Biomedical subjects

J Rivier

Publications and source records attributed to J Rivier.

At least 19 recordsLinked to original sources

PubMed2006

Enhanced pelvic responses to stressors in female CRF-overexpressing mice.

Acute stress affects gut functions through the activation of corticotropin-releasing factor (CRF) receptors. The impact of acute stress on pelvic viscera in the context of chronic stress is not well characterized. We investigated the colonic, urinary, and locomotor responses monitored as fecal pellet output (FPO), urine voiding, and ambulatory activity, respectively, in female and male CRF-overexpressing (CRF-OE) mice, a chronic stress model, and their wild-type littermates (WTL). Female CRF-OE mice, compared with WTL, had enhanced FPO to 2-min handling (150%) and 60-min novel environment (155%) but displayed a similar response to a 60-min partial restraint stress. Female CRF-OE mice, compared with WTL, also had a significantly increased number of urine spots (7.3 +/- 1.4 vs. 1.3 +/- 0.8 spots/h) and lower locomotor activity (246.8 +/- 47.8 vs. 388.2 +/- 31.9 entries/h) to a novel environment. Male CRF-OE mice and WTL both responded to a novel environment but failed to show differences between them in colonic and locomotor responses. Male WTL, compared with female WTL, had higher FPO (113%). In female CRF-OE mice, the CRF(1)/CRF(2) receptor antagonist astressin B and the selective CRF(2) receptor agonist mouse urocortin 2 (injected peripherally) prevented the enhanced defecation without affecting urine or locomotor responses to novel environment. RT-PCR showed that CRF(1) and CRF(2) receptors are expressed in the mouse colonic tissues. The data show that chronic stress, due to continuous central CRF overdrive, renders female CRF-OE mice to have enhanced pelvic and altered behavioral responses to superimposed mild stressors and that CRF(1)-initiated colonic response is counteracted by selective activation of CRF(2) receptor.

Animals↗

PubMed2006

Lack of interaction between peripheral injection of CCK and obestatin in the regulation of gastric satiety signaling in rodents.

Obestatin is a new peptide for which anorexigenic effects were recently reported in mice. We investigate whether peripheral injection of obestatin or co-injection with cholecystokinin (CCK) can modulate food intake, gastric motor function (intragastric pressure and emptying) and gastric vagal afferent activity in rodents. Obestatin (30, 100 and 300 microg/kg, i.p.) did not influence cumulative food intake for the 2h post-injection in rats or mice nor gastric emptying in rats. In rats, obestatin (300 microg/kg) did not modify CCK (1 microg/kg, i.p.)-induced significant decrease in food intake (36.6%) and gastric emptying (31.0%). Furthermore, while rats injected with CCK (0.3 microg/kg, i.v.) displayed gastric relaxation, no change in gastric intraluminal pressure was elicited by obestatin (300 microg/kg, i.v.) pre- or post-CCK administration. In in vitro rat gastric vagal afferent preparations, 20 units that had non-significant changes in basal activity after obestatin at 30 microg responded to CCK at 10 ng by a 182% increase. These data show that obestatin neither influences cumulative food intake, gastric motility or vagal afferent activity nor CCK-induced satiety signaling.

Animals↗

PubMed2005

CRF2 receptor activation prevents colorectal distension induced visceral pain and spinal ERK1/2 phosphorylation in rats.

