R H injection might result in premature luteinization of an unovulated follicle and transient secretion o f progesterone sufficient to initiate ovarian cycles. The effect of G n R H treatment on the time of first postpartum oestrus was, as expected, in line with its effect on first ovulation. This was true for b o t h trials 2 and 3. Of interest is the effect of dose level on the incidence of ovulatory anoestrus (silent heat) at the first ovulation. This is difficult to explain in the light of the available results in this study. It m a y be related, however, to a higher degree of follicular growth and hence a possibly higher pre-ovulatory oestradiol level in animals treated with a high dose (100 pg) of G n R H as compared with either the untreated animals or those receiving a low G n R H dose (50 pg). The effect o f G n R H at 100 pg/cow on reducing the number of days open by 20 days or more is consistent in trials 2 and 3. It is also similar to the reported reduction of about 24 days in the interval from calving to conception (Boiti et al., 1982) or from post-calving treatment to conception ( H u m b o l t and Thibier, 1981) resulting from G n R H injection as compared with control groups. In some other studies, however, the effect of G n R H treatment on reducing the interval from calving to conception was either small and n o t significant (Bostedt and Maurer, 1982; Ball'and Lamming, 1983) or pronounced only in cows with retained placenta (Bostedt and Maurer, 1982). In the present study, it seems that G n R H injection n o t only stimulated the pituitary function and, as a result, the resumption of :ovarian activity, b u t also resulted in regulation of the reproductive process in general. This was clear from the effect of G n R H treatment in reducing the number of services per conception. This is similar to the positive effect of G n R H reported b y Schams e t al. (1973). Other workers, however, f o u n d such effect to be small and n o t significant ( H u m b o l t and Thibier, 1981) or to be limited to cows with retained placenta (Bostedt and Maurer, 1982). The discrepancy in the results obtained in different studies may be due to the timing of G n R H treatment in relation to calving and insemination or the time when first insemination t o o k place after calving. The significant positive correlation between UI and each of POV, DO and S/C indicates the importance of rapid uterine involution in the resumption of normal functioning Of t h e reproductive system leading to conception. These relationships were discussed by Eley et al. (1981) and Peters (1984). The finding that DO showed the highest correlation with POE supports the well established view of the importance of applying an accurate and effective heat detection system for reducing calving interval (Esslemont, 1974; Buiman and Lamming, 1978; E1-Keraby and Aboul-Ela, 1982).
106 CONCLUSION
The results of this study illustrate the beneficial use of G n R H for increasing fertility in dairy cattle. Higher conception rate may be achieved through G n R H injection at the time of A.I. Postpartum treatment with a low dose of G n R H (50 pg) was less effective than a high dose (100:gg). Injection of 100 pg G n R H at either 7 or 15 days postpartum reduced the intervals from calving to complete uterine involution, first ovulation, first detected oestrus and conception and also the number of services per conception. Injection of 100 ~g G n R H on day 15 postpartum was most effective in reducing appreciably both the number of services per conception and days open which are of great importance from the economic point of view to the dairy farmer. ACKNOWLEDGEMENT
The authors wish to thank Hoechst U.K. Ltd. for the supply of 'Gonadorelin' used in this study.
