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P Barrett

Publications and source records attributed to P Barrett.

At least 73 records · Page 4Linked to original sources
PubMed1997

Short-day weight loss and effect of food deprivation on hypothalamic NPY and CRF mRNA in Djungarian hamsters.

The effect of food deprivation on hypothalamic neuropeptide Y (NPY) and corticotropin-releasing factor (CRF) gene expression in the Djungarian hamster was quantified by in situ hybridization. Hamsters housed in short days (SD) for 18 wk decreased body weight by 40% and exhibited 200% increases in both NPY and CRF mRNA when deprived of food for 24 h. Prior gonadectomy in long days (LD) affected neither basal gene expression nor the induction of gene expression by food deprivation. Gene expression in juvenile LD hamsters similar in body weight to SD animals was relatively insensitive to food deprivation of either 24- or 48-h duration or to subsequent refeeding. In juvenile hamsters, food deprivation for 24 but not 48 h decreased ob (obese) gene expression in inguinal but not epididymal white adipose tissue; ob mRNA levels were restored by refeeding. All food-deprived hamsters had reduced plasma insulin concentrations, but plasma cortisol was only elevated in SD food-deprived animals. NPY gene expression was also increased after daily dexamethasone injections in adult LD hamsters. These results suggest that the neuroendocrine consequences of food deprivation in SD Djungarian hamsters are determined by some factor other than absolute body mass such as the size of adipose tissue reserves.

Animal Feed↗
PubMed1997

Novel isoforms of Mel1c melatonin receptors modulating intracellular cyclic guanosine 3',5'-monophosphate levels.

Two cDNAs encoding novel isoforms of Xenopus laevis melatonin receptors were cloned using PCR primers specific for the X. laevis-melanophore Mel1c melatonin receptor described in a recent publication. The novel isoforms were highly homologous to the described frog Mel1c cDNA, although the C-terminal tail of both was shorter by 65 amino acid residues. Nucleotide sequences of these novel isoforms, called Mel1c(alpha) and Mel1c(beta), differed from each other by only 35 nucleotides and six amino acid residues. Studies on several animals of various Xenopus species indicate that Mel1c(alpha) and Mel1c(beta) receptors may correspond to allelic variants of the same locus. Studies on cells transfected with both receptor cDNAs showed the expression of high-affinity 2-[125I]iodomelatonin binding sites. Agonist stimulation of Mel1c(alpha) receptor was associated with the inhibition of cAMP accumulation stimulated by forskolin (IC50 approximately 10(-10) M) in HeLa, Ltk-, and human embryonic kidney 293 (HEK 293) cells. Mel1c(beta) receptor modulated cAMP in HeLa and HEK 293 cells but not in Ltk- cells. Both receptors inhibited, in a dose-dependent manner, cGMP accumulation in all three cell lines incubated with a phosphodiesterase inhibitor. This effect was localized upstream of soluble guanylyl cyclase and was blocked by pertussis toxin treatment. However, IC50 values (approximately 10(-10) M for Mel1c(beta) and 10(-9) to 10(-7) M for Mel1c(alpha)) and maximal inhibition levels showed that Mel1c(alpha) receptors are much less efficiently coupled to the cGMP pathway. Coupling differences may be explained by the fact that five of the six amino acid substitutions between Mel1c(alpha) and Mel1c(beta) receptors are located within cytoplasmic regions potentially involved in signal transduction. The existence of coupling differences is in agreement with the observation that expression of both receptors is evolutionally conserved in native tissue. In conclusion, two novel, potentially allelic, isoforms of Xenopus Mel1c melatonin receptors display identical ligand-binding characteristics, but different potencies in modulating cAMP and cGMP levels through G(i)/G(o)-dependent pathways. Furthermore, to our knowledge, this study provides the first data on the modulation of intracellular cGMP levels by cloned melatonin receptors.

Adenylyl Cyclases↗
PubMed1997

Stoma site marking: a primer.

