Rhythmic and Nonrhythmic Modes of Anterior Pituitary Gland Secretion

Because of confounding effects of subject-specific and hormone-specific metabolic clearance, the nature of anterior pituitary secretory events in vivo is difficult to ascertain. We review an approach to this problem, in which deconvolu-tion analysis is used to dissect the underlying secretory behavior of an endocrine gland quantitatively from available serial plasma hormone concentration measurements assuming one- or two-compartment elimination kinetics. This analytical tool allows one to ask the following physiological questions: (a) does the anterior pituitary gland secrete exclusively in randomly dispersed bursts, and/or does a tonic (constitutive) mode of interburst hormone secretion exist? and (b) what secretory mechanisms generate the circadian or nyctohemeral rhythms in blood concentrations of pituitary hormones? Waveform-independent deconvolution analysis of 24-h serum hormone concentration profiles of immunoreactive growth hormone (GH), luteinizing hormone (LH), follicle-stimulating hormone (FSH), prolactin, thyroid-stimulating hormone (TSH), adrenocorticotropic hormone (ACTH), and β-endorphin in normal men sampled every 10 min showed that (a) anterior pituitary gland secretion in vivo occurs in an exclusively burstlike mode for all hormones except TSH and prolactin (for the latter two, a mixed burst and basal mode pertains); (b) significant nyctohemeral regulation of secretory burst frequency alone is not demonstrable for any hormone; (c) prominent 24-h variations in secretory-burst amplitude alone are delineated for ACTH and LH; (d) TSH, GH, and β-endorphin are both frequency and amplitude controlled; (e) prolactin manifests 24-h rhythms in both secretory-burst amplitude and nadir secretory rates; (f) no significant diurnal variations occur in FSH secretory parameters; and (g) a fixed hormone half-life yields good fits of the 24-h serum hormone concentration series, which indicates that there is no need to introduce diurnal variations in hormone half-lives. In summary, the normal human anterior pituitary gland appears to release its various (glyco)protein hormones via intermittent secretory episodes that are apparently unassociated with significant basal hormone secretion, except in the case of TSH and prolactin. Hormone-specific amplitude and/or frequency control of secretory burst activity over 24 h provides the mechanistic basis for the classically recognized nyctohemeral rhythms in plasma concentrations of adenohypophyseal hormones in the human.     

Physiological control of pituitary hormone secretory-burst mass,frequency, and waveform: a statistical formulation and analysis

The present study investigates the time-varying control of pituitary hormone secretion over the day and night (D/N). To this end, we implemented an analytical platform designed to reconstruct simultaneously 1) basal (nonpulsatile) secretion, 2) single or dual secretory-burst waveforms, 3) random effects on burst amplitude, 4) stochastic pulse-renewal properties, 5) biexponential elimination kinetics, and 6) experimental uncertainty. The statistical solution is conditioned on a priori pulse-onset times, which are estimated in the first stage. Primary data composed of thyrotropin (TSH) concentrations were monitored over 24 h in 27 healthy adults. According to statistical criteria, 21/27 profiles favored a dual compared with single secretory-burst waveform. An objectively defined waveform change point (D/N boundary) emerged at 2046 (+/-23 min), after which 1) the mass of TSH released per burst increases by 2.1-fold (P < 0.001), 2) TSH secretory-burst frequency rises by 1.2-fold (P < 0.001), 3) the latency to maximal TSH secretion within a burst decreases by 67% (P < 0.001), 4) variability in secretory-burst shape diminishes by 50% (P < 0.001), and 5) basal TSH secretion declines by 17% (P < 0.002). In contrast, the regularity of successive burst times and the slow-phase half-life are stable. In conclusion, nycthemeral mechanisms govern TSH secretory-burst mass, frequency, waveform, and variability but not evidently TSH elimination kinetics or the pulse-timing process. Further studies will be required to assess the generality of the foregoing distinctive control mechanisms in other hypothalamo-pituitary axes.     

