Corticotropin-releasing hormone and proopiomelanocortin gene expression is altered selectively in the male rat fetal thymus by maternal alcohol consumption.

The present study was carried out to investigate how hormonal changes caused by chronic alcohol exposure of rats during the late period of gestation are coordinated with neuroendocrine functions of the fetal thymus, namely thymic expression of CRH and POMC genes. Alcohol consumption by pregnant dams led to a 5-fold elevation of plasma corticosterone (CORT) levels and significantly decreased fetal CORT levels. This generally inverse correlation between maternal and fetal CORT was absent in alcohol-consuming dams and their male fetuses on day 19 of gestation. These day 19 fetuses also had an attenuated plasma testosterone surge that occurred in the male control (pair-fed) fetus on day 19 of embryonic life. Furthermore, fetal alcohol exposure (FAE) resulted in a significant increase in thymic CRH and a decrease in thymic POMC expression in the male fetuses only, specifically on embryonic day 19. Thus, the strong positive correlation between CRH and POMC gene expression in the thymus of pair-fed male and female FAE fetuses was abolished in the FAE males. However, regardless of embryonic age or treatment, a strong positive correlation between thymic POMC gene expression and plasma testosterone levels in the male fetuses was detected. These data suggest that the sexually dimorphic effect of FAE on the fetal thymic POMC and CRH expression in males is driven by testosterone and may be related, therefore, to the presence of alcohol at the time of the prenatal testosterone surge in the male fetuses.

In vivo growth hormone gene expression in neonatal rat thymus and bone marrow.

The ontogenesis of GH gene expression in the pituitary gland and immune organs of rats was studied by in vitro amplification of GH mRNA using a specific reverse transcription-polymerase chain reaction. Samples were obtained after microsurgery and total RNA extraction of fetal, neonatal and adult rat tissues. Amplification was followed by analysis using Southern blot techniques and hybridization using a specific digoxigenin-labelled GH cDNA probe. The study was started on day 15 of gestation. GH gene expression occurred in the pituitary gland on day 17. GH mRNA was found in the bone marrow and thymus of neonatal rats but not in the spleen or liver. No GH transcripts were detected in the immune organs of fetal or adult rats. The transient GH expression in rat immune organs may indicate a specific function of GH in the development of the immune system during the neonatal period.

Prepro-thyrotropin-releasing hormone 178-199 increases sensitivity of AtT-20 cells to dexamethasone.

The prepro-thyrotropin-releasing hormone (ppTRH)-derived peptide, ppTRH178-199, has been proposed to inhibit ACTH release at the level of the pituitary and attenuate prolactin and behavioral responses to stress as well. The objective of this study was to elucidate a possible link between the effects of ppTRH178-199 and glucocorticoids on the inhibition of ACTH release in corticotrophs. Compared with mock-transfected cells, AtT-20 cells that were stably transfected with full-length ppTRH cDNA showed significantly increased sensitivity to dexamethasone, as measured by inhibition of ACTH release. In a group of control cells, expressing a mutated form of ppTRH cDNA lacking the ppTRH178-199 region, sensitivity to dexamethasone was not different from mock-transfected controls. Exogenous ppTRH178-199 also increased the inhibitory effect of dexamethasone in wild-type AtT-20 cells. The combined effect of dexamethasone and ppTRH cDNA in cells that express the latter was not due to increased endogenous secretion of ppTRH178-199 in response to dexamethasone, as dexamethasone was independently found to inhibit secretion of ppTRH178-199. Taken together, these data suggest that ppTRH178-199 can interact with the glucocorticoid negative feedback inhibition to regulate ACTH secretion.

Decreased in vitro sensitivity to dexamethasone in corticotropes from middle-age rats.

