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The organization of prolactin-like-immunoreactive neurons in the rat central nervous system. Light- and electron-microscopic immunocytochemical studies.

The localization and distribution of prolactin-like-immunoreactive perikarya and nerve fibers in the rat central nervous system have been studied by a preembedding immunoperoxidase method using well-characterized specific immunsera to rat prolactin. Although the localization of labeled neuronal structures in a number of brain areas correlates with the data of previous immunocytochemical studies, we found prolactin-immunoreactive neurons in various regions not previously reported. In untreated animals, the highest concentrations of prolactin-fibers were observed: (i) in the external layers of the median eminence where they exhibited close contact with blood vessels, and (ii) in the bed nucleus of the stria terminalis and in the central nucleus of the amygdala where they closely surrounded unlabeled perikarya. Dense networks of finely varicose prolactin fibers were also observed in the organum vasculosum of the lamina terminalis, in the subfornical organ, and in the dorsolateral regions of the medulla oblongata and the spinal cord. Lastly, a number of large, varicose, intensely immunoreactive fibers were found in the olfactory bulb, the cingulum, and the periventricular regions of the hypothalamus and central gray, whereas isolated fibers could be detected in the caudate nucleus and in the cerebral cortex. In animals treated with colchicine, prolactin-immunoreactive perikarya were essentially located within the periventricular and perifornical regions of the hypothalamus, and within the bed nucleus of the stria terminalis. Although corticotropin (ACTH 17-39)-immunoreactive fibers could be detected in several regions found to contain prolactin fibers, the distribution and organization of both fiber types clearly differed in numerous brain regions, and the regions containing the corresponding perikarya did not overlap.(ABSTRACT TRUNCATED AT 250 WORDS)

Adrenocorticotropic Hormone↗

Combined use of in situ hybridisation and immunocytochemistry for the investigation of prolactin gene expression in immature, pubertal, pregnant, lactating and ovariectomised rats.

We have investigated the use of in situ hybridisation together with immunocytochemistry for the study of endocrine cell function, using as an example the expression of prolactin messenger RNA (mRNA) in pituitaries of rats under various endocrinological conditions. In situ hybridisation using a 32P-labelled cRNA probe for rat prolactin was carried out on sections of 4% paraformaldehyde-fixed pituitaries from prepubertal, pubertal, pregnant, lactating and ovariectomised rats and adjacent sections were immunostained for prolactin. Northern gel analysis was performed on total RNA extracts of pregnant, lactating and control pituitaries. While in ovariectomised rat pituitaries both prolactin immunoreactivity and prolactin mRNA were decreased, no differences in prolactin immunostaining were seen between prepubertal, pubertal, pregnant or lactating rats and controls, even when the supra-optimal dilution technique was used. However, using in situ hybridisation, prolactin mRNA signal was increased in prepubertal rats, and with hybridisation and northern gel analysis the signal was reduced in pregnant rats and markedly increased in lactating rats. The combined use of in situ hybridisation and immunocytochemistry provides morphological information concerning endocrine gene expression and protein synthesis in the pituitary gland.

Animals↗

Morphometric analysis of secretory granules and prolactin levels in chromophobe pituitary adenoma.

Morphometric analysis of secretory granules was performed in 16 chromophobe adenomas: seven undifferentiated cell adenomas (group A), two of which were accompanied by mild hyperprolactinemia and nine prolactin cell sparsely granulated adenomas (group B), all associated with hyperprolactinemia. The results of morphometry were correlated with levels of prolactin in tumor and serum. The range of granule diameters was similar in both groups but granule size frequency was different resulting in a characteristic histogram for each group. Poor relationship existed between granule mean diameter and prolactin in tissue of the group B. In the same group there was virtually no linear relationship between serum prolactin on one side and granule mean diameter, granule density or tissue prolactin on the other. Also in the group B a paradoxical inverse relationship was found between granule density and tumor prolactin. Apparently, in prolactin cell adenoma a sizable part of hormone is located outside of mature secretory granules. An abundance of immature granules, misplaced exocytosis, crinophagia or defect in packaging of the hormone could all explain this paradox but the last possibility seems most plausible.

Adenoma, Chromophobe↗

Preliminary characterization of bovine pineal prolactin releasing (PPRF) and release-inhibiting factors (PPIF) activity.

