Effects of applying gamma-aminobutyric acid(B) drugs into the medial basal hypothalamus on basal luteinizing hormone concentrations and on luteinizing hormone surges in the female sheep

Prior investigations have shown that localized infusion by microdialysis of gamma-aminobutyric acid(B) (GABA(B)) agonists into the medial basal hypothalamus of male sheep rapidly increases GnRH and LH pulse amplitude. The objectives of these studies were to determine if infusion of GABA(B) agonists SKF 97541 or baclofen into the medial basal hypothalamus of female sheep would affect basal LH secretion and if infusion of a potent antagonist would alter expression of LH surges induced by injection of estrogen. Infusion of either SKF 97541 (10 or 40 microM) or baclofen (1 mM) into estrogen-treated ovariectomized ewes did not alter basal LH secretory patterns, whereas both drugs significantly elevated mean LH and LH pulse amplitude in ovariectomized ewes during the nonbreeding season. Infusion of the antagonist CGP 52432 (250 or 500 microM) did not affect expression of estrogen-induced LH surges in ovariectomized ewes. These observations support the concept that GABA(B) receptors in the medial basal hypothalamus regulate basal LH secretion but do not regulate the surge mode of LH secretion in the female sheep.     

Effect of melatonin on daily LH secretion in intact and ovariectomized ewes during the breeding season.

This study was conducted to find out whether daily LH secretion in ewes may be modulated by melatonin during the breeding season, when the secretion of both hormones is raised. Patterns of plasma LH were determined in luteal-phase ewes infused intracerebroventricularly (icv.) with Ringer-Locke solution (control) and with melatonin (100 microg/100 microl/h). Response in LH secretion to melatonin was also defined in ovariectomized (OVX) ewes without and after estradiol treatment (OVX+E2). Basal LH concentrations by themselves did not differ significantly before, during and after both control and melatonin infusions in intact, luteal-phase ewes. However, single significant (P<0.05) increases in LH concentration were noted during the early dark phase in the control and 1h after start of infusion in melatonin treated ewes. In both OVX and OVX+E2 ewes, melatonin decreased significantly (P<0.01, P<0.05, respectively) mean plasma LH concentrations as compared to the levels noted before the infusions. In OVX+E2 ewes, a single significant (P<0.05) increase in LH occurred 1h after start of melatonin treatment, similarly as in luteal-phase ewes. No significant differences in the frequencies of LH pulses before, during and after melatonin infusion were found in all treatments groups. In conclusion, melatonin may exert a modulatory effect on daily LH secretion in ewes during the breeding season, stimulating the release of this gonadotropin in the presence of estradiol feedback and inhibiting it during steroid deprivation. Thus, estradiol seems to be positively linked with the action of melatonin on reproductive activity in ewes.     

Chronic nitric oxide deficiency is associated with altered leutinizing hormone and follicle-stimulating hormone release in ovariectomized rats

Nitric oxide (NO) synthase (NOS) has been found in the gonadotrophs and folliculo-stellate cells of the anterior pituitary. Previous observations from our laboratory suggest that NO may play a role in regulating gonadotropin secretion. Because estrogen secretion by the ovary can influence gonadotropin secretion, we investigated the hypothesis that chronic in vivo NO deficiency has a direct estrogen-independent effect on luteinizing hormone (LH) and follicle-stimulating hormone (FSH) secretion. Chronic NO deficiency was induced by adding an NOS inhibitor, N-nitro-L-arginine (L-NNA, 0.6 g/l) to the drinking water of ovariectomized (OVX) rats. The control OVX rats were untreated. After 6-8 weeks, the animals were sacrificed, and the pituitaries were removed and perfused continuously for 4 hr in the presence of pulsatile gonadotropin-releasing hormone (GnRH, 500 ng/pulse) every 30 min. S-Nitroso-L-acetyl penicillamine (SNAP, an NO donor, 0.1 mM) or L-nitro-arginine methyl ester (L-NAME, an NOS inhibitor, 0.1 mM) was added to the media and perfusate samples were collected at 10-min intervals. GnRH-stimulated LH and FSH levels were significantly lower in pituitaries from OVX/NO-deficient pituitaries compared with pituitaries from the OVX control group. The addition of SNAP significantly decreased LH and FSH secretion by pituitaries from OVX control animals, but significantly increased their secretion by pituitaries from the OVX/NO-deficient animals. L-NAME also suppressed LH and FSH secretion by pituitaries from the OVX control animals and stimulated their release by pituitaries from the NO-deficient/OVX animals. Immunohistochemistry of frontal sections through the hypothalamus demonstrated that OVX/NO deficiency is associated with increased GnRH in the median eminence. We conclude that NO has a chronic stimulatory effect on LH and FSH release and the subsequent altered secretory responsiveness to NO agonist or antagonist is the result of chronic NO suppression.     

