Pup removal suppresses estrogen-induced surges of LH secretion and activation of GnRH neurons in lactating rats

During lactation, the suckling stimulus exerts profound influences on neuroendocrine regulation in nursing rats. We examined the acute effect of pup removal on the estrogen-induced surge of LH secretion in ovariectomized lactating rats. Lactating and nonlactating cyclic female rats were given an estradiol-containing capsule after ovariectomy, and blood samples were collected through an indwelling catheter for serum LH determinations. In lactating, freely suckled ovariectomized rats, estrogen treatment induced an afternoon LH surge with a magnitude and timing comparable to those seen in nonlactating rats. Removal of pups from the lactating rats at 0900, 1100, or 1300 h, but not at 1500 h, suppressed the estrogen-induced surge that normally occurs in the afternoon of the same day. The suppressive effect of pup removal at 0900 h was completely abolished when the pups were returned by 1400 h. In contrast, pup removal was ineffective in abolishing the stimulatory effect of progesterone on LH surges. Double immunohistochemical staining for gonadotropin-releasing hormone (GnRH) and c-Fos, a marker for neuronal activation, revealed a decrease, concomitantly with the suppression of LH surges, in the number of c-Fos-immunoreactive GnRH neurons in the preoptic regions of nonsuckled rats. An LH surge was restored in nonsuckled rats when 0.1 microg oxytocin was injected into the third ventricle three times at 1-h intervals during pup removal. These results suggest that the GnRH surge generator of lactating rats requires the suckling stimulus that is not involved in nonlactating cyclic female rats.     

Correlation of rat pituitary prolactin messenger ribonucleic acid and hormone content with serum levels during the estrogen-induced surge

The model of the serum PRL surge generated in the ovariectomized rat after estradiol benzoate (EB) treatment was used to study the relationship between serum and pituitary PRL levels and pituitary PRL mRNA levels. Adult ovariectomized rats were injected sc with 7 micrograms EB or vehicle at noon on day 0. Three days later (day 3), the rats were decapitated every 4 h over a 24-h period (0800 h on day 3 to 0400 h on day 4) for determination of serum and pituitary PRL and GH levels by RIA. In addition, PRL and GH mRNA content was determined using dot blot hybridization with cDNAs. Administration of EB resulted in a significant rise in serum PRL levels at 1200, 1600, and 2000 h on day 3 compared to control values. At other times, serum PRL levels in the EB group were the same as control values. EB treatment also elicited a marked increase in pituitary PRL content at all time periods examined except during (1600 and 2000 h) and after the PRL surge (2400 h on day 3) when there was a significant reduction in stored pituitary PRL. The pituitary PRL mRNA content in the EB-treated group was significantly elevated (4- to 6-fold) over control levels throughout the study. Furthermore, PRL mRNA levels in EB-treated rats were significantly higher at 2000 and 2400 h on day 3 than at other time periods. In contrast to its effects on PRL, EB treatment had a slight inhibitory effect on pituitary GH content at 2000 and 2400 h on day 3 compared to control values; otherwise, this steroid had no effect on serum GH levels and pituitary GH mRNA content. Interestingly, serum GH levels and pituitary GH mRNA content in both treatment and control groups fluctuated in a pattern consistent with circadian rhythms, with peak values occurring during the lights-on hours. These data show that estrogen has a stimulatory effect on pituitary content of PRL and its corresponding mRNA in the rat 3 days after injection. These elevated PRL mRNa levels may be necessary for the occurrence of PRL surges. Furthermore, the facts that serum PRL levels were elevated only at certain times (1200-2000 h on day 3) while PRL mRNA content was increased at all times in the EB-treated rats suggest a differential regulation between PRL release and biosynthesis.     

