Prolactin release and milk removal induced by suckling and milking in lactating ewes is prevented by L-dopa treatment

The effect of L-DOPA on milk removal and on prolactin release during suckling or milking was studied in lactating ewes. Various doses of L-DOPA (25, 50, 100 and 200 mg per animal) were injected iv 30 min before the suckling or milking period. Control ewes were injected with 0.9% NaCl solution only. Milking induced a significant long-lasting release of prolactin. An inhibition of milk removal was obtained with the dose of 200 mg of L-DOPA. An inhibition of prolactin secretion was observed related to the dose of drug administered. The inhibitory effect of 200 mg of L-DOPA on the secretion of prolactin after milking lasted for about 120 min, and thereafter a significant increase in serum prolactin level occurred. This increase in serum prolactin was not due to a "rebound" effect of L-DOPA, since the milking stimulus had to be present to induce the delayed increase in prolactin. Doses of 25 or 50 mg of L-DOPA prevented the surge of prolactin observed immediately after milking, but a long-lasting release of prolactin was obtained thereafter. The inhibitory effect of L-DOPA on prolactin release could be overridden by the suckling or milking stimuli according to the dose administered. The suckling stimulus was more effective than milking in overriding the inhibitory effect of the low dose of L-DOPA. The results indicate that milk removal and prolactin release induced by milking or suckling in lactating ewes is inhibited by an increase in monoamines at the hypothalamic-hypophyseal level.     

The response of plasma prolactin to suckling during normal and prolonged lactation in the rat

In dams which had been kept isolated from pups for 8-10 h, the magnitude of the suckling-induced prolactin rise in the plasma was studied in relation to intensity of suckling stimulus and lactational age of the mother. At midlactation the response of prolactin evoked by suckling was enhanced as litter size increased. Suckling of 2 pups induced a greater prolactin rise in dams adjusted to 2 pups than in dams adjusted to 8 pups. Suckling of 8 pups caused a greater prolactin rise in dams which had been adjusted to an 8-pup litter, than in rats with a 2-pup litter. At late and prolonged lactation the rise of prolactin in the plasma induced by the suckling stimulus of 8 pups was significantly lower than at midlactation. Injection of perphenazine after a period of suckling induced a moderate increase of plasma prolactin in dams at midlactation, and a similar increase in dams at late lactation and at day 42 of lactation. It is concluded that in the first half of lactation the number of pups, i.e. the intensity of the suckling stimulus, is an important factor in determining the magnitude of the prolactin response to suckling. The lower response of plasma prolactin to suckling in late lactation is neither caused by a decrease in suckling stimulus from the pups nor by an increase in prolactin clearance; it is probably due to a gradual reduction in prolactin synthesizing and releasing capacity of the pituitary, brought on by a desensitization of the neural or neuroendocrine system to suckling stimuli as lactation proceeds.     

Prolactin does not mediate the suppressive effect of the suckling stimulus on luteinizing hormone secretion in ovariectomized lactating rats

The plasma LH concentration in ovariectomized lactating rats is low for 14 days postpartum, while the prolactin concentration is high during this period. We examined the effect of the inhibition of increased prolactin secretion with bromocriptine (CB-154) on the LH secretion in lactating rats ovariectomized on day 2 (day 0 = day of parturition). Blood samples were collected through an indwelling atrial cannula every day. LH levels were kept low until day 9 in lactating rats injected daily with CB-154 (0.6 mg/day, s.c.). The duration of the period during which LH secretion was suppressed was shorter in lactating rats treated with CB-154 than in saline-injected controls. The replacement with ovine prolactin by means of a mini-osmotic pump (0.3 mg/day, s.c.) in CB-154-treated lactating rats restored the duration of LH suppression. In rats deprived of their pups on day 2, the LH concentration rose immediately after removal of the pups and the LH level was not significantly different between rats treated with CB-154, ovine prolactin and saline, indicating that neither the CB-154 treatment nor the high level of prolactin alone has any effect on LH secretion in rats deprived of their pups. The present results clearly demonstrate that prolactin does not mediate the suppressing effect of the suckling stimulus on LH secretion in early lactation and support our theory that the suckling stimulus controls the LH and prolactin secretion independently at the hypothalamic level.     

