Resumption of gonadotrophin release during the post-partum period in suckling and non-suckling ewes

The post-partum secretion of LH, FSH and prolactin was monitored in 15 suckling and 6 non-suckling Préalpes du Sud ewes lambing during the breeding season by measuring plasma hormone concentrations daily at 6-h intervals and also weekly at 20-min intervals for 6 h from parturition to resumption of regular cyclic ovarian activity. There was a constant phenomenon in the resumption of normal patterns of FSH and LH secretion: there was a rise in FSH values culminating on average on Day 4 post partum and returning subsequently to values observed during the oestrous cycle, and concurrently an increase in the frequency and amplitude of LH pulses more progressive in suckling than in non-suckling ewes which led to an elevation of LH mean concentrations and occurrence of an LH surge. Since neither the FSH secretory pattern nor FSH mean values differed between suckling and non-suckling ewes, the results suggested that LH pulsatile pattern was a major limiting factor for the resumption of normal oestrous cycles. Before regular oestrous cycles resumed other changes in preovulatory LH surges also occurred: (i) they increased in duration and probably in amplitude; (ii) they were preceded by an acceleration in LH pulse frequency and a large decrease in FSH values as in normal cyclic ewes; and (iii) at least in non-suckling ewes they occurred concurrently with a prolactin surge.     

Ram-induced growth of ovarian follicles and gonadotrophin inhibition in anoestrous ewes

Southdown ewes in mid-seasonal anoestrus were exposed to rams for 0 h (control group), 2 h, 24 h, 40 h, 3 days, 10 days or 20 days. Serial blood samples were then taken to determine LH and FSH levels. Ewes with greater than 24 h ram exposure were ovariectomized immediately after bleeding, and all follicles greater than 1 mm diameter were dissected from the ovaries and measured. LH basal concentrations and pulse frequency increased significantly within 2 h of ram introduction, but by 24 h fell, and then remained low. FSH concentrations fell within 2 h of ram introduction and remained low. Control group ewes (isolated) had no follicles greater than 4 mm diameter, whereas all ewes exposed to rams had large follicles, with CL or preovulatory follicles present at 40 h after ram introduction. Ram introduction was also associated with follicle recruitment (antrum formation to less than 2 mm). Follicular recruitment and development to the large follicle stage therefore occurred during a period of low plasma gonadotrophin levels and suppressed LH pulsing.     

Effect of litter size on blood haematocrit and liveweight in Booroola Merino and Merino ewes during pregnancy and the post-partum period

Blood haematocrit and liveweight were determined throughout pregnancy and the post-partum period in 217 Booroola Merino and Merino ewes in order to relate these parameters to litter size at birth. In pregnant ewes, haematocrit declined from three until five months gestation, rose immediately after parturition then declined until two months post-partum. During the third to fifth month of gestation, haematocrit decreased in proportion to litter size. Nonpregnant ewes, measured at similar intervals, did not show the same pattern. Haematocrit of nonpregnant animals was higher than that of triplet-bearing ewes at three, four and five months gestation, but was only significantly different to single- and twin-bearing ewes at five months. The liveweight of pregnant ewes increased up to parturition and then declined until two months post-partum. The liveweight of nonpregnant ewes increased over the experimental period. It was concluded that the number of foetuses a ewe carried had significant effects on the decline in haematocrit during pregnancy. Haematocrit was not a precise indicator of litter size in sheep. Haematocrit, ewe liveweight and ovulation rate together in a multiple regression only accounted for 37% of the variation in litter size.     