BACKGROUND AND AIMS: Activation of corticotropin releasing factor 1 (CRF1) receptors is involved in stress related responses and visceral pain, while activation of CRF2 receptors dampens the endocrine and some behavioural stress responses. We hypothesised that CRF2 receptor activation may influence visceral pain induced by colorectal distension (CRD) in conscious rats, and assessed the possible sites and mechanisms of action. METHODS: Male Sprague-Dawley rats were exposed to CRDs (60 mm Hg, 10 minutes twice, with a 10 minute rest interval). Visceromotor responses (VMR) were measured by electromyography or visual observation. Spinal (L6-S1) extracellular signal regulated kinase 1/2 (ERK 1/2) activation following in vivo CRD and CRF2 receptor gene expression in the T13-S1 dorsal root ganglia (DRG) and spinal cord were determined. Inferior splanchnic afferent (ISA) activity to CRD (0.4 ml, 20 seconds) was assessed by electrophysiological recording in an in vitro ISA nerve-inferior mesenteric artery (intra-arterial)-colorectal preparation. RESULTS: In controls, VMR to the second CRD was mean 31 (SEM 4)% higher than that of the first (p<0.05). The selective CRF2 agonist, human urocortin 2 (hUcn 2, at 10 and 20 microg/kg), injected intravenous after the first distension, prevented sensitisation and reduced the second response by 8 (1)% and 30 (5)% (p<0.05) compared with the first response, respectively. RT-PCR detected CRF2 receptor gene expression in the DRG and spinal cord. CRD (60 mm Hg for 10 minutes) induced phosphorylation of ERK 1/2 in neurones of lumbosacral laminae I and IIo and the response was dampened by intravenous hUcn 2. CRD, in vitro, induced robust ISA spike activity that was dose dependently blunted by hUcn 2 (1-3 microg, intra-arterially). The CRF2 receptor antagonist, astressin2-B (200 microg/kg subcutaneously or 20 microg intra-arterially) blocked the hUcn 2 inhibitory effects in vivo and in vitro. CONCLUSIONS: Peripheral injection of hUcn 2 blunts CRD induced visceral pain, colonic afferent, and spinal L6-S1 ERK 1/2 activity through CRF2 receptor activation in rats.

Animals↗

PubMed2004

Cardiovascular effects of long-term central and peripheral administration of urocortin, corticotropin-releasing factor, and adrenocorticotropin in sheep.

The neuroendocrine hormones ACTH and corticotropin- releasing factor (CRF), which are involved in the stress response, have acute effects on arterial pressure. New evidence indicates that urocortin (UCN), the putative agonist for the CRF type 2 receptor, has selective cardiovascular actions. The responses to long-term infusions of these hormones, both peripherally and centrally, in conscious animals have not been studied. Knowledge of the long-term effects is important because they may differ considerably from their acute actions, and stress is frequently a chronic stimulus. The present experiments investigated the cardiovascular effects of CRF, UCN, and ACTH in conscious sheep. Infusions were made either into the lateral cerebral ventricles (i.c.v.) or i.v. over 4 d at 5 microg/h. UCN infused i.c.v. or i.v. caused a prolonged increase in heart rate (HR) (P < 0.01) and a small increase in mean arterial pressure (MAP) (P < 0.05). CRF infused i.c.v. or i.v. progressively increased MAP (P < 0.05) but had no effect on HR. Central administration of ACTH had no effect, whereas systemic infusion increased MAP and HR (P < 0.001). In conclusion, long-term administration of these three peptides associated with the stress response had prolonged, selective cardiovascular actions. The striking finding was the large and sustained increase in HR with i.c.v. and i.v. infusions of UCN. These responses are probably mediated by CRF type 2 receptors because they were not reproduced by infusions of CRF.

Adrenocorticotropic Hormone↗

PubMed2003

The effect of urocortin on ingestive behaviours and brain Fos immunoreactivity in mice.

The influence of urocortin (UCN) on ingestive behaviours and brain neural activity, as measured immunohistochemically by the presence of Fos protein, was determined in mice. Rat UCN was administered by continuous intracerebroventricular (ICV) or subcutaneous (SC) infusion. ICV infusion of UCN (100 ng/h, 14 days) transiently reduced daily food and water intakes (days 1-4) but body weight was reduced from day 2 into the post-infusion period. Sodium intake was reduced from day 3 to the end of infusion. SC infusion of UCN caused similar but smaller reductions in food and water intakes and body weight, without change in sodium intake. In separate experiments, Fos immunoreactivity was increased in several brain nuclei known to be involved in the control of body fluid and energy homeostasis, e.g. central nucleus of the amygdala, median preoptic nucleus, bed nucleus of the stria terminalis and arcuate nucleus. Increased Fos expression was similar for ICV and SC infusions when measured on days 2-3 or 6-7 of infusion. In conclusion, increases of brain activity by UCN may be associated with stimulation of adrenocorticotrophic hormone release and sympathetic nervous activity, but increases may also indicate suppression of ingestive behaviours by stimulating central inhibitory mechanisms located in areas known to control body fluid and energy homeostasis.

