Effect of exogenous prolactin and bromocriptine on seasonal reproductive quiescence in the Bennett's wallaby (Macropus rufogriseus rufogriseus)

Two experiments were performed to investigate whether prolactin blocks the reactivation of the corpus luteum during seasonal reproductive quiescence in the Bennett's wallaby. In the first experiment three groups of non-lactating females (groups A C) were subjected in mid-April to photoperiods corresponding to those of the summer solstice from day 0 to day 13 of the experiment. Between days 14 and 52, photoperiods were reduced to correspond to those of the winter solstice (groups B and C). Animals in group A were maintained on the long photoperiods. Two milligrams ovine prolactin (groups A and C) or vehicle (group B) were administered on the mornings of days 14-22. In group A, no animal showed evidence of an active corpus luteum based on increased plasma progesterone levels. All animals in groups B and C exhibited reactivation of the corpus luteum. In group C, reactivation was significantly (P less than 0.01) delayed by a mean of 6.3 days. In the second experiment, two groups of nonlactating female wallabies in 'seasonal quiescence' were injected daily for 7 days with either 60 mg of the dopamine agonist bromocriptine or vehicle. The corpora lutea did not reactivate in either group. We conclude that exogenous prolactin is able to block the effect of short photoperiods in reactivating the quiescent corpus luteum during seasonal quiescence. However, the absence of an effect of bromocriptine suggests that if prolactin is the endogenous hormone responsible for maintaining seasonal quiescence it may not be under dopaminergic control at this time of year.     

Effects of lactation and season on plasma prolactin concentrations and response to bromocriptine during lactation in the Bennett's wallaby (Macropus rufogriseus rufogriseus)

Prolactin concentration was measured in plasma collected each week for 13 months from lactating and non-lactating Bennett's wallabies (Macropus rufogriseus rufogriseus). In non-lactating animals, prolactin concentrations decreased towards the end of the study but such changes did not appear to fit a seasonal pattern. Prolactin concentrations were low during early lactation and at a similar level to non-lactating animals, increased significantly during late pouch life (February-May), and then returned to non-lactating levels at a time coincident with permanent exit of the joey from the pouch. Temporary removal of joeys from their mothers in April was followed by a rapid decline in prolactin concentrations which remained low for 24 h until the joey was returned to its mother, whereupon prolactin concentrations increased significantly within 2 h. The effect of a single injection of bromocriptine (5 mg/kg) on lactation, embryonic diapause and plasma prolactin concentrations was examined at two stages of lactation. In November (lactational diapause), bromocriptine had no effect on prolactin concentrations but two out of four suckling joeys died on days 13 and 14 after treatment, and three out of four females gave birth on days 27, 27 and 28. Bromocriptine treatment in April (seasonal diapause) was followed by a significant reduction in prolactin concentrations and reduced growth rate of joeys belonging to treated females. New births were not observed. In view of the effect of bromocriptine on plasma prolactin concentrations in late lactation and the demonstration that domperidone (a dopamine antagonist) significantly increases plasma prolactin concentrations, it would seem that dopamine can act as a prolactin inhibitory hormone in this as in other mammalian species.     

Experimental manipulations of prolactin following removal of pouch young or bromocriptine treatment during lactational quiescence in the Bennett's wallaby

Experiments were conducted to investigate whether prolactin suppresses the corpus luteum during lactational quiescence in the Bennett's wallaby. In the first experiment, pouch young were removed from lactating wallabies (day 0) which were then treated daily for 7 days with either saline, or 8 mg domperidone or 2 mg ovine prolactin. In the saline-injected animals there was a transient peak in progesterone concentrations on day 4 and birth on day 28. The transient progesterone peak and births were significantly (P less than 0.01) delayed by 5 and 8 days in animals treated with domperidone and ovine prolactin respectively. In the second experiment, four groups of lactating wallabies were treated on day 0 with either 60 mg bromocriptine (groups C and D) or the vehicle (groups A and B). On days 0-6, groups B and D were injected daily with 2 mg ovine prolactin while groups A and C received the vehicle. In group C, three pouch young died 14-29 days after administration of bromocriptine, and there was a transient rise in progesterone on day 4 in all animals, indicating that bromocriptine resulted in immediate reactivation of the quiescent corpus luteum. New births occurred in two animals on day 28. In group D, which received bromocriptine followed by ovine prolactin for 7 days, all the original pouch young remained alive at the end of the experiment. Four of the animals from this group showed a transient progesterone peak on day 11, with births in two animals on days 35 and 36 indicating that the effects of bromocriptine were prevented whilst ovine prolactin was being administered.(ABSTRACT TRUNCATED AT 250 WORDS)     

Metabolic effects of infusion of cortisol and adrenocorticotrophin in the tammar wallaby (Macropus eugenii Desmarest).