Effects of naloxone, metaclopramide and domperidone on plasma levels of prolactin and LH following suckling in the female rabbit

Saline, naloxone, domperidone or metaclopramide was injected into lactating rabbits immediately before suckling. Blood samples were taken prior to injection (0 minutes) and then at 15, 30, 45 and 60 minutes after the start of suckling, after which the samples were assayed for plasma prolactin and LH concentrations. In all the does there was a significant increase in prolactin concentration, which was highest 15 minutes after the start of suckling, and which declined exponentially thereafter to levels significantly higher than before suckling. The increase in prolactin concentration was similar in does given saline and naloxone, but it was significantly enhanced in does given metaclopramide; with domperidone the increase was intermediate and not significantly different from that following treatment with saline. In does given saline, domperidone, and metaclopramide plasma LH concentrations declined slowly during the hour after suckling but the concentration was increased significantly in does given naloxone. The inverse correlations between prolactin and LH were low weak and were not significant.     

Effect of passive immunization against substance P in rats with hyperprolactinemia

Substance P, an undecapeptide isolated from gut and brain tissues, was reported to stimulate prolactin release. It was suggested that substance P may play a role in the control of prolactin secretion. In this investigation we studied the effects of the blockade of endogenous substance P by the administration of a specific anti-substance P serum on serum prolactin levels in rats in the evening of proestrus, in lactating rats after suckling, and in male rats with hyperprolactinemia induced by grafting 2 anterior pituitary glands under the kidney capsule. The injection of the anti-substance P serum was followed by a significant decrease of the prolactin surge induced by 30 min suckling in lactating rats, when the antiserum was administered 24 hr but not 5.30 hr earlier. Anti-substance P serum also induced a significant decrease in serum prolactin levels in pituitary grafted rats, but induced no change in the proestrous surge of prolactin and LH. These results show that substance P may be involved in the release of prolactin induced by suckling and that this peptide may have an intrapituitary role in the process of prolactin release. On the other hand, substance P does not seem to play a significant role in the proestrous peak of prolactin and LH.     

Suckling-induced prolactin release potentiates mifepristone-induced lactogenesis in pregnant rats

Suckling, starting at 19:00 h on Day 18 of pregnancy, induced a significant increase in serum prolactin concentration at 20:00 h on Day 19 of pregnancy, but no increase in mammary gland casein or lactose content. Mifepristone (2 mg/kg) injection at 08:00 h on Day 19 of pregnancy induced significant increases in casein, but not in lactose, 24 h after administration. Mifepristone alone did not induce prolactin secretion, indicating that lactogenesis was induced by placental lactogen in the absence of progesterone action. When mifepristone was injected into suckling rats, serum prolactin concentrations were higher than in the untreated suckling rats. Casein in these rats increased significantly 12 h after mifepristone administration and lactose at 24 h after. If the suckling mifepristone-treated rats were given two injections of bromocriptine (1.5 mg/kg) at 12:00 h on Days 18 and 19 of pregnancy, serum prolactin concentrations were not increased by suckling, but casein and lactose concentrations in the mammary gland showed values similar to those obtained in the mifepristone-treated non-suckling rats. Mifepristone can therefore potentiate suckling-induced prolactin release in pregnant rats, demonstrating a direct central inhibitory action of progesterone on prolactin secretion. This suckling-induced prolactin secretion, unable to induce casein or lactose synthesis in the presence of progesterone, enhanced significantly synthesis of these milk components in the absence of progesterone action (rats treated with mifepristone). Fatty acid synthase, which is stimulated by the suckling stimulus in lactating rats, was not modified by mifepristone or suckling in pregnant rats.     