Bovine pineal glands were subjected to extraction with dilute acetic acid, gel filtration on Sephadex G-25 and subsequent ultrafiltration through Diaflo membranes PM10, UM2 and UM05. Various fractions derived at each step were tested for the presence of substances which stimulate or inhibit prolactin secretion in vitro and in vivo. Both prolactin releasing (PPRF) and release-inhibiting (PPIF) activities were observed. PPRF activity was present in certain fractions derived from Sephadex G-25 and in the PM10 residue (MW congruent to less than 10,000). Whereas both the UM2 residue (MW greater than 1000) and UM05 filtrates (MW less than 500) was seen to inhibit pituitary prolactin release in vitro, the UM05 residue (MW greater than 500 and less than 1000) inhibited prolactin release in vivo, possibly by stimulating the secretion of the hypothalamic prolactin inhibiting factor. On the basis of its inactivation by trypsin it was concluded that PPIF may be a peptide or contain a peptide moiety indispensible for its biological activity. Experiments are in progress to characterize pineal prolactin-regulating activities and to elucidate further the physiological role of the pineal gland in the regulation of prolactin secretion.

Animals↗

Gonadotropin and prolactin secretion in prepubertal female rats treated with 8-hydroxy-2-(di-n-propylamino) tetralin.

The effect of serotoninergic activation on gonadotropin and prolactin release were analysed in 16-day-old intact female rats. In the first experiment, females were decapitated 30 min after i.p. administration of 100 mg/kg of 5-hydroxytryptophan (5-HTP) or vehicle; in the second experiment the rats were decapitated 15 and 30 min after i.p. injection of vehicle or some doses (0.1, 1 and 10 mg/kg) of 8-hydroxy-2-(di-n-propylamino) tetralin (8-OH-DPAT), a selective agonist of the serotonin (5-HT)1A receptors. We found that: 1) serum follicle-stimulating hormone (FSH), luteinizing-hormone (LH) and prolactin concentrations increased after 5-HTP administration; 2) serum LH and prolactin concentrations and pituitary prolactin content increased after administration of 8-OH-DPAT. Our results indicate that in prepubertal rats, activation of serotoninergic system stimulated gonadotropin and prolactin release, and that 5-HT1A receptors are involved in this effect. In addition, the simultaneous increase in serum and pituitary prolactin content suggests that 8-OH-DPAT enhances prolactin synthesis.

5-Hydroxytryptophan↗

Control of prolactin secretion.

1. Prolactin is a 21,500 Dalton single-chain polypeptide hormone but may occur in 50 kDa and 150 kDa molecular variants. 2. These large PRL variants may be secreted predominantly; this condition is termed "macroprolactinemia". It is characterized by high immunological and normal biological serum levels of prolactin, and lack of clinical symptoms of hyperprolactinemia. 3. The information on PRL is encoded on chromosome 6. Transcription can be enhanced and suppressed by a variety of hormonal factors. 4. PRL is secreted in a pulsatile fashion; it displays a circadian rhythm (with a maximum during sleep) and is stimulated by some amino acids. PRL also responds to mechanical stimulation of the breast. 5. PRL rises during pregnancy, and maintainance of hyperprolactinemia (and, thereby, physiological infertility) is dependent on the frequency and duration of breast feedings. 6. Hypothalamic regulation of prolactin mainly involves tonic inhibition via portal dopamine. The physiological importance of various stimulating factors present in the hypothalamus is still incompletely understood. In particular, there is still no place for TRH in PRL physiology. 7. PRL is released in response to stress; this response may be mediated by opioids. The low-estrogen, low-gonadotropin amenorrhea of endurance-training women is not mediated by prolactin, however. 8. Estrogens stimulate PRL gene transcription via at least two independent mechanisms. There are many clinical examples of this estrogen effect on prolactin serum levels, and also on the growth of prolactinomas. 9. Mild hyperprolactinemia remains an enigma which cannot satisfactorily be resolved by biochemical or radiological testing. The border between "normal" and "elevated" prolactin is ill-defined. The possibility of macroprolactinemia complicates this matter even further. 10. The number of drugs which suppress prolactin by acting on pituitary D2 receptors, and which are useful in the treatment of hyperprolactinemia, continues to increase. In the field of ergot alkaloids, parenteral application appears to be a logical solution to the problem of the high first-pass effect; in addition, this form of treatment is frequently better tolerated than the oral route. 11. Prolactinoma development is presently being studied employing molecular biological techniques; the question of whether tumorigenesis can be attributed to specific defects of gene regulation remains to be answered.

Adult↗

Plasma and tumor prolactin in colorectal cancer patients.

