The Octadecaneuropeptide Stimulates Somatolactin Release from Cultured Goldfish Pituitary Cells

The present study aimed to investigate the distribution of the octadecaneuropeptide (ODN) in the goldfish brain and to look for a possible effect of ODN on somatolactin (SL) release from pituitary cells. A discrete population of ODN‐immunoreactive neurones was localised in the lateral part of the nucleus lateralis tuberis. These neurones sent projections through the neurohypophyseal tract towards the neurohypophysis, and nerve fibres were seen in the close vicinity of SL‐producing cells in the pars intermedia. Incubation of cultured goldfish pituitary cells with graded concentrations of ODN (10^?9–10^?5 m ) induced a dose‐dependent stimulation of SL‐β, but not SL‐α, release. ODN‐evoked SL release was blocked by the metabotrophic endozepine receptor antagonist cyclo_1–8[DLeu⁵]OP but was not affected by the central‐type benzodiazepine receptor antagonist flumazenil. ODN‐induced SL release was suppressed by treatment with the phospholipase C (PLC) inhibitor U‐73122 but not with the protein kinase A (PKA) inhibitor H‐89. These results indicate that, in fish, ODN produced by hypothalamic neurones acts as a hypophysiotrophic neuropeptide stimulating SL release. The effect of ODN is mediated through a metabotrophic endozepine receptor positively coupled to the PLC/inositol 1,4,5‐trisphosphate/protein kinase C‐signalling pathway.     

Arachidonate Metabolism in the Anterior Pituitary: Effect of Arachidonate Inhibitors on Basal and Stimulated Secretion of Prolactin, Growth Hormone and Luteinizing Hormone. II. Hormone Release from Dispersed Pituitary Cells

In the accompanying study, we reported the effects of inhibitors of arachidonic acid metabolism on the regulation of prolactin, growth hormone (GH) and luteinizing hormone secretion by male hemipituitaries. The present work extends these investigations to primary cell cultures of the same origin. Arachidonic acid metabolism was inhibited by either 5, 8, 11, 14-eicosatetraynoic acid (ETYA), a blocker of cyclooxygenase- and lipoxygenase-catalysed pathways, or the cyclooxygenase inhibitors, indomethacin and aspirin. ETYA inhibited basal GH secretion by 60%, an effect which was reversed by micromolar concentrations of exogenous arachidonic acid. ETYA was much less effective on growth hormone-releasing factor-induced GH release, a result which contrasts with data obtained on intact glands. Growth hormone-releasing factor stimulation of adenylate cyclase was not affected by ETYA. Cyclooxygenase inhibitors decreased basal secretion to a more limited extent (?30%) and were ineffective on growth hormone-releasing factor-stimulated release. Basal prolactin secretion was reduced by 30% in the presence of ETYA and unaffected by cyclooxygenase inhibitors. As with GH, the effect was reversed by exogenous arachidonic acid. However, in contrast to growth hormone-releasing factor-stimulated GH secretion, thyrotropin-releasing hormone stimulation of prolactin release was able to overcome the inhibition by ETYA in a dose-dependent manner. Again, the insensitivity of thyrotropin-releasing hormone-stimulated prolactin release to ETYA contrasts with the data obtained in intact tissue. Moreover, ETYA inhibited (?60%) prostaglandin E₂ production; thyrotropin-releasing hormone was unable to increase the prostaglandin levels in control or ETYA-treated cells. This confirms the data obtained with cyclooxygenase inhibitors, suggesting that prostaglandins are not involved in prolactin secretion. Intracellular accumulation of Ca²⁺ by the ionophore A23187 and protein kinase C stimulation by the phorbol ester 12-O- tetradecanoyl phorbol acetate (TPA), strongly stimulated GH and prolactin release. Under these conditions, ETYA was no longer able to inhibit secretion of the hormones. As with intact glands, basal and gonadotropin-releasing hormone or TPA-induced luteinizing hormone secretion were unaffected by any of the inhibitors used. It is concluded that blockade of the arachidonic acid cascade interferes with a secretory pathway involved mainly with basal release of prolactin and GH, but not luteinizing hormone. Thyrotropin-releasing hormone, a secretagogue known to trigger phospholipase C and, hence, to stimulate Ca²⁺ mobilization and protein kinase C, overcame ETYA inhibition of prolactin secretion. Growth hormone-releasing factor, a secretagogue recognized by adenylate cyclase coupled receptors, did not overcome ETYA inhibition of GH secretion. However, both secretagogues strongly stimulated hormone release from their target cells in the presence of ETYA. The arachidonic acid cascade thus seems less important in neuromediator-induced secretion coupling processes in dispersed pituitary cells, than in the intact gland. These observations suggest that eicosanoids are more likely to mediate paracrine or autocrine modulations of secretory mechanisms, rather than to function as intracellular messengers.     

