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Adaptation to the environment is essential for survival, in all wild animal species seasonal variation in temperature and food availability needs to be anticipated. This has led to the evolution of deep-rooted physiological cycles, driven by internal clocks, which can track seasonal time with remarkable precision. Evidence has now accumulated that a seasonal change in thyroid hormone (TH) availability within the brain is a crucial element. This is mediated by local control of TH-metabolising enzymes within specialised ependymal cells lining the third ventricle of the hypothalamus. Within these cells, deiodinase type 2 enzyme is activated in response to summer day lengths, converting metabolically inactive thyroxine (T4) to tri-iodothyronine (T3). The availability of TH in the hypothalamus appears to be an important factor in driving the physiological changes that occur with season. Remarkably, in both birds and mammals, the pars tuberalis (PT) of the pituitary gland plays an essential role. A specialised endocrine thyrotroph cell (TSH-expressing) is regulated by the changing day-length signal, leading to activation of TSH by long days. This acts on adjacent TSH-receptors expressed in the hypothalamic ependymal cells, causing local regulation of deiodinase enzymes and conversion of TH to the metabolically active T3. In mammals, the PT is regulated by the nocturnal melatonin signal. Summer-like melatonin signals activate a PT-expressed clock-regulated transcription regulator (EYA3), which in turn drives the expression of the TSHβ sub-unit, leading to a sustained increase in TSH expression. In this manner, a local pituitary timer, driven by melatonin, initiates a cascade of molecular events, led by EYA3, which translates to seasonal changes of neuroendocrine activity in the hypothalamus. There are remarkable parallels between this PT circuit and the photoperiodic timing system used in plants, and while plants use different molecular signals (constans vs EYA3) it appears that widely divergent organisms probably obey a common set of design principles., (© 2014 The authors.)

Folliculostellate (FS) cells in the anterior pituitary gland appear to have multifunctional properties. FS cells connect to each other at gap junctions and thereby form a histological and functional network. We have performed a series of studies on network formation in FS cells and recently reported that FS cells markedly prolong their cytoplasmic processes and form numerous interconnections with neighboring FS cells in the presence of laminin, an extracellular matrix (ECM) component of the basement membrane. In this study, we investigated the mechanism of this extension of FS cell cytoplasmic processes under the influence of laminin and found that laminin promoted stress fiber formation within FS cells. Next, we noted that formation of stress fibers in FS cells was mediated by syndecan-4, a transmembrane proteoglycan that binds ECM and soluble factors via their extracellular glycosaminoglycan chain. We then observed that expressions of syndecan-4 and α-actinin (a microfilament bundling protein that cross-links actin stress fibers in FS cells) were upregulated by laminin. Using specific siRNA of syndecan-4, actin polymerization of FS cells was inhibited. Our findings suggest that FS cells received a signal from laminin-syndecan-4 interaction, which resulted in morphological changes, and that the formation of a morphological and functional network in FS cells was transduced by a syndecan-4-dependent mechanism in the presence of ECM.

Folliculo-stellate (FS) cells in the anterior pituitary gland appear to possess multifunctional properties. Recently, the development of transgenic rats (S100b-green fluorescent protein (GFP) rats) that express GFP specifically in FS cells in the anterior pituitary gland has allowed us to distinguish and observe living FS cells in other kinds of pituitary cells. We used S100b-GFP rats to investigate the topographic affinity of FS cells for other pituitary cells. We observed living FS cells in enzymatically dispersed anterior pituitary cells of S100b-GFP rats under a fluorescent microscope, and noted that FS cells markedly extended and contracted cytoplasmic processes and formed interconnections with neighboring FS cells. In addition, FS cells adhered to small clusters of GFP-negative cells, which were primarily hormone-producing cells, and these clusters further aggregated during the course of cytoplasmic contraction. In the presence of laminin, fibronectin, and varying types of collagen, FS cells showed marked changes in shape and specific proliferative activity; however, GFP-negative cells did not. On reverse transcription-PCR analysis and immunohistochemistry, FS cells were shown to express integrin subunits, which are the cell surface receptors for extracellular matrix (ECM). In the anterior pituitary gland, FS cells and the various types of hormone-producing cells generate a unique topography in the presence of basement membrane components and interstitial collagens. The novel characteristics of FS cells observed in the present study suggest that in the anterior pituitary gland, FS cells play important roles in determining and/or maintaining local cellular arrangement in the presence of ECM components.

To further delineate the role of estradiol in the IGF system an experiment was conducted to determine the dosage of the aromatase inhibitor, anastrozole, needed to decreases serum concentrations of estradiol-17beta (E2) in maturing boars. A second experiment was conducted to determine if administration of anastrozole to growing boars decreased serum concentrations of E2 and affected components of the serum and anterior pituitary gland (AP) IGF system vs untreated boars and barrows. In Experiment 1, 12 crossbred boars (292 days, 158 kg) were administered either 0, 1 or 10 mg/day anastrozole (n = 4/group) beginning on day 1. Blood samples were collected every 7-14 days. Mean serum concentrations of E2 were decreased (P < 0.05) in the 10 mg group vs the 0 and 1 mg groups by day 36; however, no difference (P > 0.05) existed between the 0 and 1 mg groups. In Experiment 2, 24 crossbred boars and 12 barrows (101 days, 44 kg) were stratified by litter to one of three treatment groups (n = 12): boars administered 10 mg/day anastrozole, boars administered 0 mg/day, and barrows administered 0 mg/day. Blood samples were collected and pigs were weighed on day 0 and every 14 days thereafter, then killed on day 84 when blood and APs were collected. The 10 mg/day pigs were fed the anastrozole-amended diet beginning on day 1. Mean serum concentrations of E2 did not differ (P > 0.05) between the 10 mg/day pigs and 0 mg/day pigs on day 0; however, on day 15 through to 84 mean serum concentrations of E2 were greater (P < 0.05) in 0 mg/day pigs than in the 10 mg/day pigs. Mean percentage increase in serum concentrations of IGF-I was greater (P < 0.05) in untreated boars than anastrozole-treated boars and barrows from day 58 through to 84. Mean percentage of basal IGF-I increased (P < 0.05) from day 29 through to 84 in untreated boars. Mean relative amounts of AP IGF-binding protein (IGFBP)-2 and -5 were less (P < 0.01) in 10 mg/day pigs than in the 0 mg/day pigs, but each was greater (P < 0.01) than in barrows administered 0 mg/day. These results indicate anastrozole administered at a dosage of 10 mg/day suppresses serum concentrations of E2 in pigs. Administration of anastrozole to boars reduced the percentage increase in serum concentrations of IGF-I and relative amounts of AP IGFBP-2 and -5. These data further support a role for E2 in regulating components of the IGF system in pigs.

