ISSN:0008-4212    

DOI:10.1139/y00-901

出版商:NRC Research Press    

年代:2000

数据来源: NRC

摘要:

The mammalian daily (circadian) clock is located in the suprachiasmatic nuclei of the hypothalamus. Clock function can be detected by the measurement of the circadian change in spontaneous firing rate of suprachiasmatic nuclei cells in a brain slice preparation in vitro. We investigated the effects of neuropeptide Y on this rhythm of firing rate in hamster suprachiasmatic nuclei neurons. Slices were prepared using standard techniques. On the 1st day in vitro, neuropeptide Y (200 ng/200 nL; 47 pmol) was applied as a microdrop to the suprachiasmatic nuclei region at various times. Spontaneous single-unit firing was measured for 6-12 h on the 2nd day in vitro. Peak firing rate in treated slices was compared with that of untreated control slices to measure phase shifts induced by the peptide. Neuropeptide Y induced phase advances of circa-3h when applied during the subjective day (ZT 2-10) but did not significantly alter phase when applied during the subjective night. The phase shifts to neuropeptide Y in the hamster tissue in vitro are similar in phase dependency and magnitude to shifts measured in vivo.Key words: circadian, neuropeptide Y, rhythm, suprachiasmatic.

ISSN:0008-4212    

DOI:10.1139/y99-130

出版商:NRC Research Press    

年代:2000

数据来源: NRC

摘要:

Neuropeptide Y (NPY) is an established modulator of renal function. Although NPY reduces renal blood flow and does not alter glomerular filtration rate, it enhances diuresis and natriuresis. Although initial studies on natriuresis did not detect kaliuresis, we now report that a retrospective analysis of previous studies regarding natriuresis demonstrates NPY-induced kaliuresis under several experimental conditions. Kaliuresis was observed despite a marked reduction in urinary potassium concentrations, which may explain why it has not been noted in some initial studies. In a direct comparison of NPY-induced kaliuresis and natriuresis, both effects were slow in onset (requiring >45 min to develop fully) and blocked by the cyclooxygenase inhibitor indomethacin. While natriuresis occurred solely via a Y5receptor, kaliuresis involved a Y1receptor and an additional receptor subtype, possibly Y2. The L-type Ca2+entry blocker nifedipine abolished natriuresis but did not inhibit kaliuresis. A combination of experiments with the bradykinin B2receptor antagonist icatibant, the angiotensin II receptor antagonist losartan, and the converting enzyme inhibitor ramiprilat revealed that NPY-induced natriuresis involves bradykinin while kaliuresis involves angiotensin II. We conclude that NPY-induced kaliuresis is much less pronounced than natriuresis and is mediated by distinct mechanisms.Key words: neuropeptide Y, potassium excretion, sodium excretion, angiotensin II, cyclooxygenase.

ISSN:0008-4212    

DOI:10.1139/y99-121

出版商:NRC Research Press    

年代:2000

数据来源: NRC

摘要:

Propofol is a widely used anesthetic for both induction and maintenance of anesthesia during surgery. A strong feeling of hunger has been reported during the early recovery period after propofol anesthesia. We have investigated the effect of propofol on appetite in 10 patients undergoing a craniotomy and in parallel measured neuropeptide Y (NPY), catecholamines, and serotonin levels in the cerebrospinal fluid and plasma during anesthesia. Ten patients anesthetized with a volatile agent (isoflurane) served as a control group. Plasma NPY and catecholamines levels were not affected by surgery at any time. We observed a strong increase in NPY concentrations in the cerebrospinal fluid independently of the anesthetic technique agent used, whereas catecholamines were unchanged. We found that serotonin concentrations decreased significantly in the plasma (but not in the cerebrospinal fluid) of patients treated by propofol when compared with the control group; this decrease was associated with an increase of hunger early postoperatively. We concluded that the proappetite effect of propofol is mediated through a decrease of serotonin at the peripheral level.Key words: catecholamines, serotonin, neuropeptide Y (NPY), cerebrospinal fluid, serum, appetite.

