Reduction of vasoconstriction mediated by neuropeptide Y Y2 receptors in arterioles of the guinea-pig small intestine.

Brief applications of a high-K+ solution were used to evoke transient constrictions of arterioles from the guinea-pig small intestine. Analogues of neuropeptide Y (NPY) selective for Y2-receptors reduced the constrictions, whereas NPY or a Y1-selective analogue potentiated the constrictions. We conclude that arteriolar smooth muscle has both Y1 and Y2 receptors, and suggest that Y2 receptors inhibit vasoconstriction by modulating the opening of voltage-sensitive Ca2+ channels. This may be related to the role of NPY that is present in some vasodilator nerves.     

Characterization of vascular neuropeptide Y receptors.

1. In the present study we compared neuropeptide Y (NPY) and NPY-related analogues for their ability to activate or bind to vascular NPY receptors in four experimental set-ups. Previous results have suggested the existence of different receptor subtypes, Y1 receptors requiring full-length NPY (1-36) or [Pro34]-NPY, and Y2 receptors recognizing also N-terminally truncated forms of NPY but not [Pro34]-NPY. 2. NPY 1-36 and [Pro34]-NPY dose-dependently increased arterial pressure in the anaesthetized rat with a similar magnitude and potency. NPY 2-36 was much less potent than NPY 1-36. NPY 4-36 and NPY 11-36 were inactive even at a dose as high as 10 nmol kg-1. 3. NPY 1-36, [Pro34]-NPY, NPY 2-36 and NPY 5-36 concentration-dependently increased the coronary resistance in the Langendorff preparation of the rat. NPY 1-36 and [Pro34]-NPY were equipotent, while NPY 2-36 and NPY 5-36 were about 7 and 20 times less potent. At 0.3 microM, NPY 11-36, NPY 20-36 and NPY 22-36 induced a slight contraction while NPY 23-36 was inactive. 4. NPY 1-36, [Pro34]-NPY, NPY 2-36, NPY 4-36, NPY 5-36 and NPY 11-36 evoked concentration-dependent contractions in the isolated inferior caval vein of the rat and guinea-pig. [Pro34]-NPY was more potent than NPY 1-36. NPY 2-36 was equipotent with NPY 1-36, while NPY 4-36, NPY 5-36 and NPY 11-36 were approximately 30 times less potent.(ABSTRACT TRUNCATED AT 250 WORDS)     

Emerging functions for neuropeptide Y5 receptors

The Y5 subtype of neuropeptide Y (NPY) receptors has raised considerable interest as a mediator of NPY-stimulated food intake, but with the advent of recent data, this hypothesis has come into question. Moreover, Y5 receptor-selective drugs might not be specific for food intake because additional functions in the central and peripheral nervous systems, including endogenous anti-epileptic activity, attenuation of morphine withdrawal symptoms, enhancement of diuresis and natriuresis, lowering of blood glucose and reduction of acetylcholine release in the ileum, have recently been reported to occur via Y5-like receptors. Given that mRNA for the cloned Y5 receptor is apparently restricted to the CNS, Angela Bischoff and Martin Michel discuss the possible existence of additional NPY receptor subtypes with Y5-like recognition features and their presence in peripheral tissues.     

Potentiation by neuropeptide Y of vasoconstriction in rat resistance arteries.

1. The effects of neuropeptide Y (NPY) on resistance arteries were investigated on 3rd generation mesenteric arterioles of the rat. 2. Contractions were elicited by noradrenaline (NA), 5-hydroxytryptamine (5-HT), prostaglandin F2 alpha (PGF2 alpha), depolarization (KCl substituted for NaCl) and by the calcium agonist Bay K 8644, in the absence and in the presence of NPY (100 nM), a concentration which by itself did not induce vasoconstriction. 3. NPY produced a leftward shift of the concentration-response curves to the agonists and to KCl, without any alteration of maximal contractions. 4. NPY also potentiated contractions elicited by addition of CaCl2 to KCl-depolarized vessels, but its effect on calcium-induced contractions decreased with increasing KCl concentrations (from 20 to 100 mM). 5. Calcium-induced contractions were inhibited by the calcium channel blocker nitrendipine, both in the presence and absence of NPY (100 nM). NPY increased slightly (but significantly) the sensitivity to nitrendipine (the apparent KB increased from 2.9 x 10(-10) M to 1.6 x 10(-10) M). 6. The KCl concentration necessary for the maximal effect of Bay K 8644 was decreased in the presence of NPY, and the sensitivity to the calcium channel agonist was increased. 7. Elevating the KCl concentration in the bath from 5 to 20 mM (which gives the same displacement to the left of the KCl concentration-effect curve seen in the presence of NPY) induced a parallel leftward shift of NA and 5-HT concentration-response curves. This shift was identical to the one induced by NPY on 5-HT-evoked contractions, but it was significantly smaller (P less than 0.001) than the shift of the NA concentration-response curve observed in the presence of NPY. In the latter case, NPY enhanced more markedly the contractions induced by low NA concentrations (between 10(-9) and 3 x 10(-8 M) than those induced by high concentrations (up to 3 x 10(-7) M), thus giving a shallow concentration-response curve. 8. The results strongly suggest that NPY partially depolarizes the arterioles and induces an increase in calcium entry through voltage-dependent channels, thus enhancing contractions elicited by agonists or by KCl-depolarization. In addition, they support the view that another mechanism also plays a part in the potentiation by NPY of the effects of low concentrations of NA.     