Differential modulation of gonadotropin secretion by selective estrogen receptor 1 and estrogen receptor 2 agonists in ovariectomized ewes

The objectives of this study were to determine whether activation of estrogen receptor 1 (ESR1; also known as ERalpha), or estrogen receptor 2 (ESR2; also known as ERbeta), or both are required to: 1) acutely inhibit secretion of LH, 2) induce the preovulatory-like surge of LH, and 3) inhibit secretion of FSH in ovariectomized (OVX) ewes. OVX ewes (n = 6) were administered intramuscularly 25 micrograms estradiol (E2), 12 mg propylpyrazoletriol (PPT; a subtype-selective ESR1 agonist), 21 mg diaprylpropionitrile (DPN; a subtype-selective ESR2 agonist), or PPT + DPN. Like E2, administration of PPT, DPN, or combination of the two rapidly decreased (P < 0.05) secretion of LH. Each agonist induced a gradual, prolonged rise in secretion of LH after the initial inhibition, but neither agonist alone nor the combined agonists was able to induce a "normal" preovulatory-like surge of LH similar to that induced by E2. Compared with E2-treated ewes, the beginning of the increase in secretion of LH occurred earlier (P < 0.01) in DPN-treated ewes, later (P < 0.05) in PPT-treated ewes, and at a similar interval in ewes receiving the combined agonist treatment. Like E2, PPT decreased (P < 0.05) secretion of FSH, but the duration of suppression was much longer in PPT-treated ewes. DPN did not alter secretion of FSH in this study. Modulation of the number of GnRH receptors by PPT and DPN was examined in primary cultures of ovine pituitary cells. In our hands, both PPT and DPN increased the number of GnRH receptors, but the dose of DPN required to stimulate synthesis of GnRH receptors was 10 times higher than that of PPT. We conclude that in OVX ewes: 1) ESR1 and ESR2 mediate the negative feedback of E2 on secretion of LH at the level of the pituitary gland, 2) ESR1 and ESR2 do not synergize or antagonize the effects of each other; however, they do interact to synchronize the beginning of the stimulatory effect of E2 on secretion of LH, 3) ESR1 and ESR2 may mediate at least partially the positive feedback of E2 on LH secretion by increasing the number of GnRH receptors, and 4) only ESR1 appears to be involved in the negative feedback of E2 on secretion of FSH.     

An ultrasound-aided study of temporal relationships between the patterns of LH/FSH secretion, development of ovulatory-sized antral follicles and formation of corpora lutea in ewes

To characterize the pulsatile secretion of LH and FSH and their relationships with various stages of follicular wave development (follicles growing from 3 to > or =5 mm) and formation of corpora lutea (CL), 6 Western white-faced ewes underwent ovarian ultrasonography and intensive blood sampling (every 12 min for 6 h) each day, for 10 and 8 consecutive days, commencing 1 and 2 d after estrus, respectively. Basal serum concentrations of LH and LH pulse frequency declined, whereas LH pulse duration and FSH pulse frequency increased by Day 7 after ovulation (P<0.05). LH pulse amplitude increased (P<0.05) at the end of the growth phase of the largest ovarian follicles in the first follicular wave of the cycle. The amplitude and duration of LH pulses rose (P<0.05) 1 d after CL detection. Mean and basal serum FSH concentrations increased (P<0.05) on the day of emergence of the second follicular wave, and also at the beginning of the static phase of the largest ovarian follicles in the first follicular wave of the cycle. FSH pulse frequency increased (P<0.05) during the growth phase of emergent follicles in the second follicle wave. The detection of CL was associated with a transient decrease in mean and basal serum concentrations of FSH (P<0.05), and it was followed by a transient decline in FSH pulse frequency (P<0.05). These results indicate that LH secretion during the luteal phase of the sheep estrous cycle reflects primarily the stage of development of the CL, and only a rise in LH pulse amplitude may be linked to the end of the growth phase of the largest follicles of waves. Increases in mean and basal serum concentrations of FSH are tightly coupled with the days of follicular wave emergence, and they also coincide with the end of the growth phase of the largest follicles in a previous wave, but FSH pulse frequency increases during the follicle growth phase, especially at mid-cycle.     