PACAP Stimulation of Gonadotropin-II Secretion in Goldfish Pituitary Cells: Mechanisms of Action and Interaction with Gonadotropin Releasing Hormone Signalling

Pituitary adenylate cyclase-activating polypeptide (PACAP) has recently been shown to be a hypophysiotropic factor in the goldfish. In this study, we examined the mechanisms of PACAP action on goldfish maturational gonadotropin (GTH-II) release using primary cultures of pituitary cells. The GTH-II response to mammalian PACAP1-38 (mPACAP) was inhibited by a PACAP receptor antagonist suggesting a receptor-mediated action. Addition of either an adenylate cyclase inhibitor or a protein kinase A (PKA) inhibitor reduced the mPACAP-induced GTH-II release. In addition, when GTH-II release was already stimulated by either forskolin or 8-bromo-cAMP (8Br-cAMP), mPACAP did not further increase GTH-II secretion. These results strongly implicated the involvement of an adenylate cyclase/cAMP/PKA pathway in PACAP-stimulated GTH-II release. Although mPACAP induced a rise in intracellular Ca2+ level in identified gonadotropes, results with voltage-sensitive Ca2+ channel inhibitors indicated that the GTH-II responses to mPACAP, forskolin and 8Br-cAMP did not depend upon Ca2+ entry through these channels. Two protein kinase C (PKC) inhibitors did not affect mPACAP-elicited GTH-II release, and mPACAP further increased GTH-II secretion in the presence of PKC activators. These results indicate that PKC-dependent elements are not essential for the stimulatory action of mPACAP in gonadotropes. Interestingly, while GTH-II responses to a stimulatory concentration of mPACAP were additive to responses elicited by maximal effective concentrations of two endogenous gonadotropin releasing hormones (GnRHs), a subthreshold concentration of mPACAP potentiated GnRH and PKC activator stimulation of GTH-II secretion. Similarly, submaximal concentrations of forskolin potentiated the GTH-II response to the PKC activator, tetradecanoyl phorbol acetate. These data suggest that PACAP and its cAMP-dependent signalling mechanisms provide an alternate stimulatory input to goldfish gonadotropes and may influence the effectiveness of the major neuroendocrine control exerted by the PKC-dependent GnRH signalling pathway.     

Effects in vitro of New Growth Hormone Releasing Peptide (GHRP-1) on Growth Hormone Secretion from Ovine Pituitary Cells in Primary Culture