REFERENCES Ball, P.J.H. and Lamming, G.E., 1983. Diagnosis of ovarian acyclicity in lactating dairy cows and evaluation of treatment with gonadotrophin-releasing hormone or a progesterone-releasing intravaginal device. Br. Vet. J., 139 : 522--527. Boiti, C., Bighelli, V., Simontacehi, C., Olivieri, O., Caggioni, C. and Micale, R., 1982. Postpartum progesterone profiles and reproductive performance of dairy cows treated with G n R H or progesterone. In: H. Karg and E. SchaUenberger (Editors), Factors Influencing Fertility in the Postpartum Cow. Martinus Nijhoff, The Hague, pp. 544-547. Bostedt, H., 1976. Delayed ovulation as a cause of sterility in the A.I. of cattle. Proceedings of the VIII International Congress on Animal Reproduction and Artificial Insemination, Krakow, pp. 552--555. Bostedt, H. and Maurer, G., 1982. The reproduction performance of cows after an injection of GnRH-analogue in the early puerperium -- a field study. In H. Karg and E. SchaUenberger (Editors), Factors Influencing Fertility in the Postpartum Cow. Martinus Nijhoff, The Hague, pp. 562--565. Britt, J.H., Kittok, R.J. and Harrison, D.S., 1974. Ovulation, oestrus and endocrine response after GnRH in early postpartum cows. J. Anita. Sci., 39: 915--919. Buch, N.C., Tyler, W.J. and Casida, L.E., 1955. Postpartum oestrus and involution of the uterus in an experimental herd of Holstein Friesian cows. J. Dairy Sci., 3 8 : 7 3 - - 7 9 . Bulman, D.C. and Lamming, G.E., 1978. Milk progesterone levels in relation to conception,repeat breeding and factors influencing acyclicity in dairy cows. J. Reprod. Fertil., 54: 447--458. Eley, D.S., Thatcher, W.W., Head, H.H., Collier, R.J., Wilcox, C.J. and Call, E.P., 1981. Periparturient and postpartum changes of conceptus and maternal units in Jersey cows bred for milk yield. J. Dairy Sci., 64: 312--320. E1-Keraby, F.E. and Aboul-Ela, M.B., 1982. A study of some non-genetical factors affecting the postpartum reproductive performance in Friesian cows. Trop. Anim. Prod., 7 : 307--314.
107 Esslemont, R., 1974. Economic and husbandry aspects of the manifestation and detection of oestrus in cows. Part I. Economic aspects. ADAS Q. Rev., 12: 175--184. Humbolt, P. and Thibier, M., 1981. Effect of gonadotropin releasing h o r m o n e (GnRH) treatment during the midluteal phase in repeat breeder cows. A preliminary report. Theriogenology, 16: 375--376. Karg, H., Claus, R., Gunzler, O., Rattenberger, E. and Firchner, F., 1980. Hormonal recognition o f cyclicity and infertility in cows under practical conditions. Proceedings of the l l t h World Congress of Diseases of Cattle. Israel, pp. 885--894. Kesler, D.J., Garverick, H.A., Youngquist, R.S., Elmore, R.G. and Bierschwal, C.J., 1977. Effect of days postpartum and endogenous, reproductive hormones on GnRH induced in LH release in dairy cows. J. Anita. Sci., 45: 797--803. Leidl, W . , Bostedt, H., Lamprecht, W., Prinzen, R. and Wendt, V., 1979. The influence of GnRI-I and HCG on the time o f ovulation in cows artificially inseminated. Tier~irztl. Umsch., 34: 546--555. Mori, J. and Takahashi, T., 1978. Effect of an analog of LH-RH on pregnancy rate in cows. Jpn. J. Anita. Reprod., 24: 137--138. Nakao, T., Narita, S., Tanaka, K., Haxa, H., Shirkawa, J., Noshiro, H., Saga, N., Tsunoda, N. and Kawata, K., 1983. Improvement of first-service pregnancy rate in cows with gonadotropin-releasing hormone analog. Theriogenology, 20: 111--119. Peters, A.R., 1984. Reproductive activity o f the cow in the postpartum period. I. Factors affecting the length of the postpartum acyclic period. Br. Vet. J., 140 : 76--84. Peters, A.R. and Lamming, G.E., 1984. Reproductive activity o f the cow in the postpartum period. H. Endocrine patterns and induction o f ovulation. Br. Vet. J., 140: 269--280. Sehams, D., Hofer, F., Hofmann, B., Ender, M.L. and Karg, H., 1973. Effects o f synthetic LH-RH treatment on bovine ovarian function during oestrous cycle and postpartum period. Acta Endocrinol. Suppl., 177: 296. Schels, H.F. and Mostafawi, D., 1978. The effect of GnRH on the pregnancy rate o f artificially inseminated cows. Vet. Rec., 103: 31--32. Steel, R.G.D. and Torrie, J.H., 1960. Principles and Procedures o f Statistics. McGraw Hill, New York, NY, 481 pp. Zar, J.H., 1974. Biostatistical Analysis. Prentice-Hall, Englewood Cliffs, NJ, 620 pp. Zolman, J., Convey, E.M., Britt, J.G. and Hafs, J.H., 1973. Release of the bovine luteinizing hormone b y purified synthetic gonadotropin releasing hormone. Proc. Soc. Exp. Biol. Med., 142: 189--193.