Animals↗

PubMed2002

Potent and long-acting corticotropin releasing factor (CRF) receptor 2 selective peptide competitive antagonists.

We present evidence that members of the corticotropin releasing factor (CRF) family assume distinct structures when interacting with the CRF(1) and CRF(2) receptors. Predictive methods, physicochemical measurements, and structure-activity relationship studies have suggested that CRF, its family members, and competitive antagonists such as astressin [cyclo(30-33)[DPhe(12),Nle(21),Glu(30),Lys(33),Nle(38)]hCRF((12-41))] assume an alpha-helical conformation when interacting with their receptors. We had shown that alpha-helical CRF((9-41)) and sauvagine showed some selectivity for CRF receptors other than that responsible for ACTH secretion(1) and later for CRF2.(2) More recently, we suggested the possibility of a helix-turn-helix motif around a turn encompassing residues 30-33(3) that would confer high affinity for both CRF(1) and CRF(2)(2,4) in agonists and antagonists of all members of the CRF family.(3) On the other hand, the substitutions that conferred ca. 100-fold CRF(2) selectivity to the antagonist antisauvagine-30 [[DPhe(11),His(12)]sauvagine((11-40))] did not confer such property to the corresponding N-terminally extended agonists. We find here that a Glu(32)-Lys(35) side chain to side chain covalent lactam constraint in hCRF and the corresponding Glu(31)-Lys(34) side chain to side chain covalent lactam constraint in sauvagine yield potent ligands that are selective for CRF(2). Additionally, we introduced deletions and substitutions known to increase duration of action to yield antagonists such as cyclo(31-34)[DPhe(11),His(12),C(alpha)MeLeu(13,39),Nle(17),Glu(31),Lys(34)]Ac-sauvagine((8-40)) (astressin(2)-B) with CRF(2) selectivities greater than 100-fold. CRF receptor autoradiography was performed in rat tissue known to express CRF(2) and CRF(1) in order to confirm that astressin(2)-B could indeed bind to established CRF(2) but not CRF(1) receptor-expressing tissues. Extended duration of action of astressin(2)-B vs that of antisauvagine-30 is demonstrated in the CRF(2)-mediated animal model whereby the inhibition of gastric emptying of a solid meal in mice by urocortin administered intraperitoneally at time zero is antagonized by the administration of astressin(2)-B but not by antisauvagine-30 at times -3 and -6 h while both peptides are effective when given 10 min before urocortin.

Animals↗

PubMed2002

Intracisternal urocortin inhibits vagally stimulated gastric motility in rats: role of CRF(2).

1. Corticotropin-releasing factor (CRF) acts in the brain to inhibit thyrotropin-releasing hormone (TRH) analogue, RX-77368-induced vagal stimulation of gastric motility. We investigated CRF receptor-mediated actions of rat urocortin (rUcn) injected intracisternally (ic) on gastric motor function. 2. Urethane-anaesthetized rats with strain gauges on the gastric corpus were injected i.c. with rUcn and 20 min later, with i.c. RX-77368. CRF antagonists were injected i.c. 10 min before rUcn. 3. RX-77368 (1.5, 3, 10, 30 and 100 ng, i.c.) dose-dependently increased corpus contractions, expressed as total area under the curve (AUC, mV min(-1)) to 2.6+/-2.5, 6.1+/-5.9, 9.8+/-2.6, 69.7+/-21.7 and 74.9+/-28.7 respectively vs 0.2+/-0.1 after i.c. saline. Ucn (1, 3 or 10 microg) inhibited RX-77368 (30 ng)-induced increase in total AUC by 28, 62 and 93% respectively vs i.c. saline+RX-77368. 4. The CRF(1)/CRF(2) antagonist, astressin-B (60 microg, i.c.) completely blocked i.c. rUcn (3 microg, i.c.)-induced inhibition of gastric motility stimulated by RX-77368 (30 ng). 5. The selective CRF(2) antagonist, astressin(2)-B (30, 60 or 100 microg, i.c. ) dose-dependently prevented i.c. rUCn action while the CRF(1) antagonist, NBI-27914 did not. 6. In conscious rats, rUcn (0.6 or 1 microg, i.c.) inhibited gastric emptying of an ingested chow meal by 61 and 92% respectively. rUcn action was antagonized by astressin(2)-B. 7. These data show that i.c. rUcn acts through CRF(2) receptors to inhibit central vagal gastric contractile response and postoprandial emptying.