Originally searched for and eventually isolated as factors of hypothalamic origin controlling anterior pituitary secretions, these hypophysiotropic peptides are now a chapter of physiology and medical endocrinology of their own. Defying the concept of 'neuropeptides' they and their receptors are now known to be ubiquitous and to have subtle as well as profound effects on a large number of functions of both soma and psyche. This review will be composed of brief essays on current knowledge of each of the original 'hypothalamic hormones', TRH, GnRH, somatostatin, GHRH and corticotropin releasing hormone and will close on possible and probable futures.

Adenosine receptors are widely distributed in most species and mediate a diverse range of physiological and pathological effects. Although adenosine receptors have been identified in the pituitary gland, the distribution of the individual subtypes (A(1), A(2A), A(2B), A(3)) has not been well defined. Furthermore, the effects of adenosine on pituitary trophic activity and function are not well established despite good evidence for growth- and immune-modulating properties of the nucleoside elsewhere. Recent advances have provided a more detailed description of adenosine receptor distribution and function in the anterior pituitary and this commentary reviews these observations and highlights some of the possible implications in relation to the control of the hypothalamic-pituitary-adrenal axis and the regulation of inflammation and pituitary cell growth.

A complete inventory of pituitary trophic responses depends on precise estimates of mitotic activity and apoptotic events, and accurate characterization and quantification of pituitary cell subtypes irrespective of previous and current physiological demand. For a discrete structure that has been so extensively studied, it is disappointing but perhaps not surprising that none of these measures is available and therefore that the relative contributions to changes in the proportions of pituitary cellular subpopulations of trophic activity, differentiation of pluripotent cells and variations in the secretory profiles of apparently committed cells remain almost impossible to determine. To fully appreciate the extent of this dilemma, it should be remembered that conservative estimates of the proportion of corticotrophs in the rat anterior pituitary under basal conditions vary over twofold and that it is still not clear whether the apparent threefold increase in mammotrophs during pregnancy is the result of maturation of uncommitted cells, transdifferentiation of other cells such as somatotrophs, cell division, or a mixture of all three. Equally, while it has been known for some time that adrenalectomy results in a transient increase in anterior pituitary mitotic activity and appropriately timed supraphysiological glucocorticoid replacement with a wave of apoptosis, the precise identity of the cells involved in both of these responses is open to question. Thus, although many of the physiological stimuli associated with apparent changes in the proportions of pituitary cellular subpopulations are known, the precise mechanism of the changes and the consequences of the same remain obscure. This review summarizes the limited literature on pituitary trophic activity and asks what, if anything, analysis of pituitary trophic activity using current technology can tell us.

The pineal hormone, melatonin, is uniquely defined by its role as hormonal time, but the processes whereby cells extract temporal information from the melatonin signal are not understood. Melatonin receptors are expressed in the pars tuberalis (PT) and, during fetal and perinatal life, in the pars distalis (PD). Functional studies suggest that the PT mediates the seasonal effects of melatonin on prolactin secretion, whilst the PD may be involved in photoperiodic programming of the developing gonadotrophic axis. To understand these effects at the cellular level we need to know the phenotype of melatonin-responsive cells. This review summarises current understanding in this area, and highlights present shortcomings. A case is presented for exploring the hypothesis that there is a functional association between melatonin receptor expression and cell differentiation in the anterior pituitary.

Transport of thyroxine (T(4)) into the liver is inhibited in fasting and by bilirubin, a compound often accumulating in the serum of critically ill patients. We tested the effects of chronic and acute energy deprivation, bilirubin and its precursor biliverdin on the 15-min uptake of [(125)I]tri-iodothyronine ([(125)I]T(3)) and [(125)I]T(4) and on TSH release in rat anterior pituitary cells maintained in primary culture for 3 days. When cells were cultured and incubated in medium without glucose and glutamine to induce chronic energy deprivation, the ATP content was reduced by 45% (P<0. 05) and [(125)I]T(3) uptake by 13% (NS), but TSH release was unaltered. Preincubation (30 min) and incubation (15 min) with 10 microM oligomycin reduced ATP content by 51% (P<0.05) and 53% (P<0. 05) under energy-rich and energy-poor culture conditions respectively; [(125)I]T(3) uptake was reduced by 66% (P<0.05) and 64% (P<0.05). Neither bilirubin nor biliverdin (both 1-200 microM) affected uptake of [(125)I]T(3) or [(125)I]T(4). Bilirubin (1-50 microM) did not alter basal or TRH-induced TSH release. In conclusion, the absence of inhibitory effects of chronic energy deprivation and bilirubin on thyroid hormone uptake by pituitary cells supports the view that the transport is regulated differently than that in the liver.

