Involvement of thyrotropin-releasing hormone receptor, somatostatin receptor subtype 2 and corticotropin-releasing hormone receptor type 1 in the control of chicken thyrotropin secretion.

Thyrotropin or thyroid-stimulating hormone (TSH) secretion in the chicken is controlled by several hypothalamic hormones. It is stimulated by thyrotropin-releasing hormone (TRH) and corticotropin-releasing hormone (CRH), whereas somatostatin (SRIH) exerts an inhibitory effect. In order to determine the mechanism by which these hypothalamic hormones modulate chicken TSH release, we examined the cellular localization of TRH receptors (TRH-R), CRH receptors type 1 (CRH-R1) and somatostatin subtype 2 receptors (SSTR2) in the chicken pars distalis by in situ hybridization (ISH), combined with immunological staining of thyrotropes. We show that thyrotropes express TRH-Rs and SSTR2s, allowing a direct action of TRH and SRIH at the level of the thyrotropes. CRH-R1 expression is virtually confined to corticotropes, suggesting that CRH-induced adrenocorticotropin release is the result of a direct stimulation of corticotropes, whereas CRH-stimulated TSH release is not directly mediated by the known chicken CRH-R1. Possibly CRH-induced TSH secretion is mediated by a yet unknown type of CRH-R in the chicken. Alternatively, a pro-opiomelanocortin (POMC)-derived peptide, secreted by the corticotropes following CRH stimulation, could act as an activator of TSH secretion in a paracrine way.

Identification of somatostatin receptors controlling growth hormone and thyrotropin secretion in the chicken using receptor subtype-specific agonists.

Somatostatin (SRIH) functions as an endocrine mediator in processes such as growth, immune resistance and reproduction. Five SRIH receptors (sstr1-5) have been identified in mammals, where they are expressed in both the brain and peripheral tIssues. To study the specific function of each receptor subtype, specific agonists (ag1-5) have been synthesized. The high degree of homology between mammalian and avian SRIH receptors suggests that these agonists might also be used in chickens. In this paper we describe two in vitro protocols (static incubation and perifusion system) to identify the SRIH receptors controlling the secretion of GH and TSH from the chicken pituitary. We found that basal GH or TSH secretion were never affected when SRIH or an agonist (1 microM) were added. SRIH diminished the GH as well as the TSH response to TSH-releasing hormone (TRH; 100 nM) in both systems. Our results have indicated that the SRIH actions at the level of the pituitary are regulated through specific receptor subtypes. In both the static and flow incubations, ag2 lowered the GH response to TRH, whereas stimulated TSH release was diminished by both ag2 and ag5. Ag3 and ag4 tended to increase rather than decrease the responsiveness of both pituitary cell types to TRH in perifusion studies. Our data have indicated that SRIH inhibits chicken pituitary function through sstr2 and sstr5. Only sstr2 seems to be involved in the control of chicken GH release, whereas both sstr2 and sstr5 inhibit induced GH secretion in mammals. The possible stimulatory action of ag3 and ag4 may point towards a species-specific function of sstr3 and sstr4.

Synthetic growth hormone secretagogues control growth hormone secretion in the chicken at pituitary and hypothalamic levels.

In the chicken growth hormone (GH) secretion is predominantly controlled by two hormones, thyrotropin-releasing hormone (TRH) and somatostatin (SRIH), respectively stimulating and inhibiting GH release. In view of the hypothesis of a novel GH secretagogue (GHS) in mammals, this specific species was used to further assess the exact function of two nonpeptidyl GHSs-L-692,429 and L-163,255. Both synthetic products stimulate GH secretion directly at the level of the pituitary as shown in in vitro perifusion studies. Plasma GH levels increase within 10-15 min after a single challenge of L-692,429 or L-163,255. A SRIH pretreatment dimishes this GH response. Both GH-releasing peptide mimetics decrease hypothalamic TRH concentrations, whereas SRIH levels are not affected. The novel GHS may therefore control GH secretion both at the level of the pituitary and the hypothalamus. The present article shows that nonpeptidyl mimetics also control GH secretion in nonmammalian species suggesting that the endogenous hormone may be a conserved GH stimulator in several vertebrates. The GH response to GHS in birds may be regulated both directly at the level of the pituitary and by releasing another endogenous GH stimulator (TRH) from the hypothalamus.

Distribution of somatostatin in the brain and of somatostatin and thyrotropin-releasing hormone in peripheral tissues of the chicken.

