Neuropeptide Y-family receptors Y6 and Y7 in chicken. Cloning, pharmacological characterization, tissue distribution and conserved synteny with human chromosome region

The peptides of the neuropeptide Y (NPY) family exert their functions, including regulation of appetite and circadian rhythm, by binding to G-protein coupled receptors. Mammals have five subtypes, named Y1, Y2, Y4, Y5 and Y6, and recently Y7 has been discovered in fish and amphibians. In chicken we have previously characterized the first four subtypes and here we describe Y6 and Y7. The genes for Y6 and Y7 are located 1 megabase apart on chromosome 13, which displays conserved synteny with human chromosome 5 that harbours the Y6 gene. The porcine PYY radioligand bound the chicken Y6 receptor with a K(d) of 0.80 +/- 0.36 nm. No functional coupling was demonstrated. The Y6 mRNA is expressed in hypothalamus, gastrointestinal tract and adipose tissue. Porcine PYY bound chicken Y7 with a K(d) of 0.14 +/- 0.01 nm (mean +/- SEM), whereas chicken PYY surprisingly had a much lower affinity, with a Ki of 41 nm, perhaps as a result of its additional amino acid at the N terminus. Truncated peptide fragments had greatly reduced affinity for Y7, in agreement with its closest relative, Y2, in chicken and fish, but in contrast to Y2 in mammals. This suggests that in mammals Y2 has only recently acquired the ability to bind truncated PYY. Chicken Y7 has a much more restricted tissue distribution than other subtypes and was only detected in adrenal gland. Y7 seems to have been lost in mammals. The physiological roles of Y6 and Y7 remain to be identified, but our phylogenetic and chromosomal analyses support the ancient origin of these Y receptor genes by chromosome duplications in an early (pregnathostome) vertebrate ancestor.     

Characterization of NPY receptor subtypes Y2 and Y7 in rainbow trout Oncorhynchus mykiss

We report the cloning and pharmacological characterization of two neuropeptide Y (NPY) receptor subtypes, Y2 and Y7, in rainbow trout (Oncorhynchus mykiss). These subtypes are approximately 50% identical to each other and belong to the Y2 subfamily of NPY receptors. The binding properties of the receptors were investigated after expression in human HEK-293 EBNA cells. Both receptors bound the three zebrafish peptides NPY, PYYa, and PYYb, as well as porcine NPY and PYY, with affinities in the nanomolar range that are similar to mammalian Y2. The affinity of the truncated porcine NPY fragments, NPY 13-36 and NPY 18-36 was markedly lower compared to mammalian and chicken Y2. This suggests that mammalian and chicken Y2 are unique among NPY receptors in their ability to bind truncated peptide fragments. The antagonist BIIE0246, developed for mammalian Y2, did not bind either of the two rainbow trout receptors. Our results support the proposed expansion of this gene family by duplications before the gnathostome radiation. They also reveal appreciable differences in the repertoire and characteristics of NPY receptors between fish and tetrapods stressing the importance of lineage-specific gene loss as well as sequence divergence after duplication.     

The cloned guinea pig neuropeptide Y receptor Y1 conforms to other mammalian Y1 receptors.

We have cloned the guinea pig neuropeptide Y (NPY) Y1 receptor and found it to be 92-93% identical to other cloned mammalian Y1 receptors. Porcine NPY and peptide YY (PYY) displayed affinities of 43 pM and 48 pM, respectively. NPY2-36 and NPY3-36 had 6- and 46-fold lower affinity, respectively, than intact NPY. Functional coupling was measured by using a microphysiometer. Human NPY and PYY were equipotent in causing extracellular acidification with EC50 values of 0.59 nM and 0.69 nM, respectively, whereas NPY2-36 and NPY3-36 were about 15-fold and 500-fold less potent, respectively, than NPY. The present study shows that the cloned guinea pig Y1 receptor is very similar to its orthologues in other mammals, both with respect to sequence and pharmacology. Thus, results from previous studies on guinea pig NPY receptors might imply the existence of an additional Y1-like receptor sensitive to B1BP3226.     