5—羟色胺受体介导迷走交感神经切断犬颈外血管收缩

One specific example reflecting the complexity of cardiovascular responses induced by serotonin (5-hydroxytryptamine; 5-HT) and the progress achieved in the pharmacological characterization of the receptors involved can be illustrated by the effects of 5-HT on the canine external carotid artery bed. Within this framework, it has been shown that the external carotid vasoconstrictor response to 5-HT in the dog is mediated by '5-HT1-like' receptors, which being blocked by the 5-HT1B/1D receptor antagonist GR127935, resemble 5-HT1B/1D (previously called 5-HT1D beta/1D alpha) receptors. It was proposed that these receptors could belong to the 5-HT1B, rather than the 5-HT1D, subtype on the basis of their resistance to blockade by a high dose of ritanserin (a potential 5-HT1D receptor antagonist) and the presence of mRNA for 5-HT1B(5-HT1D beta) receptors, but not for 5-HT1D(5-HT1D alpha) receptors, in vascular smooth muscle. With the advent of subtype-selective antagonists it was subsequently shown that external carotid vasoconstriction to 5-HT and sumatriptan is dose-dependently antagonized by the selective 5-HT1B receptor antagonist SB224289 (2,3,6,7-tetrahydro-1'-methyl-5-[2'-methyl-4' (5-methyl-1,2,4-oxadiazol-3-yl) biphenyl-4-carbonyl] furo [2,3-f] indole-3-spiro-4'-piperidine hydrochloride), whereas the selective 5-HT1D receptor antagonist BRL15572 (1-(3-chlorophenyl)-4-[3,3-diphenyl (2-(S,R) hydroxypropanyl) piperazine] hydrochloride) was ineffective. These findings represent the first in vivo evidence showing that vascular constriction induced by 5-HT and sumatriptan is mediated primarily via 5-HT1B, but not 5-HT1D receptors. The pharmacological profile of these receptors could be similar (isolated human temporal artery and porcine carotid arteriovenous anastomoses) to other putative 5-HT1B receptors mediating vasoconstrictor responses. In view of the putative pathophysiologic role of external carotid (and extracerebral) vasodilation in migraine, the constriction of these blood vessels by sumatriptan via 5-HT1B receptors may be, at least partly, responsible for its therapeutic efficacy in migraine.     

Structure-function studies on neuropeptide Y and pancreatic polypeptide--evidence for two PP-fold receptors in vas deferens

The biological effects of neuropeptide Y (NPY), rat pancreatic polypeptide (rPP), hybrid analogs of NPY and PP, and C-terminal fragments of NPY were studied in the field-stimulated rat vas deferens model. The results were correlated with peptide binding experiments in Y1 and PP receptor assays on rat PC-12 cells and Y2 receptors on porcine hippocampal membranes. NPY and rPP inhibited the electrically induced contractions in the vas deferens with an IC50 of 25 and 22 nM respectively. However, in contrast to NPY, rPP could not totally block muscle activity. The inhibitory action of the long C-terminal fragment of NPY, NPY-(19-36) and NPY-(11-36), indicated that NPY acts through a Y2 receptor in the vas deferens. The structural basis for the differential recognition of NPY and PP by Y2 receptors and partly also by PP receptors, could be defined with hybrid analogs of PP and NPY. The analogs, [Ile31,Gln34]PP and [Leu31,Pro33]NPY reacted in the vas deferens preparation in accordance with their relative potency in the Y2 and PP receptor assays. [Ile31,Gln34]PP, which bound to the Y2 receptor like NPY, was also able to block the part of the contractile response which was resistant to rPP. It is concluded that in the vas deferens, PP-fold peptides act through two types of receptors: Y2 and PP, and that residues in the C-terminal part of the molecules determine the differential recognition of the peptides by these receptor types.     