In vivo LH-RH output of ovariectomized rats following estrogen treatment

In the present experiment we examined the effect of estrogen upon in vivo luteinizing hormone releasing hormone (LH-RH) release from the medial basal hypothalamus of freely moving ovariectomized female rats during the period of the LH surge. Ovariectomized females (10-20 day) received two subcutaneous injections of oil or estradiol benzoate (EB; 25 micrograms/rat) at 48 and 24 h prior to push-pull perfusion and were perfused between 10:00 and 18:00 h on the 3rd, day. Unexpectedly, the activity of the LH-RH pulse generator in ovariectomized rats was characterized by a very low mean LH-RH output. Administration of EB increased the activity of the LH-RH pulse generator as indicated by a significant increase in overall mean LH-RH release of EB- versus oil-treated females. In addition, EB-treated females demonstrated a diurnal variation in LH-RH release with marginal, but significant, increases in LH-RH release during the period of the LH surge (14:10-18:00 h) as compared with levels obtained between 10:00 and 14:00 h. This increase in the overall mean release during the period between 14:10 and 18:00 h appears to be attributable to an EB-induced increase in the frequency of LH-RH release, since the amplitude did not change between the two 4-hour sampling periods.(ABSTRACT TRUNCATED AT 250 WORDS)     

Central administration of antiserum to vasoactive intestinal peptide delays and reduces luteinizing hormone and prolactin surges in ovariectomized, estrogen-treated rats

The present study investigated the role of hypothalamic VIP in the regulation of the LH and PRL surge using immunoneutralization of endogenous VIP in mature ovariectomized (OVX), estradiol benzoate (EB)-treated female Wistar rats. We compared the effect of intracerebroventricular (i.c.v.) injections of a VIP antiserum (VIP-Ab) with that of saline (Ctr) on LH and PRL profiles in two separate groups of rats following two subcutaneous EB injections on days 8 and 9 after OVX. VIP-Ab or Ctr injections were given during the second half of the dark period, i.e. at 22:00 h (day 9), and, in addition, the following morning, i.e. at 08:00 h (day 10), just before the expected onset of the LH surge. Hourly blood samples were collected between 09:00 and 18:00 h on day 10. In addition, we studied the reproducibility of EB-induced LH and PRL surges and compared the effect of Ctr and VIP-Ab treatment on sequential surges in individual OVX females, i.e. 10 and 23 days after OVX, using each animal as its own control. Although we observeda large variation in the height and timing of LH and PRL peak levels between EB-treated females, the characteristics of successive surges of individual rats were highly reproducible. This reproducibility suggests that differences in functioning of the suprachiasmatic nucleus as well as in the response of the hypothalamus to steroid feedback largely explain the normal variation in hormone responses between rats. The VIP-Ab treatment resulted in a significant delay in the time course and a strong reduction of the magnitude of the afternoon LH and PRL surge. When analyzed within individual females, the effect of VIP-Ab treatment was even more pronounced due to a reduction in variability when each animal was used as its own control. These results suggest that hypothalamic VIP is an important regulator of both the timing and the magnitude of the EB-induced LH and PRL surge in the OVX rat, and suggest that its role may be stimulatory in this respect.     

Reproductive experience reduces circulating 17beta-estradiol and prolactin levels during proestrus and alters estrogen sensitivity in female rats

The reproductive experiences of pregnancy, parturition, and lactation affect a range of neural and endocrine processes after the end of lactation. In women, previous parity results in reduced circulating prolactin (PRL) and androgen levels years after giving birth. Reductions in PRL secretion also occur in reproductively experienced, female rats. In the present study we examined the status and regulation of estradiol (E(2)) and PRL during the reproductive cycle after reproductive experience. These hormones regulate one another and have been implicated in a number of disease and aging processes. Using a rat model, the patterns of E(2) and PRL secretion, pituitary PRL content, and estrogen receptor alpha expression were characterized from 1200-1800 h on proestrus in age-matched, primiparous and nulliparous animals. The possible effect of parity on estrogen sensitivity was then examined by challenging nonlactating, ovariectomized, age-matched, multiparous and nulliparous rats with estradiol benzoate (EB; 0, 1, 5, 25, and 125 microg/kg) and measuring PRL responses 24 and 48 h later. Previous parity resulted in modest, yet significant, reductions in E(2) and PRL levels on proestrus, a limited increase in pituitary estrogen receptor alpha expression, and a significant shift in estrogen sensitivity, as measured by EB-induced PRL secretion. Nulliparous animals were more sensitive than multiparous rats to the two lower doses of EB, whereas multiparous animals were more responsive to the highest EB dose. These unique parity-induced alterations in the female's endocrine state that persist beyond lactation may impact a multitude of estrogen-mediated processes over the female's adult life span.     

Differential regulation of the nocturnal and diurnal prolactin surges in pregnant rats revealed by dopamine receptor antagonism.