THE EFFECT OF TREATMENT WITH A GnRH ANALOGUE ON POSTPARTUM REPRODUCTIVE PERFORMANCE IN FRIESIAN
99
COWS
M.B. ABOUL-ELA 1 and F.E. EL-KERABY 2
1Faculty of Agriculture, University of Mansoura, El Mansoura (Egypt) 2Animal Production Research Institute, Ministry of Agriculture, Cairo (Egypt) (Accepted 18 March 1986)
ABSTRACT Aboul-Ela, M.B. and E1-Keraby, F.E., 1986. The effect o f treatment with a GnRH analogue on p o s t p a r t u m reproductive performance in Friesian cows. Anita. Reprod. Sci., 12; 99--107. Three trials were conducted to examine the effect of GnRH injection on the reproductive performance of Friesian cows. In the first trial 100 ~g gonadotrophin releasing hormone synthetic analogue (GnRH) was given at the time o f A.I. to 32 cows while a control group received a saline placebo injection. Conception rate to first insemination was significantly higher in treated than control cows (81.3 v. 54.8%, P < 0.05). In the second trial, two groups of 19 cows each received either 100 #g GnRH or saline at 15 days postpartum. A total of 60 cows was used in the third trial in which GnRH was given at either 50 or 100 ug dose level on either days 7 or 15 postpartum while control cows were untreated. In b o t h trials GnRH treatment reduced the intervals from calving to complete uterine involution, first ovulation and first detected oestrus. The effect was most pronounced when 100 ug GnRH was given on day 7 postpartum. Number of days open and number of services per conception were appreciably reduced when 100 #g GnRH was given at either 7 or 15 days postpartum with maximal effect when given 15 days after calving.
INTRODUCTION I t is g e n e r a l l y b e l i e v e d t h a t a c a l v i n g i n t e r v a l o f 1 2 m o n t h s is a n i d e a l goal for economic dairy cattle production. Longer calving intervals have been reported for Friesian cows in Egypt (E1-Keraby and Aboul-Ela, 1982). C a l v i n g t o c o n c e p t i o n i n t e r v a l is d e p e n d e n t o n t h e r e - e s t a b l i s h m e n t o f n o r mal ovarian cycles, the occurrence of oestrous behaviour and its accurate detection, and the pregnancy rate following service (Peters, 1984). There have been many attempts to enhance the resumption of ovarian activity and oestrus early in the postpartum period by a single injection of 100--500 g g G n R H g i v e n a t d i f f e r e n t t i m e s a f t e r c a l v i n g (e.g. S c h a m s e t al., 1 9 7 3 ; B r i t t e t al., 1 9 7 4 ; B o i t i e t al., 1 9 8 2 ; B a l l a n d L a m m i n g , 1 9 8 3 ) a n d t h e s e have given variable results. Administration of GnRH at the time of A.I. has
0378-4320/86[$03.50
© 1986 Elsevier Science Publishers BoV.