The effects of gonadotrophin-releasing hormone analogue (GnRHa; buserelin) on the pituitary function and morphology of food-restricted rats were compared with those of authentic GnRH. After adult female rats had been restricted to 10 g food/day for 60 days, various doses of GnRHa (10 ng, 100 ng and 1 microgram) or GnRH (10 micrograms) were administered either daily for 7 days or twice a week for 4 weeks from day 61 of the period of underfeeding. Underfeeding brought about a decrease in the pituitary gonadotrophin content, serum levels of gonadotrophins and oestradiol, and the number and size of both LH- and FSH-positive pituitary cells. Daily and/or twice-weekly administration of authentic GnRH to underfed rats produced an increase in pituitary and serum gonadotrophin levels and the number and size of both LH- and FSH-positive pituitary cells. The administration of GnRHa daily for 7 days increased serum gonadotrophin levels, while it produced a reduction in the pituitary gonadotrophin content and number and size of both LH- and FSH-positive pituitary cells in a dose-dependent manner. Twice-weekly administration of GnRHa also produced an elevation of serum gonadotrophin levels and reduction of pituitary gonadotrophin content, although it did not affect the numbers and areas of LH- and FSH-positive pituitary cells. A GnRH loading test performed after the GnRHa treatment showed that the GnRHa treatment performed in this study did not produce down-regulation of the GnRH receptor. Thus, it can be concluded that the gonadotrophin-synthesizing activity of GnRHa is weaker than that of authentic GnRH, or that GnRHa may preferentially exert gonadotrophin-releasing activity rather than gonadotrophin-synthesizing activity in the anterior pituitary of underfed rats.

We have examined the effects of dopamine on prolactin gene expression using quantitative in-situ hybridization histochemistry in different pituitary cell (sub)populations separated according to their density on a discontinuous Percoll gradient. Administration of dopamine resulted in a drastic reduction in hybridization of 35S-labelled DNA probe complementary to prolactin mRNA in total pituitary cells and in lactotrophs with low density. In contrast, dopamine significantly stimulated mRNA accumulation in prolactin-secreting cells with high density compared with other cell layers. The combined use of Percoll gradient and quantitative in-situ hybridization is a valuable and sensitive method with which to examine prolactin-secreting cell response to a given stimulation. Prolactin-secreting cells with high and low density clearly show functional heterogeneity in their response to dopamine.

To investigate the effect of cell density on rat somatotrophs, dispersed adult rat anterior pituitary cells were plated at several densities (0.6, 1.2, 2.4 and 6.0 x 10(5) cells/cm2) or at two densities (6.07 x 10(5) and 1.21 x 10(5) cells/cm2) as high and low densities respectively. A static incubation system was used to study the release of GH and peptidyl glycine alpha-amidating enzyme (PAM; a component of secretory granules) and the cellular concentration of cyclic AMP (cAMP) in basal and stimulated cells. In addition, a perifusion system was used to characterize the sensitivity to GH-releasing factor (GRF) at both high and low densities. The high density of cells in both the static incubation and perifusion systems caused a low basal secretion rate of GH and PAM. When the cultured cells were stimulated with human GRF (hGRF) in perifusion, GH secretion from cells at high density was markedly higher than that from cells at low density. The cAMP content in the static incubation system showed a similar tendency to that observed for basal and stimulated GH secretion; that is, the basal cAMP content was increased as the cell density decreased. The cellular concentration of cAMP was increased by about threefold by 1 nmol hGFR/l and by more than tenfold by 10 nmol hGRF/l. When the medium from cells cultured at low density was replaced by that from the cells at high density, there was no change in the basal cellular cAMP content of the cells at low density.(ABSTRACT TRUNCATED AT 250 WORDS)

The precise roles of GH-releasing factor (GRF) and somatostatin (SRIF) in the orchestration of pulsatile GH secretion have not yet been fully determined. We examined the interactions of rat GRF and SRIF in the concentration ranges present in rat hypophysial-portal blood, on the secretion of GH from dispersed male rat anterior pituitary cells in monolayer culture. The effects of exposing cells to GRF and/or SRIF (0.01-1.nmol/l) for 1 h were compared with the effects of preincubation of cells with SRIF before experimental incubations. As anticipated, the stimulatory effects of 0.1-1 nmol GRF/1 were abolished by concurrent incubation with SRIF at an equimolar concentration, although SRIF, at these concentrations, did not significantly inhibit basal GH secretion. Conversely, pre-exposure to 0.1 nmol SRIF/1 for 30 or 60 min, resulted in an increase in GH secretion during a subsequent 60-min incubation period, both in the absence or in the presence of GRF (0.01-1 nmol/l). Pretreatment with GRF caused increased responsivity to GRF rather than significant sensitization of the GH response to GRF. These observations demonstrate actions of SRIF, at low and probably physiological concentrations, which are more complex than those of a pure inhibitor of GH secretion. Pre-exposure of the pituitary to SRIF enhances subsequent GH secretion, suggesting that SRIF may play an additional physiological role in amplifying the GRF signal.

