摘要:

The proopiomelanocortin (POMC) gene is expressed predominantly in corticotrophs of the pituitary anterior lobe, melanotrophs of the intermediate lobe and neurons of the arcuate nucleus of the hypothalamus. The different ontogeny of POMC mRNA as well as the complicated hormonal regulation of POMC gene expression in the three different cell types suggests a concerted interaction between several cis-acting elements in the POMC gene and transcription factors located in each of the three cell types. To investigate cell-specific elements in the POMC gene we tested two different constructs in transgenic mice. The construct –4000rPOMCLacZ, carrying 4 kb of the rat POMC promoter fused to the Escherichia coli β-galactosidase gene, showed appropriate expression in melanotrophs in 50% of the mice analyzed. β-Galactosidase activity was less evident in corticotrophs under basal environmental conditions. In brain, 7 out of 15 independently derived transgenic founders had ectopic expression of the transgene in different areas; however, none of the animals analyzed expressed β-galactosidase in neurons of the arcuate nucleus. The construct HAL*, a ‘tagged’ 10.2-kb mouse genomic fragment, was more efficiently targeted to the pituitary. Using in situ hybridization, we detected uniform expression of HAL* in melanotrophs in 100% of the 6 pedigrees analyzed and transgenic mRNA levels paralleled those of the endogenous POMC mRNA. In corticotrophs, basal expression was low but after adrenalectomy HAL* mRNA levels were comparable to those of POMC. None of the 6 pedigrees had appropriate expression of HAL* in the brain; however, 2 lines had ectopic expression in the dentate gyms of the hippocampus. These data, together with previously reported studies, suggest that accurate neuronal expression of the POMC gene requires DNA elements in addition to the sequences that are sufficient for expression in pituitary melanotrophs and corticotrophs. The consistently lower level of transgene expression in corticotrophs compared to melanotrophs under basal conditions indicates that these two pituitary cell types may also differ in their requirements for POMC gene regulatory

ISSN:0028-3835    

DOI:10.1159/000126566

出版商:S. Karger AG    

年代:1993

数据来源: Karger

摘要:

Previous studies from this and other laboratories have shown that levels of vasopressin (VP) mRNA are reduced in both hypothalamic magnocellular and extrahypothalamic nuclei of the homozygous Brattleboro rat (HOM) when compared to the normal Long-Evans (LE) and heterozygous Brattleboro rats (HET). Since extrahypothalamic VP gene expression is dependent on testosterone (T), we measured plasma T in HOM, HET and LE rats. The plasma T level of the intact HOM rat was not significantly different from intact LE or HET rats. Manipulation of circulating gonadal steroids by castration and T replacement was found to regulate the expression of VP mRNA in the bed nucleus of the stria terminalis and medial amygdala of LE and HET rats, but does not appear to modify the absence of VP mRNA in neurons in these nuclei in the HOM rat.

ISSN:0028-3835    

DOI:10.1159/000126567

出版商:S. Karger AG    

年代:1993

数据来源: Karger

摘要:

The membrane-bound dopamine transporter (DAT) protein terminates dopaminergic neurotransmission by rapid reaccumulation of dopamine into pre-synaptic nerve endings. The distribution of neurons expressing DAT mRNA was investigated in the rat hypothalamus using in situ hybridization histochemistry and oligonucleotide probes to sequences of rat DAT mRNA. DAT mRNA-containing cell bodies were demonstrated in the ventral aspect of the periventricular nucleus (A14 dopamine cell group), in the zona incerta (A13), in the dorsomedial part of the arcuate nucleus (A12), and in scattered regions of the posterior hypothalamus. There was no labelling in magnocellular neurons of the supraoptic or paraventricular nuclei, or in neurons of the ventrolateral division of the arcuate nucleus, areas previously shown to contain the dopamine-synthesizing enzyme tyrosine hydroxylase. The results suggest that hypothalamic dopamine neurons have an active uptake mechanism for dopamine, however, that DAT mRNA levels may be considerably lower than those encountered in the ventral midbrain.