Infusion of cortisol (1 mg/kg of body weight infused over 1 hr) into the tammar wallaby, Macropus eugenii Desmarest, was followed by an immediate increase in plasma corticosteroid concentration. Plasma glucose concentration began to rise 45 min later and remained above control levels for 3 hr. There were no changes in the concentrations of amino acids, free fatty acids, or urea in the plasma. It is concluded that cortisol affects plasma glucose concentration directly rather than via protein catabolism. Intravenous infusion of 0.45 IU of porcine ACTH/kg of body weight over 1 hr was followed by an immediate increase in plasma corticosteroid concentration and a rise in plasma glucose level 15 min later. The plasma glucose response to ACTH was advanced about 1 hr compared with that to cortisol. It is concluded that the effects of ACTH cannot be mediated wholly by an increase in plasma corticosteroid concentration. There were no changes in the concentration of amino acids or urea in the plasma, but the free fatty acid concentration was increased after ACTH infusion. This is consistent with the lipolytic effects of ACTH in other mammalian species.     

Seasonal patterns of circulating progesterone and prolactin and response to bromocriptine in the female tammar Macropus eugenii

Concentrations of prolactin and progesterone in the plasma of female tammars (Macropus eugenii) were measured during lactational quiescence and seasonal quiescence and during the period of natural resumption of reproduction after the summer solstice in December. Prolactin concentrations were consistently low (less than 40 ng/ml) during the period of declining day length and consistently elevated (greater than 40 ng/ml) during the period of increasing day length. Basal levels of progesterone were lowest (118.9 +/- 9.1 pg/ml) at the winter solstice and highest (244.1 +/- 21.0 pg/ml) at the summer solstice. Treatment with bromocriptine (CB154) did not depress prolactin levels at either time of the year but during February to June a single injection of 5 mg/kg body wt induced development of the quiescent corpus luteum (CL) and the diapausing embryo was reactivated. In February and March the level of progesterone increased in association with the developing CL, but after treatment at the winter solstice in June plasma progesterone did not increase although pregnancy was successfully completed. From September through November only 1 of 50 females responded to bromocriptine. None responded to bromocriptine given at the summer solstice in December, but all these animals spontaneously reactivated 2 weeks later. The difference in response of female tammars to bromocriptine treatment in the two halves of the year suggests that different endocrine controls operate in lactational and seasonal quiescence and that the latter is more complex.     

Direct effects of prolactin and dopamine on the gonadotroph response to GnRH

The intrapituitary mechanisms underlying the inhibitory actions of hyperprolactinaemia on the reproductive axis remain unclear. Previous work on primary pituitary cultures revealed combined suppressive effects of prolactin (PRL) and dopamine on the gonadotrophin response to GnRH. However, whether these effects occur directly at the level of the gonadotroph and are accompanied by changes in gene expression is still unresolved. Here, alphaT(3)-1 and LbetaT2 cells were used to investigate the effects of PRL and dopamine on gonadotrophin synthesis and release in gonadotroph monocultures under basal and GnRH-stimulated conditions. PRL receptor and dopamine receptor mRNA expressions were first determined by RT-PCR in both cell lines. Then, PRL and the dopamine agonist bromocriptine (Br), alone or in combination, were shown to block the maximal alpha-subunit and LHbeta-subunit mRNA responses to a dose-range of GnRH. The LH secretory response was differentially affected by treatments. GnRH dose-dependently stimulated LH release, with a 4-5 fold increase at 10(-8) M GnRH. Unexpectedly, PRL or Br stimulated basal LH release, with PRL, but not Br, enhancing the LH secretory response to GnRH. This effect was, however, completely blocked by Br. These results reveal direct effects of PRL and dopamine at the level of the gonadotroph cell, and interactions between these two hormones in the regulation of gonadotrophin secretion. Moreover, uncoupling between LH synthesis and release in both the basal and the GnRH-stimulated responses to PRL and dopamine was clearly apparent.     