Effects of time after ovariectomy, season and oestradiol on luteinizing hormone and follicle-stimulating hormone secretion in ovariectomized ewes.

During the breeding season, five groups of three ewes were implanted at ovariectomy with 0.36, 0.5, 1.0 and 6.0 cm oestradiol implants or implants containing no steroid. Eleven days after receiving implants, blood samples were taken every 10 min for 6 h; implants were then removed. Treatments were repeated three times during each of two consecutive breeding seasons and four times during the intervening anoestrus. In ovariectomized ewes without steroid treatment, luteinizing hormone (LH) pulse frequency increased from early to mid-breeding season, decreased to a minimum at mid-anoestrus and increased to reach a maximum at the mid-point of the second breeding season, subsequently declining. LH pulse amplitude was inversely related to frequency. Basal serum LH concentrations decreased gradually from the first breeding season to reach a minimum at mid-anoestrus and gradually increased to reach a maximum at the end of the second breeding season. Mean serum LH and follicle-stimulating hormone (FSH) concentrations were higher at the end of the second breeding season compared with the beginning of the first breeding season. All parameters of gonadotrophin secretion were decreased much more by oestradiol during the anoestrus than during the breeding season. LH pulse frequency was decreased during anoestrus and at high oestradiol concentrations during the first breeding season. Apart from LH pulse amplitude, the decreases in all parameters of gonadotrophin secretion were less during the second compared with the first breeding season. The minimum effective dose of oestradiol required to decrease mean and basal serum concentrations of LH during anoestrus was lower than in the breeding season. The minimum effective dose of oestradiol required to decrease mean serum concentrations of FSH was lower in the first compared with the second breeding season. Oestradiol depression of LH pulse amplitude and mean serum concentrations of LH and FSH showed a dose dependency during the breeding season. During anoestrus dose dependency was seen for basal concentrations of LH and mean serum concentrations of LH and FSH. We conclude that significant chronic changes in gonadotrophin secretion occur in the ewe with time after ovariectomy. Sensitivity to oestradiol also changes, and the effects of oestradiol are not always dose dependent. We suggest that the circannual pattern of LH pulse frequency and basal LH secretion are directly linked to the circannual cycle of photoperiod.(ABSTRACT TRUNCATED AT 400 WORDS)     

Effects of ovine follicular fluid on plasma LH and FSH secretion in ovariectomized ewes to indicate the site of action of inhibin

Ovariectomized ewes were given 2 ml s.c. injections of ovine follicular fluid (oFF) (N = 3) or serum (N = 3) and blood samples were collected each day for 3 days. Follicular fluid caused a significant (P less than 0.005) reduction in FSH within 1 day, but did not affect mean LH values. Two groups of 3 ewes were treated as above but sampled intensively (each 10 min for 6 h) on Days 1 (before treatment) and 4; mean plasma FSH concentration and plasma LH pulse frequency and amplitude were ascertained. Significant (P less than 0.005) reduction of FSH concentration was seen in the oFF-treated ewes. A non-specific reduction in LH pulse amplitude, but not pulse frequency, was noted in the control ewes. This experiment was repeated with 2 groups of 4 ewes that were conditioned to the experimental environment and effects on LH secretion were not observed in the controls given serum. Treatment with oFF caused a 70% reduction (P less than 0.005) in plasma FSH and a small (30%) but significant (P less than 0.005) reduction in mean LH concentrations. The latter was probably associated with a reduction in LH pulse amplitude in 3/4 animals (N.S.) with no change in LH pulse frequency. Treatment with oFF, as in Exp. 1, caused a 95% reduction in FSH values and significant (P less than 0.01) reduction (32%) of LH pulse amplitude in ovariectomized ewes that had been subjected to hypothalamo-pituitary disconnection and in which gonadotrophin secretion was reinstated with pulses of 250 ng GnRH every 2 h. These results suggest that proteins from the sheep follicular fluid, including inhibin, act at the pituitary level to inhibit FSH secretion and may have some effects on LH pulse amplitude.     