ISSN:0028-3835    

DOI:10.1159/000126568

出版商:S. Karger AG    

年代:1993

数据来源: Karger

摘要:

The hypothalamic-pituitary-adrenal (HPA) axis controls the diurnal and stress-induced release of adrenal corticosteroids into the general blood circulation. In turn, corticosteroids inhibit the HPA axis under basal conditions and during stress through occupation of their receptors (types I and II) in the brain by closing a negative feedback loop. The primary site in the brain at which corticosteroids act to inhibit the HPA axis has not been identified. High concentrations of both types of receptors are found in neurons of the hippocampal formation, a structure which has been reported by some, but not others, to control activity within the HPA axis by serving as a major negative feedback site. In many of these past studies, blood was collected after extensive handling or exposure to ether, conditions which do not favor the detection of basal hormone concentrations. To address these controversies, we tested the feedback sensitivity of the anterior pituitary hormone responsible for corticosteroid production, adrenocorticotropin (ACTH), to corticosterone (B), the main corticosteroid in rats, in total fornix- and, as controls, cortex-lesioned rats. All rats were given vascular catheters to avoid any handling-induced differences in plasma B or ACTH when sampling blood. In some experiments, fornix- and cortex-lesioned rats were adrenalectomized and given 1 of 3 doses of exogenous B provided in a subcutaneous pellet to ensure that plasma B was equal in different lesion groups. We hypothesized that if the hippocampal formation were an important site of B-mediated inhibition of the HPA axis, fornix-lesioned rats would have higher plasma B as a result of increased endogenous secretion in the morning or the evening compared to cortex-lesioned rats in rats with adrenal glands. In additionwe hypothesized that adrenalectomized fornix-lesioned rats given the same low to moderate levels of exogenous constant B would have higher basal and stress-induced ACTH than cortex-lesioned rats. Diurnal plasma B was not affected by fornix lesions in intact rats. Moreover, basal ACTH measured in the morning and the evening and stress-induced ACTH was the same in adrenalectomized fornix- and cortex-lesioned rats with constant exogenous B. We conclude, therefore, that information about occupancy of B receptors in the hippocampus carried by the fornix primarily subserves functions which do not directly regulate activity in the HPA axis.

ISSN:0028-3835    

DOI:10.1159/000126569

出版商:S. Karger AG    

年代:1993

数据来源: Karger

摘要:

Stereotaxic deletion of selected areas of the ventral noradrenergic ascending bundle (VNAB-X) by discrete bilateral injections of 6-hydroxydopamine (6-OHDA; 4 µg in 0.2 µl saline) was used to explore the role of brain catechol-amines (CA) and their interaction with corticosteroid feedback in stress responses of the ACTH-corticosterone (CORT) axis. The stereotaxic coordinates used for 6-OHDA lesions and the optimization of postlesion delays were determined by (a) radioautographic labeling of the VNAB axons after tracer injections into the dorsal A2/C2, or the ventral Al/Cl medullary areas, (b) histofiuorescence and immunocytochemical location of interrupted CA pathways versus the postlesional scar, and (c) postlesional noradrenaline and adrenaline concentrations in whole hypothalami and paraventricular nuclei (PVN) punch samples. Two sites of 6-OHDA lesions were selected; both led to striking falls in PVN concentrations of both CA. The more dorsal lesion (dVNAB-X) was the same as that of our earlier studies and interrupted pathways originating predominantly in the A2/C2 area; the second more ventral lesion (vVNAB-X) interrupted axons stemming preferentially from the A1/C1 area. Both VNAB lesions inhibited the ether stress-induced ACTH and CORT surges in rats with intact adrenals. But the blockade (overall poststress release, amplitude and swiftness of hormonal responses) by dVNAB-X was greater than by vVNAB-X. The basal ACTH level in adrenalectomized rats (ADX) was elevated 20-fold and ether stress induced a 4-fold ACTH surge. As in sham-ADX rats, vVNAB-X in ADX rats induced only moderate inhibition of the ACTH response versus ADX + sham-vVNAB-X controls. On the other hand ADX + dVNAB-X rats showed a greatly amplified ACTH stress response over the ADX-sham dVNAB-X controls. This amplification was reversed by oral CORT supplementation. The data suggest that the CA pathways of the VNAB participating, directly or indirectly, in the poststress corticotropic activation may include subsets of CA axons of different origins, whose functional roles in stress are modulated in opposite directions by the plasma corticosteroid leve