I00
been reported to improve conception rate (Nakao et al., 1983). The present study aims at investigating (a) the effect of GnRH treatment in the early postpartum period on the reproductive performance of Friesian dairy cows; (b) the effect of GnRH injection at the time of insemination on conception rate. M A T E R I A L S AND METHODS
In this study, three trials were conducted at Sakha Experimental S t a t i o n , Ministry of Agriculture, located almost in the middle of the Nile Delta. Pure Friesian cows were used and were subjected to the general management system applied on the farm. They were housed in half-shedded open yards and were fed hay, rice straw, concentrate feed mixture and green fodder, whenever available, according to their requirements for maintenance and milk production. Average daily milk yield during the first 90 days postpartum was 15.4 + 1.36 kg. After parturition, all cows were left to suckle their calves for 3 days. Thereafter, t h e y were machine milked twice daily. Cows were checked for standing oestrus twice daily at 08.00 and 15.00 h. Starting at 40 days postpartum, cows were artificially inseminated with frozen semen when seen on heat. Pregnancy diagnosis was carried out by rectal palpation at about 60 days after the last insemination. The GnRH synthetic analogue used in this study (Gonadorelin, Hoechst U.K. Ltd.) was given intramuscularly (i.m.). Trial 1. This trial involved 63 cows (second--fifth lactations) which were subjected to oestrous d e t e c t i o n and put for insemination during October and November 1983. They were r a n d o m l y chosen to receive i.m. injection of either 100 ug GnRH analogue (n = 32) or 2.5 ml saline (n = 31) as a placebo (control) at the time of insemination. Observation for standing oestrus continued for 30 days after A.I. Conception rate from first insemination was calculated from the results of pregnancy diagnosis. Differences between the two groups were statistically tested using the Fisher Exact test according to Zar (1974). Trial 2. A total of 38 cows (second--fourth lactations) which calved in July--August 1983 was used. Cows of the same parity were allocated at r a n d o m to either of two groups, 19 cows in each. Cows of group 1 received i.m. injection of 100 pg GnRH analogue each on day 15 postpartum while each cow in group 2 (control) received 2.5 ml saline as a placebo. The ovaries and uterus were examined per rectum twice weekly starting 7 days after calving to estimate dates of first ovulation and complete uterine involution. The latter was diagnosed using the procedure outlined by Buch et al. (1955). Days from parturition to first ovulation (POV), first oestrus (POE), com-
I01
plete uterine involution (UI) and conception (days open, DO) were recorded a l o n g w i t h t h e n u m b e r o f services p e r c o n c e p t i o n (S/C). D a t a w e r e statistically a n a l y s e d using g r o u p e d t-test (Steel a n d T o r r i e , 1 9 6 0 ) .