Inhibitory effect of alcohol on the established suckling-induced prolactin surge in lactating rats

The effect of acute alcohol infusion on the established suckling-induced prolactin surge in lactating rats was examined. Dams were implanted with an atrial catheter on Day 6 of lactation and blood sampling was done on Day 10. Following the separation of litters from dams for a 6-hr period, a baseline blood sample was removed via a catheter extension. Pups were weighed and returned to dams. Subsequent blood samples were obtained 10, 30, and 60 min after initiation of suckling. Dams were then infused with alcohol doses of 0, 0.5, 1.0, 2.0, or 2.5 g/kg body wt. Infusion (0.1 ml/min) was completed in approximately 30 min. Additional blood samples were obtained 10 30, 60, and 120 min after the termination of infusion. In a separate group of rats, pups were removed from the dam after the first 60 min of suckling and additional blood samples were obtained 40, 70, 90, and 150 min after removal of pups (corresponding to 10-, 30-, 60-, and 120-min samples for rats infused with various alcohol doses). Alcohol, when administered after the establishement of suckling-induced prolactin surge and resulting in blood alcohol levels equal to or greater than legal human intoxication levels, inhibited prolactin release. However, continued suckling for an extended period (120 min in the present study) overcame this inhibitory effect, even when the blood alcohol level was comparable to (2.0 g/kg group) or greater than (2.5 g/kg group) the human legal intoxication level. Furthermore, in rats with established prolactin surges, the patterns of prolactin decline that followed alcohol administration or pup removal were comparable, indicating that similar mechanism(s) may be involved.     

Time course of the insensitivity of prolactin release to morphine administration in the lactating female rat

The effect of morphine on circulating levels of prolactin and growth hormone (GH) in the lactating female model was determined at various time intervals following the termination of suckling. Morphine administration did not produce an increase in prolactin levels when dams remained suckling. Four days after suckling was terminated, 50% of the dams tested showed a morphine induced prolactin increase. The prolactin secretory response to morphine gradually returned in dams, so that after 8 days of non-suckling, all animals tested showed a morphine induced prolactin increase. Consistent with the lack of prolactin stimulation, the tuberoinfundibular dopaminergic (TIDA) neurons, were insensitive to the morphine induced inhibition of activity during lactation. In contrast, circulating levels of GH were increased in these dams following morphine administration. These results suggest that the lactating female rat is insensitive to the mu mediated stimulation of prolactin release while suckling. However, sensitivity begins to return following at least 4 days of non-suckling.     

Endocrine profiles during doe-litter separation and the subsequent pregnancy in rabbits

This study was carried out to determine the effects of a transient doe-litter separation on plasma prolactin, LH, FSH, estradiol-17β and progesterone concentrations before artificial insemination and during the subsequent pregnancy. Control does (n=12) had free access to nursing, whereas separated does (n=12) were kept away from their litters for 48 hours before artificial insemination. Both groups were inseminated on day 11 after parturition. Teat stimulation by suckling caused a high increase in prolactin concentrations in separated does (p<0.0001). Basal prolactin concentrations were observed in both groups on days 8 and 18 of pregnancy. No effect of the treatment was detected on LH and FSH concentrations during the sampling period. A rise of estradiol-17β concentrations was observed 48 hours after doe-litter separation, compared to control does and to previous values (p<0.003). Both groups showed low progesterone concentrations before artificial insemination. Pregnant rabbits in both groups showed increased progesterone concentrations on days 8 and 18 of pregnancy. Lower estradiol-17β concentrations were observed in control does on day 18 of pregnancy compared with separated rabbits (p<0.003). The results suggest that a transient separation of nursing does from their litters before artificial insemination may promote high follicular steroidogenesis activity leading to increased estradiol-l7β concentrations. This hormonal change could be a result of several stimulatory actions probably triggered by the absence of suckling episodes and may affect the luteotrophic function during the subsequent pregnancy.     

Mammogenesis and changing prolactin receptor concentrations in the mammary glands of the tammar wallaby (Macropus eugenii)

All 4 mammary glands of the tammar wallaby showed a steady increase in weight and prolactin receptor concentration during the luteal phase of the oestrous cycle to reach a peak at oestrus. Removal of the corpus luteum abolished this mammogenesis , while pregnancy, which in this species is a day or so shorter than the oestrous cycle, had no effect. This provides an explanation for the previous finding that pregnancy is not a necessary pre-requisite for lactation in marsupials and that nonpregnant animals will lactate very successfully, provided the suckling stimulus is applied at the correct stage of the oestrous cycle. During lactation, only the gland supplying the teat to which the pouch young was attached developed and showed any further increase in prolactin receptors; the other 3 glands remained small and inactive. These results indicate the importance of the suckling stimulus and milk withdrawal on the initiation and maintenance of lactation.     