Chronobiological analysis of changes in circadian rhythm of anterior pituitary hormones level during glycemia normalization by means of biostator

We compared the circadian rhythms of anterior pituitary hormones in 15 patients with noncompensated insulin-dependent diabetes on first and second day treatment with Biostator. The rhythm was evaluated by means of a least squares analysis and presented as the circle of cosinors. In noncompensated diabetes the TSH and prolactin rhythm was maintained, whereas other hormones of the anterior pituitary showed no significant rhythm. In the course of one-day normalization of glycemia by means of Biostator the TSH and prolactin rhythm was maintained, whereas the circadian rhythm of growth hormone and ACTH levels appeared with acrophase at 18.47 and 19.59 hour, respectively. The LH rhythm did not exist, whereas the FSH rhythm was dubious. One may assume that noncompensated diabetes results in the impairment of certain pituitary hormonal rhythms and these disturbances are reversible after restoring of normoglycemia.     

Changes in pituitary hormone levels induced by met-enkephalin in man—The role of dopamine

The interaction of dopamine with the effects of the opiate agonist peptide D-Ala²-MePhe⁴-met-enkephalin-O-o1 (DAMME) on anterior pituitary hormone secretion was investigated in normal male subjects. DAMME produced clear elevations in prolactin, growth hormone and thyroid-stimulating hormone, while inhibiting the release of luteinising hormone and cortisol. There was no change in follicle stimulating hormone. The elevations in prolactin and TSH were enhanced by the dopamine antagonist, domperidone, and blocked by an infusion of dopamine. Neither dopamine nor domperidone modulated the changes in growth hormone, luteinising hormone or cortisol. The data are comptible with the association of the release of prolactin and TSH by opiate peptides with decreased hypothalamic dopaminergic activity; changes in the other anterior pituitary hormones seem to involve different mechanisms.     

Effect of bombesin on basal and stimulated secretion of some pituitary hormones in humans

The effect of bombesin (5 ng/kg/min X 2.5 h) on basal pituitary secretion as well as on the response to thyrotropin releasing hormone (TRH; 200 micrograms) plus luteinizing hormone releasing hormone (LHRH; 100 micrograms) was studied in healthy male volunteers. The peptide did not change the basal level of growth hormone (GH), prolactin, thyroid-stimulating hormone (TSH), luteinizing hormone (LH) and follicle-stimulating hormone (FSH). On the contrary, the pituitary response to releasing hormones was modified by bombesin administration. When compared with control (saline) values, prolactin and TSH levels after TRH were lower during bombesin infusion, whereas LH and FSH levels after LHRH were higher. Thus bombesin affects in man, as in experimental animals, the secretion of some pituitary hormones.     

The Circadian Rhythms of Thyrotrophin and Prolactin Secretion

As with other anterior pituitary hormones, the secretion of both thyrotrophin (TSH) and prolactin (PRL) displays a circadian variation with different patterns for each hormone. In recent years there has been a substantial increase in the understanding of the neuroregulation oof TSH and PRL. However the primary events involved in the generation of their circadian rhythms remains unclear. Regulatory pathways comprise two major groups: central factors, where the control is exerted by the central nervous system via the hypothalamus and peripheral factors, which include all extra CNS mechanisms. The first group is represented mainly by neuropeptides and neurotransmitters controlling TSH and PRL release, whereas the second one comprises both physical phenomena such as variations in plasma volume or postural changes and hormonal influences arising from target glands such as the adrenal, the thyroid and the gonads.     