Effect of diet and GnRH administration on post-partum ovarian cyclicity in autumn-lambing ewes

Using autumn-lambing ewes, this study investigated (i) the effects of diet on gonadotrophin secretion and responsiveness of the hypothalamic-pituitary-ovarian axis to exogenous GnRH during the early post-partum period; and (ii) whether ovulation prior to completion of uterine involution results in an increased incidence of aberrant ovarian cycles. Thirty-two ewes rearing 1.9+/-0.12 lambs were equally allocated to two dietary treatments at lambing (22 October +/-0.2 day). Diets comprised ad libitum hay and 1.5 kg per ewe per day of one of two concentrates (11.5 MJ ME, 195 g CP per kg) containing 300 g kg(-1) cracked maize grain (M) or 300 g kg(-1) sugar beet pellets (S). Half of the ewes on each diet (G) received 25 i.v. injections of 250 ng GnRH in 2 ml 0.9% saline at 2 h intervals from days 12-14 post-partum while remaining ewes (N) were monitored for the resumption of spontaneous ovarian cyclicity. Blood samples were obtained from all ewes throughout the study (lambing to 18 December) for measurement of circulating hormone concentrations and the uteri and ovaries of all ewes were examined via laparoscopy on day 21 post-partum. There were no effects of dietary treatment on ewe daily live weight loss, lamb daily live weight gain or the immediate post-partum increase in circulating FSH concentrations. Diet did not affect insulin concentrations or LH pulse frequency on day 12 post-partum but LH pulse amplitude was lower in ewes fed concentrate M compared to concentrate S (1.4+/-0.10 versus 1.7+/-0.12 ng ml(-1), respectively, P<0.05) and this was associated with an increased interval to the resumption of spontaneous ovarian cycles (35+/-3.1 versus 26+/-2.1 day, respectively, P<0.05). Administration of exogenous GnRH increased (P<0.05) the proportion of ewes on both diets that ovulated within 20 days of parturition and advanced the onset of ovarian cyclicity in ewes fed concentrate M by 9.5 days (significance of interaction, P<0.05). Four ewes, all of which ovulated before day 22 post-partum, had extended luteal activity while in remaining ewes, duration of the first luteal phase was inversely related to the time of first ovulation (r(2)=0.16, P<0.05). Results demonstrate that (i) the onset of ovarian cyclicity is influenced by diet and can be advanced by administration of exogenous GnRH; and (ii) ovulation during the early post-partum period is associated with an increased incidence of extended luteal activity.     

Morphine, naloxone and the gonadotrophin surge in ewes

Possible endogenous opioid peptide regulation of the preovulatory gonadotrophin surge was examined in ewes during the breeding season. Intact ewes (n = 54) were synchronized by treatment for 12 days with intravaginal sponges releasing medroxyprogesterone acetate. Luteinizing hormone (LH) and follicle-stimulating hormone (FSH) secretion prior to and during the gonadotrophin surge were not affected by naloxone (0.33 mg/kg body wt per h) administered from the time of medroxyprogesterone acetate withdrawal until 30 h after the onset of oestrus (n = 6). Morphine was administered in 4 patterns: (i) 0.25 mg morphine/kg body wt per h from medroxy-progesterone acetate withdrawal until 30 h after the onset of oestrus (n = 6), (ii) 0.25 mg morphine/kg body wt per h from 24 to 48 h after medroxyprogesterone acetate withdrawal (n = 6), (iii) 0.50 mg morphine/kg body wt per h from 24 to 36 h after medroxyprogesterone acetate withdrawal (n = 6) and (iv) 0.50 mg morphine/kg body wt per h from 18 to 30 h after medroxyprogesterone acetate withdrawal (n = 6). Oestrus and the gonadotrophin surge were delayed, but not blocked, in all cases of morphine administration (P less than 0.05). Inconsistent effects of morphine on circulating oestradiol and gonadotrophin concentrations prior to the gonadotrophin surge suggest that the delays are not due to reduced gonadotrophic support of ovarian oestradiol output. Morphine may reduce responsiveness of central behavioural and gonadotrophin surge-generating centres to the oestradiol signal. The absence of effects of naloxone on gonadotrophin secretion suggest that suppression of LH secretion by opioid peptide activity is reduced after the end of the luteal phase.(ABSTRACT TRUNCATED AT 250 WORDS)     

Role of the corpus luteum of pregnancy in controlling pituitary gonadotrophin secretion during the early post-partum period in the ewe