Our research group recently presented the distribution of thyrotropin-releasing hormone (TRH) in the chicken brain. In this study we measured somatostatin (SRIH) concentrations in different brain parts and nuclei. The distribution of SRIH and TRH in peripheral tissues was also studied. Although the highest SRIH content was found in endocrine areas like diencephalon and median eminence (ME), high levels were also recorded in brain stem and several hypothalamic nuclei which do not project to the ME. SRIH immunoreactivity was also found within the pituitary. In peripheral tissues, SRIH was mainly present in gonads, thyroid and intestine. Low amounts were found in duodenum, kidney, heart and lung. SRIH concentrations were barely detectable (liver, blood cells) or undetectable (muscle, skin, spleen) in other peripheral tissues investigated. Although TRH was found in all tissues collected, it was also most abundant in brain, pituitary, thyroid and gonads. Our results suggest that also in the chicken SRIH and TRH are implicated in the control of several physiological processes like growth, reproduction and digestion.

Modulation of the growth hormone-releasing activity of thyrotropin-releasing hormone in the chicken by its gene-related peptide preproTRH((160-169))(Ps4): enhanced somatostatinergic tone?

Recent research demonstrated that endocrine actions of thyrotropin (TSH)-releasing hormone (TRH) are modulated by gene-related products within proTRH. In the present report we show that the growth hormone (GH) response to TRH is clearly inhibited after the preincubation of chicken pituitary glands with preproTRH((160-169))Ps4, whereas the TSH response is not impaired. Binding sites for(125)I-[Tyr(0)]-Ps4 were, however, not detected on chicken pituitary membranes, although (as a control) they were readily detectable on membranes from rat pituitary glands. An indirect action may therefore take place within the pituitary by modulating the action of somatostatin (SRIH), the inhibitor of GH release in the chicken. This hypothesis is strengthened by the observation that Ps4 increases the binding of(125)I-[Tyr(1)]-SRIH to chicken pituitary membranes in a dose-related way. Since Ps4 is also produced by pituitary tissue, this may reflect a local or paracrine action on the regulation of GH release.

Adrenal inhibition of corticotropin-releasing hormone-induced thyrotropin release: a comparative study in pre- and posthatch chicks.

Recent evidence indicates that corticotropin-releasing hormone (CRH) acts as a potent stimulator of thyrotropin (TSH) release in the chicken. In this study adrenal and thyroidal feedback mechanisms were studied. Administration of corticosterone 30 min prior to an ovine CRH (oCRH) challenge diminished the in vivo sensitivity of thyrotrophs to oCRH in 19-day-old chicken embryos (E19) (20 micrograms corticosterone; 2 micrograms oCRH) but not in 8-day-old chickens (C8) (40 micrograms corticosterone; 4 micrograms oCRH). At both ages studied, corticosterone (0.01 and 1 microM) did not alter the in vitro TSH response to oCRH (100 nM) indicating that an indirect mechanism is involved at the embryonic stage which is no longer present in posthatch chickens. In vitro, 3,5,3'-triiodothyronine (T3) pretreatment (0.01 and 1 microM) resulted at both ages studied in a dose-dependent drop in the in vitro oCRH-induced TSH release. As recorded previously, corticosterone treatment provoked a rise in plasma T3 in embryonic but not in posthatch chickens. The presence of an indirect adrenal feedback mechanism in chicken embryos may therefore be linked to the increase in plasma T3 which will alter the sensitivity of thyrotrophs to hypothalamic releasing factors. In conclusion, corticosterone does not directly modulate the responsiveness of thyrotrophs to CRH, but its feedback mechanism may be dependent on the evoked increase in plasma T3 which is only present in embryonic chickens. Corticosterone may in this regard play an essential role during embryonic development by coordinating thyroidal feedback mechanisms at the level of the chicken pituitary.

The drop in plasma thyrotropin concentrations in fasted chickens is caused by an action at the level of the hypothalamus: role of corticosterone.

Fasting has severe effects on thyroid metabolism in the chicken: plasma thyroxine (T4) concentrations increase, whereas 3',5,3-triiodothyronine (T3) concentrations decrease. In the present report we studied the effect of fasting at the level of: 1) the pituitary (plasma thyrotropin (TSH) concentrations; the sensitivity of thyrotrophs to corticotropin-releasing hormone (CRH) and TSH-releasing hormone (TRH)); and 2) the hypothalamus (TRH content). A regulatory role of corticosterone is discussed. One day of fasting resulted in a drop in plasma TSH concentrations. Fed and nonfed animals were treated with ovine CRH (oCRH) or TRH. The sensitivity of thyrotrophs to the respective hypothalamic hormones was increased when animals were subjected to a 1-d period of fasting. A 75% (TRH) and 50% (oCRH) increase in plasma TSH was recorded in fasted animals, whereas both secretagogues did not evoke any response in their fed counterparts. The drop in plasma TSH cannot, therefore, be attributed to a loss in sensitivity of thyrotrophs to hypothalamic stimulatory control. In an identical experiment, plasma TSH concentrations decreased, whereas hypothalamic TRH content was higher in fasted animals, suggesting a decreased hypothalamic TRH release toward the pituitary. In both fasting experiments, plasma corticosterone concentrations were increased after 1 d of fasting. Because an i.v. injection of corticosterone-elevated hypothalamic TRH contents and decreased plasma TSH concentrations, a corticosterone-induced TSH decrease during fasting is suggested through an action at the level of the hypothalamus.

