Proliferative effects of cholecystokinin in GH3 pituitary cells mediated by CCK2 receptors and potentiated by insulin. (9/4031)

1. Proliferative effects of CCK peptides have been examined in rat anterior pituitary GH3 cells, which express CCK2 receptors. 2. CCK-8s, gastrin(1-17) and its glycine-extended precursor G(1-17)-Gly, previously reported to cause proliferation via putative novel sites on AR4-2J and Swiss 3T3 cells, elicited significant dose dependent increases of similar magnitude in [3H]thymidine incorporation over 3 days in serum-free medium of 39 +/- 10% (P < 0.01, n = 20), 37 +/- 8% (P < 0.01, n = 27) and 41 +/- 6% (P < 0.01, n = 36) respectively. 3. CCK-8s and gastrin potentially stimulated mitogenesis (EC50 values 0.12 nM and 3.0 nM respectively), whilst G-Gly displayed similar efficacy but markedly lower potency. L-365,260 consistently blocked each peptide. The CCK2 receptor affinity of G-Gly in GH3 cells was 1.09 microM (1.01;1.17, n = 6) and 5.53 microM (3.71;5.99, n = 4) in guinea-pig cortex. 4. 1 microM G-Gly weakly stimulated Ca2+ increase, eliciting a 104 +/- 21% increase over basal Ca2+ levels, and was blocked by 1 microM L-365,260 whilst CCK-8s (100 nM) produced a much larger Ca2+ response (331 +/- 14%). 5. Insulin dose dependently enhanced proliferative effects of CCK-8s with a maximal leftwards shift of the CCK-8s curve at 100 ng ml(-1) (17 nM) (EC50 decreased 500 fold, from 0.1 nM to 0.2 pM; P < 0.0001). 10 microg ml(-1) insulin was supramaximal reducing the EC50 to 5 pM (P = 0.027) whilst 1 ng ml(-1) insulin was ineffective. Insulin weakly displaced [125I]BHCCK binding to GH3 CCK2 receptors (IC50 3.6 microM). 6. Results are consistent with mediation of G-Gly effects via CCK2 receptors in GH3 cells and reinforce the role of CCK2 receptors in control of cell growth. Effects of insulin in enhancing CCK proliferative potency may suggest that CCK2 and insulin receptors converge on common intracellular targets and indicates that mitogenic stimuli are influenced by the combination of extracellular factors present.  (+info)

Differential blockade of gamma-aminobutyric acid type A receptors by the neuroactive steroid dehydroepiandrosterone sulfate in posterior and intermediate pituitary. (10/4031)

Dehydroepiandrosterone sulfate (DHEAS) is a neuroactive steroid with antagonist action at gamma-aminobutyric acid type A (GABAA) receptors. Patch-clamp techniques were used to investigate DHEAS actions at GABAA receptors of the rat pituitary gland at two distinct loci: posterior pituitary nerve terminals and intermediate pituitary endocrine cells. The GABA responses in these two regions were quite different, with posterior pituitary responses having smaller amplitudes and desensitizing more rapidly and more completely. DHEAS blockade of GABAA receptors in the two regions also was different. In posterior pituitary, a site with an apparent dissociation constant of 15 microM accounted for most of the blockade, but a small fraction of blockade may be related to a site with a dissociation constant in the nanomolar range. In the intermediate lobe, DHEAS sensitivities in the nanomolar and micromolar ranges were clearly evident, in proportions that varied widely from cell to cell. Regardless of whether the GABA response of a cell was highly sensitive or weakly sensitive to DHEAS, GABA alone evoked currents that were indistinguishable in terms of amplitude, desensitization kinetics, and GABA sensitivity. Thus, the structural elements responsible for DHEAS blockade have a highly selective impact on receptor function. GABAA receptors with nanomolar sensitivity to DHEAS have not been described previously. This suggests that DHEAS may have an important role in the modulation of neuropeptide secretion, and the diverse properties of GABAA receptors in the rat pituitary provide mechanisms for selective regulation of the different peptidergic systems of this gland.  (+info)

Two gonadotropin-releasing hormone receptor subtypes with distinct ligand selectivity and differential distribution in brain and pituitary in the goldfish (Carassius auratus). (11/4031)