No difference was found between 5 intact ewes and 5 ewes from which the CL had been excised at Day 70 of pregnancy in the plasma concentration of progesterone at Day 140, and concentrations of progesterone remained below 0.2 ng/ml during the first 20 days post partum. Plasma concentrations of LH, frequency and amplitude of LH pulses were low at Day 140 and increased considerably, particularly in the CL-excised ewes, as early as Day 5 post partum. No significant differences were found between the two groups of ewes in the mean plasma concentrations of FSH for any of the 5 stages examined. Taken together, these results suggest that some factor, other than progesterone, associated with the CL of pregnancy is involved in the inhibition of pulsatile LH secretion during the early post-partum period.     

Effect of variation in embryo stage on the establishment of pregnancy, and embryo survival and growth in ewes with two embryos

Embryos at different stages of development were transferred to recipient ewes on Day 6 to investigate the effect of variation in stage of development on embryo survival and growth. Three groups of ewes received 2 embryos that were at the same stage of development, Day 4, Day 6 or Day 8. A fourth group received 1 Day-4 and 1 Day-8 embryo. At autopsy on recipient Day 34 there were no significant differences in embryo survival (Day 4, 34%; Day 6, 50%; Day 8, 46%; and Day 4 and 8, 48%). Fetuses developing from Day-8 embryos were heavier than others (Day 4, 1.10 +/- 0.06 g; Day 6, 1.15 +/- 0.06 g; Day 8, 1.41 +/- 0.08 g; P less than 0.05). In Group 4 neither survival nor growth of embryos was significantly affected by the presence of an embryo at a different stage of development. The ability of the uterus to stimulate development of a relatively retarded embryo is confirmed. Apparently the uterus has less effect in slowing the development of advanced embryos.     

Effect of gonadotrophin environment on growth and development of isolated mouse primary ovarian follicles

Collagen gels containing isolated primary follicles devoid of other ovarian tissue were transferred beneath the kidney capsule of 3 types of female recipients: cycling, ovariectomized and hypogonadal, known to have different circulating concentrations of gonadotrophins. After 10 days the gels were recovered and processed for histology or the oocytes were recovered and their diameters measured and their ability to resume meiosis was determined. The growth of isolated primary follicles was positively correlated with the concentrations of circulating gonadotrophins in the recipient mice, but the numbers of oocytes recovered, the rate of oocyte growth and resumption of meiosis did not differ in the 3 types of recipient studied. This indicates that, in the conditions provided, oocyte growth was not related to the extent of follicular development.     

Effect of human chorionic gonadotrophin on dominant follicles in cows: formation of accessory corpora lutea, progesterone production and pregnancy rates.

Thirty-four lactating Holstein cows were randomly assigned to four groups for treatment with human chorionic gonadotrophin (hCG, 1000 iu) at insemination day 0 (n = 8) or 7 (n = 9) or 14 days (n = 9) after insemination or with no hCG treatment (control, n = 8). Ultrasound imaging of the ovaries and plasma progesterone measurements were carried out to determine follicular dynamics and corpus luteum growth and function. Rates of formation of accessory corpora lutea were higher among cows treated on days 0 (three cows), 7 (seven cows) or 14 (four cows) than in the controls (one cow). Total corpus luteum diameter was greater (P less than 0.01) among hCG-treated cows than in controls 7-42 days after insemination. Concentrations of progesterone in plasma were significantly (P less than 0.05) higher in cows treated with hCG on days 7 or 14 than in those treated on day 0 or in controls, at days 18, 35 or 42 after insemination. Seven of the cows treated on day 7 became pregnant, whereas four, four and three cows treated on days 0 or 14 and control cows became pregnant, respectively. The results suggest that hCG treatment at 7 days after insemination could be used to produce accessory corpora lutea, raise plasma progesterone concentration and hence reduce the incidence of early embryonic mortality in cattle.     