Thyrotropin-releasing hormone concentrations in different regions of the chicken brain and pituitary: an ontogenetic study.

The regional distribution of thyrotropin-releasing hormone (TRH) was studied in the chicken brain. The hypothalamus and the brain stem contained the highest concentration of TRH. Lower amounts were present in the telencephalon, the optic lobes and the cerebellum. Within the hypothalamus, TRH was most abundant in the median eminence. Other important TRH sites were the nucleus paraventricularis magnocellularis, nucleus periventricularis hypothalami, nucleus ventromedialis hypothalami, nucleus dorsomedialis hypothalami and nucleus preopticus periventricularis. On the 14th day of embryonic development (E14), TRH was mostly found in the brain stem. Towards hatching, TRH concentrations increased gradually in both the hypothalamic area and the brain stem. TRH concentrations in the telencephalon, optic lobes and cerebellum remained low. Pituitaries from E14 to E16 chickens were characterized by a high TRH concentration, whereas hypophyseal TRH concentrations dropped towards hatching. Our results support the hypothesis that TRH exerts both endocrine and neurocrine actions in the chicken. On the other hand, high pituitary TRH concentrations were present when hypothalamic concentrations were low and vice versa. Therefore, the chicken pituitary may function as an important source of TRH during early in ovo development at least until the moment hypothalamic control develops.

Pre- and posthatch developmental changes in hypothalamic thyrotropin-releasing hormone and somatostatin concentrations and in circulating growth hormone and thyrotropin levels in the chicken.

Thyrotropin-releasing hormone (TRH) and somatostatin (SRIH) concentrations were determined by RIA during both embryonic development and posthatch growth of the chicken. Both TRH and SRIH were already detectable in hypothalami of 14-day-old embryos (E14). Towards the end of incubation, hypothalamic TRH levels increased progressively, followed by a further increase in newly hatched fowl. SRIH concentrations remained stable from E14 to E17 and doubled between E17 and E18 to a concentration which was observed up to hatching. Plasma GH levels remained low during embryonic development, ending in a steep increase at hatching. Plasma TSH levels on the other hand decreased during the last week of the incubation. During growth, TRH concentrations further increased, whereas SRIH concentrations fell progressively towards those of adult animals. Plasma TSH levels increased threefold up to adulthood; the rise in plasma GH levels during growth was followed by a drop in adults. In conclusion, the present report shows that important changes occur in the hypothalamic TRH and SRIH concentration during both embryonic development and posthatch growth of the chicken. Since TRH and SRIH control GH and TSH release in the chicken, the hypothalamic data are compared with plasma GH and TSH fluctuations.

Inhibition and activation of the thyroidal axis by the adrenal axis in vertebrates.

Hormones of the adrenal or interrenal axis and stress situations which induce elevated glucocorticoid plasma levels (e.g. handling and starvation), inhibit thyroid function in growing and adult vertebrates. However, data indicate that during foetal and embryonic development (mammals and birds) or during larval growth and metamorphosis (fish and amphibians), the adrenal axis may stimulate thyroid function. Recent findings have provided some information concerning this stimulatory interference of the adrenal axis. In amphibians corticotropin releasing hormone and not thyrotropin releasing hormone is thyrotropic during metamorphosis, thus providing the substrate T4 necessary for T3 production. Other data indicate that the increase in plasma T3 at metamorphic climax may be the result of an inhibition of the T3 degrading activity, rather than stimulation of the T4 into T3 converting activity, and that glucocorticoids may be responsible for this. Also, in the chick embryo glucocorticoids effectively increase plasma T3 concentration by reducing the hepatic T3 degrading activity, whereas corticotropin releasing hormone also induces an elevation in the thyrotropin plasma levels and hence raises T4 concentrations which may function as a substrate for T3 production.

Pituitary and extrapituitary action sites of the novel nonpeptidyl growth hormone (GH) secretagogue L-692,429 in the chicken.