A disregulation of the hypothalamic-pituitary-adrenal (HPA) axis due to a decline of negative feedback regulation is a consistent feature of the aging process. Hippocampus has been proposed to be a primary site responsible for this alteration in the HPA axis in aging in rat. In this study an alternative hypothesis that the decreased sensitivity of the HPA axis to glucocorticoids in aging occurs directly in pituitary corticotropes has been tested. The sensitivity of corticotropes isolated from 2- and 13-month old male Sprague-Dawley rats to dexamethasone (DEX) in vitro was examined using a modification of the combined DEX/CRH challenge test that was originally designed for investigation of relative glucocorticoid resistance in vivo. No significant difference in basal ACTH production by corticotropes from the two age groups was detected. Corticotropes from middle-aged rats showed a diminished response of ACTH to CRH stimulation. DEX treatment did not cause a significant inhibition of either basal or CRH-stimulated ACTH release in corticotropes from middle-aged rats. These findings demonstrate an age-related decrease in the sensitivity of corticotropes to glucocorticoids in vitro suggesting that there is a direct, pituitary-mediated dysregulation of the HPA axis in rat starting as early as middle age.

A novel endogenous corticotropin release inhibiting factor.

ACTH is the major regulator of the body's adaptive response to stress and the physiological stimulus for glucocorticoid secretion. A hypothalamic corticotropin release inhibiting factor (CRIF) that inhibits ACTH synthesis and secretion has long been postulated but was not characterized until recently. We have recently identified a 22 amino acid peptide, prepro-thyrotropin releasing hormone (TRH) 178-199 that inhibits basal and stimulated ACTH synthesis and secretion in vitro and stress-induced ACTH secretion in vivo. Prepro-TRH 178-199 is abundant in several brain regions, including the external zone of the median eminence, where its concentration changes in response to stress. We propose that this peptide is a physiological regulator of ACTH production: an endogenous CRIF. Because prepro-TRH 178-199 is encoded within the same precursor as TRH, its expression is likely to be negatively regulated by thyroid hormones leading to changes in endogenous glucocorticoid levels. Streptococcal cell wall (SCW)-induced inflammation, a model of rheumatoid arthritis (RA), was alleviated after long-term thyroxine treatment. Inversely, a hypothyroid milieu led to decreased basal hypothalamic-pituitary-adrenal activity, but increased expression of IL-1 beta and MIP-1 alpha, specific markers for RA in humans. These results suggest that this putative CRIF may be an important component in the development of RA and that regulation of prepro TRH may be highly relevant to the development of other autoimmune diseases that are also exacerbated by low endogenous glucocorticoid levels.

Corticotrophs and peptides.

Corticotrophs were long thought to be a static, homogeneous population of cells that respond positively to hypothalamic stimulation, are inhibited by glucocorticoid feedback and secrete a single biologically active peptide, ACTH(1-39). Our current understanding is that this is an oversimplification and corticotrophs are a dynamic and more complex group of cells. The biosynthetic precursors of ACTH and other cleavage products of proopiomelanocortin (POMC) have been found to be secreted by anterior pituitary cells, to circulate and to have biological activity. POMC and the biosynthetic intermediate, pro-ACTH, exert activity antagonistic to ACTH(1-39) on glucocorticoid secretion by adrenal cells, and other derivatives of POMC are mitogenic to adrenocortical cells. In terms of responses to hypothalamic and peripheral factors, corticotrophs are functionally heterogeneous. This is reflected in the sensitivity of individual subtypes of corticotrophs to CRH, vasopressin and glucocorticoids. There is a functional plasticity amongst the various types of corticotrophs. During gestation, in fetal sheep, changes occur in the overall ACTH-secretory responses to CRH relative to vasopressin, the proportions of total corticotrophs that respond to the respective peptides and the average secretory response of individual cells. Corticotrophs also respond to locally produced pituitary factors. Local actions of leukaemia inhibitory factor are demonstrated by the effects of immunoneutralization of the peptide in pituitary cells. Urocortin and preproTRH(178-199) are locally produced peptides with potent stimulatory and inhibitory actions on corticotrophs, respectively. The specific roles of these peptides are under investigation.