Effect of nutritionally induced liveweight differences on ovulation rates and the population of ovarian follicles in ewes

Differential feeding prior to the onset of the breeding season resulted in mean liveweight differences of 12 kg in two groups of Romney ewes. This difference was maintained while ovulation rates and the population of externally visible ovarian follicles were measured. The ovulation rate at first oestrus was significantly higher in the high liveweight group but there was no difference between groups thereafter. High liveweight ewes had consistently higher mean follicle scores at all stages of the oestrous cycle. Mean follicle score was lower following the third oestrus than following first oestrus (P < 0.01).     

The infusion of glucose in ewes during the luteal phase increases the number of follicles but reduces oestradiol production and some correlates of metabolic function in the large follicles

Short-term nutritional supplementation stimulates folliculogenesis in ewes probably by insulin-mediated actions of glucose in the follicle. The aim of this study was to determine the effect of glucose on follicle number and granulosa levels of Aromatase P450 and phosphorylated Akt and AMPK. Twelve Ile-de-France ewes were allocated to two groups; one (n=7) infused with saline and the other (n=5) with glucose (10mM/h) for 72h in the luteal phase. At the end of infusion, ovaries were collected and all follicles >1mm in diameter were dissected to recover granulosa cells. Aromatase P450 and phosphorylated Akt and AMPK were analysed by Western blotting of granulosa cell lysates. Blood plasmas collected before and during the infusions were analysed for progesterone, oestradiol, LH, FSH, glucose, insulin and IGF-I. The infusion of glucose significantly increased follicle number but, significantly reduced Aromatase P450 and phosphorylated Akt and AMPK in granulosa cells. The circulating concentration of glucose rose significantly 3h after the start of the glucose infusion and remained elevated until 27h then fell; the circulating concentration of insulin rose significantly by 3h and remained elevated. The circulating concentration of oestradiol fell significantly by 32h and remained low; the circulating concentrations of LH and FSH were unaffected. These data show that short-term infusion of glucose stimulated follicular growth but decreased Aromatase P450 in granulosa cells. The reduced levels of phosphorylated Akt and AMPK suggest that the phosphatidylinositol 3-kinase pathway has been inhibited by high concentrations of glucose. These data also suggest that there may be functional cross-talk between FSH and insulin signalling in granulosa cells.     

Plasma progesterone and gonadotrophin concentrations and ovarian activity in post-partum dairy cows

Plasma progesterone concentrations in jugular vein blood samples collected every other day after calving from 13 Friesian dairy cows indicated that ovarian cyclic activity was initiated by 16.6 +/- 1.1 (s.e.m.) days post partum, except for 1 cow which did not resume cyclic activity until Day 98 post partum. Rectal palpation of the ovaries indicated that a developing follicle was recognizable at a mean time of 15.7 +/- 2.0 days after calving. During the first oestrous cycle after parturition there was a significantly shorter period when plasma progesterone levels were elevated than during the next 2 cycles. Concentrations of progesterone, LH, FSH and prolactin were determined for 4 cows, in blood samples taken every 6 h from 2 to 36 days post partum. Tonic LH release was lower during the first 10 days than subsequently, but the lack of change in pattern for FSH suggests dissimilar control mechanisms for these hormones during this time. Three cows showed evidence of a resumption of ovarian cyclicity during the sampling period: in 2 there was an initial LH surge of a magnitude which would normally give rise to ovulation, followed 4 days later by an increase in plasma progesterone lasting only 5 and 9 days. This progesterone was considered to be of follicular origin. A second LH surge was followed by the presence of a corpus luteum.     

Effect of active immunization against oxytocin on gonadotrophin secretion and the establishment of pregnancy in the ewe