Pancreatic polypeptides affect luteinizing and growth hormone secretion in rats

We have examined the effects of third cerebroventricular (3V) injections of avian and bovine pancreatic polypeptide (APP and BPP) and the C-terminal hexapeptide amide of human PP (CHPP) on the secretion of anterior pituitary hormones in conscious ovariectomized rats. Injection of APP (2.0 micrograms; 472 pmoles) or BPP (5.0 micrograms; 1191 pmoles) decreased plasma levels of luteinizing hormone (LH) when compared to pre-injection levels in these animals or to saline-injected controls. The lower dose of BPP (0.5 micrograms; 119 pmoles) decreased plasma LH versus pre-injection levels and control animals, however, these effects diminished at later times. Plasma growth hormone (GH) also decreased following 3V injections of APP (2.0 micrograms) or BPP (5.0 micrograms). The lower dose of BPP (0.5 microgram) initially inhibited GH release, however, this effect was rapidly reversed and GH levels were significantly greater than those in controls at 60 and 120 min. Injections of BPP or APP did not alter prolactin (PRL) or thyroid stimulating hormone (TSH) secretion. Administration of 2.0 micrograms and 0.2 microgram of CHPP (2488 and 249 pmoles) produced no significant effects on plasma LH, GH, PRL or TSH. APP and BPP had no consistent effects on hormone secretion from dispersed anterior pituitary cells. The results indicate that APP and BPP exert potent central effects which inhibit LH and GH release from the pituitary gland.     

Cell swelling induced by the permeant molecules urea or glycerol induces immediate high amplitude thyrotropin and prolactin secretion by perifused adenohypophyseal cells

The permeant molecules, urea and glycerol, evoked a prompt secretory burst of TSH and PRL when added to the extracellular medium of acutely dispersed anterior pituitary cells. Secretion of both hormones was proportional to the concentration of urea or glycerol between 26 and 104 mM (r greater than 0.89, P less than 0.001). Equivalent concentrations of the impermeant molecule, mannitol, did not induce secretion. The acute TSH and PRL secretory responses to TRH, hyposmolarity, and permeant molecules were qualitatively indistinguishable. These data support our hypothesis that cell swelling and resultant plasmalemma expansion is a potent inducer of hormone secretion. Since the secretory response to permeant molecules was not reduced in a Ca2+-free medium containing 0.1 mM EGTA, an increase in Ca2+ transport across the plasmalemma to raise cytosol Ca2+ concentration does not appear involved.     

Changes in thyrotroph and somatotroph cell populations induced by stimulation and inhibition of their secretory activity

Summary The populations of cells which produce immunoreactive growth hormone (GH) and thyroid stimulating hormone (TSH) in the rat pituitary gland do not occur in fixed percentages but vary greatly under different physiological and experimental conditions. These variations can be directly correlated to the levels of stimulation and/or inhibition of the specific secretory activity. In both types of cell, sustained stimulation with trophic hormones or blockage of the feedback mechanisms induces remarkable growth in the specific cell population. Conversely, the interruption or inhibition of the stimulus thwarted the hormonal secretion and caused a massive degeneration of redundant cells. The stimulation of both GH and TSH cells is accompanied by an enhanced secretory activity as judged by their higher concentrations in serum and hypertrophy of the cytoplasmic organelles involved in synthesis and intracellular processing of the hormones. By contrast, interruption of the stimulus is followed by a variable degree of disruption of the cytoplasmic organization, including a sizable degeneration of cells. In stimulated rats, the concentrations of both GH and TSH decreased significantly in pituitary tissue due to mobilization of the hormonal stores contained in secretory granules. On the other hand, the withdrawal of stimuli blocked the hormonal release; this is reflected by the accumulation of both hormones and secretory granules in pituitary tissue. The strict correlation between the size of the GH and TSH populations with stimulation and inhibition of hormonal secretory activity reported in this investigation further supports the critical role played by the cell renewal process in endocrine secretion.     