Effect of dopamine agonists or antagonists, TRH, stress and piglet removal on plasma prolactin concentrations in lactating gilts

The effect of dopamine agonists (ergocryptine), antagonists (chlorpromazine, haloperidol, reserpine, pimozide), thyrotropin releasing hormone or stress (restraint, piglet removal) on prolactin release was studied in primiparous lactating gilts. All animals were fitted with surgically implanted jugular catheters before farrowing. The only drug treatments which resulted in a significant change in PRL concentrations in blood were thyrotropin releasing hormone (increase) and ergocryptine (decrease). The results suggest that dopamine may not be the only regulator of prolactin in lactating pigs. Further studies are needed to identify drugs which would be useful in clinical situations for treatment of lactation failure due to low prolactin secretion. In the two stress-exposed groups, there was a gradual, steady decline in the plasma concentration of prolactin which resulted from loss of suckling contact with the piglets. Thus, snare restraint does not increase prolactin secretion in lactating sows confirming the results of other studies on pigs in different physiologic states.     

Plasma prolactin concentrations during the ovarian cycle and lactation and their relationship to return of fertility post partum in the common marmoset (Callithrix jacchus)

A heterologous double-antibody radioimmunoassay was used to measure plasma prolactin concentrations in 27 marmosets. The assay was valid for the marmoset because plasma levels of prolactin were increased in response to TRH and metoclopramide and suppressed in response to bromocriptine treatment. During the cycle there were no consistent changes in plasma prolactin concentrations. During lactation mothers suckling single or twin infants had higher prolactin levels than did non-suckling females and levels were highest with twins. No statistically significant delay in the resumption of ovulation post partum was observed for the suckling and non-suckling females; conception occurred in all but one marmoset by 70 days post partum. These results show that neither the suckling stimulus nor high levels of prolactin post partum delay the return of ovulation and fertility in the common marmoset, a result in contrast to that for all other primate species so far investigated.     

Correlation between suckling-induced changes in the ultrastructure of mammotrophs and prolactin release

Effects of suckling on the structure of mammotrophs and the release of prolactin, were studied in rats on the 10th day of lactation with the use of electron microscopy and radioimmunoassay techniques. Nursing animals were separated from their young for 8 hr and subsequently united and permitted to nurse for 1, 5, 15, 30 min; or 1, 2 and 4 hr. Blood samples were obtained prior to and throughout the suckling interval and pituitary glands were processed for electron microscopy. Control animals consisted of normal lactating females and animals separated from their young for 8 hr. Normally lactating controls had high prolactin serum levels (501 +/- 95 ng/ml) and synthetically active appearing mammotrophs. An 8 hr separation from the pups induced a dramatic lowering of serum prolactin (32 +/- 5 ng/ml), an increase in secretory granule storage, and a great dilation of rough endoplasmic reticulum (RER) cisternae. Five min of renewed suckling resulted in a rise of plasma prolactin levels (605 +/- 183 ng/ml) which remained high thereafter. The major ultrastructural changes observed during the first 30 min of suckling were as follows: 1) at 1 min, the RER became cmone?); 2) AT 5 MIN, AND MUCH MORE OBVIOUSLY AT 15 AND 30 MIn, a massive discharge of secretory granules was observed; and 3) at 15 min, the collapsed RER underwent transformation for 1,2 and 4 hr) induced new hormone synthesis as suggested by the presence of hypertrophied Golgi elements and numerous immature granules. This was accompanied by a new transformation of the RER from the vesicular into a lamellar form now consisting of very slender cisternae lined with numerous ribosomes, presumably involved in the renewal of the synthetic process. The morphologic findings described correlate well with the time table of prolactin release. In addition, the dramatic early changes in the structure of the RER suggest a possible involvement of this organelle in the storage and release of a proposed rapidly releasable pool of prolactin.     