We have explored temporal changes in the magnitude of dopamine (DA) interaction (DA tone) at the anterior pituitary lactotrophs related to both the nocturnal and diurnal prolactin (PRL) surges on day 8 of pregnancy, by utilizing a competitive DA D2 antagonist, domperidone (DOM). After withdrawal of blood from pregnant rats on day 7 in order to demonstrate the presence of a PRL surge, experimental rats received DOM (100 micrograms/kg i.v. or i.a.) at various times on day 8. Blood samples were taken immediately before and following injection of DOM at 5, 15, 30 and 60 min. The peak PRL response to DOM occurred 15 min after injection. Comparisons were made between circulating PRL levels immediately prior to and at several times following DOM administration for the various times of the day, and represented as incremental increases in PRL following DOM. During times on day 8 when PRL levels were normally low (24:00, 06:00, 12:00 and 16:00 h), pregnant rats exhibited a substantial PRL response to DOM. However, during the nocturnal PRL surge (02:00, 04:00 h) the peak PRL response to DOM was significantly lower. In sharp contrast, the PRL response to DOM administered during the diurnal PRL surge (18:00 h) was significantly higher than all other times of the day tested. In a dose-response study in which 10, 100 and 1,000 micrograms/kg DOM was administered at the two critical times when the response to DOM differed greatly, 02:00 and 18:00 h, there was a significantly reduced PRL response to DOM at 02:00 h compared to 18:00 h.(ABSTRACT TRUNCATED AT 250 WORDS)     

Sexual differentiation of positive feedback: effect of hour of castration at birth on estradiol-induced luteinizing hormone secretion in immature male rats

In the male rat, a dramatic increase in hypothalamic testosterone and estradiol concentrations occurs during the first few hours of postnatal life. These experiments sought to determine whether such increases participate in the defeminization of positive estrogen feedback effects on LH secretion. Newborn male rats were castrated either in utero (0 h males), or 10 or 24 h after birth. Some males were castrated at 0 h in utero and injected at the time of surgery with 1,2.5, or 5 micrograms testosterone propionate. A group of females was ovariectomized at 0 h in utero (0 h females). The control group consisted of male and female rats sham gonadectomized at 0 h in utero which were either gonadectomized at 21 days of age or left intact. The experimental groups were challenged before puberty to determine if estrogen induced a release of LH using two different types of estrogen treatment. The first treatment consisted of an injection of 0.2 microgram estradiol benzoate (EB) on day 28 followed by a second 10 micrograms injection of EB on day 29. This treatment resulted on the afternoon of day 30 in a surge of LH in intact females. Normal males, 0 h males, or females castrated at 21 days did not have a significant LH surge. The second test consisted of the daily injection of 0.05 microgram EB on days 23-27; on day 28 the rats were injected with 2.5 micrograms EB. Zero hour male and female rats showed a large LH surge on the afternoon of day 29; sham castrated males never responded to this treatment. No sex difference was observed in the mean size of the LH surge providing the males were castrated at 0 h in utero. The effect of the hour of castration on the day of birth also was studied. Males castrated at 10 or 24 h after birth showed either no LH surge or the magnitude of the surge was greatly reduced compared to that obtained in the 0 h males (P less than 0.001). The fact that 0 h males injected with 1 microgram testosterone propionate never showed an LH surge after prepuberal treatment with estrogen suggests that 0 h is a time during which the newborn is sensitive to the defeminizing effect of androgens. These results are consistent with the idea that the testicular hyperactivity which occurs at the time of birth could influence the defeminization of the LH surge mechanisms.     

Effect of the suckling stimulus on daily LH surges induced by chronic oestrogen treatment in ovariectomized lactating rats

Effects of the suckling stimulus on the daily LH surge induced by chronic oestrogen treatment were examined in ovariectomized lactating rats. Wistar-Imamichi strain rats were kept under 14 h light:10 h darkness (lights on at 05.00 h). Litter size was adjusted to eight on day 1 (day 0 = day of parturition) and ovariectomy performed on day 2. Lactating rats deprived of their litters on day 0 served as nonlactating controls. Silicone elastomer tubing filled with oestradiol was implanted on day 6 or 15. Blood samples were collected through an indwelling cannula at 10.00 and 17.00 h on each day after implantation to detect daily LH surges. Daily LH surges occurred in the late afternoon in both lactating and non-lactating rats implanted with oestradiol on day 6 or 15. The amplitude of daily LH surges in lactating rats implanted on day 6 declined much more rapidly than in non-lactating rats implanted on day 6, but no significant difference was found in the profile of the LH surge between lactating and non-lactating rats implanted on day 15. Pituitary LH contents just before the daily LH surge (12.00-12.30 h) 4 days after implantation in lactating rats implanted with oestradiol on day 6 were significantly less than those in nonlactating rats implanted with oestradiol on day 6 or 15 and in lactating rats implanted on day 15.(ABSTRACT TRUNCATED AT 250 WORDS)     