Animals↗

PubMed2001

Time-dependent induction of anxiogenic-like effects after central infusion of urocortin or corticotropin-releasing factor in the rat.

RATIONALE: Corticotropin-releasing factor (CRF) and urocortin (Ucn) belong to the CRF-related family, share a high degree of structural homology and bind to CRF receptors. However, compared with CRF, Ucn was shown to display either weaker or similar anxiogenic-like effects in vivo. OBJECTIVE: To compare the anxiogenic-like responses of rats injected intracerebroventricularly (ICV) with different doses of either rat/human CRF (r/hCRF) or rat Ucn (rUcn) at different intervals after injection. METHODS: Rats were tested on three validated paradigms of emotional behavior [i.e. elevated plus-maze (EPM), defensive withdrawal (DW) and conflict test (CT)] 5 and 30 min after treatment. RESULTS: In the EPM test only r/hCRF, but not rUcn, produced anxiogenic-like effects at the dose of 1.0 microg, when the peptides were injected 5 min before testing. At 30 min after injection, both peptides caused a significant reduction of open arms exploration, rUcn being effective at 0.01 microg. In the DW test both peptides were equally potent in decreasing the exploratory behavior and increasing the time spent in the chamber at the dose of 1.0 microg when tested 30 min after injection. In the CT both rUcn (0.25-1.0 microg) and r/hCRF (0.75-1.0 microg) decreased significantly the responding in the punished component. However, rUcn reduced food responding also in the unpunished component possibly due to its powerful anorectic activity. CONCLUSIONS: Comparison of anxiogenic-like activities of r/hCRF and rUcn at doses up to 1.0 microg revealed striking differential effects that depended on the time of testing after ICV peptide injection, and on the paradigm of anxiety used. These results suggest that the onset of r/hCRF and rUcn actions related to behavioral responses to anxiety is likely to depend on brain peptide-specific mechanisms including binding properties to CRF-receptors, differential distribution to specific functional brain sites and the distribution and effectiveness of binding-protein interactions.

Animals↗

PubMed2001

Hypothalamic paraventricular nucleus injections of urocortin alter food intake and respiratory quotient.

Corticotropin releasing hormone (CRH) acts on the central nervous system to alter energy balance and influence both food intake and sympathetically-mediated thermogenesis. CRH is also reported to inhibit food intake in several models of hyperphagia including neuropeptide Y (NPY)-induced eating. The recently identified CRH-related peptide, urocortin (UCN), also binds with high affinity to CRH receptor subtypes and decreases food intake in food-deprived and non-deprived rats. The present experiment characterized further the feeding and metabolic effects of UCN by examining its impact after direct injections into the paraventricular nucleus (PVN) of the hypothalamus. In feeding tests (n=8), UCN (50-200 pmol) was injected into the PVN at the onset of the dark cycle and food intake was measured 1, 2 and 4 h postinjection. In separate rats (n=8), the metabolic effects of UCN were monitored using an open circuit calorimeter which measured oxygen consumption (V(O2)) and carbon dioxide production (V(CO2)). Respiratory quotient (RQ) was calculated as V(CO2)/V(O2). UCN suppressed feeding at all times studied and reliably decreased RQ within 30 min of infusion. Additional work examined the effect of UCN (50-100 pmol) pretreatment on the feeding and metabolic effects of NPY. NPY, injected at the start of the dark period, reliably increased 2 h food intake. This effect was blocked by PVN UCN administration. Similarly, UCN blocked the increase in RQ elicited by NPY alone. These results suggest that UCN-sensitive mechanisms within the PVN may modulate food intake and energy substrate utilization, possibly through an interaction with hypothalamic NPY.

Animals↗

PubMed2001

Identification of urocortin III, an additional member of the corticotropin-releasing factor (CRF) family with high affinity for the CRF2 receptor.