Trial 3. S i x t y c o w s ( s e c o n d - - f i f t h l a c t a t i o n s ) w h i c h calved d u r i n g N o v e m b e r 1 9 8 3 - - J a n u a r y 1 9 8 4 w e r e used. C o w s o f t h e s a m e p a r i t y w e r e r a n d o m l y divided i n t o five g r o u p s , 12 in each: g r o u p 1 was u n t r e a t e d as c o n t r o l ; g r o u p s 2 a n d 3 r e c e i v e d e i t h e r 50 o r 1 0 0 p g G n R H a n a l o g u e p e r c o w , respectively, o n d a y 7 p o s t p a r t u m ; while g r o u p s 4 a n d 5 r e c e i v e d e i t h e r 50 or 100 p g G n R H a n a l o g u e p e r c o w , r e s p e c t i v e l y , o n d a y 15 p o s t p a r t u m . R e c o r d s o f P O V , P O E , UI, D O a n d S/C w e r e r e c o r d e d as in trial 2. D a t a o b t a i n e d w e r e s u b j e c t e d t o s t a n d a r d analysis o f v a r i a n c e (Steel a n d T o r r i e , 1960). RESULTS
Trial 1. T r e a t m e n t w i t h G n R H at t h e t i m e o f A.I. r e s u l t e d in a significant ( P < 0.05) increase in c o n c e p t i o n r a t e f r o m first i n s e m i n a t i o n o v e r t h a t o f t h e c o n t r o l g r o u p ( 8 1 . 3 v. 54.8%). Trial 2. U t e r i n e i n v o l u t i o n was c o m p l e t e d earlier in t h e t r e a t e d t h a n t h e c o n t r o l c o w s ( T a b l e 1). T h e d i f f e r e n c e h o w e v e r ( 2 5 . 4 v. 2 8 . 2 d a y s ) , was n o t significant. First o v u l a t i o n ( P O V ) o c c u r r e d a b o u t 5 d a y s earlier in t h e G n R H t r e a t e d c o w s t h a n t h e c o n t r o l s , t h e d i f f e r e n c e b e i n g highly significant ( P < 0.01). A significant ( P < 0.01) d i f f e r e n c e o f 9.0 d a y s in P O E was f o u n d b e t w e e n t h e t w o groups. This d i f f e r e n c e was c o n s i s t e n t w i t h t h a t o f P O V . G n R H - t r e a t e d c o w s c o n c e i v e d , o n average, a b o u t 2 1 . 4 d a y s earlier t h a n t h e c o n t r o l c o w s ; a d i f f e r e n c e w h i c h was statistically significant ( P < 0.05). TABLE 1 Means and standard errors (~-+ SE) of postpartum reproductive traits as affected by GnRH treatment (100 ug on day 15 postpartum) Trait
Control
Treated
Number of cows Days to :
19
19
Complete uterine involution First ovulation First oestrus Conception Number of services per conception
28.2 32.5 36.7 86.4
-+ 2.71 +- 1.57 + 2.33 • 8.74
2.2 -+ 0.37
25.4 27.4 27.7 65.0
+ 1.72 + 1.76 + 1.82 + 4.92
1.2 + 0.15
Significance of differences
NS ** ** * *
NS = Not significant, * = Significant (P < 0.05), ** = Significant (P < 0.01).
2.0a + 0.17
1.67 ab + 0:19
+- 2 . 1 9 ab ± 2.87 ab ± 2.97 a + 5.99
12
12
29.3 24.7 33.8 70.8
50 #g on day 7
Untreated
31.5 + 2.96 35.1 + 2.18 48.3 ± 4.68 80-8a ± 5.55
2
1
reproductive
± 1.73 a ± 1.05 a -+ 2 . 0 5 b + 2.94
1.33 b ± 0.14
23.5 19.3 23.9 54.2
12
100 ~g on day 7
3
Groups
+ 1.65 b + 2.16 b ± 5.88 a ± 4.39 1.5fl ab ± 0.19
26.5 25.9 39.6 73.4
12
50 ~g on day 15
4
+ 2.09 ab + 2.41 ab + 2.64 b + 2.96 1.25 b ± 0.13
25.5 24.3 29.0 56.1
12
100 ~g on day 15
5
traits as affected by dose level and time of GnRH treatment
N 8 = N o t s i g n i f i c a n t , * S i g n i f i c a n t ( P < 0 . 0 5 ) , * * = S i g n i f i c a n t ( P < 0 , 0 1 ). 1 M e a n s h a v i n g t h e s a m e s u p e r s c r i p t a r e n o t s i g n i f i c a n t l y (P < 0 . 0 5 ) d i f f e r e n t ,
Number of services per conception
Complete uterine involution First ovulation First oestrus Conception
Number of cows Days to :
Trait
M e a n s 1 a n d s t a n d a r d el-~ors (~ ± S E ) o f p o s t p a r t u m
TABLE 2
*
NS ** ** **
Significance of variation among groups
O
103
