Dopaminergic control of prolactin mRNA accumulation in the pituitary of the male rat

Dopaminergic control of the expression of the prolactin gene was investigated by administration of biomoergocryptine (CB154) to male rats. The effects of the drug on the following parameters were measured: (i) circulating levels of GH and PRL; (ii) synthesis of GH and PRL measured by pulse labeling pituitary fragments in vitro; (iii) GH and PRL mRNA activities; and (iv) content of PRL mRNA. After 1 day of CB154 administration, serum PRL fell to undetectable levels whereas it took 3 days to observe a 50% reduction in PRL synthesis. This effect was accounted for by a parallel decrease in PRL mRNA activity and content. GH synthesis and GH mRNA were not affected by the treatment. Our results show that the dopaminergic inhibition of PRL production involves regulation at a pre-translational level.     

Impaired induction of prolactin secretion from the anterior pituitary by suckling in streptozotocin-induced diabetic rat.

Prolactin (PRL) secretion in streptozotocin-induced diabetic rats postpartum was examined to elucidate the reason for the reduced milk secretion of diabetic mothers. Pregnant Wistar rats were given citrate buffer (control group) or streptozotocin only (DM group) or with insulin (insulin group). Growth of pups was significantly lower in the DM group than in the control group, but similar in the insulin and control groups. Suckling-induced PRL secretion was significantly lower in the DM group than in the control group, and intermediate in the insulin group. TRH-induced PRL secretion was significantly lower in the DM group than in the control group, but the same in the insulin and control groups. Histologically, the mammary glands in the DM group were relatively less developed than those in the control group. The results suggest that reduced milk secretion in diabetic mothers is due to impaired induction by suckling of PRL secretion from the anterior pituitary as well as poor development of the mammary gland.     

Stimulation of anterior pituitary galanin and prolactin gene expression in suckling rats

Recent evidence suggests that galanin, may regulate prolactin (PRL) secretion during lactation. In this article, we describe the regulation of anterior pituitary galanin and PRL gene expression during pregnancy and after parturition in the rat. Expression of galanin and PRL in the anterior pituitary were significantly higher at d 20 of pregnancy compared to diestrus. One day after parturition, galanin mRNA levels increased a further 4.5-fold. This post partum increase in gene expression was not observed for PRL. The increase in galanin gene expression was maintained above the diestrous level for at least 10 d after parturition. PRL mRNA expression, on the other hand, was largely unchanged after parturition. Although the increase in galanin gene expression 1 d after parturition was independent of suckling, subsequently, galanin, gene expression was significantly higher in nursing mothers. Anterior pituitary galanin gene expression was 12-fold higher in nursing mothers compared with those that were not, 3 d after parturition. Similarly, PRL gene expression was significantly lower in mothers who were not suckling their pups 3 d after parturition. Initiation of suckling alone was insufficient to stimulate galanin and PRL expression. Despite suckling for 2 d, removal of the suckling stimulus subsequently resulted in a rapid decrease in galanin gene expression. Hence, the stimulatory effect of suckling on galanin expression requires a sustained suckling stimulus. In conclusion, the data support the hypothesis that anterior pituitary galanin plays an important role during lactation, likely acting to amplify lactotroph stimulation through paracrine and autocrine mechanisms.     

Effect of estrogen on prolactin mRNA in the rat pituitary. Analysis by in situ hybridization and immunohistochemistry

The effect of estrogen on prolactin (PRL) synthesis at a single-cell level was studied by in situ hybridization and immunohistochemistry. Long-term estrogen treatment increased PRL-containing cells from 10-20% of total cell population to 80-90%, as revealed by immunohistochemistry. PRL mRNA containing cells also increased in a similar fashion. Moreover, cytoplasmic PRL mRNA expressed as the number of silver grains per cell increased 4- to 5-fold by estrogen. These results suggest that long-term estrogen treatment causes not only PRL cell proliferation but also an increase in PRL mRNA in a single cell.     