Components of the melatonin message in the response to photoperiod of the tammar wallaby (Macropus eugenii)

Female tammars experiencing long-day photoperiod (LD 15:9) are in a reproductive state termed seasonal quiescence. After a change to LD 12:12, a sequence of endocrine events, dependent on a melatonin message being interpreted during the next 3 days, leads to the disappearance of a morning pulse of prolactin on day 4, and to reactivation of the quiescent corpus luteum by day 8. Elements of the message were investigated in three experiments. In experiment 1, tammars on LD 15:9 were injected with melatonin 2.5 hr before lights out on 0-5 successive days. All those receiving four or five daily injections, and six of ten receiving two or three injections, reactivated. In experiment 2, tammars on LD 15:9 were injected with melatonin or oil on 5 consecutive days to achieve a lengthening of the period of elevated melatonin from 9.0 hr to 9.5 hr, 10 hr, 10.5 hr, and 11.5 hr respectively. One of five tammars experiencing 10 hr and seven of ten experiencing the longer periods reactivated. In experiment 3, tammars on LD18:6 received melatonin 6 hr, 6 + 3 hr or 3 hr before lights out for ten successive days. Between injections the melatonin concentrations returned to basal levels. The treatments thus provided a skeletal increase in the profile of melatonin from 6 to 9 hr or 6 to 12 hr. All those treated at 6 + 3 hr reactivated and three of five treated at 6 hr only reactivated. We conclude that the tammar stores information about the change in the maximum duration of elevated melatonin experienced each night and, after three successive periods in which the duration is greater than 10.5 hr, it reactivates. Since reactivation cannot occur until completion of the third dark phase, the processing center must be active during the fourth day, prior to the abolition of the prolactin peak.     

Thyrotropin and prolactin response to intraspinal TRH administration in man.

The effect of intraspinal (i.s.) TRH administration of Prolactin (Prl) and thyrotropin stimulating hormone (TSH) serum levels was studied in order to verify the existence of a ventricular route in man for releasing factor delivery to the anterior pituitary, which has been previously reported in rats. Ten young male subjects were given 200 microgram thyrotropin releasing hormone (TRH) i.s. injections and Prl and TSH were measured by radioimmunoassay (RIA) before and at various times after TRH administration. In the same subjects, an i.v. TRH test was also performed. After i.s. TRH, a prompt Prl increase (peak values at 10-30 min and return to baseline within 150 min) and a delayed increase (3-5 h following TRH injection) were observed in 7 and 5 subjects respectively, while an early elevation in serum TSH occurred in 6 subjects and a late one in other 6. In two subjects, a biphasic response of both tropins was present. Prl and TSH response to i.v. TRH was within the normal range in all cases; no late rise of the 2 hormones was observed. A kinetic experiment with 125I-TRH was also carried out to elucidate the mode of i.s. vs i.v. TRH action. These results confirm in man data reported in animals which suggest that TRH can be transported from the cerebrospinal fluid (CSF) to the portal system and the hypophysis.     

Heterogeneity of prolactin and TSH response to TRH in hypotonadotropic hypogonadism.

To evaluate prolactin and TSH secretion in isolated gonadotropin deficiency, thyrotropin-releasing hormone (TRH), in a dose of 500 microgram, was administered iv to fifteen male subjects with this disorder. In 4 out of 8 untreated patients, TRH-mediated prolactin release was significantly blunted and this attenuated response was not improved in one patient after treatment with testosterone for 1 year. In 7 patients who were treated with testosterone for 2 to 8 years, four failed to show a normal prolactin response after TRH injection. TRH-induced TSH secretion, on the other hand, was attenuated in two out of 8 untreated and in two of 8 treated patients with hypogonadotropic hypogonadism. The decreased TSH reserve was not necessarily associated with the poor prolactin response to TRH. It was concluded that heterogeneity exists in TRH-mediated prolactin and TSH release in "isolated" gonadotropin deficiency syndrome.     