The within-group individual variation was greater in DO compared with those of other intervals studied, with the GnRH-treated cows showing smaller individual variation than the control cows ( C V - - 4 5 . 2 v. 33.8%). In the control group, this was partially due to animals (n = 5) that had to be inseminated three times before t h e y conceived, b u t it was mainly due to three animals that did n o t show oestrus for periods of 45 days or more after t h e y manifested their first oestrus. Rectal examination of these animals revealed that they did n o t have a palpable corpus luteum. On the other hand, only one of the treated cows had a period of anoestrus, with no palpable corpus luteum detected, for a b o u t 40 days. The GnRH-treated cows required, on average, 1.2 inseminations per conception, whereas the control animals required, on average, 2.2 inseminations to conceive (Table 1), The difference between the two groups was statistically significant ( P < 0.05). Trial 3. Longer UI was observed in untreated cows than in all treated groups (Table 2). The overall differences among groups were, however, n o t statistically significant. Among the treated groups, complete uterine involution occurred earliest (23.5 days) in cows receiving 100 pg G n R H at 7 days postpartum. Significant ( P < 0.01) reduction in POV occurred due to G n R H injection, as c o m p a r e d with untreated cows. First postpartum ovulation occurred markedly earlier (at 19.3 days) in group 3 (Receiving 100/~g G n R H on day 7 postpartum) than in the other treated groups, in which the averages of POV were similar (Table 2). Similar to the first ovulation, first detected oestrus was recorded significantly ( P < 0.01) earlier in all treated groups compared with the untreated control. Slightly longer POE was observed in cows treated on day 15 p o s t p a r t u m (groups 4 and 5) than in the corresponding groups receiving the same dose of G n R H on day 7 postpartum (groups 2 and 3, respectively). It is worth noting that incidence of the first ovulation not being associated with signs of oestrus (silent heat) was 33.3% in cows receiving 100 pg G n R H while it was 58.3% in both untreated cows and those receiving 50 #g GnRH. Overall differences in the number of days open (DO) were statistically significant ( P < 0.01). The latest animals to conceive were those of the control group (80.8 days) b u t t h e y did n o t significantly differ in DO from the two groups receiving 50 pg G n R H (groups 2 and 4), the latter having closely similar values (70.8 and 73.4 days). On the other hand, conception occurred significantly earlier in the cows receiving 100 pg G n R H (groups 3 and 5) than in other groups. Time of G n R H treatment after parturition, at high or low dose, did not have a significant effect on DO. The control animals required the greatest n u m b e r of services per conception (2.0). G n R H treatment at a dose of 50 pg either on day 7 or 15 p o s t p a r t u m insignificantly reduced S/C. An appreciable and significant ( P < 0 . 0 5 ) reduction in S/C occurred in
104 c6ws treated with 100 #g GnRH, as compared to the untreated group. The lowest S/C was t h a t of cows receiving 100 #g on day 15 postpartum. The effect of time of GnRH injection after calving on S/C was very small and insignificant.