Intracerebroventricular administration of ghrelin rapidly suppresses pulsatile luteinizing hormone secretion in ovariectomized rats.

Ghrelin, an endogenous growth hormone (GH) secretagogue, is shown to increase food intake, which action is similar to that of orexin, also a hypothalamic peptide. Since orexin suppresses pulsatile LH secretion in ovariectomized (OVX) rats, the present study was undertaken to investigate whether ghrelin also suppresses LH secretion. Effects of intracerebroventricularly injected ghrelin (0.1 nmol/0.3 microl) were examined in OVX rats treated with a small dose of 17beta-estradiol (E(2)). After ghrelin injection, pulsatile LH secretions which were ongoing in these E(2)-treated OVX rats were significantly suppressed for about 1 h, whereas GH secretion increased, peaking at 30 min. The main parameter suppressed by ghrelin was the pulse frequency, not the pulse amplitude, suggesting the hypothalamus as the site of ghrelin action. This study provides evidence that ghrelin acts not only in the control of food intake but also in the control of LH secretion.     

Different neuroendocrine systems modulate pulsatile luteinizing hormone secretion in photosuppressed and photorefractory ewes.

The objective of this study was to determine whether two photoperiod regimens that induce anestrus in the ewe-short-day photorefractoriness (SDPR) and long-day photosuppression (LDPS)--act by different neuronal mechanisms. In separate experiments, ovary-intact (INTACT), ovariectomized (OVX), and ovariectomized estradiol-treated (OVX + E) ewes were subjected to three different photoperiodic regimens that resulted in reproductive quiescence: (1) exposure to long days (16L:8D), which caused photosuppression (INTACT, n = 9; OVX, n = 6; OVX + E, n = 5; (2) prolonged exposure to short days (10L:14D)), which caused photorefractoriness (INTACT, n = 10; OVX, n = 6; OVX + E, n = 5); (3) exposure to natural photoperiod, which induced seasonal anestrus (INTACT, n = 11; OVX, n = 6; OVX + E, n = 5). Effect of photoregimen was monitored by measuring progesterone or LH. Drug challenges were made after two sequential estrous cycles were missed in INTACT ewes, after mean LH concentrations dropped below 1 ng/ml in OVX + E ewes, and after LH interpulse intervals increased in OVX ewes. Effects of drug on LH pulse pattern were determined by taking blood samples at 12-min intervals for 8 h after i.v. diluent injection; then for 8 h after i.v. injection of cyproheptadine, a serotonin antagonist (3 mg/kg); and again 7 days later after i.v. injection of diluent or pimozide, a dopamine antagonist (0.25 mg/kg). Cyproheptadine had little effect except to decrease (p = 0.05) mean LH in INTACT anestrous ewes and decrease (p less than 0.01) pulse amplitude in OVX + E SDPR ewes. Pimozide did not affect LH pulse frequency in LDPS ewes. However, pimozide increased LH pulse frequency (p less than 0.005) and mean concentrations (p less than 0.005) in SDPR OVX + E ewes, whereas it suppressed LH pulse frequency (p less than 0.05) and amplitude (p less than 0.03) in SDPR INTACT and SDPR OVX ewes. The results suggest that (1) the role of the dopaminergic system differs in SDPR and LDPS ewes, and that different neuronal systems may effect SDPR and LDPS, (2) the effect of pimozide in SDPR ewes is altered by ovarian steroids, and (3) the serotonergic system has relatively little role in regulating pulsatile LH secretion in any of the three different states of anestrus.     

Central actions of neuropeptide-Y may provide a neuromodulatory link between nutrition and reproduction.