Extraction of rat pituitary prolactin for radioimmunoassay

The bulk of Prolactin (PRL) in rat pituitary gland stored as big molecular forms in secretory granules is mostly excluded from radioimmunoassay (RIA) determinations because secretory granules remain intact after tissue homogenization. Also big PRL is little immunoreactive and must be deaggregated to monomers to allow a complete detection by RIA. Different dissociating agents, used to render PRL monomers soluble, were tested at various temperatures and pH conditions. Incubations of pituitary homogenates in buffers at neutral pH yield consistent levels of PRL, but the sole alkalinization of the media increases significantly the radioimmunoassayable PRL content. No significant increase was detected with EDTA and thiols. The 2.5 M urea was the most effective extraction agent increasing about 7-fold the quantification of PRL by RIA. Extraction of PRL with urea was enhanced at pH 9.0 and at 4 C and this combination constitutes the method of choice for a complete extraction of pituitary PRL.     

Effect of suckling on prolactin receptor immunoreactivity in the hypothalamus of the rat

Previous studies showed that expression of prolactin (PRL) receptor is increased in numerous hypothalamic nuclei in mid-lactating rats. The increase in PRL receptor expression could be initated by neurohormonal changes during proestrus or pregnancy, or by the suckling stimulus during lactation. The present study investigated whether the PRL receptor expression in numerous hypothalamic nuclei is altered by the suckling stimulus. Three groups (n = 4) of rats on lactation day 10 were used: a continuously suckled group, a nonsuckled group (pups removed for 12 h) and a resuckled group (pups removed for 12 and then resuckled for 9 h). Animals were perfused with 2% paraformaldehyde and brains were sectioned (20 microm) for the immunofluorescence study. Immunoreactivity was semiquantitatively analyzed by counting the immunoreactive cells and measuring the immunostaining intensity in a specific area. Neurons expressing PRL receptors were observed in numerous hypothalamic areas with the highest number being in the arcuate, paraventricular and supraoptic nuclei. The PRL receptor immunofluorescence in several nuclei was significantly decreased in the nonsuckled group, and recovered in the resuckled group. These areas included the ventromedial preoptic, ventrolateral preoptic, lateroanterior hypothalamic, ventrolateral hypothalamic and ventromedial hypothalamic nuclei. PRL receptor immunoreactivity in other areas was not significantly altered by the suckling stimulus. These results demonstrate that expression of PRL receptor in hypothalamic nuclei was differentially affected by the suckling stimulus. PRL receptors in those nuclei which were significantly altered by suckling stimulus may play more critical roles during lactation than those areas which were less sensitive to the suckling stimulus.     

Plasma prolactin in the rat during suckling without prior separation from pups

The relation between suckling and plasma prolactin (Prl) was studied in the rat, without prior separation of the dam from its pups. When the pups were replaced by a hungry foster litter, upon renewed suckling plasma Prl showed episodic increases and decreases in individual rats. When, subsequent to litter removal, similar rats were injected with perphenazine, a significant increase of plasma Prl was observed. This indicates that a decline of plasma Prl during suckling was not caused by exhaustion of Prl stores in the pituitary. In 22 individual rats blood was sampled every other minute while observations were made on nursing behaviour of the dams. During apparent suckling, increases as well as decreases of plasma Prl occurred. However, in most cases suckling did not affect plasma Prl, i.e. it remained stable at a high or a relatively low level. On the other hand, a considerable rise of plasma Prl was frequently observed when a dam was away from the nest. The data indicate that in the physiological situation Prl secretion from the pituitary is not directly related suckling activity, though episodes of suckling are essential to maintain a high Prl secretory capacity of the pituitary gland.     

Comparison of plasma profiles of oxytocin and prolactin following suckling in the rat

The purpose of these studies was to determine the effect of suckling on the plasma oxytocin (OT) concentration profile in conscious primiparous rats during midlactation. Comparisons were made with plasma prolactin (PRL) levels obtained in the same rats. OT levels in the majority of rats exhibited a single peak during the first 5-30 min, then fell rapidly during the course of a 45-min period of suckling. The plasma OT levels were sustained over a longer period in mothers suckling 8 rather than 6 pups; the amplitudes of the OT response were similar, however. By contrast, plasma PRL profiles indicated that a steady secretion of the hormone occurred throughout the suckling period, with suckling of 8 pups resulting in significantly higher plasma levels than suckling of 6 pups. A considerably greater increase in the peak plasma OT concentration resulted when hungry foster litters of 6 pups were suckled after the mothers' own 6 pups had been suckled. Plasma PRL levels during the two sucklings, though, were similar. The rapid onset of the OT response to suckling was seen more clearly in urethane-anesthetized rats following mammary nerve stimulation. Plasma OT levels rose to a peak within 5 s after the onset, then fell to prestimulus levels by the end of the 65-second stimulation period. These results suggest that different regulating mechanisms are involved in the secretory responses of OT and PRL to suckling and that different thresholds of activation are likely to exist for the two hormones.     