The purpose of pre-operative stoma site marking is to select an appropriate location in an area of the abdomen for surgical placement of a stoma. A poorly located stoma may result in pouching problems, increase potential for leakage, and place undue hardship and emotional trauma on the patient. The type of ostomy surgery must be known to select the appropriate abdominal quadrant to be marked. The ideal stoma site is located below the umbilicus, within the rectus muscle, away from scars, creases, bony prominences, umbilicus, and belt line, on the summit of the infraumbilical fat mound, and visible to the patient. It is vital for the clinician to be aware of any extenuating circumstances of the patient that may alter the site selection. Optimal stoma site placement will promote self-care and rehabilitation of the patient.

Anthropometry↗
PubMed1997

Pregnancy, delivery, and postpartum experiences of fifty-four women with ostomies.

OBJECTIVE: The pregnancy, labor, and delivery experiences of women with ostomies are described. DESIGN: Patient survey. SETTINGS AND SUBJECTS: Distribution of surveys to women with these experiences: patients of the Cleveland Clinic Foundation, referrals by networking of ET nurses, and distribution of surveys at a national conference of the United Ostomy Association and through local ostomy associations. INSTRUMENT: A 12-point questionnaire was adopted for each type of diversion: ileostomy, urostomy, continent ileostomy, colostomy, and pelvic reservoir. METHOD: The survey was distributed to attendees at a national United Ostomy Association conference and local United Ostomy Association meetings. Surveys were distributed to other EI nurses in the mideastern United States region and to qualified patients at Cleveland Clinic Foundation. MAIN OUTCOME MEASURE: The subjective experiences of pregnancy, labor and delivery of women with ostomies are described. RESULTS: Seventy-five questionnaires were distributed and 54 women responded. Eight women (15%) had fertility problems. Stoma-related problems during the second or third trimester were reported by 68.5%. Most problems were corrected without medical intervention. CONCLUSIONS: The presence of an ostomy should not be a deterrent to successful pregnancy and delivery.

Aged↗
PubMed1996

Melatonin suppresses the induction of AP-1 transcription factor components in the pars tuberalis of the pituitary.

In ovine pars tuberalis cells which express high affinity Mel 1a melatonin receptors, the ability of melatonin to directly stimulate or inhibit AP-1 transcription factor gene expression was studied. Effects of melatonin upon mRNA expression by forskolin, serum and IGF-1 were also investigated. Northern analysis showed melatonin had no direct stimulatory nor inhibitory effect upon transcription or translation. Melatonin was able to significantly inhibit forskolin-stimulated induction of c-fos and jun B mRNA whilst forskolin had no effect upon c-jun or jun D. Induction of c-Fos translation by forskolin was also inhibited by melatonin. Serum induced c-fos and c-jun, but melatonin was unable to affect these changes. Similarly IGF-1 stimulated c-fos and melatonin had no effect upon this induction. From these results it can be concluded that melatonin has no independent effects on expression of the AP-1 genes, rather its primary function is to inhibit cell activities through cyclic AMP-dependent routes of gene activation.

Animals↗
PubMed1996

Are nuclear receptors involved in pituitary responsiveness to melatonin?

Evidence suggests that the pineal hormone melatonin modulates prolactin (PRL) secretion in part through direct effects on the anterior pituitary. However, high-affinity membrane receptors for melatonin are only found in the pars tuberalis (PT) of the anterior pituitary, whereas lactotrophs are confined to the pars distalis (PD). This study therefore sought to determine whether melatonin might have direct effects on the PD, through alternate pathways. Such a possibility had been suggested by recent reports of melatonin binding to members of the retinoid-related orphan nuclear receptor family (ROR/RZR). Expression of ROR(alpha)/RZR(alpha) isoforms was observed by reverse transcription PCR in the ovine PT and PD. Correspondingly unidentified nuclear proteins from these tissues showed binding to consensus DNA response elements for members of the ROR/RZR family. In contrast nuclear 2-[125I]iodomelatonin (IMEL) binding was not detectable in PD or PT extracts even at high ligand and tissue concentrations. Nevertheless, the conditions used allowed membrane and nuclear IMEL binding to be observed in PT and liver extracts, respectively. Overall these findings do not support the possibility of direct effects of melatonin on the adult PD, and by implication they reinforce the view that the melatonin-responsive PT is an intermediary in the control of PRL secretion.