The Y1 antagonist BIBP 3226 inhibits potentiation of methoxamine-induced vasoconstriction by neuropeptide Y

We investigated the interaction of neuropeptide Y (NPY) with the α₁-adrenoceptor agonist, methoxamine, in control of mean arterial pressure, renovascular resistance and mesenteric vascular resistance in anaesthetized rats. Infusion of 3.0 but not 0.3μg/kg/min NPY enhanced the elevations of all three haemodynamic parameters caused by bolus injections of methoxamine (10–100μg/kg). These enhancements largely involved a prolongation of the methoxamine effects. While infusion of the Y₁ NPY receptor-selective antagonist, BIBP 3226 (10μg/kg/min), alone did not alter methoxamine-induced vasoconstriction, it inhibited the potentiation by NPY. We conclude that NPY can potentiate methoxamine-induced vasoconstriction in vivo. This is mediated predominantly, if not exclusively, via the Y₁ receptor. Endogenously released NPY does not appear to reach sufficient concentrations to cause tonic systemic vasoconstriction or potentiation thereof in the anaesthetized rat. Received: 30 May 1997 / Accepted: 25 July 1997     

Evidence for co-transmitter role of neuropeptide Y in the pig spleen.

1. The possible involvement of neuropeptide Y (NPY) in relation to noradrenaline (NA) and adenosine triphosphate (ATP) mechanisms in the sympathetic nervous control of the vascular tone and capsule contraction in the blood perfused pig spleen was investigated in vivo. 2. Local injections or infusions of NA, NPY and alpha-, beta-methylene ATP (mATP) caused vasoconstriction (perfusion pressure increase) and capsule contraction (increased venous blood flow). ATP only evoked vasodilatation. NPY was about 50 fold more potent than NA as a vasoconstrictor, and the NPY response was more long-lasting. Reserpine treatment did not change the effects of NPY. 3. Electrical stimulation of the splenic nerves in control animals caused a frequency-dependent, guanethidine-sensitive output of both NPY-like immunoreactivity (-LI) and NA, suggesting co-release. The output of NPY-LI relative to NA was enhanced at high frequency stimulation. Furthermore, alpha-adrenoceptor blockade by phentolamine enhanced both the output of NPY-LI and NA while inhibition of the neuronal uptake of NA with desipramine reduced the low frequency stimulation-evoked overflow of NPY-LI. Preganglionic denervation did not change the output of NPY-LI or NA. 4. Reserpine treatment reduced both the splenic content of NA and NPY-LI. Preganglionic denervation inhibited the reserpine-induced depletion of the NPY content but not of NA in terminal areas. The stimulation-evoked NPY overflow was markedly enhanced, especially at low-frequency stimulation after reserpine, and the plasma levels of NPY-LI in the venous effluent were then in the nmolar range (i.e. where exogenous NPY induced vasoconstriction). The perfusion-pressure increase upon stimulation in reserpine-treated, preganglionically-denervated animals was highly correlated (r = 0.91) to the NPY overflow. The functional 0.5 Hz responses were reduced after reserpine, while at higher frequencies the functional effects were of similar magnitude to controls but longer-lasting. 5. Tyramine induced a release of NA but not of NPY-LI. Furthermore, the increase in perfusion pressure induced by tyramine was absent after reserpine. 6. After tachyphylaxis to the vasoconstrictor effects of mATP, the nerve stimulation-evoked, functional response as well as the NA and NPY-LI overflow were unchanged. After reserpine treatment, both the perfusion-pressure increase and NPY-LI overflow to nerve stimulation were reduced after mATP tachyphylaxis. 7. In conclusion, release of NPY rather than ATP may explain the long-lasting, non-adrenergic, splenic functional responses in reserpinized animals upon sympathetic stimulation. However, NA is most likely the main splenic transmitter when low-frequency stimulation is used under control conditions.(ABSTRACT TRUNCATED AT 400 WORDS)     

5-羟色胺受体介导迷走交感神经切断犬颈外血管收缩(英文)