Preferential increase in pituitary prolactin versus vasoactive intestinal peptide as a function of estradiol benzoate dose in the ovariectomized rat

Vasoactive intestinal peptide (VIP) is synthesized in various tissues, including the anterior pituitary gland, where it may stimulate the release of PRL. Because estrogen plays a central role in the regulation of PRL, it becomes important to determine the effects of this steroid on both pituitary VIP and PRL. To study this, pituitary VIP and PRL and plasma PRL were assayed in ovariectomized rats after treatment with estradiol benzoate (EB; 0.007, 0.07, 0.7, 7 or 70 microgram/rat). Pituitary and plasma TSH were also determined as well as VIP content in the medial basal hypothalamus, suprachiasmatic region, cerebral cortex, and jejunum. Oil-treated rats served as controls. Injection of 0.7 or 7 microgram EB resulted in a significant increase in pituitary PRL without changing plasma PRL levels or pituitary VIP content compared to values in the control group. Only treatment with 70 microgram EB produced a significant increase in both pituitary VIP and PRL as well as in plasma PRL compared to control values. EB treatment at any of the doses used had no significant effect on pituitary and plasma TSH or VIP content in any of the other tissues examined. These data show that pituitary PRL and VIP are differentially regulated in response to estrogen. The increases in pituitary VIP and basal plasma PRL after treatment with the highest dose of EB suggest that pituitary VIP may be involved in the development of estrogen-induced hyperprolactinemia. These data also show that the regulations of pituitary VIP and TSH are independent of each other in the estrogen-treated rat.     

Effect of adrenalectomy on mating-induced prolactin surges and pseudopregnancy in the female rat

In the estrous female rat, mating stimulation induces an acute surge of prolactin (PRL) within 20 min after mating followed by the onset of twice-daily PRL surges which persist for an 8- to 13-day period of acyclicity called pseudopregnancy. In Experiment 1, we examined whether the release of adrenal hormones after mating modulates mating-induced PRL secretion during the first 38 h after mating. Ovariectomized females were adrenalectomized (Adx) or sham-operated (Sham) and were implanted with jugular vein catheters 2 days later. They were given estrogen and progesterone and mated 6 days after the last surgery until they received 15 intromissions or 15 mounts-without-intromission from a male. Blood samples were collected beginning 20 min before mating at 23:00 h and continuing for 38 h. Plasma PRL concentrations were measured using radioimmunoassay. Mating that included intromissions induced an acute (20-min) PRL response which was higher in Adx than in Sham animals, and advanced in the Adx animals in the onset of the first daily PRL surge to 10:00 h, some 18 h before the surge was observed at 04:00 h in the Sham-mated animals. A small but measurable nocturnal surge was observed in Adx and Sham groups 18-24 h later at 04:00-10:00 h. In Experiment 2, Adx- and Sham-cycling animals received 5 (5I) or 7 (7I) intromissions from a male 12-16 days after surgery. Adx animals receiving 5I showed a significantly higher incidence of pregnancy or pseudopregnancy (%P/PSP) than did Sham 5I animals, while there was no difference in %P/PSP in the 7I groups. We conclude that adrenal gland secretions normally suppress plasma PRL concentrations immediately post-mating and before the onset of the nocturnal mating-induced PRL surge and also inhibit pseudopregnancy when females receive a subthreshold number of intromissions normally required for its induction.     