The corticotropin-releasing factor (CRF) family of neuropeptides includes the mammalian peptides CRF, urocortin, and urocortin II, as well as piscine urotensin I and frog sauvagine. The mammalian peptides signal through two G protein-coupled receptor types to modulate endocrine, autonomic, and behavioral responses to stress, as well as a range of peripheral (cardiovascular, gastrointestinal, and immune) activities. The three previously known ligands are differentially distributed anatomically and have distinct specificities for the two major receptor types. Here we describe the characterization of an additional CRF-related peptide, urocortin III, in the human and mouse. In searching the public human genome databases we found a partial expressed sequence tagged (EST) clone with significant sequence identity to mammalian and fish urocortin-related peptides. By using primers based on the human EST sequence, a full-length human clone was isolated from genomic DNA that encodes a protein that includes a predicted putative 38-aa peptide structurally related to other known family members. With a human probe, we then cloned the mouse ortholog from a genomic library. Human and mouse urocortin III share 90% identity in the 38-aa putative mature peptide. In the peptide coding region, both human and mouse urocortin III are 76% identical to pufferfish urocortin-related peptide and more distantly related to urocortin II, CRF, and urocortin from other mammalian species. Mouse urocortin III mRNA expression is found in areas of the brain including the hypothalamus, amygdala, and brainstem, but is not evident in the cerebellum, pituitary, or cerebral cortex; it is also expressed peripherally in small intestine and skin. Urocortin III is selective for type 2 CRF receptors and thus represents another potential endogenous ligand for these receptors.

Amino Acid Sequence↗

PubMed2001

Urocortin II: a member of the corticotropin-releasing factor (CRF) neuropeptide family that is selectively bound by type 2 CRF receptors.

Here we describe the cloning and initial characterization of a previously unidentified CRF-related neuropeptide, urocortin II (Ucn II). Searches of the public human genome database identified a region with significant sequence homology to the CRF neuropeptide family. By using homologous primers deduced from the human sequence, a mouse cDNA was isolated from whole brain poly(A)(+) RNA that encodes a predicted 38-aa peptide, structurally related to the other known mammalian family members, CRF and Ucn. Ucn II binds selectively to the type 2 CRF receptor (CRF-R2), with no appreciable activity on CRF-R1. Transcripts encoding Ucn II are expressed in discrete regions of the rodent central nervous system, including stress-related cell groups in the hypothalamus (paraventricular and arcuate nuclei) and brainstem (locus coeruleus). Central administration of 1-10 microg of peptide elicits activational responses (Fos induction) preferentially within a core circuitry subserving autonomic and neuroendocrine regulation, but whose overall pattern does not broadly mimic the CRF-R2 distribution. Behaviorally, central Ucn II attenuates nighttime feeding, with a time course distinct from that seen in response to CRF. In contrast to CRF, however, central Ucn II failed to increase gross motor activity. These findings identify Ucn II as a new member of the CRF family of neuropeptides, which is expressed centrally and binds selectively to CRF-R2. Initial functional studies are consistent with Ucn II involvement in central autonomic and appetitive control, but not in generalized behavioral activation.

Amino Acid Sequence↗

PubMed2001

Identification of potent and selective neuropeptide Y Y(1) receptor agonists with orexigenic activity in vivo.