A group of 14 ewes was actively immunized against oxytocin coupled to carrier protein, and comparisons of the reproductive status of these animals were made against ewes immunized against carrier protein only (N = 5) and untreated controls (N = 6). The last two groups were indistinguishable and were therefore combined as a single control group for analysis of the results. Oestrous cycle lengths were significantly extended in oxytocin-immunized ewes (P less than 0.005) with 42% of cycles lasting greater than 18 days. Cloprostenol treatment in the mid-luteal phase resulted in apparently normal luteal regression and re-ovulation, but luteal phase FSH concentrations and follicular phase LH concentrations were elevated in the immunized ewes, although surge levels of both hormones were unaffected. Measurements of free oxytocin concentrations in the blood suggested that these were significantly raised in treated animals. Progesterone concentrations in peripheral plasma were not altered by treatment. Mating resulted in a conception rate of 91% in control ewes compared with only 28% in oxytocin-immunized animals (P less than 0.01). There was no evidence of any conceptus material in the uteri of non-pregnant immunized ewes 25 days after service. Some had re-ovulated, whereas the ovaries of others contained mature corpora lutea which had been maintained. Ovarian histology appeared normal. We conclude that active immunization against oxytocin influences gonadotrophin secretion and reduces fertility. The site(s) of action for both of these effects needs to be determined.     

Development of dominant follicles and length of ovarian cycles in post-partum dairy cows

The resumption of ovarian activity after normal calvings was studied in 18 lactating Friesian cows. Since, in 17 cows, first post-partum ovulation occurred without overt oestrous behaviour being detected, the resultant cycles were called 'ovarian cycles'. The mean (+/- s.d.) length of the ovarian cycles was 21.0 +/- 8.7 days. The duration of cycles tended to be normal (18-24 days) or long (greater than or equal to 25 days) when the ovulatory dominant follicles were identified before Day 10 post partum; they were consistently short (9-13 days) when dominant follicles identified after Day 20 post partum ovulated. When such follicles were detected between Days 10 and 20 post partum, long, normal and short ovarian cycles were detected. The number of waves of follicular growth with associated dominant follicles observed during the ovarian cycles tended to be related to cycle length; short cycles had 1 dominant follicle, normal cycles predominantly 2, and long cycles mostly 3 dominant follicles. The mean (+/- s.d.) duration of 13 oestrous cycles studied was 23.1 +/- 2.1 days. Of these cycles, 7 had 3 and 6 had 2 dominant follicles. The oestrous cycles with 3 dominant follicles had a mean (+/- s.d.) duration of 24.0 +/- 1.2 days and the respective dominant non-ovulatory follicles reached maximum sizes on Days 8 and 18, respectively; oestrous cycles with 2 dominant follicles were 22.2 +/- 2.6 days in duration, and the dominant non-ovulatory follicle reached maximum size by Day 8. Ovarian follicular development during the first 45 days of pregnancy was characterized by the growth and regression of successive dominant follicles, each lasting 10-12 days. These results show that the first ovarian cycle was predominantly short when the ovulatory dominant follicle was first detected after Day 20 post partum.     

LH release and luteal function in post-partum acyclic ewes after the pulsatile administration of LH-RH

The administration of LH-RH in a pulsatile regimen (100 ng i.v./h for 48 h) to acyclic ewes 26-30 days post partum increased plasma LH concentrations, and both the frequency and amplitude of plasma LH pulses. In 12/14 ewes these increases were followed by plasma LH surges similar to the preovulatory surges observed in 10 control cyclic ewes. Subsequent luteal function in the post-partum ewes was deficient. Plasma progesterone was detected in 7/12 post-partum ewes showing plasma LH surges. The concentrations were lower (1.3 +/- 0.2 ng/ml) and detected for shorter periods (3-10 days) than in cyclic ewes (2.4 +/- 0.2 ng/ml, 12/15 days). In the post-partum ewes the increases in plasma LH concentrations before the LH surge were higher but of shorter duration than in the cyclic ewes. The inadequate luteal function in the post-partum ewes could therefore have been due to inappropriate LH stimulation of the ovary before the LH surge.     