Impaired prolactin responsiveness to a dopamine antagonist in long-term survivors of acute lymphoblastic leukaemia

Cranial irradiation is known to affect the neuroendocrine control of GH secretion. The aim of the present study was to clarify the effect of leukaemia treatment on neuroendocrine control of PRL secretion in long-term survivors of acute lymphoblastic leukaemia (ALL). Pituitary LH, FSH and PRL responses to GnRH and to the dopamine receptor antagonist metoclopramide were evaluated in 18 boys 1.6-9 years after discontinuation of medication for ALL. All the boys had been subjected to prophylactic cranial irradiation and 8 to testicular irradiation. Eight of the boys had supranormal gonadotropin levels either basally or after GnRH. All had normal basal PRL levels, but in 14 of the 18 boys the metoclopramide-releasable PRL levels were subnormal. The decreased PRL responses cannot be explained by the hypergonadotropism, since exaggerated rather than decreased responses are frequently found in patients with primary gonadal failure. We conclude that abnormal regulation of PRL secretion is a frequent consequence of treatment of ALL, probably being due to decreased pituitary lactotrop mass.     

Changes of pituitary thyrotropin releasing hormone (TRH) receptor level and prolactin response to TRH during the rat estrous cycle.

The plasma PRL and TSH responses to TRH injected iv at different stages of the estrous cycle in normal rats under Surital anesthesia were maximal during the afternoon of proestrus and morning of estrus and lowest on diestrus I. As calculated from the areas under the plasma response curves, a 10-fold difference was found between the maximal and minimal PRL responses while a 2-fold difference was measured for TSH. The plasma PRL and TSH responses to TRH showed a correlation with the binding of [3H]TRH to anterior pituitary gland, a 3-fold difference being observed between the minimal binding measured on the morning of diestrus II and the maximal value found on the evening of proestrus. Contrary to findings with LHRH and LH, repeated injections of a small dose (10 ng) of TRH in the afternoon of proestrus abolished PRL and TSH responses to subsequent injection of the neurohormone.     

Release of thyrotropin and prolactin by a thyrotropin-releasing hormone (TRH) precursor, TRH-Gly: conversion to TRH is sufficient for in vivo effects

The mechanism by which TRH-Gly (pGlu-His-Pro-Gly), a biosynthetic precursor of thyrotropin-releasing hormone (TRH, pGlu-His-Pro-NH2), stimulates pituitary thyrotropin (TSH) and prolactin release has been studied in urethane-anesthetized 2-month-old (250 g) male Sprague-Dawley rats and in vitro with GH3 cells, a rat anterior pituitary tumor cell line. We used specific radioimmunoassays to measure TRH-Gly and TRH levels in rat cortex, hypothalamus, medulla, eyes and whole blood as a function of the intracisternal (IC) dose of TRH and TRH-Gly administered 40 min prior to sacrifice. IC injection of 1.0 mg of TRH-Gly led to a significant (p less than 0.005) increase in the TRH levels in hypothalamus, medulla and blood. The relative potency of IC and intracardiac (IK) TRH and TRH-Gly release of rat TSH was compared by radioimmunoassay and further refined using estimates based on in vivo kinetics of TRH-Gly alpha-amidation. The binding of TRH-Gly to the plasma membrane receptors for TRH on GH3 cells was also investigated. In regard to TSH release, TRH-Gly given IC had only 0.042% of the potency of TRH given IC and was consistent with its rate of IC alpha-amidation. IK TRH-Gly had 0.16% of the potency of IK TRH of TSH release and was also consistent with its rate of intravascular conversion to TRH. The mean peak TSH response occurred at 20 min after IC TRH-Gly or IC TRH injection but the post-peak decline was slower for IC TRH-Gly.(ABSTRACT TRUNCATED AT 250 WORDS)     

Changes in the prolactin response to thyrotropin-releasing hormone (TRH) during the menstrual cycle of normal women.