Relation between various reproductive traits The data obtained in trial 3 were used to calculate the within-groups correlation coefficients between different traits studied. Out of these, the highest correlation was obtained between POV and POE (r= 0.697) which is, obviously, n o t unexpected. UI was positively correlated with each of POV and S/C ( r = 0 . 3 3 0 and 0.355, respectively), but its strongest correlation (r= 0.464) was t h a t with DO. On the other hand, poor and insignificant correlation was f o u n d between UI and POE (r = 0.166). The correlation between DO and POE (r = 0.544) was stronger than that between DO and POV (r = 0.41). Furthermore, both POV and POE were insignificantly correlated with S/C (r= 0.219 and 0.221, respectively). DISCUSSION The improved conception rate resulting from GnRH injection at the time of A.I. in the first trial supports the findings of Mori and Takahashi (1978), Schels and Mostafawi (1978) and Nakao et al. (1983). GnRH injection causes a luteinizing h o r m o n e (LH) surge (Zolman et al., 1973), and enhances ovulation (Leidl et al., 1979) and thus eliminates the occurrence of delayed ovulation, relative to the time of A.I. It has been reported t h a t in 10% or more of cows,: ovulation might be delayed in relation to oestrus, resulting in a mistiming of A.I. and low conception rate (Bostedt, 1976; Karg et al., 1980). The effect of postpartum t r e a t m e n t with GnRH on uterine involution was consistent in both trials 2 and 3. The effect was more pronounced when a 100 #g dose of GnRH was given 7 days postpartum. GnRH enhances uterine involution in cows with retained placenta (Bostedt and Maurer, 1982) and also in cows after normal parturition without retained placenta (Britt et al., 1974). The role of G n R H in uterine involution, however, remains unelucidated. It has been reported that GnRH injection in the early postpartum period enhances the resumption of ovarian function and the occurrence of first ovulation (Schams et al., 1973; Britt et al. 1974). The results of b o t h trials 2 and 3 support this view. Although GnRH given at either high or low dose and either at 7 or 15 days postpartum significantly enhanced the occurrence of first ovulation, the effect was most pronounced when a dose of 100 #g was given on day 7 postpartum. The responsiveness of the anterior pituitary to GnRH stimulation has been reported to be regained by day 7 or 8 postpartum in lactating dairy cows (Kesler et al.,
105 1977). The results of b o t h trials 2 and 3 (Tables 1 and 2) indicate that there was a time lapse which ranged from 9.3 to 17.7 days on average, between GnRH treatment and the occurrence of first ovulation. Peters and Lamming (1984) suggested that the induced LH release resulting from G n R H injection might result in premature luteinization of an unovulated follicle and transient secretion o f progesterone sufficient to initiate ovarian cycles. The effect of G n R H treatment on the time of first postpartum oestrus was, as expected, in line with its effect on first ovulation. This was true for b o t h trials 2 and 3. Of interest is the effect of dose level on the incidence of ovulatory anoestrus (silent heat) at the first ovulation. This is difficult to explain in the light of the available results in this study. It m a y be related, however, to a higher degree of follicular growth and hence a possibly higher pre-ovulatory oestradiol level in animals treated with a high dose (100 pg) of G n R H as compared with either the untreated animals or those receiving a low G n R H dose (50 pg). The effect o f G n R H at 100 pg/cow on reducing the number of days open by 20 days or more is consistent in trials 2 and 3. It is also similar to the reported reduction of about 24 days in the interval from calving to conception (Boiti et al., 1982) or from post-calving treatment to conception ( H u m b o l t and Thibier, 1981) resulting from G n R H injection as compared with control groups. In some other studies, however, the effect of G n R H treatment on reducing the interval from calving to conception was either small and n o t significant (Bostedt and Maurer, 1982; Ball'and Lamming, 1983) or pronounced only in cows with retained placenta (Bostedt and Maurer, 1982). In the present study, it seems that G n R H injection n o t only stimulated the pituitary function and, as a result, the resumption of :ovarian activity, b u t also resulted in regulation of the reproductive process in general. This was clear from the effect of G n R H treatment in reducing the number of services per conception. This is similar to the positive effect of G n R H reported b y Schams e t al. (1973). Other workers, however, f o u n d such effect to be small and n o t significant ( H u m b o l t and Thibier, 1981) or to be limited to cows with retained placenta (Bostedt and Maurer, 1982). The discrepancy in the results obtained in different studies may be due to the timing of G n R H treatment in relation to calving and insemination or the time when first insemination t o o k place after calving. The significant positive correlation between UI and each of POV, DO and S/C indicates the importance of rapid uterine involution in the resumption of normal functioning Of t h e reproductive system leading to conception. These relationships were discussed by Eley et al. (1981) and Peters (1984). The finding that DO showed the highest correlation with POE supports the well established view of the importance of applying an accurate and effective heat detection system for reducing calving interval (Esslemont, 1974; Buiman and Lamming, 1978; E1-Keraby and Aboul-Ela, 1982).