Aminergic regulation of neuroendocrinological functions: theoretical background and some clinical examples

Remarkable progress has been made during recent years in the central regulation of the hypothalamic releasing and inhibiting factors and the respective anterior pituitary hormones. There are two nearly universal inhibitory organizations: short tuberoinfundibular dopamine (TIDA) neurons and somatostatinergic system originating from the periventricular hypothalamus and terminating to the median eminence. It is now known that e.g. dopamine, noradrenaline and acetylcholine enhance while 5-hydroxytryptamine and GABA inhibit somatostatin secretion. These transmitters are also involved in the regulation of all releasing factors and pituitary hormones. Clinical applications have been developed based on the regulation of prolactin and growth hormone. Inhibitory TIDA neurons are undoubtedly the major determinants of prolactin secretion. Hyperprolactinaemia is one of the most common endocrinological side-effects of the drugs antagonizing dopaminergic transmission. Expectedly, dopaminergic drugs (bromocryptine, lergotrile, piribedil, dopamine and levodopa) are quite effective in reducing high prolactin levels regardless of the reason. The secretion of growth hormone is predominantly under dual dopaminergic control: hypothalamic stimulation and pituitary inhibition. The former masters the function of the normal gland, while the peripheral inhibitory component takes over in acromegalic gland. Hence dopaminergic drugs are able to reduce elevated growth hormone levels in 30-50% of the acromegalic patients. In normal man, dopamine agonists increase growth hormone levels. An analogous situation can be seen in Cushing's disease regarding ACTH secretion.     

Localization of Carboxyl Ester Lipase in Human Pituitary Gland and Pituitary Adenomas

Carboxyl ester lipase (CEL) is an enzyme that hydrolyzes a wide variety of lipid substrates, including ceramides, which are known to show inhibitory regulation of pituitary hormone secretion in experimental models. Because no studies on CEL expression in human pituitary and pituitary adenomas have been reported in the literature, we investigated CEL expression in 10 normal pituitary glands and 86 well-characterized pituitary adenomas [12 FSH/LH cell, 17 α-subunit/null cell, 6 TSH cell, 21 ACTH cell, 11 prolactin (PRL) cell, and 19 GH cell adenomas] using IHC, immunoelectron microscopy, Western blotting, and quantitative RT-PCR. In normal adenohypophysis, CEL was localized in GH, ACTH, and TSH cells. In adenomas, it was mainly found in functioning GH, ACTH, and TSH tumors, whereas its expression was poor in the corresponding silent adenomas and was lacking in FSH/LH cell, null cell, and PRL cell adenomas. Ultrastructurally, CEL was localized in secretory granules close to their membranes. This is the first study demonstrating CEL expression in normal human pituitary glands and in functioning GH, ACTH, and TSH adenomas. Considering that CEL hydrolyzes ceramides, inactivating their inhibitory function on pituitary hormone secretion, our findings suggest a possible role of CEL in the regulation of hormone secretion in both normal and adenomatous pituitary cells. (J Histochem Cytochem 58:881–889, 2010)     

The effects of tunicamycin on anterior pituitary hormone producing cells in the rat

An electron microscopic study was performed to clarify the effects of tunicamycin, a glycosylation inhibitor, on rat anterior pituitary cells. Tunicamycin (10, 50, and 100 micrograms/250 g B.W.) was intraperitoneally injected into rats, which were sacrificed 24 hrs later. Protein hormone producing GH and prolactin cells, and ACTH cells which are known to have a glycosylated precursor, showed no recognizable ultrastructural changes. TSH cells and gonadotrophs, both of which secrete glycoprotein hormones consisting of alpha and beta subunits, showed remarkable dilatation of the rough endoplasmic reticulum, and decreased numbers of secretory granules. These results suggest that the role of glycosylation in TSH cells and gonadotrophs may have a different biological significance from that in ACTH cells.     

Opioid components of the clockwork that governs luteinizing hormone and prolactin release in male rats

Daily rhythms of secretion have been described for luteinizing hormone (LH) and prolactin (PRL) from the anterior pituitary of rats. Using selective opioid antagonists, we found that mu and kappa opioid receptor ligands regulate LH and PRL secretion and, of particular interest, that the magnitude of opioidergic effects varies with the time of day. In addition, incomplete temporal overlapping of the LH and PRL responses to the antagonists suggests that different endogenous opioid pathways, with different temporal profiles of peptide release, may control each of these hormones.     