Neuropeptide Y (NPY) binds to a family of G-protein coupled receptors termed Y(1), Y(2), Y(3), Y(4), Y(5), and y(6). The use of various receptor subtype-selective agonists and antagonists has facilitated identification of the receptor subtypes responsible for mediating many of the biological effects of NPY. For example, the potent orexigenic activity of NPY is believed to be mediated by both the Y(1) and Y(5) receptor subtypes. Several selective Y(5) receptor agonists that stimulate food intake in rodents are available, but no selective Y(1) receptor agonist has been reported. We have identified several NPY analogs that bind the NPY Y(1) receptor with high affinity and exhibit full agonist activity, measured as inhibition of forskolin-stimulated cAMP production in cells expressing the cloned NPY Y(1) receptor. [D-Arg(25)]-NPY, [D-His(26)]-NPY, Des-AA(10--17)[Cys(7,21),Pro(34)]-NPY, Des-AA(11--18)[Cys(7,21),D-Lys(9)(Ac)]-NPY, Des-AA(11--18)[Cys(7,21),D-Lys(9)(Ac),Pro(34)]-NPY, Des-AA(11--18)[Cys(7,21),D-Lys(9)(Ac),D-His(26)]-NPY and Des-AA(11--18)[Cys(7,21),D-Lys(9)(Ac),D-His(26), Pro(34)]-NPY bind the NPY Y(1) receptor with K(i) values of 0.9 +/- 0.2, 2.0 +/- 0.3, 0.2 +/- 0.05, 0.7 +/- 0.1, 0.2 +/- 0.01, 2.2 +/- 0.3, and 1.2 +/- 0.3 nM, respectively, and inhibit forskolin-stimulated cAMP production with EC(50) values of 0.2 +/- 0.02, 0.5 +/- 0.04, 0.3 +/- 0.03, 0.5 +/- 0.05, 0.4 +/- 0.16, 5.3 +/- 0.32, and 5.1 +/- 0.97 nM, respectively. These peptides are highly selective for the NPY Y(1) receptor relative to the NPY Y(2), Y(4), and Y(5) receptors. [D-Arg(25)]-NPY, [D-His(26)]-NPY and Des-AA(11--18)[Cys(7,21), D-Lys(9)(Ac),D-His(26),Pro(34)]-NPY stimulate food intake dose-responsively in Long-Evans rats for at least 4 h after intracerebroventricular administration. Although the involvement of Y(1) receptors in several physiological activities, such as vasoconstriction and anxiolysis, remains to be investigated, adequate tools are now available.

Animals↗

PubMed2000

SST3-selective potent peptidic somatostatin receptor antagonists.

A family of octapeptide derivatives of somatostatin cyclized via a disulfide bridge (des-AA(1,2,4,5,12,13)[d-2Nal(8)]-somatostatin-14, ODN-8) was identified that has high affinity and selectivity for the human sst(3) somatostatin receptor subtype transfected in CCL39 cells. The binding affinity of carbamoyl-des-AA(1,2,4,5,12, 13)[d-Cys(3),Tyr(7),d-Agl(8)(Me,2-naphthoyl)]-somatostatin-14 (sst(3)-ODN-8) is equal to that of somatostatin-28 for sst(3) and less than one-thousandth that for the other four somatostatin receptor subtypes. Compound sst(3)-ODN-8 potently reverses the somatostatin-28-induced inhibition of forskolin-stimulated cAMP production (pK(B) = 9.07) and reverses the somatostatin-28-induced stimulation of phospholipase C activity (pK(i) = 9.22) in sst(3)-transfected CCL39 cells. [(125)I-Tyr(7)]sst(3)-ODN-8 selectively labels sst(3)-expressing cells with subnanomolar binding affinity (K(D) = 0.27 nM). With the use of this radioligand, sst(3)-expressing human tumors, particularly inactive pituitary adenomas, can be identified with receptor autoradiography; moreover, areas of the human lymphoreticular system express sst(3) binding sites selectively displaced by nanomolar concentrations of sst(3)-ODN-8. Based on the structure-activity relationship of selected analogs substituted at positions 3, 7, and 8, we hypothesize that the basis for sst(3) selectivity, high affinity, and possibly antagonism resides in the ring size of the analog and the unique conformational and structural character of the N-methylated amino-2-naphthoyl side chain of aminoglycine at position 8 and not in the Tyr(7) substitution or in the d-configuration at position 3. The family of labeled and unlabeled sst(3)-ODN-8 analogs represents highly innovative, potent, and specific sst(3)-selective antagonist tools for the study of sst(3)-mediated physiological and pathophysiological conditions that may suggest novel clinical applications.

Adenylyl Cyclases↗

PubMed2000

(R)-tert-Butoxycarbonylamino-fluorenylmethoxycarbonyl-glycine from (S)-benzyloxycarbonyl-serine or from papain resolution of the corresponding amide or methyl ester.