One specific example reflecting the completity ofcardiovascular responses induced by serotonin ( 5-hydroxytryptamine; 5-HT) and the progress achieved inthe pharmacological characterization of the receptorsinvolved can be illustrated by the effects of 5-HT on thecanine external carotid artery bed. Within thisframework, it has been shown that the external carotidvasoconstrictor response to 5-HT in the dog is mediatedby '5-HT_1-like' receptors, which being blocked by the5-HT_(1B/1D) receptor antagonist GR127935, resemble 5-HT_(1B/1D)(previously called 5-HT_(1B/1D) ) receptors. It     

Comparative structural requirements of brain neuropeptide Y binding sites and vas deferens neuropeptide Y receptors

A series of fragments and analogues of neuropeptide Y (NPY), both human (hNPY) and porcine (pNPY), were synthesized and tested for their affinities at brain NPY receptor binding sites and their potencies in inhibiting the electrically stimulated twitch response of rat vas deferens. Results with N- and C-terminal fragments suggest that amino acid residues in the N-terminal portion of the molecule are mostly important for recognition of brain and vas deferens NPY receptors, in addition to being relevant for the maintenance of adequate receptor affinity. On the other hand, C-terminal amino acid residues appear to be responsible for triggering receptor activation in the rat vas deferens preparation, because full intrinsic activity is maintained with fragments up to NPY18-36. C-terminal fragment NPY25-36 and N-terminal fragment NPY1-15 were devoid of affinity for [3H]NPY brain receptor sites and showed no activity in the rat vas deferens preparation. Similarly, N-terminal fragment hNPY1-24CONH2 showed no affinity toward [3H]NPY brain receptor sites and no inhibition of the twitch response in the rat vas deferens preparation at concentrations up to 1.0 microM. On the contrary, this fragment appears to selectively increase the amplitude of the twitch response to electrical stimulation at low micromolar concentrations, an effect opposite to that of NPY and all other NPY fragments and analogues studied here. The exact mechanism mediating this contractile action of hNPY1-24CONH2 remains to be established. Modifications of the tyrosine residue in position 20 led to the development of two analogues, [D-Tyr20]hNPY and [D-Trp20]hNPY, which show an apparent preference for the vas deferens NPY receptor. On the other hand, substitutions of the tyrosine residue in position 21 by a phenylalanine ([Phe21]hNPY) or a methylated tyrosine residue ([Tyr-O-Me21]hNPY) produced analogues demonstrating an apparent preference for the brain receptor site. This suggests that modifications of tyrosine residues at positions 20 and/or 21 may eventually lead to the development of NPY analogues distinguishing between the most abundant class of sites present in the brain and vas deferens, respectively.     

Modulation of neuropeptide Y receptors for the treatment of obesity

Background: Neuropeptide Y (NPY) has been demonstrated to have critical roles in the physiological control of appetite and energy homeostasis through NPY Y1, Y2, Y4 and Y5 receptors. A number of synthetic ligands for NPY receptor subtypes have been developed to date, with Y5 receptor antagonists and Y2 and Y4 receptor agonists advancing into clinical trials. Methods: A survey of the scientific and patent literature since mid-2006 is presented. Conclusion: In addition to the specific modulation of respective NPY receptor subtypes, recent investigations have revealed that modulation of multiple NPY receptor subtypes produces additive or even synergistic anti-obesity effects. Development of reliable small molecule Y1, Y2 and Y4 receptor ligands would greatly accelerate investigations and drug discovery.     

Amygdalar neuropeptide Y Y1 receptors mediate the anxiolytic-like actions of neuropeptide Y in the social interaction test

The effects of intra-amygdalar neuropeptide Y infusions were assessed in rats using the social interaction test. Neuropeptide Y administered into the central nucleus of the amygdala did not alter behavior, while injections into the basolateral nucleus of the amygdala produced an increased social interaction time. Furthermore, the anxiolytic-like effect was antagonized by co-administration of the potent neuropeptide Y Y1 receptor antagonist ((R)-N-[[4-(aminocarbonylaminomethyl)-phenyl]methyl]-N2-(diphen ylacetyl)-argininamide trifluoroacetate) 3304, but not with the inactive enantiomer ((R)-N-[[4-(aminocarbonylaminomethyl)-phenyl]methyl]-N2-(diphen ylacetyl)-argininamide trifluoroacetate) 3457. Therefore, neuropeptide Y produces an anxiolytic-like effect in the social interaction test through neuropetide Y Y1 receptors located in the basolateral amygdala.     