Central injection of neuropeptide-Y (NPY) has been shown to attenuate secretion of LH in ovariectomized rats, rabbits, and monkeys. Several investigators have reported elevated concentrations of NPY in the central nervous system of undernourished animals. The relationship between nutrition and reproduction positions NPY as a potential neuromodulator involved in nutritionally induced changes in secretion of LH. Three experiments were conducted with the following objectives: 1) to examine the effects of NPY on secretion of LH in ovariectomized (OVX) ewes and the influence of estradiol-17 beta (E) on these effects; 2) to determine whether NPY may act through direct effects on the pituitary to influence secretion of LH; and 3) to determine changes in concentrations of NPY in laterocerebro-spinal fluid (CSF) of food-restricted ewes compared to well-fed ewes. In Experiment 1, OVX ewes with s.c. implants of E (OVX + E, n = 4) or no steroid treatment (OVX, n = 4) were fitted with intracerebroventricular (i.c.v.) and jugular cannulae. One of 4 doses of porcine NPY (pNPY; 0, 0.5, 5, or 50 micrograms) was injected i.c.v. and blood samples were collected every 10 min for 4 h prior to and following i.c.v. injection. Blood serum was assayed for LH. The experiment was replicated four times such that each ewe received each dose of pNPY. Mean concentrations of LH as well as frequency and amplitude of pulses of LH were attenuated in response to i.c.v. injection of pNPY in a dose-related manner in both OVX and OVX + E ewes.(ABSTRACT TRUNCATED AT 250 WORDS)     

Pituitary receptors for gonadotropin-releasing hormone in relation to changes in pituitary and plasma luteinizing hormone in ovariectomized-hypothalamo pituitary disconnected ewes. I. Effect of changing frequency of gonadotropin-releasing hormone pulses

Studies were undertaken to determine if changes in the amplitude of luteinizing hormone (LH) pulses that occur in response to changes in the frequency of gonadotropin-releasing hormone (GnRH) pulses are due to an alteration in the number of GnRH receptors. Ewes were ovariectomized (OVX) and the hypothalamus was disconnected from the pituitary (HPD). Ewes were then given pulses of GnRH at a frequency of 1/h or 1/3 h. Two control groups were included: OVX ewes not subjected to HPD, and HPD ewes that were not OVX. At the end of one week of treatment, blood samples were collected to determine the amplitude of LH pulses. The treated ewes were killed just before the next scheduled pulse of GnRH, and the content of LH and number of GnRH receptors were measured in each pituitary. The amplitude of LH pulses was highly correlated with the amount of LH in the pituitary gland (r = 0.71, p less than 0.01), and both LH content and pulse amplitude (mean + SEM) were higher in ewes receiving GnRH once per 3 h (189.7 +/- 39.3 microgram/pituitary, 10.3 +/- 1.1 ng/ml, respectively) than in ewes receiving GnRH once per h (77.8 +/- 11.4 microgram/pituitary, 5.2 +/- 1.3 ng/ml). The pituitary content of LH was highest in the OVX ewes (260.2 +/- 57.4 micrograms/pituitary) and lowest in the nonpulsed HPD ewes (61.7 +/- 51.2 micrograms/pituitary). The number of GnRH receptors was similar in all groups, and was not correlated with any other variable.(ABSTRACT TRUNCATED AT 250 WORDS)     

Pulsatile plasma profiles of FSH and LH before and during medroxyprogesterone acetate treatment in the bitch

The aim of this study was to investigate the effect of medroxyprogesterone acetate (MPA) on pulsatile secretion of gonadotropins in the bitch. Five intact Beagle bitches were treated with MPA in a dose of 10mg/kg body weight subcutaneously at intervals of 4 weeks for a total of 13 injections, starting during anestrus. The 6-h plasma profiles of luteinizing hormone (LH) and follicle stimulating hormone (FSH) were determined before, and 3, 6, 9, and 12 months after the start of MPA treatment. After 6 months of MPA treatment basal plasma LH concentration was transiently increased significantly. Basal plasma FSH concentration and the area under the curve above the zero level (AUC0) for FSH were significantly higher after 3 months of MPA treatment than before or after 9 and 12 months of treatment. MPA treatment did not significantly affect pulse frequency, pulse amplitude, or AUC above the baseline for either LH or FSH. During treatment 58 significant LH pulses were identified, and although each LH pulse coincided with an increase in plasma FSH concentration, in 17 cases the amplitude of the increase was too small to be recognized as a significant FSH pulse. In conclusion, MPA treatment did not suppress basal plasma gonadotropin levels in the bitches. On the contrary, it caused a temporary rise in the basal concentration of both FSH and LH, which may have been due to a direct effect of MPA on the ovary. In addition, several LH pulses were not accompanied by a significant FSH pulse, suggesting that MPA treatment attenuated the pulsatile pituitary release of FSH.     