Fifteen consecutive patients with recently diagnosed colorectal cancer were studied for plasma and tumor tissue prolactin content. In eight patients (four men and four postmenopausal females), preoperative high plasmatic prolactin was found (mean 1553 nmol; range 516-3677 nmol). In three of them, prolactin was also present in the tumor cells. All plasma prolactin levels returned to normal after tumor resection and remained so during a three-month follow-up. The tumor stage by Duke distribution was similar for both high and normal plasmatic prolactin patients. The role of prolactin in the pathogenesis of colorectal cancer, and as a marker of the tumor, remains to be established. This is the first time that prolactin has been detected in human colon cancer.

Aged↗

Routine prolactin measurement is not necessary in the initial evaluation of male impotence.

The authors determined the prevalence of hyperprolactinemia in impotent men in a community setting and assessed the cost of case detection with routine estimation of serum prolactin. They recruited 299 consecutive patients with impotence and determined the hormonal levels (prolactin, luteinizing hormone, follicle-stimulating hormone, and testosterone). Pituitary gland imaging was done when the prolactin level was elevated. Simultaneous prolactin and testosterone levels were available for 212 patients. Three patients (1.4%) had elevated prolactin levels but none had pituitary tumor. Two of these had low testosterone levels. Overall, 51 patients (24.1%) had low testosterone levels. Cost of selective prolactin estimation in patients with low testosterone levels resulted in a net saving of $2,574 per case detected. The authors conclude that the prevalence of hyperprolactinemia in impotence is low. Routine measurement of prolactin levels in impotence is not indicated. Selective determination in patients with low testosterone reduces the cost of diagnostic evaluation.

Costs and Cost Analysis↗

Prolactin pulsatility in polycystic ovarian disease.

Secretion patterns of LH and prolactin were studied in 5 nonobese (group A) and 7 obese (group B) women with polycystic ovaries (PCO) by taking blood samples at 15-min intervals for 6 h. Serum LH level showed a distinct pulsatility in all patients. There was no significant difference in the mean pulse rate of LH between group A (6.2 +/- 0.45, mean +/- SE) and group B (5.8 +/- 0.24). A pulsatile secretion pattern in prolactin was also found in 11 of the 12 PCO patients studied, the pulse rate being 3.2 +/- 1.0 in group A and 3.9 +/- 0.45 in group B. 79% of prolactin pulses coincided with those of LH but their amplitude was significantly lower than that of LH. In the total group of PCO patients a positive correlation was found between the serum concentrations of prolactin and free estradiol (r = 0.85), and prolactin and estrone (r = 0.78) indicating that estrogens participate in the regulation of prolactin secretion in polycystic ovarian disease. Four patients showed a steep decrease in the plasma prolactin level at the beginning of the study period suggesting a sensitive stress response.

Adult↗

Reciprocal effects of prolactin and thyroxine on the growth rate of rat pups from weaning to puberty.

Prolactin and thyroxine were added to the daily diet of rat pups from weaning to puberty (days 20--40). While prolactin significantly retarted the pups' growth, thyroxine stimulated growth and even compensated for the depression caused by prolactin. The significant retardation in the growth of the prolactin-treated pups around days 30--35 of age indicates that at that period they are extremely prolactin-dependent, thus confirming previous reports that exogenous prolactin inhibits its own endogenous secretion. On the other hand, the possible growth promotion caused by thyroxine is mild and does not initiate a feedback mechanism. This suggests that prolactin is indispensable for the normal development of the weaned, immature rat.

Animals↗

Prolactin-secreting macroadenoma in a prepubertal girl.

Prolactin-secreting adenoma, which usually presents with amenorrhea and galactorrhea syndrome, is quite rarely diagnosed in the prepubertal age group. We reported a rare case of a prepubertal prolactin-secreting adenoma and discuss its clinical, radiological and histological features. An 8-year-old girl presented with headache, progressive visual deterioration and precocious puberty. The serum prolactin level was 57.8 ng/ml. Computerized tomography and magnetic resonance imaging revealed an invasive suprasellar tumor. The tumor was partially resected through an interhemispheric approach in a first operation, and residual tumor was resected through the right pterional approach in a second operation. The histological diagnosis was a prolactin-secreting adenoma with high cellular pleomorphism. The Ki-67 labeling index was 5.7%, indicating aggressive biological behavior. Postoperatively, the patient was prescribed bromocriptine as maintenance therapy, and the serum prolactin level became normalized. There is a tendency for diagnosis of a prepubertal prolactin-secreting adenoma to be delayed because there are no endocrinological manifestations. Therefore, the tumor tends to become larger and invasive. Although it is rarely experienced, a prolactin-secreting adenoma should be considered in the differential diagnosis of a large, invasive parasellar lesion in the prepubertal age group.

Bromocriptine↗

Possible individual and gender differences in the small increases in plasma prolactin levels seen during clozapine treatment.