Evidence for involvement of neuropeptide Y receptors in the regulation of food intake: studies with Y1-selective antagonist BIBP3226

1. Experiments were conducted to evaluate the effects of the novel non-peptide neuropeptide Y Y₁ receptor antagonist, BIBP3226 (N²-(diphenylacetyl)-N-[(4-hydroxy-phenyl)methyl]-D-arginine amide) on spontaneous, fasting-induced and NPY-induced food intake in rats. In addition to consumption of regular chow, the effects of BIBP3226 on consumption of highly palatable sweetened mash were monitored in a 1 h test on first exposure and after familiarization with novel food.
2. BIBP3226 (10.0 nmol, i.c.v.) had no effect on the consumption of regular chow, but reduced significantly the intake of highly palatable diet and the food intake stimulated by fasting (24 h). Neuropeptide Y (NPY, 1.0 nmol, i.c.v.) significantly increased the consumption of regular rat chow. This orexigenic effect of NPY was blocked by BIBP3226 (10.0 nmol, administered i.c.v. 5 min before NPY) at 30  min and 4  h, but not at 1 and 2  h. When animals were pretreated with diazepam (0.5 mg kg⁻¹, i.p., 20 min before NPY), BIBP3226 failed to suppress NPY-induced feeding.
3. An NPY Y₁ and Y₃ receptor agonist, [Leu³¹,Pro³⁴]NPY and a Y₅ receptor agonist human peptide YY_3–36 (hPYY_3–36, both 30 pmol), microinjected into the paraventricular nucleus of the hypothalamus (PVN) increased the consumption of regular rat chow. BIBP3226 (0.4 nmol, into the PVN) completely blocked the stimulatory effect of [Leu³¹,Pro³⁴]NPY but not that of hPYY_3–36. BIBP3226 (0.4 nmol) alone failed to modify the consumption of the regular chow. Higher doses of BIBP3226 (1.0 and 2.0 nmol) injected into the vicinity of the PVN reduced the consumption of the sweetened mash.
4. These results suggest that both the NPY Y₁ and Y₅ receptors in the PVN are involved in the regulation of food intake. The stimulatory effect of exogenous NPY is probably mediated through an NPY receptor subtype that is not identical with the Y₁ receptor (possibly Y₅ receptor). However, the NPY Y₁ receptors may mediate the effect of endogenous NPY in conditions of increased energy demand or on intake of highly palatable diets.

     

Pharmacological characterization of prostanoid receptors mediating vasoconstriction in human umbilical vein

1. This study was undertaken to characterize pharmacologically the prostanoid receptor subtypes mediating contraction in human umbilical vein (HUV). 2. HUV rings were mounted in organ baths and concentration-response curves to U-46619 (TXA(2) mimetic) were constructed in the absence or presence of SQ-29548 or ICI-192,605 (TP receptor antagonists). U-46619 was a potent constrictor (pEC(50): 8.03). SQ-29548 and ICI-192,605 competitively antagonized responses to U-46619 with pK(B) values of 7.96 and 9.07, respectively. 3. Concentration-response curves to EP receptor agonists: PGE(2), misoprostol and 17-phenyl-trinor-PGE(2) gave pEC(50) values of 5.06, 5.25 and 5.32, respectively. Neither pEC(50) nor maximum of PGE(2) and 17-phenyl-trinor-PGE(2) concentration-response curves were modified by the DP/EP(1)/EP(2) receptor antagonist AH 6809 (1 micro M). However, ICI-192,605 produced a concentration-dependent antagonism of the responses to all the EP receptor agonists. The pA(2) estimated for ICI-192,605 against PGE(2) or misoprostol were 8.91 and 9.22, respectively. 4. Concentration-response curves to FP receptor agonists: PGF(2)(alpha) and fluprostenol gave pEC(50) values of 6.20 and 5.82, respectively. ICI-192,605 (100 nM) was completely ineffective against PGF(2)(alpha) or fluprostenol. In addition, lack of antagonistic effect of AH 6809 (1 micro M) against PGF(2)(alpha) was observed. 5. In conclusion, the findings obtained with TP-selective agonist and antagonists provide strong evidence of the involvement of TP receptors promoting vasoconstriction in HUV. Furthermore, the action of the natural and synthetic EP receptor agonists appears to be mediated via TP receptors. On the other hand, the results employing FP receptor agonists and antagonists of different prostanoid receptors suggest the presence of FP receptors mediating vasoconstriction in this vessel.     