Effects of opiate antagonists on serotonin turnover and on luteinizing hormone and prolactin secretion in estrogen- or morphine-treated rats

Previous findings indicate that estradiol benzoate (EB) acutely activates serotonergic projections in the medial preoptic area that are stimulatory to prolactin (PRL) secretion. Because opioid agonists also stimulate both PRL release and serotonin (5-HT) turnover, the present experiments tested whether endogenous opioid neurons may mediate estrogen feedback effects. In experiment 1, ovariectomized rats received either 50 micrograms EB or oil vehicle, and either saline or a long-acting opiate receptor blocker, nalmetrene (10 mg/kg), simultaneously, 3 h prior to decapitation. The pargyline method was used to determine 5-HT turnover; hence members of each treatment group received either saline or pargyline (75 mg/kg i.p.) 30 min prior to decapitation. Plasma PRL and luteinizing hormone (LH) concentrations were determined by radioimmunoassays, and 5-HT concentrations from microdissected, individual brain nuclei were measured by liquid chromatography with electrochemical detection. Nalmetrene blocked both the acute elevation of PRL and the increase in 5-HT turnover in the medial preoptic nucleus and ventromedial nucleus induced by EB. Nalmetrene alone also enhanced LH release and 5-HT turnover in the bed nucleus of the stria terminalis, and these effects were prevented by EB. A second study tested whether the opioid agonist, morphine, mimics estrogen effects on PRL and preoptic 5-HT turnover. Animals received saline, morphine (10 mg/kg i.p.) and/or naloxone (5 mg/kg i.p.) 30 min prior to decapitation. Simultaneously, half in each group received pargyline as above.(ABSTRACT TRUNCATED AT 250 WORDS)     

Evidence for the role of alpha-MSH in the induction of pseudopregnancy in the rat

In a previous study we have demonstrated that cervical stimulation (CS) induces alpha-MSH release. The present experiments were undertaken to (1) examine the pattern of serum alpha-MSH during CS-induced pseudopregnancy (PSP) and (2) assess the possibility that alpha-MSH contributes to the induction and maintenance of PSP. Throughout PSP serum alpha-MSH fluctuated in a cyclic manner demonstrating two daily surges which occurred between 12.00 and 13.00 h (diurnal surge) and between 24.00 and 04.00 h (nocturnal surge). Chronic exposure of animals to alpha-MSH administered via minipumps (24 micrograms/day, starting on the morning of estrus), induced PSP as determined by deciduoma formation and persistence of a characteristic diestrous vaginal cytology. Furthermore, insertion of an alpha-MSH-containing minipump at diestrus 1 (D1) resulted in progesterone and prolactin (PRL) levels on the afternoon of diestrus 2 (D2) similar to those levels found on day 2 of PSP. Uterine weight was significantly decreased in alpha-MSH-treated rats and pseudopregnant rats as compared with cyclic D2 controls. alpha-MSH was found to release PRL indirectly, through stimulation of adrenal progesterone. This effect, however, necessitates the presence of the ovaries as a source of estradiol (EB) since it is demonstrable in intact and acutely ovariectomized rats, but not in chronically ovariectomized animals. EB treatment of chronically ovariectomized rats is capable of restoring the sequence. These results indicate that, as demonstrated for PRL, cervical stimulation initiates rhythmic daily surges of alpha-MSH secretion which are maintained through PSP.(ABSTRACT TRUNCATED AT 250 WORDS)     

The hypothalamic paraventricular nucleus has a pivotal role in regulation of prolactin release in lactating rats

The affect of paraventricular nucleus (PVN) lesions on PRL secretory response to suckling was studied in adult female rats. Basal levels of PRL were similar in the control and lesioned groups. Substantial decreases in PRL levels occurred after separation of pups from their mothers in the control as well as lesioned animals. When mothers and pups were reunited, the circulating PRL concentrations of the control groups rose immediately from basal values of 50-100 micrograms/liter to reach peaks of 450-550 micrograms/liter. PVN lesions significantly decreased the suckling-induced rise of PRL levels. Furthermore, PVN lesions abolished the high amplitude, episodic pattern of PRL release in continuously lactating rats. These findings are consistent with the view that PVN neurons produce PRL releasing factor(s), which is (are) required for normal secretory patterns of PRL in lactating rats.     