ISSN:0008-4212    

DOI:10.1139/y99-122

出版商:NRC Research Press    

年代:2000

数据来源: NRC

摘要:

The design of non-peptide, Y1-selective antagonists of neuropeptide Y (NPY) as pharmacological tools is in progress and is increasingly important as therapeutic applications are expected. Starting from the potent histamine H2agonist and weak NPY Y1antagonist arpromidine, 16 imidazolylpropylguanidine derivatives were synthesized and tested for Y1antagonistic activity (inhibition of NPY-stimulated Ca2+increase in human erythroleukemic cells), where the pheniramine-like moiety of arpromidine was replaced with 2-pyridylaminoalkyl, benzyl-(2-pyridyl)aminoalkyl, and phenyl-(2-pyridyl)alkylaminoalkyl partial structures derived from mepyramine. The pA2values of the most active compounds are in the range of 6.2-6.5. Quantitative structure-activity relationships (QSAR) were investigated by fragment regression analysis. Results indicate that a tetramethylene spacer between the guanidino group and the amino nitrogen is optimal. For an at least moderate degree of Y1antagonistic activity, a second benzyl or phenyl group must be present in addition to the 2-pyridyl ring. At this second group, hydrophobic substituents such as 3,4-di-Cl and 4-Br further enhance Y1antagonism. The most active derivative additionally bears a 5-Br substituent at the 2-pyridyl moiety. Structure-activity relationships suggest that the compounds might be able to partially imitate the role of NPY when interacting with Y1receptors and thus behave as moderate non-peptide NPY Y1antagonists.Key words: neuropeptide Y Y1antagonists, imidazolylpropylguanidines, quantitative structure-activity relationships.

ISSN:0008-4212    

DOI:10.1139/y99-120

出版商:NRC Research Press    

年代:2000

数据来源: NRC

摘要:

We have evaluated 3 newly developed neuropeptide Y receptor antagonists in various in vitro binding and bioassays: BIBO3304 (Y1), T4[NPY33-36]4(Y2), and CGP71683A (Y5). In rat brain homogenates, BIBO3304 competes for the same population of [125I][Leu31,Pro34] peptide YY (PYY) binding sites (75%) as BIBP3226, but with a 10 fold greater affinity (IC50of 0.2 ± 0.04 nM for BIBO3304 vs. 2.4 ± 0.07 nM for BIBP3226),while CGP71683A has high affinity for 25% of specific [125I][Leu31,Pro34]PYY binding sites. Both BIBO3304 and CGP71683A (at 1.0 µM) were unable to compete for a significant proportion of specific [125I]PYY3-36/Y2sites. The purported Y2antagonist T4[NPY33-36]4competed against [125I]PYY3-36binding sites with an affinity of 750 nM. These results were confirmed in HEK 293 cells transfected with either the rat Y1, Y2, Y4, or Y5receptor cDNA. BIBO3304, but not CGP71683A, competed with high affinity for [125I][Leu31,Pro34]PYY binding sites in HEK 293 cells transfected with the rat Y1receptor cDNA, whereas the reverse profile was observed upon transfection with the rat Y5receptor cDNA. Additionally, both molecules were inactive at Y2and Y4receptor subtypes expressed in HEK 293 cells. Receptor autoradiographic studies revealed the presence of [125I][Leu31,Pro34]PYY/BIBO3304-insensitive sites in the rat brain as reported previously for BIBP3226. Finally, the selective antagonistic properties of BIBO3304 were demonstrated in a Y1bioassay (rabbit saphenous vein; pA2value of 9.04) while being inactive in Y2(rat vas deferens) and Y4(rat colon) bioassays. These results confirm the high affinity and selectivity of BIBO3304 and CGP71683A for the Y1and Y5receptor subtypes, respectively, while the purported Y2antagonist, T4[NPY33-36]4possesses rather low affinity for this receptor.Key words: NPY receptor antagonist, receptor subtypes, bioassays, receptor binding assays, autoradiographic studies, receptor distribution.