The enantiospecific synthesis of (R)-Boc-(Fmoc)-aminoglycine 7 was achieved. (S)-Cbz-serine 1 was reacted with diphenylphosphoryl azide in the presence of triethylamine to yield cyclic (S) carbamate 2. The ring nitrogen of 2 was protected with a Boc group (3). The cyclic carbamate of 3 was hydrolyzed with benzyltrimethylammonium hydroxide to yield the (R)-enantiomer of alcohol 4. The oxidation of 4 with pyridinium dichromate yielded the enantiomerically pure (97% ee) (R)-Boc-(Cbz)-aminoglycine 5, which was converted to 7 with retention of optical purity. Similarly, starting from (S)-Boc-serine 9, cyclic (S) carbamate 10 was obtained. The ring nitrogen of 10 was protected with a Cbz group (11) with retention of configuration. The cyclic carbamate of 11 was base hydrolyzed to yield 12, the (S)-enantiomer alcohol. Independently, Boc-(Fmoc)-aminoglycine amide 13 and Boc-(Fmoc)-aminoglycine methyl ester 14 were resolved using papain. The stereochemistry of the isolated acid was determined to be (R) by coelution on HPLC of its derivative with Marfey's reagent and that of an authentic sample (7) obtained by enantiospecific synthesis.

Amides↗

PubMed2000

Somatostatin receptor gene expression in neuroblastoma.

Somatostatin receptor expression is a favorable prognostic factor in human neuroblastoma. Somatostatin receptors have been demonstrated in vitro by pharmacologic analysis of tumor tissue and in vivo by diagnostic radioreceptor scintigraphy. However, which receptor subtypes (sst(1), sst(2), sst(3), sst(4), and sst(5)) are expressed in these tumors has not yet been delineated. We used RT-PCR to analyze expression of the five somatostatin receptor genes in 32 neuroblastoma tumor specimens. All 32 tumor specimens expressed mRNA for c-abl and sst(1); sst(2) mRNA was detected in 27/32 samples and somatostatin mRNA was detected in 30/32 tumor specimens. The remaining receptor subtypes, sst(3), sst(4), and sst(5) were variably expressed. Receptor protein for sst(1) and sst(2) was visualized in tumor neuroblasts as well as in endothelial cells of tumor vessels using immunostaining with specific anti-receptor antibodies. The effect of high expression of somatostatin receptors on cell proliferation was examined in SKNSH neuroblastoma cells transfected with sst(1) and sst(2). SS(14) binding to wild-type SKNSH cells was undetectable; but the native peptide bound with high affinity to the SKNSH/sst(1) and SKNSH/sst(2) neuroblastoma cell lines. Pharmacologic analysis of binding with two long-acting analogues, CH275 and octreotide, confirmed selective expression of sst(1) and sst(2) in stably transfected SKNSH cells. Formation of neuroblastoma xenograft tumors in nude mice was significantly delayed for both SKNSH/sst(1) (P<0.001) and SKNSH/sst(2) (P<0.05) cells compared to wild-type SKNSH. We conclude that: (1) Somatostatin receptors, sst(1) and sst(2), are expressed in the majority of neuroblastomas at diagnosis; and (2) upregulation of functional sst(1) or sst(2) in neuroblastoma cell lines suppresses tumorigenicity in a xenograft model. These observations suggest that somatostatin receptors may be a useful therapeutic target in neuroblastoma.

Animals↗

PubMed2000

The inhibitory effect of hormones associated with stress on Na appetite of sheep.