Hysterectomy-induced alterations in prolactin secretion of lactating rats

The contribution of the uterus to the regulation of PRL secretion in lactating dams and cycling female rats was investigated. Lactating animals were hysterectomized or sham operated 2 days after parturition, and the number of pups was adjusted to eight. Blood samples for PRL RIA were obtained through intra-atrial cannulae implanted 2 days before experimentation. In order to study the PRL secretory profile in undisturbed freely lactating rats, blood samples were taken every 2 h for 24 h starting at 1400 h. During early lactation (days 7-8), hysterectomy did not alter the PRL secretory profile compared to that of sham-operated controls. On days 14-15 post partum, PRL secretion followed a characteristic bimodal pattern showing two PRL surges at 1800 h and 0600 h. After hysterectomy, the early morning PRL surge disappeared and PRL secretion showed an unimodal daily rhythm reaching its peak at 1800 h. The possible effect of the absence of the uterus on suckling-induced PRL release at various stages of lactation was studied. On days 7-8, suckling stimuli after 4 h of pup deprivation induced robust PRL release. Hysterectomy did not significantly alter PRL release at this earlier stage of lactation. In control groups, the suckling-induced PRL secretory response markedly declined as the postpartum period advanced. On the other hand, the hysterectomized animals retained significantly greater responsiveness to suckling during the second half of lactation. These data indicate an inhibitory influence of the uterus on PRL secretion. The onset of this uterine effect is considerably delayed, and its influence became prominent only at a later phase of lactation. The effect of length of pup deprivation preceding the suckling stimulus, in combination with hysterectomy, was also investigated. Hysterectomy significantly increased suckling-induced PRL release after 4 and 24 h separation compared to the sham-hysterectomized animals. When the separation was longer than 48 h, the inducibility of PRL release by suckling declined and was not influenced by hysterectomy. In order to study the possible influence of the uterus on PRL secretion during the estrous cycle, regularly cycling female rats were hysterectomized at diestrus 1. Twelve days later the animals were cannulated, and serial blood samples were taken during the subsequent proestrus. Hysterectomy did not alter the PRL surge which occurred on the afternoon of proestrus indicating that the uterus does not have a major function in regulating PRL secretion on proestrus. In conclusion, hysterectomy significantly delayed the extinction of suckling-induced PRL release revealing the active role of the uterus in the regulation of this neuroendocrine reflex.     

Prolactin release and milk ejection in rats suckling underfed pups

Mother rats chronically exposed to underfed pups (obtained daily from nonlactating mothers) show increased milk production which is associated with increased consumption of food and water. The purpose of the present study was 2-fold: to examine whether the magnitude of maternal PRL discharge is related to the nutritional state of the pups and to differentiate between the role of the mother and that of the pups themselves in the regulation of milk intake. PRL release in response to reunion with pups (preceded by 4-h separation from pups) was measured between days 12 and 14 postpartum; each mother was tested twice, once with hungry pups (deprived of food for 24 h) and once with satiated (normally fed) pups. The magnitude of PRL discharge elicited by hungry pups in lactating as well as in thelectomized rats was twice as large as the discharge elicited by satiated pups. For each type of stimulus pups, the magnitude of PRL release was smallest in thelectomized rats, largest in rats that had nursed normally fed pups before testing, and intermediate in rats that had nursed underfed pups. The frequency of milk ejection was considerably reduced in mothers that were chronically exposed to underfed pups compared with mothers that had nursed normally fed pups. Regardless of the mother's previous history, and independent of the milk ejection frequency or maternal food and water intake, hungry pups ingested more milk than did satiated pups over the same suckling period even when both types of pups were suckled simultaneously by the same female. Milk ingestion in satiated pups was not enhanced by the vigorous sucking exerted by hungry littermates. It is suggested that hungry pups are able to meet their increased nutritional demands both directly, by withdrawing a larger portion of the milk available in the mammary glands, and indirectly, by enhancing maternal PRL secretion which in turn may promote maternal consummatory behavior and milk production.     

Evidence for a physiological role for oxytocin in the control of prolactin secretion

The presence of oxytocin (OT) in neuronal elements of the external layer of the median eminence and in hypophysial portal plasma suggests a role for the peptide in the control of anterior pituitary function. We have reported previously that OT stimulates PRL release in vitro; therefore, we attempted to establish evidence for a physiological PRL-releasing role for OT. Plasma OT levels rose significantly just before the PRL surges occurring during a suckling stimulus in lactating rats (10 min after pup reinstatement vs. 15 min for PRL) and 48 h after estrogen injection in ovariectomized (OVX) rats (at 1200 h vs. 1300 h). Dispersed anterior pituitary cells harvested from lactating female rats and OVX estrogen-primed rats released PRL in a specific, significant, and dose-related fashion when perifused in vitro with incubation medium containing 10(-7)-10(-9) M OT, doses similar to levels found previously in hypophysial portal plasma. Infusion of antiserum specific for OT into lactating females before pup reinstatement and into estrogen-primed OVX rats 2 h before the expected release of endogenous OT delayed and significantly reduced subsequent PRL surges compared to levels in saline-or normal rabbit serum-infused rats; however, PRL release was not completely abolished. These data indicate that OT plays a physiological role in the hypothalamic control of PRL secretion and further suggest the importance of multiple factors in coordinated regulation of PRL release.     