In vitro, animal studies and acute short-term clinical studies suggest clozapine releases prolactin but the effect is much smaller than that of typical antipsychotics. Repeated early morning trough measures of plasma clozapine and prolactin levels on each subject were studied during the course of a double-blind dose-response clozapine study. After a 4-week 10 mg/day haloperidol trial and a one-week washout, treatment- refractory schizophrenics were successively randomized to 100, 300,or 600 mg/day of clozapine for a 16- week treatment. The statistical analyses included 35 subjects (19 females and 16 males). The within-subject correlation of prolactin levels was 0.32 with clozapine levels and 0.75 with haloperidol levels. An increment of 100 ng/ml in clozapine level yielded an average increment of 0.45 ng/ml of prolactin levels in females and of 0.15 ng/ml in males. An increment of 1 ng/ml in haloperidol level yielded an average increment of 2.6 ng/ml of prolactin levels in females and of 1.5 ng/ml in males. At least one fourth of patients demonstrated a significant and strong (r > 0.6) correlation between clozapine and prolactin levels. This study suggests that clozapine has effects on prolactin levels but effects are small and may be more evident in some individuals, particularly females.

Adult↗

N5 endometrial stromal cell line: a model system to study decidual prolactin gene expression.

Prolactin gene expression in extrapituitary tissues, such as decidua and lymphocytes, is regulated by a distinct promoter approximately 6 kb upstream of the pituitary prolactin gene transcription start site. Here we describe studies in a human endometrial stromal cell line, N5, that was immortalized by transfection with an SV40 mutant and which expresses the prolactin gene driven by the extrapituitary promoter. The N5 cells have phenotypic features of primary cultures of decidualized human endometrial stromal cells and secrete low levels of prolactin and insulin-like growth factor binding protein-1 (IGFBP-1), both of which are markers of decidualized endometrial stromal cells. As in primary cultures of endometrial stromal cells, treatment of N5 cells with progesterone and estradiol alone or in combination with prostaglandin E2 stimulated the synthesis and release of prolactin. Transient transfection of the N5 cells with an expression vector containing - 2927/ + 66 bp of the decidual prolactin promoter coupled to a luciferase reporter gene resulted in a 20 to 25-fold increase in luciferase activity, a magnitude similar to that which occurs in primary cultures of endometrial stromal cells decidualized in vitro by treatment with progesterone and estradiol. Luciferase expression levels were similar in untreated N5 cells and N5 cells treated with progesterone and estradiol. Taken together, these results indicate that the N5 human endometrial stromal cell line has phenotypic characteristics of normal decidualized stromal cells and is a useful model to study regulation of decidual prolactin gene expression.

Bucladesine↗

Epithelial regulation of prolactin effect on amnionic permeability.

The permeability of human amnion to tritiated water is reduced in the presence of both human and ovine prolactin. The cellular composition of amnion is such that the action of prolactin on this membrane probably occurs by way of the epithelium lining the fetal surface. The present study sought to confirm an epithelial site of action of prolactin on the permeability of amnionic membrane to tritiated water. In addition, radioautography and competition experiments were conducted to determine a possible receptor-mediated mechanism for prolactin action. Membrane permeability to tritiated water was found to be equivalent for both intact membranes and membranes enzymatically stripped of the lining epithelial cells. However, when ovine prolactin was presented to the fetal surface of amnion, only intact membrane displayed decreased permeability to tritiated water. Although localization of iodine 125-labeled prolactin to the light cell population of amniotic epithelium was observed, positive evidence of a receptor-mediated mechanism could not be established. The results indicate that the permeability of human amnion to water is influenced principally by cells of the epithelium in response to prolactin.

Autoradiography↗

An enzyme-linked immunosorbent assay for rat prolactin.

A sensitive and specific competitive enzyme-linked immunosorbent assay (ELISA) for rat prolactin was developed using reagents from the National Institute of Arthritis, Diabetes, Digestive Diseases and Kidney. In this assay soluble prolactin and prolactin adsorbed to a solid-phase support compete for rabbit anti-prolactin antibody binding sites. Therefore, a high concentration of soluble prolactin in the sample will result in a low concentration of antibody immobilized to the adsorbed prolactin. The immobilized antibody-prolactin complex is detected and quantified using goat anti-rabbit immunoglobulin G covalently conjugated to the enzyme horseradish peroxidase. Assay parameters were optimized by investigating the concentration of reagents and the reaction kinetics in each of the assay steps. The assay can be performed in 24 h. A sensitivity range of 0.06 to 6 ng in the region of 90 to 10% binding was obtained. Near 50% binding (0.6 ng), the intraassay coefficient of variation (CV) was 4.2% and the interassay CV was 7.6%. The correlation between radioimmunoassay and the ELISA was 0.868. Selected applications of the assay are described. The assay should prove a useful alternative to the radioimmunoassay in those instances where steps involving the use of 125I become limiting, for example, iodination facility and gamma counter availability or prolonged reagent storage.