Continuous perifusion of pituitary cells was used to study the effects of a newly synthesized GHRP (GHRP-1 or KP 101) on growth hormone (GH) secretion from ovine pituitary cells and these have been compared to effects of growth hormone-releasing factor (GRF) and the original growth hormone-releasing peptide (GHRP-6). GH was continuously released at a constant rate during perifusion and secretion was increased by KP 101, GHRP-6 and GRF in a dose-dependent manner. The half-maximal effective dose of KP 101 and GHRP-6 was 10^?7 M, an order of magnitude higher than that for GRF. The maximal effects of KP 101 and GHRP-6 were similar but significantly less than the maximal effect of GRF. Blockade of calcium channels with Cd²⁺ (2 mM) totally and reversibly abolished the releasing effects of all three peptides. Like GHRP-6, the GH release induced by KP 101 was not affected by a GRF antagonist ([Ac-Tyr¹, D-Arg²]-GRF 1–29, 1 μM) which significantly reduced the effect of GRF on GH release. For each peptide, the response to a second application (1 h after the first application) was lower than the first response. When GRF (or KP 101, GHRP-6) was applied first and then KP 101 or GHRP-6 (or GRF) given 1 h later, the second response was not attenuated. Only a small additive effect on the release of GH by GRF was obtained by the co-administration of either KP 101 or GHRP-6. This result was achieved with maximal doses of the peptides, but not with half-maximal doses. These data indicate that KP 101 has similar potency and GH releasing properties to GHRP-6 but both are less potent than GRF. There is no synergistic effect of the peptides with GRF and only a small additive effect of KP 101 or GHRP-6 on GRF-stimulated GH release from ovine pituitary cells in vitro. KP 101 stimulates GH release by a mechanism that involves a common step employed also by GRF and GHRP-6, an increase in calcium influx. In addition, our data strongly suggest that KP 101 like GHRP-6 do not act through the GRF receptor and that there is no cross-desensitization the GRF elicited response.     

Abnormal Growth Hormone Release Following Luteinizing Hormone Releasing Hormone in Anorexia Nervosa

Abstract: We studied the luteinizing hormone (LH), follicle stimulating hormone (FSH) and growthhormone (GH) secretion following an i.v. injection of 0.1 nig of luteinizing hormone releasing hormone (LHRH) in patients with anorexia nervosa, who showed the GH secretion afterthyrotropin releasing hormone (TRH). Five out of 11 patients had an elevated plasma GH level in a fasting state. The administration of LHRH resulted in a significant increase in the plasma GH concentrations in 3 of the 11 patients. Three other patients also showed anelevation of the plasma GH concentration to 7.0, 18.4 and 29.6 ng/ml, which were 212, 175 and 191% of the preinjection levels, respectively. There is a positive correlation between the basal and peak plasma GH levels after LHRH. These increases, however, were related to neither the plasma gonadotropin responses to LHRH nor the plasma GH responses to TRH. The basal levels of plasma LH were reduced in 8 patients and normal responses to LHRH were observed in only one patient. Although plasma FSH was undetectable in 5 patients, the FSH response to LHRH appeared normal in 9 patients. These results indicate thatan elevation of the plasma GH level after LHRH is not confined to patients with a GH secreting pituitary tumor but observed in subjects with anorexia nervosa and further suggest that the GH responsiveness to non-specific hypothalamic releasing hormones may be due to the impaired hypothalamic control in anorexia nervosa.     

Dexamethasone Increases Growth Hormone (GH)-Releasing Hormone (GRH) Receptor mRNA Levels in Cultured Rat Anterior Pituitary Cells

To examine the effects of glucocorticoid (GC) on growth hormone (GH)-releasing hormone (GRH) receptor gene expression, a highly-sensitive and quantitative reverse-transcribed polymerase chain reaction (RT-PCR) method was used in this study. Rat anterior pituitary cells were isolated and cultured for 4 days. The cultured cells were treated with dexamethasone for 2, 6, and 24  h. GRH receptor mRNA levels were determined by competitive RT-PCR using a recombinant RNA as the competitor. Dexamethasone significantly increased GRH receptor mRNA levels at 5  nM after 6- and 24  h-incubations, and the maximal effect was found at 25  nM. The GC receptor-specific antagonist, RU 38486 completely eliminated the dexamethasone-induced enhancement of GRH receptor mRNA levels. Dexamethasone did not alter the mRNA levels of β -actin and prolactin at 5  nM for 24  h, whereas GH mRNA levels were significantly increased by the same treatment. The GH response to GRH was significantly enhanced by the 24-h incubation with 5  nM dexamethasone. These findings suggest that GC stimulates GRH receptor gene expression through the ligand-activated GC receptors in the rat somatotrophs. The direct effects of GC on the GRH receptor gene could explain the enhancement of GRH-induced GH secretion.     