In-situ hybridization with synthetic oligonucleotide probes was used to determine the mRNA content of corticotrophin-releasing factor (CRF) and proenkephalin A mRNA in the paraventricular nucleus, and of pro-opiomelanocortin (POMC) mRNA in the anterior pituitary gland of male rats immediately after, and during recovery from, chronic high-dose prednisolone treatment. Levels of transcripts for mRNA for both CRF and POMC were markedly reduced after the treatment, but there was a rapid return to control values for CRF mRNA within 18 h of steroid withdrawal. In untreated animals, the stressful stimulus of i.p. hypertonic saline increased CRF and proenkephalin A mRNA within 4 h with no significant difference in response seen whether the tissues were removed at 13.00 or 20.00 h. The increase in POMC mRNA did not reach statistical significance in these animals. Although prednisolone resulted in a marked reduction of basal CRF mRNA, the stress-induced increment of CRF mRNA remained comparable with that found in untreated animals. On the day following cessation of prednisolone treatment at 09.00 h, basal and stress levels of CRF mRNA were significantly higher in rats killed at 20.00 h than at 13.00 h. Proenkephalin A mRNA transcripts were below quantifiable levels of detection in control or non-stressed prednisolone-treated animals at all the time-points studied. Stress, however, resulted in the accumulation of proenkephalin A mRNA in control animals. This response was inhibited by prednisolone treatment and only returned 18 h after withdrawal. Prednisolone treatment reduced POMC mRNA below the levels detected in untreated animals, with no detectable response to stress.(ABSTRACT TRUNCATED AT 250 WORDS)

Circulating GH concentrations were suppressed 24 h after intracerebroventricular (i.c.v.) injection of native or recombinant DNA-derived chicken GH (rcGH) (10 micrograms in 4 microliters vehicle) into the right lateral ventricles of conscious 6-week-old cockerels. Plasma concentrations of LH were suppressed within 1 h of i.c.v. administration of GH at doses of 1 or 10 micrograms, and remained suppressed for 24 h in birds injected with the highest concentration of GH. In contrast, concentrations of plasma prolactin were increased 24 h after i.c.v. administration of GH (10 micrograms). The i.v. administration of rcGH (at concentrations of 10 or 100 micrograms/kg) did not mimic the effects of i.c.v. injection of GH. These results demonstrate central effects of GH on pituitary function in the domestic fowl.

The past 2 years have seen dramatic progress in our understanding of the molecular basis of prolactin gene expression in the pituitary gland. The identification of the pituitary-specific factor Pit-1/GHF-1 has been a major advance, and it leads to new questions, such as the basis for the tissue-specificity of Pit-1/GHF-1 expression itself. It clearly cannot be the only factor involved, as it is found in three pituitary cell types, yet prolactin is expressed in only one of these. It is probable that a network of trans-acting factors, some tissue-specific and some ubiquitous but hormonally activated, interact to determine the overall pattern of expression of the prolactin gene, and the dissection of this system should provide some valuable insights into endocrine gene regulation in general.

The mode of action of a recently isolated gonadal protein, termed FSH-suppressing protein (FSP) or follistatin, on basal and gonadotrophin-releasing hormone (GnRH)-stimulated release of FSH and LH and on pituitary cell content of FSH and LH was examined in rat pituitary cell cultures and compared with the previously reported effects of inhibin. Pituitary cells were cultured for 3-9 days in the presence of graded doses of FSP and the basal release rates and changes in cell contents of FSH and LH determined during this period. FSP suppressed both the basal release rate and the cell content of FSH with median inhibitory concentrations (IC50) of 135 and 161 pmol/l respectively. The corresponding effects of FSP on LH basal release rate and LH cell content (IC50 = 200 pmol/l) were limited compared with the effects on FSH. The effect of FSP on GnRH-stimulated release of FSH and LH during 4 h was determined in cells which had been preincubated with FSP for 3 days, and the GnRH-stimulated release of FSH and LH analysed as a percentage of the respective gonadotrophin available for release. FSP antagonized GnRH action with dose-related increases in the GnRH median effective (stimulatory) concentrations for FSH and LH release (EC50 values = 56 and 400 pmol/l respectively) and a suppression in the maximum release of FSH and LH by excess GnRH (IC50 values = 142 and 150 pmol/l respectively). The effect of FSP on FSH cell content after 3 days in culture was insensitive to the neutralizing effects of an inhibin antiserum.(ABSTRACT TRUNCATED AT 250 WORDS)

Placental lactogen has been detected in the field vole, Microtus agrestis, and the bank vole, Clethrionomys glareolus, using a co-culture technique. In field voles this activity could be detected from about day 8 of pregnancy to shortly before term, and stimulated both mouse and vole mammary gland to secrete in vitro. Partial immunological cross-reaction was detected in a radioimmunoassay system between rat prolactin and either extracts of vole pituitaries or media on which vole pituitaries had been cultured; vole placental lactogen showed no cross-reaction with rat prolactin. One site of origin for this hormone is probably the trophoblastic giant cells.

Plasma and testicular testosterone concentrations in the frog, Rana esculenta, were studied by radioimmunoassay and showed similar seasonal fluctuations. The increase in testicular androgen during November preceded that occurring in the plasma by 2 months. Pituitary products and gonadotrophin releasing hormone, and the responsiveness of the testis to these substances play an important role in determining the hormone profile.