Regulation of ghrelin secretion during pregnancy and lactation in the rat: possible involvement of hypothalamus

We investigated the plasma concentration of ghrelin peptide during pregnancy and lactation in rats. Plasma ghrelin levels on days 10 and 15 of pregnancy were significantly lower than those of the non-pregnant rats. Thereafter, the plasma ghrelin levels on day 20 of pregnancy sharply increased to levels comparable with those in non-pregnant rats. Ghrelin peptide concentrations in the stomach did not change significantly during pregnancy. In the hypothalamus, ghrelin mRNA levels were significantly lower on day 15 of pregnancy than in the non-pregnant rats. Also, plasma ghrelin levels were significantly lower in lactating dams than non-lactating controls on days 3 and 8 of lactation. We examined the possible involvement of prolactin and oxytocin in the regulation of plasma ghrelin concentrations during lactation. Although plasma prolactin levels were decreased by the administration of bromocriptine, plasma ghrelin levels did not differ significantly between vehicle- and drug-treated lactating rats. Administration of haloperidol produced a marked increase in plasma prolactin levels as compared with the non-lactating controls. However, plasma ghrelin levels were not significantly different between vehicle- and drug-treated rats. Administration of an oxytocin antagonist into the lateral ventricle significantly inhibited the increase in the plasma oxytocin level induced by acute suckling. However, plasma ghrelin levels did not significantly between the groups. These observations indicated that the decrease in serum ghrelin is caused by a loss of the contribution of hypothalamic ghrelin. Furthermore, the present results suggested that the suckling stimulus itself, but the release of prolactin or oxytocin, is the factor most likely to be responsible for the suppression of ghrelin secretion during lactation.     

Endocrine profiles during doe-litter separation and the subsequent pregnancy in rabbits

This study was carried out to determine the effects of a transient doe-litter separation on plasma prolactin, LH, FSH, estradiol-17beta and progesterone concentrations before artificial insemination and during the subsequent pregnancy. Control does (n=12) had free access to nursing, whereas separated does (n=12) were kept away from their litters for 48 hours before artificial insemination. Both groups were inseminated on day 11 after parturition. Teat stimulation by suckling caused a high increase in prolactin concentrations in separated does (p < 0.0001). Basal prolactin concentrations were observed in both groups on days 8 and 18 of pregnancy. No effect of the treatment was detected on LH and FSH concentrations during the sampling period. A rise of estradiol-17beta concentrations was observed 48 hours after doe-litter separation, compared to control does and to previous values (p < 0.003). Both groups showed low progesterone concentrations before artificial insemination. Pregnant rabbits in both groups showed increased progesterone concentrations on days 8 and 18 of pregnancy. Lower estradiol-17beta concentrations were observed in control does on day 18 of pregnancy compared with separated rabbits (p < 0.003). The results suggest that a transient separation of nursing does from their litters before artificial insemination may promote high follicular steroidogenesis activity leading to increased estradiol-17beta concentrations. This hormonal change could be a result of several stimulatory actions probably triggered by the absence of suckling episodes and may affect the luteotrophic function during the subsequent pregnancy.     

Effect of haloperidol, suckling, oxytocin and hand milking on plasma relaxin and prolactin concentrations in cyclic and lactating pigs

Prolactin secretion was stimulated in 5 cyclic gilts during the luteal phase (Day 10-13) with 5 mg haloperidol given i.v. Stimulation of prolactin secretion was also attempted by inducing milk let-down by suckling (4 sows), or by the injection of 1 mg oxytocin i.v. followed by hand milking (3 sows). Plasma prolactin concentrations increased significantly 1-2 h after haloperidol injection, and in 3 of 4 sows during suckling (P = 0.001); plasma relaxin concentrations did not change significantly at these times. No change was observed in plasma prolactin or relaxin concentrations at 15 min or 1-2 h after oxytocin injection and hand milking. Plasma relaxin concentrations ranged from below the sensitivity of the assay (100 pg/ml) to 450 pg/ml in lactating sows and from 100 to 2000 pg/ml in cyclic gilts. The results suggest that in cyclic gilts treated in the luteal phase with a dopaminergic receptor blocker, and in lactating sows during suckling, elevations in plasma prolactin concentrations were not accompanied, during the same period, by detectable changes in relaxin concentrations.     