ISSN:0008-4212    

DOI:10.1139/y99-119

出版商:NRC Research Press    

年代:2000

数据来源: NRC

摘要:

This investigation describes the relative potencies of four peptide agonists, namely, peptide YY (PYY), [Leu31,Pro34]PYY (Pro34PYY), neuropeptide Y (NPY), and [Leu31,Pro34]NPY (Pro34NPY), as antisecretory agents in human, rat, and mouse gastrointestinal preparations. The inhibition of agonist responses by the Y1-receptor antagonist BIBP 3226 was also tested in each preparation. An unexpectedly pronounced preference for PYY and Pro34PYY was observed in functional studies of two human epithelial lines stably transfected with the rat Y1receptor (Y1-7 and C1Y1-6). NPY and Pro34NPY were at least an order of magnitude less effective than PYY in these functional studies but were only marginally less potent in displacement binding studies using membrane preparations of the same clonal lines. The orders of agonist potency obtained in Y1-7 and C1Y1-6 epithelia were compared with those obtained from a single human colonic adenocarcinoma cell line (Colony-6, which constitutively expresses Y1receptors) and also from mucosal preparations of rat and mouse descending colon. Similar peptide orders of potency were obtained in rat and mouse colonic mucosae and Colony-6 epithelia, all of which exhibited PYY preference (although less pronounced than with Y1-7 and C1Y1-6 epithelia) and significant sensitivity to the Y1receptor antagonist, BIBP 3226. We have compared the pharmacology of these five mammalian epithelial preparations and provide cautionary evidence against the reliance upon agonist concentration-response relationships alone, in the characterization of NPY receptor types.Key words: Y receptors, neuropeptide Y, gastrointestinal epithelia, ion transport.

ISSN:0008-4212    

DOI:10.1139/y99-123

出版商:NRC Research Press    

年代:2000

数据来源: NRC

摘要:

Aiming to develop a functional assay for the human NPY Y5receptor based on adenylyl cyclase activity, HEC-1B cells, in which cAMP synthesis can be efficiently stimulated with forskolin, were selected for the transfection with the pcDNA3-Y5-FLAG and the pcDEF3-Y5vectors. After optimization of the transfection procedure, the binding of [3H]propionyl-NPY to transiently and stably expressed Y5receptors was determined. The affinities of NPY, NPY derivatives, and rPP (pNPY >= p(Leu31Pro34)NPY = p(2-36)NPY >= p(D-Trp32)NPY > p(13-36)NPY > rPP) were in accordance with the NPY Y5receptor subtype. For [3H]propionyl-pNPY approximately 1.7 × 105and 1 × 106binding sites per transiently and stably transfected cell, respectively, were determined. TheKDvalues were 2.4 ± 0.4 and 1.7 ± 0.2 nM, respectively. Due to the high expression of the receptor protein, both stably and transiently transfected cells can be conveniently used in routine radioligand binding studies. By contrast, functional assays were only feasible with HEC-1B cells stably expressing the Y5receptor. In these cells, 10 nM pNPY inhibited the forskolin-stimulated cAMP synthesis by 75%. This effect was partially antagonized by the Y5antagonistN-{trans-[4-(2-naphthylmethylamino)- methyl]cyclohexylmethyl}naphthalene-2-sulfonamide. Although the genetic variability of cancer cells is in principle incompatible with a stable phenotype, both ligand binding characteristics and functionality of the Y5receptor remained unchanged for more than 30 passages.Key words: human NPY Y5receptor, HEC-1B cells, stable expression, radioligand binding, cAMP assa

ISSN:0008-4212    

DOI:10.1139/y99-125

出版商:NRC Research Press    

年代:2000

数据来源: NRC

摘要:

Neuropeptide Y (NPY) and melanocortin (MC) peptides have opposite effects on food intake: NPY-like peptides and MC receptor antagonists stimulate feeding and increase body weight, whereas melanocortins and NPY antagonists inhibit food intake. In this study we tested whether the orexigenic effect of the selective MC4receptor antagonist HS014 (1 nmol) could be inhibited by three different NPY antagonists, (R)-N2-(diphenylacetyl)-N-[(4-hydroxy-phenyl)methyl]- D-argininamide (BIBP3226), (R)-N-[[4-(aminocarbonylaminomethyl)-phenyl]methyl]-N2-(diphenylacetyl)-argininamide-trifluoroacetate (BIBO3304), and decapeptide [D-Tyr27,36D-Thr32]NPY27-36, after icv administration in freely feeding male rats. All three NPY receptor antagonists inhibited the orexigenic effects of HS014 partially and with markedly different potency. [D-Tyr27,36D-Thr32]NPY27-36was active only in subconvulsive dose. The NPY Y1selective antagonist BIBP3226 was more effective in inhibiting the effect of HS014 than BIBO3304 despite in vitro data indicating that BIBP3226 is about 10 times less potent than BIBO3304 at NPY Y1receptor. An enantiomer of BIBO3304, BIBO3457, failed to inhibit HS014-induced feeding, indicating that the effects of BIBO3304 were stereoselective. These results suggest that stimulation of food intake caused by weakening of melanocortinergic tone at the MC4receptor is partially but not exclusively related to NPY Y1receptor activation.Key words: neuropeptide Y, NPY Y1receptor antagonist, BIBO3304, BIBP3226, [D-Tyr27,36D-Thr32]NPY(27-36), 1229U91, food intake, MC4receptor antagonist, HS014.

ISSN:0008-4212    

DOI:10.1139/y99-124

出版商:NRC Research Press    

年代:2000

数据来源: NRC

摘要:

FMRFamide and related peptides (RFamides) were found to inhibit the association binding of iodinated human pancreatic polypeptide ([125I]hPP) to Y5-like neuropeptide Y (NPY) receptor in rodent tissues. An allosteric regulation of the activity of the rodent kidney PP-sensitive neuropeptide Y (NPY) receptor by RFamides was indicated by potency decrease with particle concentration in the inhibition of the association binding of125I-labeled human pancreatic polypeptide (hPP) by RFamides at rabbit kidney membranes. The competition by C-terminal hexapeptide of hPP (LTRPRY.NH2) did not show such affinity change. The steady-state binding of hPP showed little sensitivity to any of the RFamides tested. The Y1-selective binding of [125I][Leu31,Pro34]hPYY (at 2 nM hPP) was much less sensitive to RFamides than the binding of [125I]hPP, albeit with some differences across tissue or cell types. The binding of Y2-selective agonist125I-labeled human peptide YY (3-36) was quite insensitive to RFamides. The presence of a unique component in the inhibition of hPP binding by RFamides was further indicated by a degree of antagonism with phospholipase C inhibitor U-73122, and by an only limited cooperation with a N5-amiloride compound, and with alkylator chloroethylclonidine. Change of the chirality of individual residues in the FMRFamide molecule produced a significant reduction of inhibitory potency only with D-Phe in the C-terminal position. Substitution of the (C-3) L-Met by L-Leu greatly increased the inhibitory potency of RFamides relative to otherwise identical congeners. RFamides could act both as ligands of membrane neighbors of the PP receptor, and as competitors of Y5-like NPY receptor epitopes that accommodate the C-terminal aspects of agonist peptides.Key words: Y1receptor, Y2receptor, Y5receptor, RFamide, allosteric interaction, hydrophobic pocket, amino acid chirality.

ISSN:0008-4212    

DOI:10.1139/y99-126

出版商:NRC Research Press    

年代:2000

数据来源: NRC