ISSN:0028-3835    

DOI:10.1159/000126570

出版商:S. Karger AG    

年代:1993

数据来源: Karger

摘要:

Adrenalectomized rats replaced with constant, physiological levels of corticosterone via a subcutaneous pellet (Pellet) have normal basal morning ACTH but exhibit enhanced and prolonged ACTH responses to stress vs. sham-operated (Sham) rats. It has not been determined if the lack of either stress-induced or circadian increases in corticosterone, both of which are missing in Pellet rats, may account for this enhanced response. To test the extent to which stress-associated increases in corticosterone alone can normalize stress-induced hypersecretion of ACTH, we approximated endogenous secretion by injecting additional corticosterone in Pellet rats via an indwelling subcutaneous cannula, 5 min before hypoxia stress (10% O2). A corticosterone dose of 666 µg/kg (Pellet+B), but not 333 µg/kg (Pellet+Low B), produced plasma corticosterone levels comparable to those in Shams and normalized stress-induced but not post-stress plasma ACTH. Administration of the type II corticosteroid receptor antagonist RU 38486 30 min before corticosterone reversed this inhibition. We conclude that enhanced ACTH responses to stress in Pellet rats result in large part from lack of type II receptor-mediated feedback inhibition by corticosterone increases during stress, although prior circadian increases in corticosterone may also be require

ISSN:0028-3835    

DOI:10.1159/000126571

出版商:S. Karger AG    

年代:1993

数据来源: Karger

摘要:

Lipocortin 1 (LC1), a mediator of anti-inflammatory steroid action in some peripheral tissues, may contribute to the acute inhibitory effects of these steroids on hypothalamo-pituitary-adrenal (HPA) function. Accordingly, in the present study we have used an in vitro model to examine the potential role of this protein in the regulation of the release of corticotrophin (ACTH) from the anterior pituitary gland. Hypothalamic extracts (0.05-0.4 HE/ml), the 41 amino acid corticotrophin-releasing factor (CRF-41, 1-100 nM), the adenyl cyclase stimulator, forskolin (0.1 µM–10 mM), and the L-Ca2+ channel opener, BAY K8644 (0.01-10 mM), all caused concentration-dependent increases in the release in vitro of immunoreactive (ir)-ACTH from segments of rat anterior pituitary tissue. The secretory responses to submaximal concentrations of these secretagogues were overcome by preincubation of the tissue with dexamethasone (0.1 and 1 µM). LC1 was readily detectable by Western blotting in protein extracts of freshly excised pituitary tissue; a small proportion of the protein was found to be attached to the outer surface of the cell membranes where it was retained by a Ca2+-dependent mechanism. Exposure to dexamethasone (0.1 µM) in vitro did not affect the total LC1 content of the pituitary tissue but, over a 2-hour period, it caused a progressive two- to fivefold increase in the amount of LC1 attached to the outer surface of the cell; this response developed in parallel with the inhibitory effects of the steroid on ir-ACTH release. Both the dexamethasone-induced ‘externalization’ of LC1 by the pituitary tissue and the concomitant steroid-induced inhibition of peptide release were blocked by cycloheximide (1.0 µg/ml) but not by actinomycin D (0.5 µg/ml). A stable N-terminal lipocortin 1 fragment, LC11–188 (10 pg-10 ng/ml), attenuated (p < 0.01) the release of ir-ACTH evoked by HE (0.1 HE/ml), CRF-41 (1 nM), forskolin (1 mM) and BAY K8644 (1 nM). Conversely, inclusion of the anti-LC1 antibody in the medium substantially overcame the inhibitory effects of dexamethasone (0.1 µM) on the release of ir-ACTH evoked by the secretagogues whilst a control isotype matched antibody was without effect. The results suggest that LC1 plays a key role in effecting the acute inhibitory actions of glucocorticoids on the secretion of ir-ACTH by the rat anterior pi