Plasma Concentration of Nine Hormones and Neurotransmitters During Usual Activities or Constant Bed Rest for 34 H

Thyroid-stimulating hormone (TSH), Cortisol, melatonin, prolactin, luteinizing hormone (LH), delta-sleep-inducing peptide (DSIP), its phosphorylated form (P-DSIP), heart rate, and body temperature were measured every half hour during two 24-h periods in five normal men. τ-Amino-butyric acid (GABA) and 3-methoxy-4-hydroxyphenylglycol (MHPG) were measured less frequently. The first period, the “activity” condition, included usual daily activities. The second period, or “rest” condition, consisted of fasting, constant bed rest during 34 h, and partial light deprivation. Compared with the “rest” condition, the “activity” condition increased heart rate, temperature, LH, and TSH in most subjects, and Cortisol in two of five subjects. It retarded the onset of nocturnal Cortisol and melatonin secretion. The temporal pattern and the absolute values of the concentrations of DSIP, P-DSIP, MHPG, GABA, and prolactin showed no or minimal changes during the two conditions. In spite of the influence of the “activity” versus “rest” condition on several hormones, the mean concentrations as well as the temporal organization of their secretion into plasma were quite stable within each subject, whereas they varied much more between individuals. TSH, Cortisol, and melatonin values were also stable within an 8-month period in one subject who was studied on four occasions. The results illustrate that the patterns of hormones rhythms and their reactivity to changes in the environment are, to a large extent, specific to each subject.     

HORMONE SECRETION BY CELLS DISSOCIATED FROM RAT ANTERIOR PITUITARIES

A new procedure has been developed for dissociating anterior pituitary tissue and producing a viable suspension of single cells. The procedure involves incubation of small tissue blocks in 1 mg/ml trypsin (15 min), followed by incubation in 8 µg/ml neuraminidase and 1 mM EDTA (15 min), followed by mechanical dispersion. Cell yields are ~55%, based on recovered DNA. By electron microscopy five types of secretory cells (somatotrophs, mammotrophs, thyrotrophs, gonadotrophs, and corticotrophs) plus endothelial and follicular cells can be identified and are morphologically well preserved up to 20 h after dissociation. Throughout this period, the cells incorporate linearly [³H]leucine into protein for up to 4 h at a rate 90% greater than hemipituitaries, and they synthesize, transport intracellularly, and release the two major pituitary secretory products, growth hormone and prolactin. Immediately after dissociation the cells' ability to respond to secretogogues (high K⁺ and dibutyryl cyclic AMP) is impaired, but after a 6–12-h culture period, the cells apparently recover and discharge 24% and 52%, respectively, of their content of prelabeled growth hormone over a 3-h period in response to these two secretogogues. This represents a stimulation of 109% and 470% over that released by cells incubated in control medium. The results demonstrate that function and morphologic integrity are preserved in this cell system. Therefore it is suitable for the study of various aspects of pituitary secretion and its control.     

Transforming growth factor-beta and activin inhibit basal secretion of prolactin in a pituitary monolayer culture system

Incubations of rat anterior pituitary cells with transforming growth factor (TGF)-beta 1 for 48 hr suppressed the secretion of basal prolactin (PRL) in a dose-dependent manner (ED50, 100 pg/ml). Activin, a gonadal hormone processing cysteine distribution similar to TGF beta, also suppressed basal PRL secretion, but it was less effective (ED50, 4 mg/ml). Treatment with TGF beta 1 significantly suppressed basal PRL secretion from the pituitary after 24 hr and up to 72 hr of incubation. TGF beta 1 also inhibited thyrotropin-releasing hormone-mediated PRL secretion and activin inhibited thyrotropin-releasing hormone-mediated PRL secretion slightly, but significantly. In addition, we also measured the secretion of growth hormone by cultured pituitary cells treated with TGF beta 1 or activin for 24 to 72 hr. TGF beta 1 and activin showed an opposite effect on growth hormone secretion; TGF beta stimulated and activin inhibited basal secretion of growth hormone. These results suggest that TGF beta 1 is a potent inhibitor of basal secretion of PRL by the pituitary, and both TGF beta 1 and activin play a multifunctional role in basal secretion of pituitary hormones.     