The role of nerve cells of the arcuate nucleus and endogenous opioid peptides in the regulation of GH and prolactin secretion has been investigated. Electrical stimulation of the medial-basal hypothalamus (MBH) for 10 min raised plasma levels of both hormones in male rats anaesthetized with pentobarbitone sodium. Plasma hormone levels increased within 5 min after the termination of the stimulus, while no marked changes were found during stimulation. The GH response to the electrical stimulus was substantially reduced in rats with arcuate lesions induced by neonatal treatment with monosodium-L-glutamate (MSG). By contrast, the size of the prolactin response was not altered by MSG treatment. The opiate receptor antagonist naloxone (10 mg/kg, i.v.) failed to influence GH secretion induced by electrical stimulation in either control or MSG-treated animals. The post-stimulus rise of plasma prolactin levels was attenuated by naloxone in control rats, while the same dose of the drug was ineffective in rats which had been exposed to MSG. We conclude that endogenous opioids participate in the increase of prolactin release upon electrical stimulation of the MBH but are not involved in the GH secretory response. Arcuate neurones are important in the maintenance of the GH response to electrical stimulation. By contrast, lesioning of the arcuate nucleus failed to affect the prolactin secretory response elicited by MBH stimulation. However, prolactin release in MSG-treated rats appeared less susceptible to the inhibitory action of naloxone, suggesting a possible supersensitivity towards endogenous opioids.

Two experiments were conducted with ovariectomized and hypothalamo-pituitary disconnected (HPD) ewes to ascertain the pattern of inputs, to the pituitary gland, of gonadotrophin-releasing hormone (GnRH) necessary for the full expression of an oestrogen-induced LH surge. The standard GnRH replacement to these sheep was to give pulses of 250 ng (i.v.) every 2h; at the onset of experimentation, pulses were given hourly. In experiment 1, groups of sheep (n = 7) were given an i.m. injection of 50 micrograms oestradiol benzoate, and after 10 h the GnRH pulse frequency or pulse amplitude was doubled. Monitoring of plasma LH concentrations showed that a doubling of pulse frequency produced a marked increase in baseline values, whereas a doubling of amplitude had little effect on the LH response. In a second experiment, ovariectomized HPD sheep that had received hourly pulses of GnRH for 16 h after an i.m. injection of oil or 50 micrograms oestradiol benzoate were given either a 'bolus' (2.25 micrograms GnRH) or a 'volley' (500 ng GnRH pulses 10 min apart for 30 min, plus a 500 ng pulse 15 min later). Both groups then received GnRH pulses (250 ng) every 30 min for the next 13 h. Oestrogen enhanced the LH responses to the GnRH treatments, and the amount of LH released was similar in ovariectomized HPD ewes given oestrogen plus bolus or volley GnRH treatments and ovariectomized hypothalamo-pituitary intact ewes given oestrogen. These results suggest that the oestrogen-induced LH surge is initiated by a 'signal' pattern of GnRH secretion from the hypothalamus.

Alpha-Melanocyte-stimulating hormone was shown to act synergistically with testosterone to stimulate the sebaceous, prostate and the seminal vesicles in hypophys-ectomized-castrated rats. The sebaceous glands differed from the other three organs in that alpha-MSH not only acted synergistically, but also had a significant effect which was independent of the presence of exogenous testosterone. The response of the brown adipose tissue to testerone, considerably reduced by hypophysectomy, was not restored by alpha-MSH. The Harderian and lachrymal glands were also pituitary-dependent and their weights in hypophysectomized-castrated rats were not restored by alpha-MSH.

Seven morphologically and tinctorially distinct types of cells (types 1--7) have been distinguished in the pars anterior of the pituitary gland of the musk shrew (Suncus murinus L.). On the basis of their responses to various experimental stimuli, these cell types were correlated with the secretion of various trophic hormones. Type 1 cells exhibited conspicuous changes after thyroidectomy or inactivation of the thyroid gland and hence appeared to be the source of TSH. Types 2 and 3 cells responded to gonadectomy and administration of androgens, which suggests that they were associated with gonadotrophin secretion. The granules of the type 2, but not the type 3 cells could be extracted with 10% trichloroacetic acid, which may indicate that type 2 and 3 cells secrete FSH and LH respectively. After the administration of either reserpine or oestrogen, the type 4 cells underwent hypertrophy and hyperplasia, which suggests that they were the likely source of prolactin. Type 6 cells, which are distinguishable from type 4 cells by their thinly dispersed erythrosinophilic granulation, showed conspicuous changes after unilateral adrenalectomy, administration of metyrapone or exposure to stress and may therefore be responsible for secretion of ACTH. Type 5 cells tinctorially resembled the somatotrophic cells of other mammalian species and did not respond to any of the experimental treatments used in the present study. It is therefore possible that these cells have a somatotrophic function. The possible significance of type 7 cells has been discussed previously.

Relationships among the release of prolactin, the effect of oestrogens and the proliferation of prolactin-secreting cells were studied under several experimental conditions. Administration of sulpiride or oestradiol released prolactin and stimulated cell proliferation in the anterior pituitary gland of adult male rats. Clomiphene completely abolished the rise in cell proliferation, but did not interfere with the sulpiride-induced release of prolactin. Treatment with oestradiol plus sulpiride significantly increased serum prolactin concentrations and the mitotic index compared with the sum of the stimulation produced by both drugs separately. Bromocriptine abolished the stimulatory effect of oestradiol on the serum prolactin concentration and on cell proliferation. In oestradiol- and/or sulpiride-treated rats, 80% of the cells in mitoses were lactotrophs. The remaining 20% did not stain with antisera against any of the pituitary hormones. The number of prolactin-secreting cells in the anterior pituitary gland significantly increased after the administration of oestradiol or sulpiride. The results demonstrate that treatment with sulpiride and/or oestradiol increases the proliferation and the number of lactotrophs in the anterior pituitary gland of the rat.