5-Hydroxytryptamine receptors mediating vasoconstriction in pulmonary arteries from control and pulmonary hypertensive rats.

1. We investigated 5-hydroxytryptamine (5-HT)-receptor mediated vasoconstriction in the main, first branch and resistance pulmonary arteries removed from control and pulmonary hypertensive rats. Contractile responses to 5-HT, 5-carboxamidotryptamine (5-CT, non-selective 5-HT1 agonist), and sumatriptan (5-HT1D-like receptor agonist) were studied. The effects of methiothepin (non-selective 5-HT1 + 2-receptor antagonist) and ketanserin (5-HT2A receptor antagonist) and GR55562 (a novel selective 5-HT1D receptor antagonist) on 5-HT-mediated responses were also studied. Basal levels of adenosine 3':5'-cyclic monophosphate ([cyclic AMP]i) and guanosine 3':5'-cyclic monophosphate ([cyclic GMP]i) were determined and we assessed the degree of inherent tone in the vessels under study. 2. 5-HT was most potent in the resistance arteries. pEC50 values were 5.6 +/- 0.1, 5.3 +/- 0.1, 5.0 +/- 0.2 in the resistance arteries, pulmonary branch and main pulmonary artery, respectively (n = at least 5 from 5 animals). The sensitivity to, and maximum response of, 5-HT was increased in all the arteries removed from the chronic hypoxic (CH) rats. In CH rats the pEC50 values were 5.9 +/- 0.2, 6.3 +/- 0.2, 6.4 +/- 0.2 and the increase in the maximum response was 35%, 51% and 41% in the resistance arteries, pulmonary branch and main pulmonary artery, respectively. Sumatriptan did not contract any vessel from the control rats whilst 5-CT did contract the resistance arteries. In the CH rats, however, they both contracted the resistance arteries (responses to sumatriptan were small) (pEC50: 5-CT; 5.4 +/- 0.2) and the pulmonary artery branches (pEC50: sumatriptan, 5.4 +/- 0.2; 5-CT, 5.4 +/- 0.2). 5-CT also caused a contraction in the main pulmonary artery (pEC50: 6.0 +/- 0.3). 3. Ketanserin (1 nM-1 microM) caused a competitive antagonism of the 5-HT response in all vessels tested. In control rats, the estimated pKb values for ketanserin in resistance arteries, pulmonary branches and main pulmonary artery were 8.3, 7.8 and 9.2, respectively. Methiothepin (1 nM-1 microM) inhibited responses to 5-HT in the first branch (estimated pKb value: 7.8) and main pulmonary artery. In CH rats, the estimated pKb values for ketanserin in resistance arteries, pulmonary branches and main pulmonary artery were 7.7, 8.3 and 9.6, respectively. Methiothepin also inhibited contractions to 5-HT in the pulmonary artery branch and main pulmonary artery with estimated pKb values of 7 and 9.5, respectively. In control animals, GR55562 had no effect on responses to 5-HT in any of the vessels tested. In the CH rats the estimated pKb values for GR55562 were 6.5, 7.8 and 7.0 in the pulmonary resistance arteries, first branch and main pulmonary artery, respectively. 4. Large pulmonary arteries from controls demonstrated inherent tone and this was increased three fold in the CH rats. The resistance arteries from controls demonstrated little inherent tone though this was enhanced in those from the CH rats. 5. [Cyclic AMP]i was 259 +/- 23 pmol mg-1 protein in the pulmonary artery branches removed from control rats and decreased to 192 +/- 11 pml mg-1 protein in the CH rats (P < 0.01, n = 8). [Cyclic GMP]i also decreased in the pulmonary artery branches (from 550 +/- 15, control to 462 +/- 31 pmol mg-1 protein in CH vessels, n = 8, P < 0.01) and in the main pulmonary arteries (from 566 +/- 33, control to 370 +/- 25 pmol mg-1 protein in CH vessels, n = 8, P < 0.001). No changes in either [cyclic AMP]i or [cyclic GMP]i were observed in the resistance arteries. 6. The results suggest that the increased vasoconstrictor response to 5-HT in CH rat pulmonary arteries is due to an increase in 5-HT2A-receptor mediated contraction combined with an increase in r5-HT1B-like receptor-mediated contraction. An increase in vascular tone and decreased levels of [cyclic GMP]i in the large pulmonary arteries may contribute to the observed increase in activity of r5-HT1B-like receptor     

Evidence for two different P2X-receptors mediating vasoconstriction of Ap5A and Ap6A in the isolated perfused rat kidney.