Animals↗
PubMed1996

Differential regulation of melatonin receptors in sheep, chicken and lizard brains by cholera and pertussis toxins and guanine nucleotides.

G-proteins define both the pharmacological characteristics and the signalling pathways of G-protein-coupled receptors. Melatonin receptors have been shown to belong to this class of receptors through their sensitivity to modulators of G-protein function. This study reveals that 2-125I-iodomelatonin (125I-MEL) binding to different target tissues is differentially affected by agents which disrupt the G-protein cycle. GTP gamma S, pertussis (PTX) and cholera (CTX) toxins each reduce 125I-MEL binding to ovine pars tuberalis (oPT) and lizard brain membranes, whereas chicken brain is affected only by GTP gamma S (guanosine 5'-O-(3-thiotriphosphate)) and CTX. In contrast, high affinity binding of 125I-MEL in the ovine hippocampus was not affected by any of these agents. This finding, together with the fact that neural binding sites of the sheep brain were found to have markedly lower molecular mass than those of the oPT on native gel electrophoresis (365 vs 525 kDa), suggests that the neural 125I-MEL binding sites in sheep may not be G-protein coupled. Pharmacologically, however, the binding sites in the hippocampus and oPT could not be distinguished using 11 analogues of melatonin. Therefore, these data support the notion not only of multiple forms of melatonin receptor/G-protein complex, but of high affinity binding sites for 125I-MEL which do not display sensitivity to guanine nucleotides.

Animals↗
PubMed1996

The ovine pars tuberalis secretes a factor(s) that regulates gene expression in both lactotropic and nonlactotropic pituitary cells.

The purpose of this study was to determine whether the cells of the ovine pars tuberalis (PT) secrete a factor(s) that can influence the activity of cells in the pars distalis (PD). By Northern blotting of total RNA isolated from PD cells that had been stimulated in the presence of cycloheximide (10 micrograms/ml), PT cell-conditioned medium was shown to induce a significant increase in the expression of the early response gene, c-fos, above both PD cell-conditioned and nonconditioned medium control levels (P < 0.05). Although forskolin (5 microM) induced a weak increase in c-fos expression in PD cells, the effect of PT medium conditioned in the presence of forskolin enhanced this expression more than additively (P < 0.05); furthermore, this effect was reversed by melatonin. These results are consistent with the release of a factor(s) from the PT, which for simplicity we have called tuberalin. This factor was released from PT cells in a time-dependent and cycloheximide-sensitive manner and was resistant to heating at 100 C for 10 min. Tuberalin activity could be size-fractionated using molecular size cut-off filters to produce activity in both the 1- to 10-kDa and more than 10-kDa size ranges. The activities in both of these fractions were sensitive to trypsin degradation and, therefore, appeared to be peptidergic. However, it was not clarified whether the biological activities were due to one or two components. Tuberalin also induced c-fos expression in other cell types, including GH3 and NIH3T3 cells. Dual labelling of PD cells by in situ hybridization using riboprobes for c-fos and PRL demonstrated that both the less than and more than 10-kDa fractions of tuberalin activated c-fos expression in some, but not all, lactotrophs in PD cell cultures, suggesting that a primary function of the PT is to regulate the activity of lactotrophs. This was supported further by enhanced secretion of PRL from PD cells in the presence of either PT-conditioned medium or PT cells in coculture. In addition, PT-conditioned medium was found to increase c-fos in a second cell type, which did not hybridize positively for PRL, indicating the existence of other endocrine interactions between the PT and PD.