Oxytocin plasma level in the lactating sows--effect of suckling

Oxytocin concentration in the peripheral blood was measured by RIA during suckling period in lactating sows (n = 8). Blood samples were taken from the jugular vein around the clock for every 2 h on day 5, 10, 15, 20, 25, 30 and on day 35 of lactation. Besides that blood samples were collected more frequently during suckling periods. Oxytocin plasma concentration was very low and in most cases it was on a border of sensitivity of our method (3 pg/ml). Marked but short-lasting rise of oxytocin was observed only during a period of initial massage of the udders by the piglets. This rise observed in all studied pigs was higher (p less than 0.01) compared to the values before the massage on the onset of lactation only, and was 14.6 +/- 4.2 pg/ml and 6.4 +/- 1.2 pg/ml on day 5 and day 10 of lactation, respectively. In all other studied days in a few cases only suckling stimulated the release of oxytocin over its basic concentration. Mean values (+/- SEM) of oxytocin in blood samples collected during massage of udder on day 15, 20, 25, 30 and day 35 were 3.7 +/- 0.5, 4.2 +/- 0.8, 4.9 +/- 1.1, 3.2 +/- 0.4 and 3.0 +/- 0.6 pg/ml plasma, respectively. There was no relationship between the size of the litters and neither basic level of oxytocin nor its blood concentration during suckling (r = 0.13).     

Initiation of fatty acid synthesis in rat mammary glands.

The rate of fatty acid synthesis from [6-14C]glucose in mammary tissue remained low until parturition at 22 days of gestation and increased 10-fold at 1 day post partum. Administration of progesterone on days 20 and 21 or removal of pups at parturition abolished this increase. In the latter case, administration of prolactin, corticosterone or oxytocin had no stimulatory effect; tissue from suckled glands in which the ducts had been ligated at parturition also showed no increased rate in 24 h. Foetoplacentectomy on day 18 did not stimulate fatty acid synthesis but subsequent suckling by foster pups did. Whereas lactose synthesis is initiated by withdrawal of progesterone from the circulation, a further stimulus related to removal of milk by suckling is required to initiate fatty acid synthesis.     

Changes in concentration of serum prolactin during social and reproductive development of the spotted hyaena (Crocuta crocuta).

A homologous radioimmunoassay system developed for humans was used to measure prolactin concentrations in spotted hyaenas. Concentrations of prolactin showed a significant (P < 0.05) decrease in lactating females, which is consistent with the infrequent suckling pattern of this species. This lack of hyperprolactinaemic conditions during lactation may explain the ability of females to resume reproductive activity soon after the loss of a litter, or even during lactation. Prolactin concentrations did not increase significantly during dispersion in male spotted hyaenas. This conforms to the pattern observed for cortisol, but differs from that for androgen, which fluctuates significantly with social suppression. Although comparative data from other species provide some circumstantial evidence for hyperprolactinaemic conditions during male dispersal, no obvious deductions regarding the recorded inverse relationship between prolactin and cortisol concentrations in mature males could be made.     

The suckling-induced rise of plasma prolactin in lactating rats: its dependance on stage of lactation and litter size

The response of plasma prolactin in vigorous suckling was measured in lactating rats which had been isolated for 10-12 h from their offspring. Plasma prolactin was investigated during suckling at various stages of lactation. The results demonstrate that prolactin responds maximally to suckling already in the first days of lactation. In the second half of the lactation period, the prolactin rise in the plasma induced by suckling decreases gradually; this is not due to a reduced suck-intensity of older pups. A relationship is found between the height of the suckling-induced prolactin rise and litter size. The data suggest that during suckling in the first weeks of lactation the pituitary secretes large amounts of prolactin at a constant rate. It is speculated that in the first minutes of suckling, receptors may clear considerable amounts of released prolactin from the circulation.