Effect of transient dopamine antagonism on thyrotropin-releasing hormone-induced prolactin secretion in the serotonin-blocked, estrogen-treated ovariectomized rats

Recent studies showed that a brief interruption of dopamine (DA) action markedly increased the thyrotropin-releasing hormone (TRH)-stimulated prolactin (PRL) release. It is thus of interest to delineate whether the estrogen-induced afternoon PRL surge involves the same mechanism. Long-term ovariectomized rats pretreated with polyestradiol phosphate (PEP, 0.1 mg/rat s.c.) for 6 days were used in this study. They also received either p-chlorophenylalanine (PCPA, 250 mg/kg i.p.) or ketanserin (Ket, 10 mg/kg i.p.), two serotonergic drugs known to inhibit the estrogen-induced afternoon PRL surge. Then the animals were either treated with a DA antagonist, domperidone (Domp, 0.01 mg/rat i.v.), or vehicle at 16.00 h on the sampling day. Ten minutes later, the ones receiving Domp were injected with a DA agonist, 2-bromo-alpha-ergocryptine (CB154, 0.5 mg/rat i.v.), followed 50 min later by the administration of TRH (1 microgram/rat i.v.). Plasma samples taken through indwelling intraatrial catheters were assayed for PRL by radioimmunoassay. The estrogen-induced afternoon PRL surges were completely blocked in both PCPA- and Ket-treated animals. A significant PRL surge with similar amplitude, however, was induced by either Domp or TRH, although pretreatment with Domp did not cause any potentiating effect on the action of TRH. On the other hand, Domp induced only a small rise of PRL secretion and TRH was totally ineffective in rats untreated with PEP. It is concluded that both DA antagonism and TRH stimulation can induce significant PRL release in the afternoon of estrogen-treated, serotonin-blocked rats.(ABSTRACT TRUNCATED AT 250 WORDS)     

Effect of pulsatile gonadotropin-releasing hormone on the release of luteinizing hormone and follicle-stimulating hormone in vitro by anterior pituitaries from lactating and cycling rats

The ability of pituitaries from lactating animals to secrete LH and FSH in response to gonadotropin-releasing hormone (GnRH) was studied in vitro using a pituitary incubation system. Hemipituitaries were exposed to GnRH for 6 min during each hour of incubation. LH release by anterior pituitaries (APs) from day 5 postpartum rats nursing eight pups, in response to pulsatile exposure to GnRH, was significantly less than that released by APs from diestrous cycling females. Even though the amount of LH released by APs increased as lactation progressed, LH release by APs from day 15 postpartum rats nursing eight pups was still less than LH release by APs from diestrous females. In contrast pituitaries from lactating females nursing two pups released amounts of LH similar to that released by pituitaries from diestrous females, whereas females deprived of their litters for 48 h showed a greater response than diestrous females. Generally, there was a good quantitative relationship between the amount of LH released in vitro and plasma LH concentrations for all the intact groups studied. The ability of lactation to suppress the postcastration rise in serum LH also was demonstrated in vitro as pituitaries from ovariectomized or intact females nursing eight pups released similar amounts of LH on days 5 and 10 postpartum. However, by day 15 postpartum, even though serum LH concentrations were still very low, pituitaries from ovariectomized lactating females released LH in vitro at a rate similar to pituitaries from nonlactating rats. Serum FSH concentrations were not suppressed but similar in intact and cycling females. Also, the total amount of FSH released in vitro in response to GnRH by pituitaries from lactating and cycling females did not differ significantly, even though LH release differed greatly among these groups of animals. However, the patterns of GnRH-stimulating FSH secretion differed among intact lactating, ovariectomized lactating, and nonlactating females. Pituitary LH concentrations were similar on day 5 postpartum and diestrus and on day 15 postpartum and proestrus. Pituitary FSH concentrations on day 5 postpartum were similar to those during diestrus and proestrus and had increased 2-3 times by day 15 postpartum. Generally, there was no correlation between the amount of LH or FSH released by pituitaries in response to GnRH and pituitary gonadotropin content. In summary, the inability of pituitaries from lactating rats to respond adequately to large doses of GnRH in vitro suggests that the suckling stimulus indirectly suppresses pituitary responsiveness to GnRH. This suppression differentially affects basal LH secretion, but not basal FSH secretion, and may be the direct result of inadequate GnRH stimulation in vivo.     