Arachidonate Metabolism in the Anterior Pituitary: Effect of Arachidonate Inhibitors on Basal and Stimulated Secretion of Prolactin, Growth Hormone and Luteinizing Hormone. I. Hormone Release from Incubated or Perifused Pituitary Fragments

The potential involvement of arachidonic acid metabolites in the regulation of adenohypophyseal secretion was analysed on pituitary glands from male rats incubated in the presence of various inhibitors with different mechanisms of action: two inhibitors of phospholipase A₂ (parabromophenacylbromide, PB and compound CB 874), an inhibitor of cyclooxygenase- and lipoxygenase-catalysed pathways (5, 8, 11, 14-eicosatetraynoic acid, ETYA) and an inhibitor of cyclooxygenase (ε-lysyl acetylsalicylate, ASP). Under conditions which minimize side effects of the drugs, all inhibitors reduced prostaglandin synthesis and release, without affecting the metabolic integrity of the tissues (assessed by their intracellular adenosine triphosphate levels). All agents tested (PB, ETYA, ASP) suppressed prolactin secretion induced either by thyrotropin-releasing hormone or vasoactive intestinal peptide. Basal prolactin secretion was sensitive to phospholipase A₂ inhibitors. Similar inhibitions were obtained with ETYA and CB 874 on growth hormone secretion under basal conditions as well as after stimulation by growth hormone-releasing factor, thyrotropin-releasing hormone, or vasoactive intestinal peptide. In contrast, luteinizing hormone secretion, stimulated or not by gonadotropin-releasing hormone, was not sensitive to any of the agents used. It is concluded that, in intact male hemipituitaries, arachidonic acid metabolism is involved in the stimulation of prolactin and growth hormone secretion by neuropeptides. In contrast, luteinizing hormone release does not seem to depend on that mechanism. It has been verified that the inhibitors of arachidonic acid metabolism do not directly interfere with adenylate cyclase, or with the activation of protein kinase C, two enzymes which are involved in the regulation of secretory mechanisms.     

Acetylcholine Stimulates Growth Hormone Secretion in the Neonatal Rat Pituitary

The direct effect of acetylcholine on pituitary growth hormone secretion during the postnatal period of the rat was studied using a superfusion system. Acetylcholine elicited a dose related stimulatory effect on growth hormone (GH) secretion in the pituitaries from 2-day old rats. M, muscarinic agonist McN A343 mimicked the GH releasing effect of acetylcholine, nicotine was ineffective. The GH release elicited by acetylcholine diminished with postnatal development, it was small by the end of the third postnatal week and was not demonstrable in the adult pituitaries. The effect of acetylcholine was potently antagonized by pirenzepine (M₁ antagonist) and 4-DAMP (M₃ and M₁ antagonist) but not by methoctramine (M₂ antagonist). It is concluded that unlike in the adult, in the newborn rat the cholinergic regulation of pituitary GH secretion plays a prominent role directly at pituitary level most likely via M₁ receptors.     

Corticotrophin-Releasing Hormone Stimulates Neurohypophyslal Hormone Release through an Interaction with the Intermediate Lobe of the Pituitary

Corticotrophin-releasing hormone is found co-localized with oxytocin in the magnocellular-neurohypophysical system but its function in this context is unknown. We tested its effects on neurohypophysical hormone secretion in vitro , in the presence and absence of the intermediate lobe of the pituitary. Corticotrophin-releasing hormone caused significant, calcium-dependent secretion of oxytocin and vasopressin from neural lobes in contact with intermediate lobes, i.e. neurointermediate lobes. This effect was inhibited by the dopamine agonist, bromocriptine. Corticotrophin-releasing hormone had no effect on isolated neural lobes in the absence of the intermediate lobe, but α- and γ-melanocyte-stimulating hormone produced an increase in secretion that was comparable in pattern and magnitude to the effect of corticotrophin-releasing hormone on neurointermediate lobes. These findings suggest that corticotrophin-releasing hormone released with oxytocin may act in a paracrine fashion to stimulate release of intermediate peptides which, in turn, can directly evoke release of oxytocin and vasopressin from neural lobe terminals.     