In the goldfish (Carassius auratus) the two endogenous forms of gonadotropin-releasing hormone (GnRH), namely chicken GnRH II ([His5, Trp7,Tyr8]GnRH) and salmon GnRH ([Trp7,Leu8]GnRH), stimulate the release of both gonadotropins and growth hormone from the pituitary. This control is thought to occur by means of the stimulation of distinct GnRH receptors. These receptors can be distinguished on the basis of differential gonadotropin and growth hormone releasing activities of naturally occurring GnRHs and GnRHs with variant amino acids in position 8. We have cloned the cDNAs of two GnRH receptors, GfA and GfB, from goldfish brain and pituitary. Although the receptors share 71% identity, there are marked differences in their ligand selectivity. Both receptors are expressed in the pituitary but are differentially expressed in the brain, ovary, and liver. Thus we have found and cloned two full-length cDNAs that appear to correspond to different forms of GnRH receptor, with distinct pharmacological characteristics and tissue distribution, in a single species.  (+info)

Modulation of estrogen action in the rat pituitary and mammary glands by dietary energy consumption. (12/4031)

We are investigating the mechanisms through which estrogens induce development of prolactin (PRL)-producing pituitary tumors and mammary carcinomas in rats and how these mechanisms are affected by dietary energy consumption. The hypothesis under examination is that dietary energy restriction inhibits tumorigenesis in estrogen-responsive tissues by altering cellular responsiveness to estrogenic hormones. In the Fischer 344 (F344) rat strain, a 40% restriction of energy consumption virtually abolishes development of estrogen-induced pituitary tumors. Inhibition of pituitary tumorigenesis in the F344 strain by energy restriction results from modulation of estrogen regulation of cell survival, not cell proliferation. In contrast, energy restriction has no inhibitory effect on estrogen-induced pituitary tumor development in the ACI rat strain. However, energy restriction markedly inhibits induction of mammary carcinomas in female ACI rats treated with 17beta-estradiol. Data presented herein indicate that dietary energy restriction modulates the responsiveness of specific cell populations to estrogenic hormones and thereby inhibits estrogen-induced tumorigenesis in a manner specific to both rat strain and tissue.  (+info)

L-type Ca2+ channels and K+ channels specifically modulate the frequency and amplitude of spontaneous Ca2+ oscillations and have distinct roles in prolactin release in GH3 cells. (13/4031)

GH3 cells showed spontaneous rhythmic oscillations in intracellular calcium concentration ([Ca2+]i) and spontaneous prolactin release. The L-type Ca2+ channel inhibitor nimodipine reduced the frequency of Ca2+ oscillations at lower concentrations (100nM-1 microM), whereas at higher concentrations (10 microM), it completely abolished them. Ca2+ oscillations persisted following exposure to thapsigargin, indicating that inositol 1,4,5-trisphosphate-sensitive intracellular Ca2+ stores were not required for spontaneous activity. The K+ channel inhibitors Ba2+, Cs+, and tetraethylammonium (TEA) had distinct effects on different K+ currents, as well as on Ca2+ oscillations and prolactin release. Cs+ inhibited the inward rectifier K+ current (KIR) and increased the frequency of Ca2+ oscillations. TEA inhibited outward K+ currents activated at voltages above -40 mV (grouped within the category of Ca2+ and voltage-activated currents, KCa,V) and increased the amplitude of Ca2+ oscillations. Ba2+ inhibited both KIR and KCa,V and increased both the amplitude and the frequency of Ca2+ oscillations. Prolactin release was increased by Ba2+ and Cs+ but not by TEA. These results indicate that L-type Ca2+ channels and KIR channels modulate the frequency of Ca2+ oscillations and prolactin release, whereas TEA-sensitive KCa,V channels modulate the amplitude of Ca2+ oscillations without altering prolactin release. Differential regulation of these channels can produce frequency or amplitude modulation of calcium signaling that stimulates specific pituitary cell functions.  (+info)

IL-8 mRNA expression by in situ hybridisation in human pituitary adenomas. (14/4031)