Naloxone enhances LH but not FSH release during various phases of the estrous cycle in the ewe

Suffolk x whiteface ewes were infused with 0.5 mg/kg/hr naloxone hydrochloride (NAL) for 6 hrs during the early, mid and late luteal and early follicular phases of the estrous cycle. Basal serum luteinizing hormone (LH) concentration was increased by NAL during each trial in the luteal phase and LH pulse amplitude was proportionately increased by 158%, 164% and 350% during the early luteal, mid luteal and early follicular phases, respectively. The apparent NAL induced increase (92%) in LH pulse amplitude during the late luteal phase was not significant. NAL only affected LH pulse frequency during the early follicular phase, when it was decreased. Mean serum follicle stimulating hormone (FSH) concentration was not affected by NAL. The results of this study indicate that endogenous opioid peptides (EOPs) may partially mediate the suppressive influence of estradiol-17 beta (E2) on LH pulse amplitude and also the stimulatory effect of E2 on LH pulse frequency in the early follicular phase. The data may suggest that NAL enhances the amplitude of pulses of gonadotropin releasing hormone (GnRH) by counteracting E2 inhibitory effects on LH release at the level of the pituitary. Alternately, some component of E2 feedback may be an EOP mediated component at the level of the hypothalamus.     

Modulatory role of substance P on gonadotropin and prolactin secretion in the rabbit.

Substance P (SP) is present in large quantities in the brainstem and hypophysiotropic areas of the brain, but its roles in gonadotropin and prolactin secretion are controversial. The aim of this study was to measure luteinizing hormone (LH), follicle-stimulating hormone (FSH), and prolactin (PRL) release from the pituitary after either intracerebroventricular (ICV) injection or infusion of SP or its C- and N-terminal fragments in intact (INT) and ovariectomized (OVX) conscious rabbits. A single injection of SP into the 3rd cerebral ventricle (3CVT) in INT and OVX rabbits augmented plasma LH concentrations, especially when SP was applied during the initial phase of an LH peak. Injection of SP during the declining phase of LH release was not effective. Injection of SP into the 3CVT was followed by increased plasma PRL concentrations in OVX but not in INT rabbits. Both SP 1-11 and SP 1-7 failed to alter LH, FSH, and PRL secretion when the peptides were slowly infused into the 3CVT, although ICV infusion of SP 6-11 did cause a delayed increase in LH release. The results support a stimulatory role of SP on LH and prolactin release. The results further indicate that although the stimulatory effect of SP on LH is ovarian steroid-independent, in the absence of ovarian steroids, SP is stimulatory only during the rising phase of an LH pulse. A dual role of SP-ergic transmission in modulating LH secretion is discussed.     

Neuroendocrine control of FSH secretion: IV. Hypothalamic control of pituitary FSH-regulatory proteins and their relationship to changes in FSH synthesis and secretion

The current dogma is that the differential regulation of luteinizing hormone (LH) and follicle-stimulating hormone (FSH) synthesis and secretion is modulated by gonadotropin-releasing hormone (GnRH) pulse frequency and by changes in inhibins, activins, and follistatins both at the pituitary and at the peripheral level. To date no studies have looked at the overlapping function of these regulators in a combined setting. We tested the hypothesis that changes in GnRH pulse frequency alter the relative abundance of these regulators at the pituitary and peripheral levels in a manner consistent with changes in pituitary and circulating concentrations of FSH; that is, an increase in FSH will be accompanied by increased stimulatory input (activin) and/or reduced follistatin and inhibin. Ovariectomized ewes were subjected to a combination hypothalamic pituitary disconnection (HPD)-hypophyseal portal blood collection procedure. Hypophyseal portal and jugular blood samples were collected for a 6-h period from non-HPD ewes, HPD ewes, or HPD ewes administered GnRH hourly or every 3 h for 4 days. In the absence of endogenous hypothalamic and ovarian hormones that regulate gonadotropin secretion, 3-hourly pulses of GnRH increased pituitary content of FSH more than hourly GnRH, although these differences were not evident in the peripheral circulation. The results failed to support the hypothesis in that the preferential increase of pituitary content of FSH by the lower GnRH pulse frequency could be explained by changes in the pituitary content of inhibin A, follistatin, or activin B. Perhaps the effects of GnRH pulse frequency on FSH is due to changes in the balance of free versus bound amounts of these FSH regulatory proteins or to the involvement of other regulators not monitored in this study.     