Serum prolactin (PRL) and thyrotropin (TSH) levels were measured after iv administration of 200 microng of synthetic thyrotropin-releasing hormone (TRH) in 20 normal women ages 18 to 34. Ten women received TRH on days 7 to 8 of the menstrual cycle and 10 women received TRH on days 21-22. Although there was no difference in the dose of TRH relative to body weight in the two groups of women, the peak PRL level after TRH stimulation was greater in the women studied on day 21-22 (48.5+/-5.7 ng/ml, mean+/-SE) than on day 7-8 (35.2+/-4.2 ng/ml) of the cycle (P less than 0.05). In contrast, TSH rose to a greater degree in the preovulatory phase (13.8+/-1.8 micronU/ml) than the luteal phase (7.7+/-0.7 micronU/ml of the cycle (P less than .01). Studies of the PRL and TSH response after TRH administration should take the phase of the menstrual cycle into account.     

Fibroblast growth factor-2 and epidermal growth factor modulate prolactin responses to TRH and dopamine in primary cultures

Fibroblast growth factor-2, (FGF-2) and epidermal growth factor (EGF) are expressed in most tissues of the organism including pituitary, FGF-2 increases PRL levels and PRL mRNA in GH3 cells and primary cultures, and it has been involved in the lactotroph proliferation and hyperplasia. EGF also increases PRL levels in vitro. However, the effects of these two factors in the responses of lactotroph cells to TRH and dopamine (DA) remain to be clarified. In the present work we have studied the modulator activity of FGF-2 and EGF on in vitro PRL in responses to TRH and DA in primary cultures from in vivo vehicle-or estrogen (E2)-treated rats. We have found that FGF-2 (2×10⁻¹¹ M) prevents the EGF-induced dose-dependent increase in PRL levels in control cells, and reversed the EGF-stimulating effects in cells from E2-treated rats. Both FGF-2 (2×10⁻¹¹ M) and EGF (6.6×10⁻⁹ M) significantly increase (>30% and >120%, respectively) the PRL levels in response to TRH (10⁻⁶ 10⁻⁵ M). FGF-2 blocked the inhibitory effects of low doses of DA (10⁻⁹ M). EGF was unable to do so, although markedly increased (>200%) the post-DA PRL rebound. In cells from in vivo E2-treated rats, FGF-2 increased (>50%) the PRL secretion in response to TRH, while EGF reduced responses to high doses of TRH (10⁻⁶, 10⁻⁵ M). In addition, FGF-2 reversed and EGF increased the inhibitory effects of DA. Both FGF-2 and EGF completely blocked the post-DA PRL rebound, in these cells. Taken together our data suggest that FGF-2 and EGF are important regulators of lactotroph responsiveness to TRH and DA in vitro, although their actions are highly dependent on estrogenic milieu.     

Effects of the estrous cycle stage on the prolactin secretory response to dopamine in vitro

Dopamine (DA) will both stimulate and inhibit prolactin (PRL) secretion from the anterior pituitary gland in vitro and in vivo. The present study was designed to determine if there are selected times during the estrous cycle of the rat when one function is favored over the other. Anterior pituitary glands collected on diestrus-1 (D1), diestrus-2 (D2), the morning of proestrus (Pro-AM), the afternoon of proestrus (Pro-PM), and estrus (E) were enzymatically dissociated and placed in monolayer culture. On the fourth day in culture, cells were challenged for 10, 20, 30, 60, 120, 180, or 240 min with media alone or media containing either 100 pM or 1 μM DA. The concentration of PRL in the media was determined by radioimmunoassay. Regression analysis revealed that in the absence of DA, PRL secretion from cultured cells differed significantly depending on the stage of the estrous cycle during which they were obtained. Cells obtained during the morning of diestrus-2 secreted PRL at the greatest rate compared to other stages of the cycle. When all stages were compared, the rates of PRL secretion were: D2>E>D1>Pro-AM>Pro-PM (each significantly different from the others,P<0.01). By 20–30 min of exposure to 100 pM DA, the rate of PRL secretion from cells obtained during each stage of the cycle was significantly enhanced. This enhanced secretion persisted in cells obtained during D2 and Pro-PM but was short-lived in cells obtained during other stages. No inhibition of PRL secretion was induced by this dose of DA. PRL secretion was inhibited when treated with 1 μM DA in cells obtained at all stages of the estrous cycle. Inhibition was more prolonged in cells obtained on D1, D2, and Pro-AM. DA was least effective as an inhibitor of PRL secretion in cells obtained during Pro-PM and E. Prior to inhibiting PRL secretion in cells obtained during Pro-PM, 1 μM DA rapidly stimulated PRL secretion. This effect persisted for 60 min. These data suggest that in the absence of DA, the dynamics of PRL secretion from anterior pituitary cells in vitro differ depending on the stage of the estrous cycle during which the cells were obtained. Moreover, the in vivo environment of the cell determines the direction and magnitude of the PRL-secretory response to DA.     