Modulation of functional capacity of small ovarian follicles in the post-partum cow by prolactin

This study was undertaken to examine the possibility that the prolonged anovulatory period frequently experienced by the post-partum cow is due to a disruption of function at the ovarian level promoted by the high, suckling-induced, blood prolactin concentrations. Fifteen cows, less than 35 days post partum, were allocated to three groups (1, 3 and 5) and given no hormonal treatment, prostaglandin plus pregnant mare serum gonadotrophin (PMSG) treatment or injected with 2-bromo-alpha-ergocryptine to reduce circulating prolactin levels. Ten synchronized cyclic cows were allocated to two groups (2 and 4) and given prostaglandin or prostaglandin plus PMGS treatment. All cows were ovariectomized 1 or 2 days after treatment of Graafian follicles less than 9 mm in diameter were selected after dissection from the ovaries. The follicles were cultured for 18 h with or without prolactin (1 microgram/ml) and steroid accumulation in the culture medium estimated. The follicles were then separated into theca and granulosa which were incubated for 40 min with LH (1 microgram/ml) or FSH (5 micrograms/ml). Cyclic AMP concentrations were estimated as an indication of tissue responsiveness to gonadotrophins. The secretion of oestradiol-17 beta, progesterone, testosterone or androstenedione during 18 h culture did not differ between follicles isolated from post-partum or cyclic cows. The presence of prolactin in the culture medium had no overall effect on steroid secretion although some specific effects within each group were noticed. Incubation with LH increased cyclic AMP levels in the theca but the granulosa did not respond. Likewise FSH increased cyclic AMP levels in granulosa preparations but not in theca. There were no differences in response between post-partum and cyclic cows, but exposure of the follicles to prolactin in vitro did significantly reduce the LH-induced increase in cyclic AMP levels in isolated theca. We have concluded that endogenous prolactin may modify but does not inhibit the resumption of ovarian function following parturition in the beef cow.     

Differences in the reproductive endocrine status of ewes in the early post-partum period and during seasonal anoestrus

More (P less than 0.05) post-partum acyclic ewes (8/9) showed evidence of pulsatile LH release than did seasonally anoestrous ewes (2/8). Mean plasma prolactin concentrations were higher (P less than 0.05) in the post-partum ewes than in the seasonally anoestrous ewes. Hypothalamic and pituitary cytosol oestrogen receptor number, median eminence GnRH content and pituitary LH, FSH and prolactin contents were similar in the two groups of ewes. The number of nuclear-bound oestrogen receptors was greater (P less than 0.01) in pituitaries from seasonally anoestrous ewes than in pituitaries from post-partum ewes. These data suggest that the basis of acyclicity in seasonally anoestrous ewes and in post-partum ewes is probably due to their inability to generate LH pulse frequencies similar to that in the follicular phase of the oestrous cycle. The higher LH pulse frequency in post-partum ewes, compared to that in seasonally anoestrous ewes, suggests that pregnancy may partly negate the reduction in LH secretion that is associated with photoperiod-induced acyclicity. The lower number of nuclear-bound oestrogen receptors in post-partum ewes suggests that there may be lower plasma oestrogen levels and less ovarian follicular growth than in non-suckling anoestrous ewes.     

Effect of human chorionic gonadotrophin and indomethacin on ovulation, steroidogenesis and prostaglandin synthesis in preovulatory follicles of PMSG-primed immature rats

Immature rats were treated with PMSG followed 56 h later by 10 i.u. hCG. Follicles were removed at intervals after hCG injection. Transient increases in progesterone, testosterone and oestradiol synthesis were first evident 1 h after hCG, but values peaked at 3-5 h and returned to control levels by 10 h. Increased synthesis of PGE-2 and PGF-2 alpha was not evident until 3 h and peaked at more than 10 h after hCG. Ovulation began between 8 and 10 h after hCG and 83% of animals had ovulated within 12 h. Doses of 90 or 1800 micrograms indomethacin given together with hCG substantially inhibited ovulation and PG synthesis, but only the higher dose inhibited the hCG-induced elevation of progesterone and testosterone synthesis; hCG-induced oestradiol synthesis was not affected by either dose of indomethacin. We conclude that the peak of PG synthesis after hCG treatment related closely to the timing of ovulation; the steroidogenic response to hCG was not blocked by doses of indomethacin sufficient to inhibit synthesis of PGE-2 and PGF-2 alpha by more than 80%.