Several cytokines have been shown to be expressed in normal and adenomatous pituitary tissue. Recently, interleukin-8 (IL-8) mRNA was identified by reverse transcription (RT)-PCR in each of a series of 17 pituitary tumours examined. We have investigated further the presence of IL-8 mRNA, using in situ hybridisation in two normal human anterior pituitary specimens and 25 human pituitary adenomas. IL-8 mRNA was not identified in either of the two normal pituitary specimens. Only three of the 25 adenomas were positive for IL-8 mRNA. In these three tumours, which included two null cell adenomas and one gonadotrophinoma, the majority of tumour cells (>90%) were positive for IL-8 mRNA. The remaining 22 adenomas were completely negative. There was no difference in tumour size or type between the IL-8 positive and the IL-8 negative tumours, and immunocytochemistry for von Willebrandt factor showed that the two groups were also similar in their degree of vascularisation. In conclusion, IL-8 mRNA was found in 12% of pituitary adenomas studied and was histologically identified within the tumour cells. In situ hybridisation is a more appropriate technique for assessing cytokine mRNA production by human pituitary tumours because RT-PCR may be too sensitive, identifying very small, possibly pathologically insignificant, quantities of mRNA that could be produced by supporting cells such as fibroblasts, endothelial cells or macrophages.  (+info)

Effect of long-term food restriction on pituitary sensitivity to cLHRH-I in broiler breeder females. (15/4031)

The effect of long-term food restriction on the sensitivity of the pituitary to exogenously administered chicken luteinizing hormone releasing hormone I (cLHRH-I) was investigated in three groups of broiler breeder females fed ad libitum, fed a restricted quantity of food or fed a restricted quantity of food to obtain an intermediate body weight between those of the first two groups. At 16 weeks of age, basal FSH release was higher in ad libitum fed birds, culminating in ovarian development and subsequent oestradiol production by the small follicles. At this age, LH secretion was independent of ovarian feedback factors. In all groups, cLHRH-I was most active in releasing LH in intact and ovariectomized animals and, to a lesser extent, in releasing FSH in ovariectomized birds. At 39 weeks of age, basal FSH concentrations were similar among intact animals of all groups, whereas LH concentrations differed among groups, with higher values in the restricted birds. This food effect was enhanced in ovariectomized birds. Furthermore, the high response to cLHRH-I in the ovariectomized, restricted birds compared with the ad libitum, ovariectomized group suggests an improved sensitivity of the hypothalamic-pituitary axis. In conclusion, birds fed ad libitum showed the highest responsiveness to ovarian factors and to cLHRH-I in releasing FSH in the period before sexual maturity. No effect of amount of feeding could be observed for LH. However, during the egg laying period, LH release by cLHRH-I was highly dependent on amount of feeding and on ovarian feedback regulation. This finding indicates that the amount of feeding can modify the sensitivity of the pituitary to cLHRH-I, and possibly to gonadal hormones, during the laying period.  (+info)

Annual cycle in LH and testosterone release in response to GnRH challenge in male woodchucks (Marmota monax). (16/4031)

Testosterone and LH concentrations were determined in serum samples obtained before and 15 min after injections of GnRH (1 microgram kg-1) administered at 4-7 week intervals over 20 months to groups of male woodchucks (n = 6-7) born and maintained in Northern Hemisphere (boreal) versus Southern Hemisphere (austral) simulated natural photoperiods, beginning at 18-24 months of age. Nadir and peak unstimulated testosterone (0.1 +/- 0.01 and 7.0 +/- 0.1 ng ml-1, respectively) and LH (0.8 +/- 0.2 and 8.1 +/- 1.1 ng ml-1, respectively) concentrations did not differ in boreal versus austral males. In the five boreal and five austral males that were confirmed to be photoentrained, basal (pre-GnRH) concentrations of LH and testosterone were lowest in summer, increased simultaneously in late autumn or early winter, and declined in the spring. GnRH stimulated some LH release throughout the year except for a 1-4 month period in the summer. The initial annual increase in the LH response to GnRH occurred in early autumn, and in 17 of 20 cycles it occurred 1-2 months before the initial increase in basal LH was detected. In the three free-running males not entrained to the photoperiod, the endocrine patterns were similar but were advanced by several months. The results demonstrate that in woodchucks there is a late autumn increase in LH secretion associated with the onset of testicular recrudescence, and an early autumn increase in pituitary response to GnRH before a detectable increase in serum testosterone.  (+info)