The effects of adrenocortical and gonadal steroids on the secretion in vitro of ACTH by adenohypophysial segments and corticotrophin releasing factor (CRF) by isolated hypothalami were studied in the rat. Corticosterone (1.25 X 10(-6) mol/l), betamethasone (2.5 X 10(-8) mol/l) and progesterone (2.5 X 10(-7) mol/l) reduced the hypothalamic extract-induced secretion of ACTH by pituitary tissue in vitro but aldosterone (2 X 10(-7) mol/l), testosterone, androsterone, androstenedione (2 x 10(-7) mol/l), oestradiol, oestriol, and oestrone (10(-6) mol/l) did not. Corticosterone (2.5 +/- 10(-9) mol/l), aldosterone (2 X 10(-8) mol/l) and betamethasone (2 X 10(-10) mol/l) inhibited and oestradiol, oestriol and oestrone (10(-8) - 10(-6) mol/l) potentiated the production of CRF by isolated hypothalami which occurred when acetylcholine or 5-hydroxytryptamine were added to the incubation medium but progesterone (2.5 X 10(-7) mol/l), testosterone, androsterone and androstenedione (2 X 10(-7) mol/l) had no effects. The results indicate that hypothalamo-pituitary-adrenocorticotrophic activity may be modified not only by glucocorticoids but also by other steroids.

The TRH-degrading activity of rat serum in vitro is five times more potent than that of human serum. In rats, it is significantly reduced in hypothyroidism (thiouracil-induced) and significantly increased in hyperthyroidism (T3 or T4-induced). This suggests a possible role in the regulation of adenohypophysial-thyroid function which is probably, in turn, dependent on thyroid hormone, rather than TSH, levels.

The relative responses to testosterone of the prostate, preputial, Harderian, and lachrymal glands, the seminal vesicles and the brown adipose tissue, have been compared in litter-mate, castrated and hypophysectomized-castrated rats, with or without treatment with bovine GH. The responses of both prostate and preputial glands, though not the seminal vesicles, were reduced by hypophysectomy, and restored by GH. Growth hormone had some independent action on the preputial glands, but most of its effect was synergistic with testosterone. The respinse of brown adipose tissue to testosterone was similarly reduced by hypophysectomy, but was not restored by the GH. The weights of the Harderian and lachrymal glands were not affected by castration, but they were considerably reduced by hypophysectomy. Growth hormone had a slight restorative effect on the Harderian glands, but none on the lachrymal glands.

We have investigated whether the priming effect of LH-RF can be elicited by electrical stimulation of the medial preoptic area, or by i.v. infusion or multiple i.v. injections of the synthetic decapeptide. All experiments were carried out on animals anaesthetized with sodium pentobarbitone at 13.30 h. In pro-oestrous rats, the LH response to the second of two electrical stimuli, 15 min in duration and separated by 60 min, was significantly greater than the response to the first stimulus. When synthetic LH-RF was infused at a constant rate for 90 min, plasma LH increased gradually for the first 45-60 min after which it increased markedly. This enhanced secretion of LH did not occur in rats which were infused with the same total dose of LH-RF, either 15 or 75 ng/100 g body wt, over periods of 45 min or less. When a dose of 15 ng LH-RF/100 g body wt was administered in six divided doses by i.v. injections, each separated by 15 min, there was a marked increase in plasma LH after 75 min. The profile of the mean plasma LH concentration in rats subjected to preoptic stimulation for 90 min was similar to that in rats infused for 90 min with LH-RF, but the variation in response was much greater in the stimulated rats. These results indicate that the priming effect can be elicited by endogenous as well as synthetic LH-RF, and that whether LH-RF reaches the pituitary at a constant rate or in a pulsatile manner the factor is capable of significantly increasing the responsiveness of the gonadotrophs. The relevance of these findings with respect to the development of the spontaneous preovulatory LH surge is discussed. A priming effect could not be elicited by constant LH-RF infusion in dioestrous rats; this supports the view that steroid hormones, especially oestradiol-17phi, determine the magnitude of the effect. The LH response in male rats subjected to i.v. infusion of LH-RF was much lower than in females. Pre-treatment with oestradiol benzoate did not increase the response significantly, suggesting that this sex difference cannot be ascribed simply to low levels of plasma oestrogen in the male.

Surgical disconnection of the ovine hypothalamus from the pituitary gland (hypothalamo-pituitary disconnection; HPD) has provided a useful experimental model for studying the control of gonadotrophin secretion. The objective of the present study was to define the characteristics of prolactin secretion using stimuli acting through the hypothalamus or directly on the pituitary gland in HPD ewes. Prolactin responses to either a stressful stimulus or the dopaminergic antagonists metoclopramide (20 mg i.v.) or chlorpromazine (50 mg i.v.) seen in intact animals (sham-HPD) were completely abolished by HPD. Injection of TRH (100 micrograms i.v.) caused an immediate release of prolactin in both groups of ewes. In the HPD ewes plasma prolactin concentrations remained raised for at least 3 h after TRH injection, whereas in sham-HPD ewes prolactin concentrations began to decline after 20 min. Administration of bromocriptine (1 mg i.v.) 10 min after TRH inhibited the prolonged response to TRH in HPD ewes. The results support the hypothesis that prolactin exerts a short-loop feedback effect on its own secretion at the hypothalamic level.

Corticosteroid feedback mechanisms were investigated at the hypothalamic level using the rat hypothalamus in vitro and the pituitary level using basal hypthalamic-lesioned rats. Both fast and delayed corticosteroid feedback effects were demonstrated at the level of the hypothalamus and pituitary gland with doses of corticosteroids within or near the physiological range. These two phases of feedback were separated temporally by a 'silent period' during which no feedback was apparent. Studies on the mechanism of action of corticosteroids at the hypothalamic level showed that the fast feedback mechanism acts by inhibition of release whilst the delayed feedback mechanism acts by inhibition of both synthesis and release. The fast feedback action of corticosterone does not appear to act by excitation of neuroinhibitory pathways since neither picrotoxin nor phentolamine prevented the feedback action of corticosteroids in vitro. Corticosterone inhibition of corticotrophin releasing factor release was overcome by depolarization of the membrane with K+ suggesting that the mechanism of action of the fast feedback of corticosteroids is by membrane stabilization.