Carbamazepine Effects on Preoptic GABA Release and Pituitary Luteinizing Hormone Secretion in Rats

In vivo effects of carbamazepine (CBZ) on the neuroendocrine preopticopituitary feedback system were studied by local application of CBZ through a push—pull cannula into the preoptic area and measurement of local effects on γ-aminobutyric acid (GABA) and distant effects on a subsequent biologic response: luteinizing hormone (LH). Perfusion with 8 and 12 μg CBZ/ml cerebro-spinal fluid caused a reduction in preoptic GABA release and concomitant suppression of plasma LH levels. These results suggest a GABA component to the mechanism(s) of action of CBZ: (a) CBZ reduces extracellular available GABA concentration, and (b) owing to the known inhibitory role of preoptic GABA in pituitary LH secretion, an increase of postsynaptic GABAergic transmission by CBZ itself could be inferred.     

Cyclosporine Effects on in vitro Responsiveness of Anterior Pituitary Hormone Release to Dopamine and Thyrotropin-Releasing Hormone in Young Female Rats

Endocrine side effects of the immunosuppressive drug cyclosporine (CyA) include changes in anterior pituitary hormone secretion. The aim of the present study was to examine the effects of CyA on the responsiveness of in situ and ectopic anterior pituitary prolactin (PRL), growth hormone (GH) and luteinizing hormone (LH) release response to dopamine (DA) and thyrotropin-releasing hormone (TRH) treatment in young female rats, and to evaluate the possible PRL participation in these effects. Thirty day old rats were rendered hyperprolactinemic by transplanting an anterior pituitary gland of a littermate donor, under the kidney capsule, and were then injected with CyA or vehicle for 2 or 8 days. Sham-operated rats were used as controls and treated in the same way. CyA treatment prevented the increase in plasma PRL levels which occurred in controls after pituitary grafting. In vitro basal PRL release of in situ pituitaries from either sham-operated and/or pituitary-grafted animals was decreased by CyA treatment at any point studied. Basal in vitro secretion of GH was only decreased in the in situ pituitaries from grafted animals after 2 days of CyA therapy. The presence of an ectopic pituitary lead to an increase in the in vitro basal LH secretion from in situ pituitaries as compared to those from sham-operated rats. Basal LH release rates were not changed by CyA treatment, although the LH release in vitro did increase in the in situ pituitaries from sham-operated animals treated with the drug for 2 days. DA addition to the incubation media decreased the in vitro release of PRL, GH and LH from the in situ pituitaries of sham-operated and pituitary-grafted animals treated with vehicle. In CyA treated animals, DA decreased in vitro PRL release from the in situ pituitaries of animals, independently of the presence or absence of an ectopic pituitary. Reductions of the in vitro GH and LH release after DA treatment were higher in the in situ pituitaries from grafted animals on day 8 of CyA or vehicle treatment. TRH increased the in vitro release of the three hormones with differential effects related to the length of the treatment with CyA and/or the presence of an ectopic pituitary. In vitro release of PRL and GH by ectopic pituitaries was inhibited by previous treatment with CyA and this effect was decreased proportional to the duration of the treatment with the drug, while LH secretion was not modified. Addition of DA to the incubation media resulted in a marked reduction of in vitro PRL and GH release, but only at day 8 of vehicle treatment on GH release did DA addition to media further decrease the release of both hormones from ectopic pituitaries from animals treated for 2 or 8 days with the drug, whereas LH secretion was not modified. TRH addition to the incubation media of ectopic pituitaries surprisingly reduced PRL and GH secretion on day 8 of CyA treatment or after surgery. The results of these studies suggest that CyA can act directly at the hypophyseal level modifying pituitary responsiveness to external stimuli. CyA seems to exert its main effects on lactotroph activity while its effects on somatotrophs and gonadotrophs are less.     