ISSN:0028-3835    

DOI:10.1159/000126572

出版商:S. Karger AG    

年代:1993

数据来源: Karger

摘要:

The mechanism by which adrenal medulla transplants to the brain of Parkinson’s patients ameliorate the symptoms of this neurodegenerative disease is unknown. It has been postulated but not proven that these transplants secrete a neurotrophic factor(s) responsible for the activation of the dopamine (DA) neurons. DA-releasing protein (DARP) was identified by immunocytochemistry in cells of the adrenal gland but not in tissues such as the liver or spleen. In the adrenal gland the immunopositive DARP cells were distributed both in the cortex as well as in the medulla. In the former, the distribution was rather uniform covering a broad area of the zona glomerulosa and fasciculata without positive cells in the zona reticularis. In the adrenal medulla the DARP signal was found in segregated cells forming clusters similar to the homotypic distribution of norepinephrine and epinephrine cells. The immunopositive cells for DARP were oval or spindle in shape and the staining was localized in the cytoplasm surrounding the nucleus. In addition, in this paper we present evidence confirming that the adrenal gland is the source of a protein that after immunopurification in nanogram amounts releases DA from corpus striatum fragments superfused in vitr

ISSN:0028-3835    

DOI:10.1159/000126573

出版商:S. Karger AG    

年代:1993

数据来源: Karger

ISSN:0028-3835    

DOI:10.1159/000126574

出版商:S. Karger AG    

年代:1993

数据来源: Karger

摘要:

We recently demonstrated that the ability of administered ovine prolactin (oPRL) to suppress postcastration LH secretion exhibited a clear dose dependency. In the present study, we determined whether this dose-related suppression of mean LH levels resulted from differential, dose-related effects of oPRL on LH pulse amplitude and pulse frequency. Adult male rats were orchiectomized and adrenalectomized, implanted with an atrial cannula and a 50% corticosterone pellet, and injected every 12 h with oPRL or its polyvinylpyrrolidone (PVP) vehicle beginning at time 0. Increasing doses of oPRL (600, 2,400 and 9,600 µg/injection) suppressed mean LH titers in a dose-dependent manner at 48 h postcastration. The mean maximal LH increments (Δ LH) to two LHRH challenges at two doses (5 and 25 ng LHRH/100 g body weight) were unaffected by oPRL administration. The 600 µg oPRL dose significantly suppressed mean LH values by markedly increasing the inter-peak interval (42.6 ± 6.7 min) compared with controls (26.6 ± 0.2 min) since the pulse amplitude was unaffected (2.8 ± 0.4 vs. 2.6 ± 0.4 ng/ml, respectively). The two higher oPRL doses suppressed both LH pulse frequency and pulse amplitude. Hence, elevated PRL levels first suppress LH pulse frequency and then, at higher concentrations, pulse amplitude as well. Presuming that LHRH pulses result from ensemble firing of all or a significant proportion of the LHRH neurons projecting to the median eminence, the present data suggest that the neurons first affected by elevated PRL levels are the ones responsible for the frequency of this coordinated firing. Possible candidates for this neuronal mechanism would be the LHRH-LHRH collateral connections and/or other hypothalamic intern

ISSN:0028-3835    

DOI:10.1159/000126575

出版商:S. Karger AG    

年代:1993

数据来源: Karger