A role for the regulation of cellular Ca2+ homeostasis in the dopaminergic control of prolactin secretion was investigated in rat anterior pituitary glands. Withdrawal of dopamine stimulated the uptake of 45Ca2+ into hemipituitary tissue by 48% after 3 min. Radioisotope desaturation from tissue prelabelled with 45Ca2+ was significantly retarded in the presence of dopamine. Withdrawal of dopamine rapidly stimulated 45Ca2+ efflux from prelabelled tissue by 79% and was accompanied by a three- to fourfold rise in prolactin secretion. The 45Ca2+ efflux response to dopamine withdrawal was reduced in tissue prelabelled in the presence of dopamine. Agonist displacement with metoclopramide mimicked the effect of dopamine withdrawal on 45Ca2+ efflux and prolactin secretion. These observations demonstrate that the stimulation of prolactin release by dopamine withdrawal is accompanied by a redistribution of cellular Ca2+ and support the hypothesis that dopamine inhibits secretion by decreasing Ca2+ influx in the mammotroph cell.

The effect of the 5-hydroxytryptamine (5-HT) uptake blockers on the surges of LH and prolactin has been investigated in pro-oestrous rats and various experimental models used frequently to study the effects of steroids on LH and prolactin secretion in female rats. The steroid models were: rats ovariectomized on dioestrus, injected immediately with oestradiol benzoate (OB) and at 12.00 h on the next day (presumptive pro-oestrus) with progesterone (model 1); long-term ovariectomized rats injected with a single injection of OB and 72 h later with either progesterone (model 2) or OB (model 3); long-term ovariectomized rats injected daily with OB (model 4). The uptake blockers alaproclate (3-30 mg/kg) and zimelidine (20 mg/kg) were injected and blood samples withdrawn from previously implanted intra-atrial cannulae. Plasma LH and prolactin concentrations were determined by radioimmunoassay. The present study confirmed that a surge of LH occurs at about 17.00-18.00 h of the presumptive day of pro-oestrus in model 1, at about 5 h after (approximately 17.00 h) the injection of either progesterone or the second injection of OB in models 2 and 3, and diurnally in model 4, and the simultaneous occurrence of a prolactin surge in models 2 and 4. A surge of prolactin at the same time as the LH surge was shown to occur also in models 1 and 3. Alaproclate (30 mg/kg) administered at 15.00 h delayed significantly the peak of the prolactin surge in the pro-oestrous rat and models 1, 3 and 4, and in the latter the magnitude of the prolactin surge was also significantly reduced. By contrast, the peak of the prolactin surge in model 2 was significantly prolonged by alaproclate. Alaproclate had no significant effect on either the timing or the magnitude of the LH surge in the pro-oestrous rat, and models 3 and 4. The peak of the LH surge was delayed by alaproclate in model 1 and abolished in model 2, providing further evidence for the possible importance of interactions between 5-HT and progesterone in neuroendocrine control. Zimelidine had no significant effect on either the LH or prolactin surge in the pro-oestrous rat and in models 1 and 2. These results show that normal 5-HT uptake is necessary for the normal timing and/or magnitude of the spontaneous and steroid-induced prolactin surge but is not essential for the normal timing and magnitude of the spontaneous surge of LH and the LH surge in some but not all steroid models.(ABSTRACT TRUNCATED AT 400 WORDS)

Mink anterior pituitaries were incubated in Medium 199 for up to 9 or 13 days. Biological activity of prolactin and GH was determined. Daily concentrations of prolactin and GH in the incubation medium were also measured by radioimmunoassay and radioreceptor assay. When females were kept under short days for several weeks before the experiment, a significant decrease in prolactin secretion by the anterior pituitary was observed as compared with that in females maintained under long days. In contrast, secretion of GH was not modified by the photoperiodic history of the animals. Pineal gland denervation by ablation of the superior cervical ganglia a few months before the experiment, or addition of melatonin to the incubation medium of anterior pituitaries from intact or ganglionectomized females, did not modify the secretion of prolactin and GH. The pituitary gland does not therefore seem to be a direct target site for melatonin in transducing the duration of daylength on the hypothalamo-pituitary axis.

The effects of selective agonists and antagonists of type 1 (V1) and type 2 (V2) vasopressin receptors on the secretion of ACTH in vitro by segments of adenohypophysial tissue and in vivo in rats pretreated with pentobarbitone and chlorpromazine were studied in the presence and absence of the 41 amino acid-containing peptide, corticotrophin-releasing factor-41 (CRF-41). The non-selective vasopressin receptor agonist, arginine vasopressin (AVP) and the V1-receptor agonist, felypressin caused dose-related increases in ACTH release in vivo and in vitro but the V2-receptor agonist, desmopressin was only weakly active in this respect. Their actions in vitro were antagonized competitively by the V1-receptor antagonist, d(C2H5)2-AVP, but were unaffected by the V2-receptor antagonist, d(CH2)5-D-Iso2-Thr4-AVP. Arginine vasopressin, felypressin and desmopressins in concentrations considerably lower than those necessary to elicit directly the release of ACTH, potentiated, in a dose-related manner, the activity of CRF-41 in vitro. The potentiating effects were not antagonized by the V2-receptor antagonist or by low concentrations of the V1-receptor antagonist. At a higher concentration, the V1-receptor antagonist reduced, but did not abolish, the potentiating effects of AVP and its analogues. However, at this concentration, it also exhibited weak intrinsic activity and, like the agonists, potentiated the response to CRF-41. The results suggest that the direct effect of AVP on ACTH release is mediated by V1-like receptors. The vasopressin receptors involved in the potentiation of CRF-41 activity appear to be different.