3T3 Cells↗
PubMed1996

Regulation of the Mel 1a melatonin receptor mRNA and protein levels in the ovine pars tuberalis: evidence for a cyclic adenosine 3',5'-monophosphate-independent Mel 1a receptor coupling and an autoregulatory mechanism of expression.

The Mel 1a receptor gene has recently been cloned and sequenced, and its product has been found to represent a new class of the G protein-coupled receptor superfamily. A unique aspect of its endogenous hormonal ligand, melatonin, is the prolonged duration (16 h) required to initiate appropriate photoperiodic responses in the animal. This study provides the first data on the transcriptional and translational regulation of this receptor in primary cultures of its native tissue, the pars tuberalis of the ovine pituitary. Using RNAse protection assays we show Mel 1a mRNA expression is rapidly induced through changes in intracellular cAMP levels. Forskolin enhances Mel 1a mRNA expression, whereas melatonin reverses this effect. RNA stability is also a factor, as Mel 1a mRNA expression decreases rapidly in the presence of actinomycin D. Although the expression of receptor protein is sensitive to forskolin stimulation, the lag between induction of mRNA and protein suggests that other factors (e.g. RNA splicing) contribute to functional receptor expression. Furthermore we observed that both mRNA and receptor protein levels increase spontaneously during primary culture, and these rises could be blocked by melatonin. A spontaneous rise in Mel 1a mRNA can also occur in serum-depleted cultures. In the absence of corresponding changes in cAMP, these results suggest not only that the Mel 1a receptor can autoregulate its own expression by an undefined pathway, but also they provide the first evidence that melatonin can act via a cAMP-independent signal transduction pathway to repress transcription in this tissue.

Animals↗
PubMed1995

Assessment of arylamine N-acetyltransferase (NAT1) activity in mononuclear leukocytes of cystic fibrosis patients.

The clearance of sulphamethoxazole (SMX), a compound metabolised primarily by the N-acetyltransferase NAT1, is increased in cystic fibrosis (CF) patients. We assessed the activity and kinetic properties of NAT1 in lysates of peripheral blood mononuclear leukocytes (MNL) from CF (n = 17) and control (n = 22) subjects using SMX and p-aminobenzoic acid (PABA) as test substrates. The Km and Vmax values of both substrates in MNL from CF patients and control subjects were not significantly different. The acetylation of PABA (100 microM) by intact MNL from CF patients (n = 4) was not different from the observed in intact MNL from controls (n = 9) (25 +/- 3 pmol h-1 per 10(6) MNL vs 27 +/- 4 pmol h-1 per 10(6) MNL). These results suggest that there are not systemic changes in this enzyme in CF. The increased metabolic clearance of SMX may therefore be related to factors other than alterations in the level of activity of the N-acetyltransferase NAT1.

4-Aminobenzoic Acid↗
PubMed1995

Regional distribution of iodomelatonin binding sites within the suprachiasmatic nucleus of the Syrian hamster and the Siberian hamster.

The pineal hormone melatonin is a potent regulator of seasonal and circadian rhythms in vertebrates. In order to characterize potential target tissues of melatonin, the distribution of iodomelatonin (IMEL)-binding sites was examined within neurochemically and anatomically defined subdivisions of the suprachiasmatic nucleus (SCN), a structure necessary for seasonal and circadian rhythms in mammals. Studies were carried out in both the adult Syrian (Mesocricetus auratus) and Siberian (Phodopus sungorus) hamster. The retinoreceptive zone of the SCN was identified anatomically by immunocytochemical (ICC) visualization of cholera toxin B subunit tracer (ChTB-ir) following its intra-ocular injection. Photically-responsive SCN cells were identified by immunostaining for the protein product of the immediate-early gene c-fos (Fos-ir) following exposure of the animal to light. The non-photoresponsive zone of the SCN was identified using in situ hybridization (ISH) for arginine vasopressin (AVP) mRNA, whilst sites of IMEL-binding in the SCN were identified by in vitro film autoradiography using the specific ligand 2-[125I]-iodomelatonin. To compare directly the distribution of IMEL-binding sites and one of the functional zones of the nucleus, alternate serial coronal sections through the SCN were processed for autoradiography for IMEL and one of the following: ICC for ChTB-ir or Fos-ir, or ISH for AVP mRNA. Overall, the regional distribution of the various markers within the SCN was comparable in the two species. The retinorecipient (ChTB-ir) and photically-responsive (Fos-ir) zones of the SCN mapped together to the middle and caudal thirds of the nucleus, predominantly in its ventro-lateral division.(ABSTRACT TRUNCATED AT 250 WORDS)