106 CONCLUSION
The results of this study illustrate the beneficial use of G n R H for increasing fertility in dairy cattle. Higher conception rate may be achieved through G n R H injection at the time of A.I. Postpartum treatment with a low dose of G n R H (50 pg) was less effective than a high dose (100:gg). Injection of 100 pg G n R H at either 7 or 15 days postpartum reduced the intervals from calving to complete uterine involution, first ovulation, first detected oestrus and conception and also the number of services per conception. Injection of 100 ~g G n R H on day 15 postpartum was most effective in reducing appreciably both the number of services per conception and days open which are of great importance from the economic point of view to the dairy farmer. ACKNOWLEDGEMENT
The authors wish to thank Hoechst U.K. Ltd. for the supply of 'Gonadorelin' used in this study.
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107 Esslemont, R., 1974. Economic and husbandry aspects of the manifestation and detection of oestrus in cows. Part I. Economic aspects. ADAS Q. Rev., 12: 175--184. Humbolt, P. and Thibier, M., 1981. Effect of gonadotropin releasing h o r m o n e (GnRH) treatment during the midluteal phase in repeat breeder cows. A preliminary report. Theriogenology, 16: 375--376. Karg, H., Claus, R., Gunzler, O., Rattenberger, E. and Firchner, F., 1980. Hormonal recognition o f cyclicity and infertility in cows under practical conditions. Proceedings of the l l t h World Congress of Diseases of Cattle. Israel, pp. 885--894. Kesler, D.J., Garverick, H.A., Youngquist, R.S., Elmore, R.G. and Bierschwal, C.J., 1977. Effect of days postpartum and endogenous, reproductive hormones on GnRH induced in LH release in dairy cows. J. Anita. Sci., 45: 797--803. Leidl, W . , Bostedt, H., Lamprecht, W., Prinzen, R. and Wendt, V., 1979. The influence of GnRI-I and HCG on the time o f ovulation in cows artificially inseminated. Tier~irztl. Umsch., 34: 546--555. Mori, J. and Takahashi, T., 1978. Effect of an analog of LH-RH on pregnancy rate in cows. Jpn. J. Anita. Reprod., 24: 137--138. Nakao, T., Narita, S., Tanaka, K., Haxa, H., Shirkawa, J., Noshiro, H., Saga, N., Tsunoda, N. and Kawata, K., 1983. Improvement of first-service pregnancy rate in cows with gonadotropin-releasing hormone analog. Theriogenology, 20: 111--119. Peters, A.R., 1984. Reproductive activity o f the cow in the postpartum period. I. Factors affecting the length of the postpartum acyclic period. Br. Vet. J., 140 : 76--84. Peters, A.R. and Lamming, G.E., 1984. Reproductive activity o f the cow in the postpartum period. H. Endocrine patterns and induction o f ovulation. Br. Vet. J., 140: 269--280. Sehams, D., Hofer, F., Hofmann, B., Ender, M.L. and Karg, H., 1973. Effects o f synthetic LH-RH treatment on bovine ovarian function during oestrous cycle and postpartum period. Acta Endocrinol. Suppl., 177: 296. Schels, H.F. and Mostafawi, D., 1978. The effect of GnRH on the pregnancy rate o f artificially inseminated cows. Vet. Rec., 103: 31--32. Steel, R.G.D. and Torrie, J.H., 1960. Principles and Procedures o f Statistics. McGraw Hill, New York, NY, 481 pp. Zar, J.H., 1974. Biostatistical Analysis. Prentice-Hall, Englewood Cliffs, NJ, 620 pp. Zolman, J., Convey, E.M., Britt, J.G. and Hafs, J.H., 1973. Release of the bovine luteinizing hormone b y purified synthetic gonadotropin releasing hormone. Proc. Soc. Exp. Biol. Med., 142: 189--193.