The activation of various P2-receptor subtypes in rat renal vasculature by P1, P5-diadenosine pentaphosphate (ApsA) and P1, P6-diadenosine hexaphosphate (Ap6A) were studied by measuring their effects on perfusion pressure during continuous perfusion in a rat isolated perfused kidney. Permanent perfusion with Ap5A and Ap6A elicited both a transient and sustained vasoconstriction with both vasoconstrictions to be different: the transient vasoconstriction can be elicited with concentrations > or = 10 nM, whereas the sustained vasoconstriction is observed with concentrations > or = 1 nM. ApsA and Ap6A act via the same receptors as alpha,beta-methylene ATP (alpha,beta-meATP). The rank order of potency for transient vasconstriction was alpha,beta-meATP = ApsA>Ap6A>B,gamma-meATP, and for sustained vasoconstriction alpha,beta-meATP = Ap5A > beta,gamma-meATP > or = Ap6A. Suramin, a non-selective P2-receptor antagonist, and pyridoxal-phosphate-6-azophenyl-2;4-disulphonic acid (PPADS) a highly selective P2X-receptor antagonist antagonized both the transient and the sustained vasoconstriction. Taken together the results of the agonist profile of Ap5A and Ap6A and comparing its findings to literature it can be demonstrated that the transient but not the sustained vasoconstriction is mediated via the P2X1-receptor which is present in rat renal vasculature. It is demonstrated that the agonist profile of the sustained vasoconstriction induced by ApsA and Ap6A does not fit to any currently known P2X- or P2Y-receptor subtype. We conclude a yet unidentified P2X-receptor or chimeric P2X-receptor may contribute to the effects on rat renal vasculature produced by Ap5A and Ap6A and which may play an important role in glomerular perfusion pressure and blood pressure control.     

Agonists for neuropeptide Y receptors Y1 and Y5 stimulate different phases of feeding in guinea pigs

1. The stimulatory effect of neuropeptide Y (NPY) on food intake is well established but the roles of the receptor subtypes Y(1) and Y(5) have been difficult to define. We have studied the effects of two novel Y(1)-preferring and two Y(5)-preferring agonists on feeding in guinea pigs. 2. The Y(1)-preferring receptor agonists [Arg(6),Pro(34)]pNPY and [Phe(7),Pro(34)]pNPY had high affinity for the Y(1) receptor (K(i) values 0.07 and 0.04 nM, respectively) and nanomolar affinity for the Y(5) receptor. Administration of either compound into the third brain ventricle increased food intake equally to NPY. 3. The Y(5) agonist [Ala(31),Aib(32)]pNPY displayed a moderate affinity for the Y(5) receptor (K(i) 7.42 nM) and a low affinity for Y(1) (K(i) 1.7 micro M). This compound had only a modest effect on feeding. 4. The other Y(5)-preferring peptide [cPP(1-7),NPY(19-23),Ala(31),Aib(32),Gln(34)]hPP had a higher affinity at the Y(5) receptor (K(i) 1.32 nM) and also at the Y(1) receptor (K(i) 85 nM). It potently stimulated feeding: the food consumption after administration of this peptide was two-fold compared to NPY. 5. Our results support the view that both the receptor subtypes Y(1) and Y(5) are involved in the stimulation of feeding. As the action profiles of the Y(1) and Y(5) agonists on feeding parameters were different, it seems that they influence different phases of eating.     

Modulatory effect of neuropeptide Y on acetylcholine-induced oedema and vasoconstriction in isolated perfused lungs of rabbit.