Synchronous Pulsatile Release of Luteinizing Hormone and Immunoreactive Beta-Endorphin from the Human Pituitary in vitro

An in vitro perifusion system employing very frequent (30 s) perifusate collectionswas utilized to investigate the relationship between pulsatile release of luteinizing hormone (LH) and immunoreactive β-endorphin (iβ-END) during individual perifusions of adult human anterior hemipituitaries. Each of six hemipituitaries released LH and iβ-END in a distinctlypulsatile fashion, with pulses occurring approximately every 3.2 min for each. Power spectral analysis revealed that pulsatile release of both LH and iβ-END occurred in a rhythmic pattern, with a periodicity of 3.1 and 3.2 min, respectively, and that the periodicity of pulsatile LH and iβ-END release was correlated within individual perifusions. Moreover, the relative amplitudes (% change) of the synchronous LH and iβ-END pulses were correlated. The effluent fractions from two of the perifusions were also assessed for thyrotropin, and it was determinedthat thyrotropin pulses were synchronized to both LH and iβ-END pulses. These studies confirm that LH and iβ-END are released from human anterior pituitaries in vitro in an intrinsically pulsatile fashion, and demonstrate that the LH and iβ-END pulses tend to occur rhythmically and in synchrony and proportion with each other. Furthermore, correlation of thyrotropin pulses to both the LH and iβ-END pulses suggests a common fundamental intrapituitary pulse generating mechanism.     

Effects of Estrogen and Ectopic Pituitary Transplants on the Responsiveness of Pituitary Prolactin Release to Luteinizing Hormone-Releasing Hormone and Dopamine

The responsiveness of prolactin release to regulatory inputs depends on the functional state of the lactotrophs. In the present study, we have examined the effects of luteinizing hormone-releasing hormone (LHRH; 10^–7 or 10^–6 M) on the release of prolactin in vitro from hyperplastic pituitaries of estrogen-treated male Fischer rats, ectopic pituitary transplants and in situ pituitaries of grafted and control rats. The effects of dopamine (10^–8 or 10^–7 M) in this system were also examined. The extent of inhibition of prolactin release by dopamine was not related to the amounts of prolactin secreted under basal conditions or to plasma prolactin levels. LHRH significantly suppressed prolactin secretion in all groups but its effect was most pronounced in the ectopic pituitary transplants and in the hyperplastic pituitaries of animals after chronic exposure to estrogen followed by a period of recovery. Thus, the effects of LHRH on prolactin release appear to be related to the secretory activity and/or to the absolute or relative number of the lactotrophs.     

Seasonal Effect of Gonadotrophin Inhibitory Hormone on Gonadotrophin‐Releasing Hormone‐induced Gonadotroph Functions in the Goldfish Pituitary

We have shown that native goldfish gonadotrophin inhibitory hormone (gGnIH) differentially regulates luteinsing hormone (LH)‐β and follicle‐stimulating hormone (FSH)‐β expression. To further understand the functions of gGnIH, we examined its interactions with two native goldfish gonadotrophin‐releasing hormones, salmon gonadotrophin‐releasing hormone (sGnRH) and chicken (c)GnRH‐II in vivo and in vitro. Intraperitoneal injections of gGnIH alone reduced serum LH levels in fish in early and mid gonadal recrudescence; this inhibition was also seen in fish co‐injected with either sGnRH or cGnRH‐II during early recrudescence. Injection of gGnIH alone elevated pituitary LH‐β and FSH‐β mRNA levels at early and mid recrudescence, and FSH‐β mRNA at late recrudescence. Co‐injection of gGnIH attenuated the stimulatory influences of sGnRH on LH‐β in early recrudescence, and LH‐β and FSH‐β mRNA levels in mid and late recrudescence, as well as the cGnRH‐II‐elicited increase in LH‐β, but not FSH‐β, mRNA expression at mid and late recrudescence. sGnRH and cGnRH‐II injection increased pituitary gGnIH‐R mRNA expression in mid and late recrudescence but gGnIH reduced gGnIH‐R mRNA levels in late recrudescence. gGnIH did not affect basal LH release from perifused pituitary cells and continual exposure to gGnIH did not alter the LH responses to acute applications of GnRH. However, a short 5‐min GnIH treatment in the middle of a 60‐min GnRH perifusion selectively reduced the cGnRH‐II‐induced release of LH. These novel results indicate that, in goldfish, gGnIH and GnRH modulate pituitary GnIH‐R expression and gGnIH differentially affects sGnRH and cGnRH‐II regulation of LH secretion and gonadotrophin subunit mRNA levels. Furthermore, these actions are manifested in a reproductive stage‐dependent manner.     