Chickens were used as a model to further analyze the efficacy and specificity of L-692,429, a novel nonpeptidyl mimic of growth hormone (GH)-releasing peptide-6 (GHRP-6), which is a specific GH-releasing secretagogue in mammals. Actions at the level of the pituitary and the hypothalamus were studied. Pituitaries isolated from 1-day-old (C1) chicks responded in a dose-dependent manner to L-692,429 (ED50 = 10 nM). Using equimolar concentrations of thyrotropin-releasing hormone (TRH), human GH-releasing hormone (hGHRH1-29), and L-692,429 (10 nM), L-692,429 had 20-25% the in vitro potency of the two endogenous releasing factors. There was an additive effect between hGHRH1-29 (10 nM) and L-692,429 (10 or 100 nM) on GH release from C1 pituitaries but no such additive effect was observed when pituitaries were exposed to both TRH (10 nM) and L-692,429 (100 nM). An acute challenge with 50 microg L-692,429 resulted in increased plasma GH levels within 5 min, which remained elevated for up to 15 min (C1 chickens). This increase in GH was accompanied by a drop in hypothalamic TRH content by 5 min. Hypothalamic somatostatin (SRIH) content did not change. Plasma corticosterone concentrations were increased following L-692,429 treatment, whereas plasma alpha-subunit, T4, and T3 levels were unchanged. To confirm the role of the decreased hypothalamic TRH concentrations in the GH-releasing activity of L-692,429 in the chicken, chickens (C1) were pretreated with normal rabbit serum (NRS) or a TRH antiserum (1/50) 1 h prior to the L-692,429 challenge. Both groups showed an increase in circulating GH but the increase was within 5 min inhibited by the TRH antiserum pretreatment, whereas no differences were noted in plasma corticosterone levels. It is concluded that in the chicken the GH secretagogue L-692,429 has a dual action site: (1) directly at the level of the pituitary and (2) centrally through an increase in hypothalamic TRH release.

Evidence of a thyrotropin-releasing activity of ovine corticotropin-releasing factor in the domestic fowl (Gallus domesticus).

Ovine corticotropin-releasing factor (oCRF) administered to 19-day-old chicken embryos (E19) increased plasma concentration of pituitary glycoprotein alpha-subunit concentrations within 15 min for at least 4 hr. Follicle stimulating hormone levels were unchanged, while plasma luteinizing hormone concentrations only began to increase 1 hr after the oCRF treatment. Calculation of circulating thyrotropin (TSH) indicator values revealed a rapid elevation in TSH plasma levels following oCRF. Concentrations of thyroxine (T4), 3,5,3'-triiodothyronine (T3), 3,3',5'-triiodothyronine (rT3), and corticosterone increased from 1 hr postinjection. Hypothalamic outer ring deiodinating type II increased and hepatic inner ring deiodinating type III fell after 2 and 4 hr, explaining at least in part the plasma T3 increase at the end of the experiment. In a second experiment, using E18 chicks, a comparison was made between the effects of a single injection of 2 micrograms oCRF and 20 mlU bovine TSH. Both hormones increased T4, T3, and rT3 plasma concentrations, supporting the hypothesis of a TSH-releasing activity for oCRF in the embryonic chicken. The proposed TSH-mediated effect of CRF on thyroid function was further confirmed in two in vitro experiments in which oCRF did not directly influence the thyroidal T4 secretion but, when applied to pituitaries, clearly increased the alpha-subunit release. In chickens CRF is concluded to not only control the adrenal axis, but also to participate in the coordination of avian TSH release.

Plasma thyroid hormone levels and iodothyronine deiodinase activity following an acute glucocorticoid challenge in embryonic compared with posthatch chickens.

Embryonic chickens (Day 18 of incubation) and 8-day-old posthatch chicks were subjected to an acute glucocorticoid challenge by a single iv injection of corticosterone (B), dexamethasone, or porcine adrenocorticotropin. Plasma samples were analyzed for changes in T4, T3, rT3, alpha-subunit, LH, GH, and B levels; iodothyronine deiodinase activity was measured in liver, kidney, and hypothalamus at several time points after injection. The effects of the different treatments were broadly similar within one age group, but differed clearly between pre- and post-hatch animals. In 18-day-old embryos glucocorticoids increased plasma T3 and decreased plasma T4, rT3, and the calculated TSH index, within hours after injection. These changes were accompanied by an immediate (1-4-24 hr after injection) decrease in hepatic inner ring deiodinating type III enzyme (IRD-III) activity and a delayed (24-48 hr after injection) increase in hepatic outer ring deiodinating type I enzyme (ORD-1) activity. Glucocorticoid challenge in 8-day-old chicks similarly decreased plasma T4 and the TSH index but tended to also lower plasma T3. Hepatic ORD-I activity decreased within 1 hr after injection, while the already very low hepatic IRD-III activity was for the most part unaffected. Acute increases in glucocorticoids lower thyroidal T4 secretion in both pre- and posthatch chickens but have clearly different effects on peripheral thyroid hormone deiodination and hence on circulating T3 at both developmental stages.