Afferent Pathways↗
PubMed1995

Melatonin receptors couple through a cholera toxin-sensitive mechanism to inhibit cyclic AMP in the ovine pituitary.

The nature of melatonin receptor-G-protein coupling in ovine pars tuberalis (PT) cells of the pituitary was addressed using cholera (CTX) and pertussis (PTX) toxins. ADP-ribosylation of ovine PT membrane proteins using 32P-NAD in the presence of CTX radiolabelled several substrates including 44, 51, and 60 kD proteins. Each were clearly distinct from the 40 kD substrate radiolabelled in the presence of PTX. Acute incubation of PT membranes with either toxin reduced the number of high affinity binding sites for 125I-MEL, although the magnitude of the inhibition was much greater for CTX (56%) than for PTX (20%). A CTX-sensitive component also mediates the inhibition of forskolin-stimulated cyclic AMP accumulation as pre-treatment of PT cells with CTX (5 micrograms/ml) for 16 h blocked this response. Gs alpha is a major substrate for ADP-ribosylation by CTX, and 16 h pre-treatment of PT cells with CTX (5 micrograms/ml) caused a down-regulation of Gs alpha. Northern analysis showed only one major transcript of Gs alpha of about 2 kb, which would encompass all of the known splice variants of the Gs gene. Screening of a cDNA library from ovine PT for Gs-related genes and sequencing of clones, combined with RT-PCR of PT mRNA, revealed no novel products. On this basis it is concluded that the CTX substrate is unlikely to be a novel splice variant or related gene product of the Gs class of G-protein.(ABSTRACT TRUNCATED AT 250 WORDS)

Adenosine Diphosphate Ribose↗
PubMed1995

Hypothalamic NPY and prepro-NPY mRNA in Djungarian hamsters: effects of food deprivation and photoperiod.

Two catabolic states leading to loss of body weight were compared in the Djungarian hamster (Phodopus sungorus campbelli). Hypothalamic neuropeptide Y (NPY) and gene expression for NPY and corticotropin-releasing factor (CRF) were examined after withdrawal of food for 48 h or exposure to short photoperiod for 10 or 20 wk. Food deprivation was accompanied by increases in both NPY and prepro-NPY mRNA in the hypothalamic arcuate nucleus (ARC). Increases in gene expression were limited compared with published data from the rat and were inversely related to predeprivation body weight. Exposure to short photoperiod for 20 wk reduced body weight by 39%, but the activity of the NPY-ergic system was not affected; peptide concentration and gene expression were similar in short photoperiod hamsters and long photoperiod controls. The hypothalamic NPY-ergic system of the Djungarian hamster is sensitive to weight loss due to imposed manipulations of energy balance, but the catabolism observed in short photoperiod gives rise to a body weight that is appropriate to the season encoded by the photoperiod. CRF gene expression was not affected by food deprivation or short photoperiod.

Animals↗
PubMed1995

Detection of human growth hormone receptors on IM-9 cells and peripheral blood mononuclear cell subsets by flow cytometry: correlation with growth hormone-binding protein levels.