Nutrient Sensing Overrides Somatostatin and Growth Hormone‐Releasing Hormone to Control Pulsatile Growth Hormone Release

Pharmacological studies reveal that interactions between hypothalamic inhibitory somatostatin and stimulatory growth hormone‐releasing hormone (GHRH) govern pulsatile GH release. However, in vivo analysis of somatostatin and GHRH release into the pituitary portal vasculature and peripheral GH output demonstrates that the withdrawal of somatostatin or the appearance of GHRH into pituitary portal blood does not reliably dictate GH release. Consequently, additional intermediates acting at the level of the hypothalamus and within the anterior pituitary gland are likely to contribute to the release of GH, entraining GH secretory patterns to meet physiological demand. The identification and validation of the actions of such intermediates is particularly important, given that the pattern of GH release defines several of the physiological actions of GH. This review highlights the actions of neuropeptide Y in regulating GH release. It is acknowledged that pulsatile GH release may not occur selectively in response to hypothalamic control of pituitary function. As such, interactions between somatotroph networks, the median eminence and pituitary microvasculature and blood flow, and the emerging role of tanycytes and pericytes as critical regulators of pulsatility are considered. It is argued that collective interactions between the hypothalamus, the median eminence and pituitary vasculature, and structural components within the pituitary gland dictate somatotroph function and thereby pulsatile GH release. These interactions may override hypothalamic somatostatin and GHRH‐mediated GH release, and modify pulsatile GH release relative to the peripheral glucose supply, and thereby physiological demand.     

Large Molecular Weight Immunoreactive Corticotrophin-Releasing Hormone has Bioactivity on Superfused Ovine Pituitary Cells

Human placental extracts fractionated with Sephadex G-50 produced three peaks of corticotrophin-releasing hormone immunoreactivity, a large molecular weight peak (M_r30,000), an intermediate peak (4,758 < M_r < 10,000) and a low molecular weight peak coeluting with the 41-residue hormone. All three peaks of immunoreactivity stimulated the release of β-endorphin-like immunoreactivity from ovine pituitary cells superfused in vitro. No response was observed from unstimulated cells superfused in parallel. Gel chromatography indicated that intermediate and small molecular weight forms of human corticotrophin-releasing hormone immunoreactivity remained intact after contact with the ovine pituitary cells, whereas the large molecular weight material dissociated to produce 41-residue hormone immunoreactivity. The secreted β-endorphin immunoreactivity was shown by gel chromatography to comprise both β-lipotrophin-like and the 31-residue β-endorphin-like immunoreactivity. The data show that the intermediate and low molecular weight forms of placental corticotrophin-releasing hormone immunoreactivity are bioactive and suggest that the intermediate form is a hormone precursor, possibly procorticotrophin-releasing hormone_125–196, and the small form is identical to the hypothalamic hormone. The results with the larger molecular weight material indicate that it is likely to be a complex of the mature 41-residue hormone and a binding protein.     

Prolonged Hormone Secretion from Neuroendocrine Cells of Aplysia is Indepentdent of Extracellular Calcium