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.     

Effect of transport on pulsatile LH release in ovariectomized ewes with or without prior steroid exposure at different times of year

The initial aim of the present study was to test whether the stress of transport suppresses LH pulsatile secretion in ewes. In a pilot experiment in the late breeding season, transport resulted in an unexpected response in three out of five transported, ovariectomized ewes pretreated with oestradiol and progesterone. Before transport, seasonal suppression of LH pulses had occurred earlier than anticipated, but LH pulsatility suddenly restarted for the period of transport. This finding was reminiscent of unexplained results obtained in ovariectomized ewes infused centrally with high doses of corticotrophin-releasing hormone after pretreatment with low doses of oestradiol with or without progesterone. Hence, an additional aim of the present study was to examine whether these latter results with corticotrophin-releasing hormone could be reproduced by increasing endogenous corticotrophin-releasing hormone secretion by transport. Subsequent experiments used groups of at least eight ovariectomized ewes at different times of the year with or without prior exposure to steroids to assess whether these unexpected observations were associated with season or the prevailing endocrine milieu. In the mid-breeding season, transport for 4 h in the absence of steroid pretreatment for 8 months reduced LH pulse frequency from 7.5 +/- 0.3 to 6.3 +/- 0.4 pulses per 4 h (P < 0.05) and LH pulse amplitude from 2.6 +/- 0.5 to 1.8 +/- 0.3 ng ml-1 (P < 0.05). Similarly, in the mid-breeding season, 34 h after the cessation of pretreatment with oestradiol and progesterone, transport suppressed LH pulse frequency from 6.1 +/- 0.4 to 5.5 +/- 0.3 pulses per 4 h (P < 0.05) with a tendency of effect on amplitude (6.2 +/- 2.7 to 2.61 +/- 0.6 ng ml-1; P = 0.07; note the large variance in the pretransport data). During mid-anoestrus, evidence of a suppressive effect of transport was only observed on LH pulse amplitude (4.7 +/- 0.6 versus 3.0 +/- 0.5 pulses per 4 h; P < 0.05) in ovariectomized ewes that had not been exposed to ovarian steroids for 4 months. Repetition of the pilot experiment with 12 ewes during the transition into anoestrus resulted in one ewe with LH pulses seasonally suppressed but increased by transport; 11 ewes had a distinct pulsatile LH pattern which was decreased by transport in six ewes. In anoestrus, there was no effect of transport on LH pulse frequency or amplitude in intact ewes, or those ovariectomized 2-3 weeks previously, with or without prior oestradiol and progesterone treatment. However, basal concentrations of cortisol were greater in anoestrus than in the breeding season, and the increment in cortisol during transport was similar in anoestrus and the breeding season but greater during the transition into anoestrus (P < 0.05). Progesterone concentrations increased from 0.31 +/- 0.02 ng ml-1 before transport to 0.48 +/- 0.05 ng ml-1 during the second hour of transport (P < 0.05). In conclusion, transport reduced LH pulse frequency and amplitude in ovariectomized ewes that had not been exposed to exogenous steroids for at least 4 months. In most animals, the previously observed increase in LH pulsatility induced by exogenous CRH was not reproduced by increasing endogenous CRH secretion by transport. However, in four ewes, transport did increase LH pulsatility, but only during the transition into anoestrus in ewes with seasonally suppressed LH profiles after withdrawal of steroid pretreatment.     