Endothelin-3 inhibits prolactin and stimulates LH, FSH and TSH secretion from pituitary cell culture

The influence of endothelin-3 (ET-3) on anterior pituitary hormone secretion was investigated over a wide range of concentrations (from 10(-14) to 10(-6) M) and incubation times (from 4 to 48 hours). ET-3 elicited a concentration-dependent inhibition of prolactin (PRL) secretion and stimulated the release of luteinizing hormone (LH), follicle stimulating hormone (FSH) and thyroid stimulating hormone (TSH) from primary monolayer cultures of anterior pituitary cells derived from female rats. The responsiveness of different pituitary cells to ET-3 differs markedly in terms of onset and duration: the maximal inhibition of PRL secretion occurred after 12 hours and the stimulation of LH, FSH and TSH reached the maximum after 4, 48 and 48 hours of incubation, respectively. These data corroborate the concept that ET-3 has an important role as a neuroendocrine modulator. Moreover, the data presented suggest different intracellular mechanisms underlying ET-3 actions.     

Ultrastructural localization of prolactin,growth hormone and luteinizing hormone by immunocytochemical techniques in the bovine pituitary

Summary The technique of ultrastructural immunocytochemistry involving the unlabeled antibody and the soluble peroxidase-antiperoxidase complex was used to identify and describe the prolactin (P) cells, somatotropic (STH) cells and luteinizing hormone (LH) cells in the bovine anterior pituitary gland. This method was used to localize the three hormones at the electron microscopic level. Staining of varying intensity was found on the secretory granules and on the small granules and vesicles within the Golgi complex. No stain was found in nuclei, on mitochondria or in the endoplasmic reticulum.     

Cortisol feedback state governs adrenocorticotropin secretory-burst shape,frequency, and mass in a dual-waveform construct: time of day-dependent regulation

Quantification of in vivo pituitary hormone secretion requires simultaneous appraisal of implicit 1) secretory-burst waveform, mass, and stochastic pulse timing; 2) basal secretion; 3) biexponential elimination kinetics; and 4) random experimental error (Keenan DM, Licinio J, and Veldhuis JD. Proc Natl Acad Sci USA 98: 4028-4033, 2001). The present study extends this analytic formalism to allow for time of day-dependent waveform adaptation (burst-shape change) at statistically determinable boundary times. Thereby, we test the hypothesis that diurnal mechanisms and glucocorticoid negative feedback jointly govern distinctive facets of the burstlike secretion of ACTH. To this end, we reanalyzed intensively (10 min) sampled 24-h plasma ACTH concentration profiles collected previously under feedback-intact and drug-induced cortisol depletion in nine healthy adults. Akaiki information criterion-based model comparison favored dual (rather than single) secretory-burst representation of 24-h ACTH release in both the intact and low-cortisol setting in eight of nine subjects. Under feedback-intact conditions, analytically predicted waveform changepoints (median clock times 0611 and 1739) flanked an interval of elevated ACTH secretory-burst mass (P < 10-3). Experimental hypocortisolemia did not alter day/night boundaries, but 1) stimulated day ACTH secretory-burst mass (P < 10-3); 2) accelerated day ACTH secretory-burst frequency (P < 10-3); and 3) forced skewness of day ACTH secretory bursts toward more rapid initial release (P < 0.05). In contrast, the basal ACTH secretion rate and regularity of interpulse-interval lengths were invariant of day/night segmentation and cortisol availability. In conclusion, unknown diurnal factors and systemic cortisol concentrations codetermine ACTH secretory-burst waveform, frequency, and mass, whereas neither mechanism regulates basal ACTH release or regularity of the burst-renewal process.