A multi-column perifusion system was used to investigate the dynamics of the dose-response relationships of ACTH release by ovine pituitary cells when stimulated by both corticotrophin-releasing hormone (CRF) and arginine vasopressin (AVP) given alone and in combination. A dose-response relationship was obtained when 10-min pulses were given at 60-min intervals over the range of 0.002-2000 nmol CRF/1 and 1-2000 nmol AVP/1, with a minimum effective concentration of 0.02 nmol CRF/1 or 1 nmol AVP/1. When AVP was given together with CRF, the expected potentiation of the ACTH response occurred when compared with the summed response of these secretagogues given separately. At the higher concentrations of CRF and AVP used, the ACTH responses to repeated pulses decreased with time during the experiment. The rate of this loss of responsiveness was significantly correlated to the size of the response to the first pulse (for CRF: r = 0.89, P less than 0.01; for AVP: r = 0.95, P less than 0.01), being greatest when the response was potentiated by adding the secretagogues together (for CRF plus AVP: r = 0.95, P less than 0.01). Reduced availability of receptors or changes in intracellular transduction processes may contribute to this desensitization. Reduced levels of secretable ACTH do not appear to be implicated because desensitization to pulses of one secretagogue did not cause equivalent desensitization to the other. In addition, cells stimulated continuously with submaximal levels of either secretagogue showed desensitization while more ACTH was still available for release to higher levels of stimulant.(ABSTRACT TRUNCATED AT 250 WORDS)

The aim of this study was to determine whether the priming effect of LH-RF depends upon RNA and protein synthesis. In in-vivo studies saline, actinomycin D, or cycloheximide was administered i.p. 3-5-4h before the first i.v. injection of synthetic LH-RF into pro-oestrous rats anaesthetized with sodium pentobarbitone at 13.30 h. The LH-response to the second injection of LH-RF (given 60 min after the first) was markedly reduced by the inhibitors, but the response to the first injection was not significantly affected. Studies with cycloheximide given i.v. showed that the inhibition of protein synthesis up to the second injection of LH-RF reduced the magnitude of the priming effect, the reduction being greatest when the inhibitor was administered up to 30 min after the first LH-RF injection. Pituitary incubation studies showed that the priming effect could also be elicited in vitro and that it could be significantly reduced by actinomycin D, cycloheximide and puromycin. As in vivo, the inhibitors had relatively little effect on the LH-response to the first exposure to LH-RF. The protein synthesized after an injection of LH-RF may be new LH, and/or a protein(s) concerned with 'activation' of the receptor or release components of the LH-secretory apparatus.

We have detected significant amounts of corticotrophin-releasing factor-41 (CRF-41) in the rat hypothalamo-neurointermediate lobe system using a radioimmunoassay and reversed-phase high-performance liquid chromatography. Total amounts of CRF-41 in extracts of median eminence (ME), supraoptic nucleus (SON), paraventricular nucleus (PVN) and neurointermediate lobe (NIL) from control animals were 1076 +/- 132, 196 +/- 44, 22 +/- 7 and 147 +/- 50 fmol respectively (means +/- S.E.M., n = 6). In animals given 340 mmol NaCl/l instead of tap water to drink for 12 days, no significant changes occurred in the CRF-41 content of the ME, SON or PVN, but CRF-41 content increased more than twofold in the NIL (362 +/- 58 fmol). Plasma concentrations of CRF-41 and ACTH in control animals were 23 +/- 6 and 51 +/- 8 pmol/l respectively. After saline treatment no significant change in plasma CRF-41 was detected (20 +/- 8 pmol/l) but concentrations of circulating ACTH were decreased (15 +/- 2 pmol/l). The CRF-41 content of both the ME and the NIL was significantly depleted after intracerebroventricular injection of colchicine (414 +/- 81 and 34 +/- 7 fmol respectively). These data suggest that NIL CRF-41 is of hypothalamic origin and can be regulated by an osmotic stimulus.

Pituitary glands taken from intact rats on day 2 of dioestrus and incubated with LHRH show a biphasic pattern of LH and FSH release. Initially the release of the gonadotrophins is low (first-phase or lag-phase response), but increases during further incubation with LHRH (second-phase or primed-state response). Removal of the influence of an unidentified ovarian factor either by ovariectomy or prolonged incubation in medium only leads to an increased (lag-phase) response to LHRH. The development of the increased response after prolonged incubation was prevented by the addition of cycloheximide to the media, implicating that this process is dependent upon the synthesis of protein. Steroid-free material (bovine follicular fluid or rat ovarian extracts) prevented the development of this process. In addition, it was shown that steroid-free rat ovarian extracts were also able to induce the development of a lag phase in pituitary glands from ovariectomized rats. Finally, it was found that steroid-free ovarian extracts reversed the self-priming effect of LHRH. The biological activity which reduced the responsiveness of the pituitary gland towards stimulation by LHRH was eliminated after the use of protein-denaturating techniques such as increased temperature or addition of methanol. The presence of this activity in ovaries, did not vary during the oestrous cycle, contrary to inhibin-like activity. Hence the ovarian factor responsible for the low lag-phase response is a protein which is probably not identical to inhibin. It is concluded that a non-steroidal ovarian factor reduces the responsiveness of the anterior pituitary gland to LHRH.(ABSTRACT TRUNCATED AT 250 WORDS)