We have developed a method using flow cytometry to identify fluorescein-conjugated GH receptors (GHR) on IM-9 lymphocytes and circulating peripheral blood mononuclear cell subsets. Binding to IM-9 cells and peripheral blood mononuclear cells was concentration dependent and could be competitively blocked by the addition of unlabeled human GH, but not by the addition of rat or bovine GH or human insulin or PRL. Using two-color flow cytometric analysis, fluorescein-conjugated human GHR were readily detected on more than 90% of B lymphocytes and monocytes, but only variably on T lymphocytes. B Lymphocytes and monocytes had approximately 6000 GHR/cell. Using two-color flow cytometry, we identified GHR on circulating B lymphocytes in subjects with GH deficiency (n = 9), precocious puberty (n = 6), and Turner syndrome (n = 5) and in seven subjects with miscellaneous disorders, including familial short stature, bone dysplasia, Crohn disease, congenital adrenal hyperplasia, and acromegaly. The percentage of B lymphocytes expressing GHR in subjects with GH deficiency (mean +/- SD, 95 +/- 9%), precocious puberty (91 +/- 15%), and Turner syndrome (84 +/- 15%) was not different from that in normal volunteers (90 +/- 12%; n = 14). In 10 subjects, serum GH-binding protein levels were assayed simultaneously with B lymphocyte GHR. GH-binding protein was normal in all (mean, 1255 pmol/L; range, 773-1809). There was a good correlation between GHR expression on B lymphocytes and GH-binding protein levels (r = 0.75; P = 0.01). We postulate that GHR found on circulating B lymphocytes may contribute to the pool of receptors identified in serum as GH-binding proteins. Two-color flow cytometry appears to be an effective method for the detection of GHR on circulating peripheral blood mononuclear cell subsets. The evaluation of GHR on circulating B lymphocytes may prove to be a useful means of evaluating GH-GHR interactions in subjects with growth disorders.

Adult↗
PubMed1995

Endocrine cells of the human gastrointestinal tract have no proliferative capacity.

There is compelling evidence that the epithelial cell lineage of the gastrointestinal tract are derived from a common stem cell precursor, but the details of the subsequent cellular hierarchies remain uncertain. In this context, it is important to know the arrangement of cell proliferation that gives rise to the final cell populations. In rodents, a number of studies have been performed examining the possible proliferative capacity of endocrine cells, but a wide range of technical problems makes interpretation of these data difficult. Continuous labelling studies suggest there is potential for proliferation in endocrine cells but flash labelling studies have not been conclusive. In man there are no data on this issue. We have taken advantage of the ability to perform double immunostaining for operational markers of proliferation (Ki67 antigen) and endocrine cell phenotype (chromogranin expression). We demonstrate that there are no double-labelled cells in the normal stomach, small intestine or colon of fetal, neonatal or adult humans. Moreover, no double-labelled cells are found in pathological states associated with endocrine cell hyperplasia (gastritis, ulcerative colitis). These data indicate that the normal endocrine cells of the human gut have no proliferative capacity and that, in this cell lineage, population expansion precedes differentiation.

Adult↗
PubMed1994

Functional expression of 2-amino-4-phosphonobutyrate (APB) receptors in Xenopus laevis oocytes by injection of poly(A)+ RNA from quail brain.

The glutamate analogue 2-amino-4-phosphonobutyrate (APB) is known to activate a subtype of metabotropic glutamate receptor in the central nervous system, including the retina. In the present study, APB receptors were studied using the Xenopus oocyte expression system. No endogenous APB sensitivity was detected in control oocytes. In contrast, microinjection of mRNA, extracted from quail brain, into Xenopus oocytes resulted in the functional expression of APB receptors after 3-5 days incubation. Application of 50 microM-1 mM APB to injected oocytes voltage clamped at a holding potential of -60 mV produced a sustained outward current which was associated with a significant decrease in membrane conductance; the reversal potential was around -11 mV. The response to APB was dose-dependent and non-desensitizing. This is the first demonstration of the expression of a conductance-decreasing receptor mechanism in Xenopus oocytes.

Animals↗