Central nervous system regions involved in the estrogen-induced afternoon prolactin surge. II. Implantation studies

In the intact cycling female rat, there is a surge of plasma PRL during the afternoon of proestrus. Ovariectomy on diestrous day 1 eliminated the PRL surge completely, and injection of 100 micrograms polyestradiol phosphate, a long-acting estrogen, not only maintained the surge, but amplified and prolonged it. Bilateral implantation of an estradiol (E2)-containing cannula [diluted 1:4 with cholesterol (C)] in the medial preoptic area (MPO), but not in the cerebral cortex (CC), also maintained the surge. In long term ovariectomized rats, bilateral implantation of E2-containing cannulae (1:5, 1:10, 1:20, and 1:200) in the corticomedial amygdala (CMA) or ventromedial nucleus of the hypothalamus or a singular implantation in the third ventricle failed to induce a PRL surge 12 days later. Similar singular implantation in the anterior pituitary (AP) increased the basal levels of plasma PRL compared to those in C-implanted controls, but no surge was evident. Only bilateral implantation of E2 in the MPO induced a small but significant rise of plasma PRL at 1700 and 1900 h. Using higher concentration implants with a higher E2 concentration (1:4) in the MPO and sampling at shorter intervals, significant afternoon PRL surges were induced on days 2-4. However, systemic effects of E2, i.e. vaginal cornification and uterine weight enlargement, were also evident. Similar implants in other brain regions had the same results. Further increasing the E2 to C dilution from 1:10 to 1:200 eliminated the systemic effect of E2 implants, while the PRL surge-inducing ability persisted. It appears that the highest diluted E2 implants (1:150 and 1:200) gave the highest PRL response and persisted for the greatest number of days. Using the highly diluted E2 implants (1:100 and 1:200) in various brain regions, the MPO and the ventromedial hypothalamus were the most sensitive areas in inducing the PRL surge; the other areas studied, including suprachiasmatic nuclei, CMA, AP, and CC were ineffective. In conclusion, highly diluted E2 implants in the brain appear to be effective in obtaining a specific effect on the afternoon PRL surge; the CMA, suprachiasmatic nuclei, AP, and CC are not estrogen feedback sites for the induction of PRL release, and the MPO was the most sensitive area of the estrogen action in the brain regions examined for the induction of the afternoon PRL surge.     

Plasma prolactin concentrations in parental male and female rats: effects of exposure to rat young

The effects of pup presentation on the PRL responses in parental male rats were measured and compared with those in parental virgin and lactating female rats. Blood samples were collected from rats through indwelling intraatrial cannulas after a suckling challenge, i.e. presentation of rat young. Lactating rats showed full parental behavior and characteristic large surges in plasma PRL levels within the first 5-10 min on each day that rat young were presented (days 4, 8, and 12 of lactation). When pups were not presented, PRL rises did not occur. In contrast to the pattern of PRL responses shown by lactating mothers, parental ovariectomized nulliparous female and parental intact male rats failed to show specific increases in PRL in response to pup presentation. Plasma PRL levels in these groups, as in nonparental female and male rats, occasionally rose in response to blood collection rather than to pup presentation alone. Treatment of nulliparous female as well as male rats with estradiol and progesterone Silastic implants for 21 days before the initiation of behavioral testing significantly reduced the latencies of both nulliparous females and males to respond to foster young from about 5 to 2 days. The PRL responses of these steroid-primed groups were quite different. The steroid-primed females exhibited a pattern of PRL responses to pups identical to that found in lactating rats. The steroid-primed parental males, in contrast, failed to show specific increases in plasma PRL levels in response to young. These data demonstrate a sex difference in the hormonal, but not behavioral, responses of male and female rats to young and are suggestive of possible sex differences in the hypothalamic and/or peripheral regulation of pup-induced PRL secretion.