Stress is a large stimulus of Na appetite in rabbits, rats, and mice. This study investigated the influence of some peptides implicated in stress, i.e., adrenocorticotropin (ACTH), corticotropin-releasing factor (CRF), and the recently discovered member of the CRF family, urocortin, on the ingestive behavior of sheep. Intracerebroventricular infusion of these peptides over 4 days decreased the need-free Na intake of Na-repleted sheep. Intracerebroventricular infusion of urocortin, however, did not alter Na intake of Na-depleted sheep. Systemic infusion of ACTH increased, whereas systemic infusion of either urocortin or CRF decreased, Na intake of Na-repleted sheep. The increase in Na intake caused by the peripheral infusion of ACTH was blocked by concurrent i.v. infusion of urocortin, substantiating the inhibitory role of this peptide on Na appetite. Central administration of all peptides and i.v. administration of urocortin or urocortin and ACTH combined decreased food intake. Water intake was not directly influenced by the peptides. Rather, decreased water intake, when observed, was secondary to decreased food intake, as determined by pair-feeding experiments. Whereas systemic infusion of ACTH mimics the increase in Na intake observed in several different stressful situations, CRF and urocortin actually inhibit Na intake, indicating a direct central action overriding any effect of these peptides on ACTH release. Indeed, the inhibition of Na intake by urocortin occurred despite its stimulation of ACTH release and the subsequent increase in peripheral level of cortisol. Thus it would appear that hormones associated with stress have both excitatory and inhibitory influences on Na intake. Presumably, other physiological processes entrained by stress also will be important in determining the quantitative outcome on Na appetite.

Adrenocorticotropic Hormone↗

PubMed2000

Behavioral effects of central administration of the novel CRF antagonist astressin in rats.

Astressin, a novel corticotropin releasing factor (CRF) antagonist, has been found to be particularly potent at inhibiting the hypothalamo-pituitary-adrenal axis. The aim of the present study was to determine the effects in rats of astressin in attenuating the anxiogenic-like response produced by social stress and intracerebroventricular (ICV) CRF administration on the elevated plus-maze, and ICV CRF-induced locomotor activation in the rat. Astressin significantly reversed the anxiogenic-like response induced by both social stress and ICV rat/humanCRF (r/hCRF) on the elevated plus-maze, but failed to block the effects of r/hCRF-induced locomotor activity in a familiar environment. When these results were compared to previous studies performed with the same paradigms using other CRF antagonists, astressin showed effects similar to those of D-PheCRF(12-41) on plus-maze performance. However, contrary to alpha-helicalCRF(9-41) and D-PheCRF(12-41), astressin had no effect on CRF-induced locomotor activity. These results suggest that astressin may have a unique anti-CRF profile compared to previously tested antagonists.

Aggression↗

PubMed2000

Intracisternal injection of somatostatin receptor 5-preferring agonists induces a vagal cholinergic stimulation of gastric emptying in rats.

We previously showed that the somatostatin receptor 5 (sst(5))-preferring agonist BIM-23052 injected intracisternally (i.c. ; 0.8 nmol/rat) stimulated gastric emptying of a non-nutrient meal in conscious rats. In this study, we investigated the neural pathways and specificity of BIM-23052 action. BIM-23052 (0.4, 0.8, and 1.2 nmol/rat i.c.) stimulated gastric transit; values of gastric emptying were 65.5 +/- 6.5, 77.4 +/- 5.3, and 77.7 +/- 1.9%, respectively, compared with 43.2 +/-3.2% in i.c. saline group. Intravenous injection of BIM-23052 (0.8 nmol/rat) had no effect. BIM-23052 (0.8 nmol/rat i.c.) action was prevented by subdiaphragmatic vagotomy or atropine. Medullary thyrotropin-releasing hormone (TRH) is known to play a physiological role in the vagal stimulation of gastric motor function. TRH receptor antisense oligodeoxynucleotides injected i.c. with a regimen that prevented TRH (0.3 nmol/rat i.c.)-induced enhanced gastric emptying did not influence BIM-23052 stimulatory action. Somatostatin-28 (0.2-1.2 nmol/rat i.c.), which possesses a higher affinity than somatostatin-14 for sst(5), and the cyclic octapeptide des-AA(1,2,4,5,12,13)[D-Trp(8)]somatostatin (0.2-1.2 nmol/rat i.c.), an oligo-somatostatin analog that shares similar brain actions as somatostatin-28, induced a dose-related stimulation of gastric emptying. Somatostatin-14 and the preferring peptide agonists for sst(1), CH-275; sst(2), DC-32-87; sst(3), BIM-23056 and L-796,778; and sst(4), L-803,087 had no significant effect on gastric emptying when injected i.c. at 0.8 nmol/rat. These results show that BIM-23056 injected i.c. acts in the brain independently from medullary TRH to induce a vagal cholinergic stimulation of gastric emptying through the sst(5) receptor subtype.

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