1. The modulatory role of neuropeptide Y (NPY) on pulmonary oedema induced by acetylcholine and capsaicin was investigated. The effects of NPY on the haemodynamic response to acetylcholine, phenylephrine and substance P were also investigated. 2. Isolated, ventilated, exsanguinated lungs of the rabbit were perfused with a constant flow of recirculating blood-free perfusate. The double/arterial/venous occlusion method was used to partition the total pressure gradient (delta Pt) into four components: the arterial gradient (delta Pa), the pre- and post-capillary gradients (respectively delta Pa' and delta Pv') and the venous pressure gradient (delta Pv). Endothelial permeability was evaluated by measuring the capillary filtration coefficient (Kf,c). 3. Acetylcholine (10(-8) M to 10(-4) M) and substance P (SP, 10(-10) M to 10(-6) M) induced a concentration-dependent increase in the Kf,c. Capsaicin (10(-4) M) and 5-hydroxytryptamine (5-HT) (10(-4) M) also increased this parameter. NPY (10(-8) M) completely inhibited the effects of acetylcholine and capsaicin on the Kf,c, without preventing the effects of substance P and 5-HT. 4. Acetylcholine induced concentration-dependent vasoconstriction in the precapillary segment. The effect was inhibited by NPY and aspirin, an inhibitor of cyclo-oxygenase, while ketanserin, a 5-HT2 receptor antagonist, and SR140333, a new NK1 antagonist, had no protective effect. Phenylephrine increased delta Pa at high concentration, an effect also inhibited by NPY and aspirin. Substance P had no significant haemodynamic effect. When injected together with NPY, substance P (10(-6) M) induced a significant increase in the total pressure gradient.(ABSTRACT TRUNCATED AT 250 WORDS)     

Ligand internalization by cloned neuropeptide Y Y5 receptors excludes Y2 and Y4 receptor-selective peptides

In human embryonic kidney-293 (HEK-293) cells, the cloned human neuropeptide Y Y5 receptor saturably internalized agonists, with the rank order of neuropeptide Y-(19-23)-[Gly1,Ser3,Gln4,Thr6,Ala31,Aib32,Gln34]human pancreatic polypeptide (neuropeptide Y-Aib-pancreatic polypeptide)>human neuropeptide Y>porcine peptide YY>[Pro34]human peptide YY>[Leu31,Pro34]human peptide YY>human peptide YY-(3-36). Human pancreatic polypeptide competed [125I]neuropeptide Y binding and internalization in neuropeptide Y Y5 receptor-expressing cells, but itself showed no internalization. The internalization was strongly dependent on temperature. The surface binding, and especially the internalization, of human neuropeptide Y were highly sensitive to the clathrin network inhibitor phenylarsine oxide, and to the cholesterol-complexing antibiotic filipin III. The internalized ligands were present in particles corresponding to secondary endosomes in Percoll gradients, but especially in particles banding with the acid hexosaminidase lysosomal marker. At any temperature tested, internalization of the neuropeptide Y Y5 receptor driven by human neuropeptide Y in HEK-293 cells was much slower than the internalization of the neuropeptide Y Y1 receptor reported in the same cells, or in Chinese hamster ovary (CHO) cells. The neuropeptide Y Y5 receptor subtype could be the metabotropic receptor responding to protracted challenges by neuropeptide Y-like peptides, and its density could be little sensitive to concentration of extracellular agonists.     

G-Protein coupling and signalling of Y1-like neuropeptide Y receptors in SK-N-MC cells

Summary We have studied [¹²⁵I]neuropeptide Y-binding sites and neuropeptide Y-mediated second messenger responses in human SK-N-MC neuroblastoma cells with special reference to the role of G-proteins. Neuropeptide Y stimulated two second messenger responses in SK-N-MC cells, inhibition of cAMP accumulation and mobilization of Ca²⁺ from intracellular stores. Both effects were completely abolished by pretreatment with pertussis toxin. Binding of [¹²⁵I]neuropeptide Y to intact cells or SK-N-MC cell membranes was rapid, reversible, characterized by high affinity and low capacity, and had pharmacological characteristics of a homogeneous population of Y₁-like neuropeptide Y receptors. In permeabilized cells, [¹²⁵I] neuropeptide Y binding was inhibited by GTP S in a concentration-dependent manner. Saturation experiments in the absence and presence of GTPS demonstrated a reduction in the number of high-affinity [¹²⁵I]neuropeptide Y-binding sites without a decrease in affinity of the remaining sites. Pretreatment of intact cells with ertussis toxin completely abolished the inhibition of [¹²⁵I]neuropeptide Y binding by GTPS. Moreover, pertussis toxin treatment reduced the number of high-affinity [¹²⁵I]neuropeptide Y binding sites. We conclude that the agonist ligand [¹²⁵I]neuropeptide Y identifies functional neuropeptide Y receptors in SK-N-MC cells; however, the number of specific [¹²⁵I]neuropeptide Y-binding sites may not necessarily reflect the number of neuropeptide Y receptors, because the former is affected by the functional state of cellular G-proteins.