The endogenous stimulatory rhythm regulating prolactin secretion is present in the lactating rat

Prolactin (PRL) secretion in the female rat is regulated by an endogenous stimulatory rhythm (ESR), which is normally under the tonic inhibition of dopamine (DA). The ESR consists of a nocturnal (N) component which peaks at approximately 03.00 h, and a diurnal (D) component which peaks at approximately 17.00 h. This ESR has been shown to be present in ovariectomized and cervically stimulated rats. We have proposed that the ESR is continuously present in the female rat and that any suppression of the tonic inhibitory influence on PRL secretion can reveal its existence. In this study, the effects of the DA-lowering stimulus of suckling was investigated at different times of day in lactating rats. In addition, the pattern of PRL secretion in freely lactating rats throughout a 24-hour period was studied. Female rats were separated from their pups for 6 h prior to reunion at either 03.00 (coincident with the N component), 12.00 (control) or 17.00 h (coincident with the D component) at various stages of lactation. Blood samples were collected from intra-atrial cannulae immediately before separation of pups and dams, immediately before reunion of pups and dams (0 time), and 15, 30, 60 and 120 min following reunion of pups and dams. Four days following parturition, dams suckled at either 03.00 or 17.00 h secreted significantly greater PRL than rats suckled at 12.00 h. Peak levels of PRL were 60-, 90- and 25-fold greater than 0 time levels, at 03.00, 17.00 and 12.00 h, respectively.(ABSTRACT TRUNCATED AT 250 WORDS)     

Opiate modulation of the anterior pituitary hormone response during suckling in the rat

We have investigated the influence of endogenous opiates on hormone responses during suckling in the rat. In complementary experiments, opiate receptors were blocked by naloxone (NAL) or endogenous opiate release from the pituitary was inhibited by dexamethasone (DEX). Serial blood samples from unanesthetized suckled rats were then assayed for plasma PRL, beta-endorphin-like immunoreactivity (beta-END-LI), TSH, and GH levels. Identical studies were also done in saline-treated (control) suckled rats and in unsuckled rats exposed to control objects. Whereas suckling caused a rise in plasma PRL, beta-END-LI, and GH, introduction of plastic control objects did not elevate hormone levels. NAL blocked the GH rise and depressed TSH levels, but did not significantly inhibit the PRL or beta-END-LI response. DEX prevented the beta-END-LI rise and blocked the GH rise, but did not inhibit TSH. DEX enhanced PRL release during suckling. These results demonstrate that 1) the responses of beta-END-LI, PRL, and GH are not an artifact of the sampling procedure; 2) PRL release during suckling is independent of beta-END-LI release by the pituitary; and 3) suckling stimulates the release of ACTH, beta-END, and beta-lipotropin from the anterior pituitary. Our results are consistent with both a role of pituitary beta-END in the control of GH and a role of corticosterone in the control of PRL during suckling.     

Microtubules are mobilized in the lactating rat anterior pituitary gland during suckling

Microtubules in the lactating rat anterior pituitary gland were depolymerized into their constituent tubulin dimers by exposure of the pituitary to 4 C. The tubulin then was quantified with the [3H]colchicine binding assay, which was adapted for use with individual rat pituitary glands. The binding of [3H]colchicine to the tubulin fraction contained in high speed supernatants of lactating rat pituitary glands proved to be specific and saturable, and was pH, temperature, and time dependent. The amount of [3H]colchicine bound was linear over the range of protein concentrations tested (2-22 micrograms). To determine whether suckling affected the levels of microtubules in the anterior pituitary, tubulin levels were measured in groups of lactating rats after they were suckled (six pups per litter) for 0, 5, 10, 15, and 30 min following 4-5 h of nonsuckling. The tubulin concentration in the anterior pituitary progressively increased from 4 to 22 mumol during the 30 min of suckling; the increase was statistically significant (P less than 0.05) by the 10th min. Plasma PRL levels analyzed from trunk blood rose from 5 to 75 ng/ml during the 30-min suckling period. These results indicate that a mobilization of microtubules occurs in the anterior pituitary at the same time that PRL is being transformed and released into the circulation.