Ovarian antral follicular dynamics and their associations with peripheral concentrations of gonadotropins and ovarian steroids in anoestrous Finnish Landrace ewes

Daily transrectal ultrasonography of ovaries was done in seven Finn ewes during three 17-day periods from May to July. Blood samples were collected each day for estimation of the serum follicle-stimulating hormone (FSH), oestradiol and progesterone concentrations, and also every 15 min for 6 h, halfway through each period of ultrasonographic examination, to determine the patterns of gonadotropic hormone secretion. Four ewes ceased cycling from March to mid-April (ewes entering anoestrus early) and three in May (ewes entering anoestrus late). In all ewes cyclicity resumed during the period from mid-August to mid-September. The growth of ovarian antral follicles to periovulatory sizes of >/=5 mm in diameter was seen at all stages of anoestrus. An average of four waves of follicular development (follicles growing from 3 to >/=5 mm in diameter before regression) with a periodicity of 4 days were recorded during each of the three scanning periods. There was a close temporal relationship between days of follicular wave emergence and peaks of successive FSH fluctuations. Ewes entering anoestrus late exceeded ewes that became anoestrus early in numbers of large (>/=5 mm in diameter) ovarian antral follicles and maximum follicle diameter. Peak concentrations of transient FSH increases were higher (P<0.05) in ewes entering anoestrus late than in ewes entering anoestrus early. The secretion of luteinising hormone, (LH; mean and basal level, and LH pulse frequency, but not amplitude) was lowest during the month of June in all ewes. Oestradiol production was markedly suppressed throughout anoestrus. Peaks of progesterone secretion appeared to occur at regular intervals and were associated with the end of the growth phase of the largest follicles of sequential waves. In conclusion, the growth of ovarian follicles to ostensibly ovulatory diameters is maintained throughout anoestrus in Finn ewes and periodic emergence of follicular waves is correlated with an endogenous rhythm of FSH secretion. The present study also provides evidence for the inverse relationship between the time of the onset of seasonal anoestrus and the number and size of antral follicles developing throughout anoestrus in Finn ewes, and indicates that differences exist in both the secretion of and ovarian responsiveness to gonadotropic hormones among early and late anoestrous ewes.     

Evidence that the corpus luteum of pregnancy contributes to the control of tonic secretion of LH in the ewe

Concentrations of LH and FSH were measured in blood samples collected from the jugular vein at 20-min intervals for 7 h (09:00-16:00 h) on Days 60, 80, 100 and 120 of pregnancy in 5 intact ewes and 5 from which the CL had been excised on Day 70. In the 5 intact ewes, plasma LH concentrations remained low and unchanged between Days 60 and 120. During this period, pulsatile release of LH occurred irregularly and infrequently. Removal of the CL resulted in an increase in the basal values of LH and in the frequency and amplitude of LH pulses. Concentrations of FSH were relatively constant in all stages of pregnancy examined and were similar in both groups of ewes. These results show that (1) LH concentrations are low during the second half of pregnancy; and (2) LH, but not FSH, increases after CL excision, presumably by removing some luteal factor inhibitor of LH secretion.     

Follicle populations,ovulation rates and plasma profiles of LH,FSH and prolactin in Scottish Blackface ewes in high and low levels of body condition

Scottish Blackface ewes in high body condition (mean score = 2.86) had a higher mean ovulation rate (1.8 v. 0.9; P < 0.05) and more large (⪖ 4 mm diameter) follicles (4.6 v 2.2; P < 0.05) than ewes in low condition (mean score = 1.84) but similar numbers of small (1–4 mm diameter) follicles (6.3 v 6.0; NS). There was little difference in LH profiles with body condition but FSH and prolactin concentrations were significantly greater, during both luteal and follicular phases of the cycle, in ewes in high condition.Despite the relationships between body condition and ovulation rate and between condition and hormone concentrations, within the high condition groups, there was no significant difference in FSH levels with ovulation rate. Prolactin levels were higher in ewes with a single ovulation than in ewes with two or three ovulations. There was a trend towards a higher mean LH pulse frequency in the luteal phase and a higher mean LH pulse amplitude in the follicular phase in ewes with multiple ovulations compared with ewes with a single ovulation. During oestrus, only circulating prolactin concentrations differed with body condition, being significantly higher in ewes in high condition, but mean LH concentrations were higher and FSH concentrations lower in ewes with multiple ovulations. Subsequent luteal function, as measured by circulating progesterone concentrations, was normal in all ewes. It is concluded that body condition affected the size of the large follicle (⪖ 4 mm diameter) population through changes in FSH and possibly pulsatile LH secretion and prolactin secretion during the luteal and follicular phases of the cycle and that the number of follicles that were potentially ovulatory was probably determined during the luteal phase of the cycle. However, their ability to undergo the final stages of development and to ovulate may be related to the amount of LH secreted during the follicular phase.