The role of Ca²⁺ from extracellular and intracellular sources in stimulating neurosecretion was investigated in four experiments using neuroendocrine bag cells of the marine mollusk Aplysia . (i) Bag cells were treated with either an extracellular calcium chelator (BAPTA) or Co²⁺-substitution within 30  s after onset of an electrical afterdischarge to prevent influx of Ca²⁺ from extracellular fluid. These treatments shortened the duration of the afterdischarge, but did not significantly affect the overall pattern or total amount of egg laying hormone (ELH) secretion, suggesting that extracellular Ca²⁺ is not required for maintenance of ELH release. (ii) Substitution of Ba²⁺ for Ca²⁺ has previously been shown to support bag cell afterdischarges that trigger transient elevations in intracellular Ca²⁺. We showed that this treatment also stimulates ELH secretion, suggesting that Ca²⁺ release from intracellular stores can stimulate ELH secretion. (iii) To raise intracellular Ca²⁺ levels in the absence of an afterdischarge, the calcium ionophore X537A was used to transport Ca²⁺ across plasma and organelle membranes. When this treatment was combined with extracellular calcium chelators so that the only source of Ca²⁺ was from intracellular compartments, ELH secretion was stimulated. Taken together, these findings are consistent with the hypothesis that release of Ca²⁺ from intracellular stores is sufficient to stimulate ELH secretion.     

Effects of Glucose on Thyrotrophin-Releasing Hormone, Growth Hormone-Releasing Hormone, Somatostatin and Luteinizing Hormone-Releasing Hormone Release from Rat Hypothalamus in vitro

Increasing concentrations of D-glucose (1 to 25 mM) inhibited somatostatin, thyrotrophin-releasing hormone (TRH) and growth hormone-releasing hormone (GHRH) release from incubated adult rat hypothalami in a stereospecific manner. In contrast, the effects of D- and L-glucose on luteinizing hormone-releasing hormone release were virtually identical. Increasing concentrations of D-glucose also inhibited somatostatin release following depolarization with high K⁺, but had no obvious effect on depolarization-induced TRH or GHRH release when compared with L-glucose.
In conclusion, D-glucose exerts a potent, dose-related modulatory action on the release of rat hypothalamic TRH and GHRH as well as somatostatin in vitro. Further studies are required to establish any physiological relevance of glucose in the modulation of these hypothalamic neuropeptides.     

Stimulatory Effect of Calcitonin Gene-Related Peptide on Adrenocorticotropin Release from Rat Anterior Pituitary Cells

In the present study, we examined the direct regulatory effect of rat calcitonin gene-related peptide (CGRP) on adrenocorticotropin (ACTH) release from rat cultured anterior pituitary cells. CGRP significantly increased ACTH release at concentrations of 10⁻⁸–10⁻¹¹ M. The ACTH release was gradually increased by CGRP concentrations lower than 10⁻¹⁰ M, and was decreased at concentrations higher than 10⁻⁹ M, presenting a bell-shaped dose-response curve. As well as having an additive effect on corticotropin-releasing factor-induced ACTH release, CGRP stimulated the accumulation of intracellular cAMP. The CGRP-induced ACTH release was inhibited by a protein kinase A inhibitor, suggesting that its stimulatory effect on the ACTH release was mediated via an adenylate–cyclase–protein kinase system. CGRP-like immunoreactive nerve fibers have been reported to innervate the anterior pituitary, so that the stimulatory effect of CGRP on the ACTH release suggests that this peptide may be involved in neural regulation of hormone secretion in the anterior pituitary.     

脂多糖抑制离体培养大鼠垂体前叶细胞释放S-100b蛋白

目的观察脂多糖(LPS)对大鼠垂体前叶滤泡星形细胞分泌S-100b蛋白的影响。方法将原代培养大鼠垂体前叶细胞分为对照组和LPS处理组,经LPS不同剂量处理后以作者科室建立的敏感和特异的S-100蛋白ELISA检测方法测定培养上清中和细胞裂解液中S-100b蛋白水平。结果大鼠垂体前叶细胞培养上清中S-100b蛋白水平随细胞生长过程逐渐增高,尤其在培养前12 h增高明显。培养上清中乳酸脱氢酶水平无明显变化。LPS可剂量依赖性地抑制培养上清中S-100b蛋白水平增高。在培养12 h时,对照组细胞的胞浆中S-100b蛋白水平显著增加,LPS可完全抑制胞浆中S-100b蛋白水平上升。结论LPS抑制大鼠垂体前叶滤泡星形细胞分泌S-100b蛋白,提示感染时LPS可能通过改变S-100b蛋白分泌而影响垂体前叶细胞的功能。