The effects of age,season, and gender on serum cortisol levels in the tammar wallaby,Macropus eugenii

Serum cortisol levels were measured in a total of 73 tammar wallabies maintained in a captive population at Macquarie University, NSW, Australia. Previous studies of corticosteroids in marsupials have generally involved low sample numbers, a diverse array of analytical techniques, and a variety of sampling conditions. We have conducted a substantive, longitudinal study of serum cortisol levels using a radioimmunoassay protocol, and data have been analysed with respect to age, sex, and seasonality. There were no apparent effects of age or sex on serum cortisol levels, although an inverse but non-significant relationship was observed between males and females. However a significant difference in serum cortisol levels was observed between seasons, with mean serum cortisol significantly higher in summer than in autumn. These data will serve as a reference for 'normal' ranges of serum cortisol levels, particularly in the female tammar wallaby. As deviations from these values can indicate compromised animal health and well-being, this information will assist wildlife managers in assessing and monitoring the health status of individuals in captive and free-ranging populations.     

Effects of vasoactive intestinal polypeptide, TRH, and dopamine on prolactin secretion in estrogen-primed postmenopausal women

Prolactin secretion is influenced by at least three important hypothalamic neurotransmitters: TRH, vasoactive intestinal polypeptide and dopamine. The purpose of this study was to determine whether, in estradiol-primed postmenopausal women, the PRL response to TRH, vasoactive intestinal polypeptide, and dopamine differed. Ten postmenopausal women were studied during treatment with estradiol benzoate at a dose of 0.625 mg per day during 15 days. TRH (200 micrograms iv), saline infusion, vasoactive intestinal polypeptide (75 micrograms infused iv during 15 min), coadministration of vasoactive intestinal polypeptide and TRH, and dopamine (4 micrograms.kg-1.min-1 iv for 3 h) were administered for 5 consecutive days before and during the last 5 days of estradiol benzoate treatment. Before estradiol benzoate administration, the PRL, response to TRH was significantly greater than that of vasoactive intestinal polypeptide. Estradiol benzoate treatment increased significantly the PRL release induced by TRH (p less than 0.01), but did not modify the response to vasoactive intestinal polypeptide. At the end of estradiol benzoate treatment, the maximal increase in PRL after the combined (vasoactive intestinal polypeptide + TRH) test was greater than that obtained with TRH alone and occurred earlier (p less than 0.04). Dopamine suppressed PRL secretion to a similar extent after estradiol benzoate treatment.(ABSTRACT TRUNCATED AT 250 WORDS)     

Pituitary response to a dopamine antagonist at different times of the day in normal women

In order to determine whether or not pituitary responsiveness to the dopaminergic antagonist clebopride changes during the nyctohemeral cycle, 10 healthy women with regular cycles were given 1 mg of clebopride orally at 09.00 h and 24.00 h with at least a 5 day interval between each test. In addition, 5 of the women were given a placebo instead of clebopride at midnight to evaluate the spontaneous hormonal changes. During the 24.00 h test the women had significantly higher (P less than 0.05) mean TSH basal levels. Serum prolactin (Prl) increased significantly (P less than 0.001) after clebopride administration while these changes did not occur when placebo was used instead of clebopride at midnight. The Prl response to clebopride was qualitatively similar at 09.00 h and at 24.00 h. Clebopride given at midnight induced a significant increase (P less than 0.05) in serum TSH while this change did not occur when the drug was given at 09.00 h or when placebo was given at midnight. The administration of clebopride resulted in no discernible alternations in serum LH, FSH or GH in either the 09.00 h or the 24.00 h tests. Thus, Prl responses to clebopride were similar in the morning and at midnight, TSH significantly increased after clebopride at midnight whereas this did not occur when the drug was given in the morning, and no significant changes were induced in LH, FSH or GH at the times studied.