Animals↗

Studies of the neural mechanisms by which hypothyroidism decreases prolactin secretion in the rat.

These studies report that chronic hypothyroidism in the rat is accompanied by decreased serum prolactin. An investigation of those mechanisms by which hypothyroidism decreases prolactin secretion revealed the following findings: (1) activation of tyrosine hydroxylase in the median eminence (ME) and increase in the turnover-rate of dopamine (DA) in the ME; (2) a decreased content and in vitro release of prolactin by pituitaries from hypothyroid rats; (3) decreased [3H]spiroperidol binding to pituitary homogenates obtained from hypothyroid rats, and (4) normal increase of serum prolactin following administration of haloperidol. Comparison of the effects of hypothyroidism on dopaminergic terminals of the striatum with those of the ME was made and the following tentative conclusions proposed. Hypothyroidism presumably increases the release of DA into the pituitary portal system. A deficiency of thyroid hormone desensitizes the pituitary lactrotrope to inhibition by DA, decreases the total pituitary prolactin content and presumable also reduces the peripheral catabolism of prolactin. The overall net effect of hypothyroidism is, therefore, a decrease in serum prolactin levels which can increase normally following haloperidol. The mechanism by which a deficiency of thyroid hormone alters the function of the tuberoinfundibular and striatal dopaminergic systems is unknown.

Animals↗

Evidence that the medial and dorsal raphe nuclei mediate serotonergically-induced increases in prolactin release from the pituitary.

Electrolytic lesions of the medial (MR) or dorsal (DR) raphe nucleus significantly antagonized serum prolactin elevations produced by 5-hydroxytryptophan (5-HTP) in rats pretreated with fluoxetine or citalopram, (serotonin (5-HT) uptake blockers). Lesioned animals in which total blockade of serum prolactin elevations was observed also had total blockade of 5-HT accumulation in the median eminence. However, the increase in serum prolactin levels produced by 5-HTP plus 5-HT reuptake blockade in lesioned rats was not significantly different from sham-operated rats if as little as 15-20% of control median eminence accumulation was present. Serum prolactin elevations produced by quipazine, a direct acting 5-HT agonist, were not significantly affected by MR lesions. On the basis of these results, we suggest that: (1) serum prolactin elevations following 5-HT reuptake blockade plus 5-HTP are correlated with 5-HT concentration in the median eminence; (2) lesion-induced antagonism of 5-HTP-induced prolactin elevation is critically dependent upon complete blockade of median eminence 5-HT accumulation; and (3) 5-HT neurons arising from cell bodies located in the MR and DR are necessary for endogenous serotonergically mediated effects on prolactin secretion in the rat.

Animals↗

Pharmacological studies on stress-induced renin and prolactin secretion: effects of benzodiazepines, naloxone, propranolol and diisopropyl fluorophosphate.

Stress-induced renin and prolactin secretion was investigated using a conditioned emotional response paradigm. Three minutes after placement in a chamber the rats received an electric shock to their feet via the grid floor, then were immediately returned to their home cage. This procedure was repeated for 3 consecutive days. On the fourth day, instead of receiving an electric shock, they were removed after 3 min and sacrificed by decapitation. Control rats were treated identically with the exception that shock was not administered at any time. There was a significant increase in plasma renin activity and prolactin level in the stressed rats. The administration of the antianxiety drugs chlordiazepoxide (10 mg/kg i.p.) or midazolam (0.125-2 mg/kg i.p.) blocked the stress-induced increase in prolactin levels but not the stress-induced rise in plasma renin activity. Administration of the beta-blocker propranolol (1 mg/kg i.p.) inhibited, but did not completely block, stress-induced rise in plasma-renin activity and had no effect on stress-induced prolactin secretion. The opiate antagonist naloxone (0.1-10 mg/kg i.p.) and the acetylcholinesterase inhibitor diisopropyl fluorophosphate (0.5 mg/kg i.p.) did not block stress-induced renin or prolactin secretion. It is concluded that stress-induced prolactin secretion is regulated by a benzodiazepine-mediated mechanism and that stress-induced renin but not prolactin secretion is mediated in part via beta-receptors.

Animals↗