Chicken neuropeptide Y receptor Y2: structural and pharmacological differences to mammalian Y2(1)

Here we report the molecular cloning of the chicken (Gallus gallus) neuropeptide Y (NPY) receptor Y2, the first non-mammalian Y2 receptor. It displays 75-80% identity to mammalian Y2 and has a surprisingly divergent cytoplasmic tail. Expression of the receptor protein in a cell line showed that the receptor did not bind the mammalian Y2 selective antagonist BIIE0246. Furthermore, porcine [Leu(31), Pro(34)]NPY, which binds poorly to mammalian Y2, exhibited an unexpectedly high affinity for chicken Y2. In situ hybridisation revealed expression in the hippocampus. Thus, the chicken Y2 receptor exhibits substantial differences with regard to sequence and pharmacological profile in comparison to mammalian Y2 receptors, while the expression pattern in the central nervous system resembles that observed in mammals.     

Potent and selective tools to investigate neuropeptide Y receptors in the central and peripheral nervous systems: BIB03304 (Y1) and CGP71683A (Y5)

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 (IC50 of 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 microM) were unable to compete for a significant proportion of specific [125I]PYY3-36/Y2 sites. The purported Y2 antagonist T4[NPY33-36]4 competed against [125I]PYY3-36 binding sites with an affinity of 750 nM. These results were confirmed in HEK 293 cells transfected with either the rat Y1, Y2, Y4, or Y5 receptor cDNA. BIBO3304, but not CGP71683A, competed with high affinity for [125I][Leu31,Pro34]PYY binding sites in HEK 293 cells transfected with the rat Y1 receptor cDNA, whereas the reverse profile was observed upon transfection with the rat Y5 receptor cDNA. Additionally, both molecules were inactive at Y2 and Y4 receptor 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 Y1 bioassay (rabbit saphenous vein; pA2 value 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 Y1 and Y5 receptor subtypes, respectively, while the purported Y2 antagonist, T4[NPY33-36]4 possesses rather low affinity for this receptor.     

Cloning and characterization of a zebrafish Y2 receptor

The NPY receptors belong to the superfamily of G-protein coupled receptors and in mammals this family has five members, named Y1, Y2, Y4, Y5, and Y6. In bony fish, four receptors have been identified, named Ya, Yb, Yc and Y7. Yb and Y7 arose prior to the split between ray-fined fishes and tetrapods and have been lost in mammals. Yc appeared as a copy of Yb in teleost fishes. Ya may be an ortholog of Y4, but surprisingly no unambiguous receptor ortholog to any of the mammalian subtypes has yet been identified in bony fishes. Here we present the cloning and pharmacological characterization of a Y2 receptor in zebrafish, Danio rerio. To date, this is the first Y2 receptor outside mammals and birds that has been characterized pharmacologically. Phylogenetic analysis and synteny confirmed that this receptor is orthologous to mammalian Y2. We show that the receptor is pharmacologically most similar to chicken Y2 which leads to the conclusion that Y2 has acquired several novel characteristics in mammals. Y2 from zebrafish binds very poorly to the Y2-specific antagonist BIIE0246. Our pharmacological characterization supports our previous conclusions regarding the binding pocket of BIIE0246 in the human Y2 receptor.     

The novel neuropeptide Y Y(1) receptor antagonist J-104870: a potent feeding suppressant with oral bioavailability

Neuropeptide Y (NPY) is known to induce robust feeding through the action of NPY receptors in the hypothalamus. Among the subtypes of NPY receptors, Y(1) receptors may play a key role in feeding regulation. In the present study, we demonstrated that a novel Y(1) antagonist, J-104870, shows high selectivity and potency for the Y(1) receptor with an anorexigenic effect on NPY-mediated feeding. J-104870 displaced [(125)I]peptide YY (PYY) binding to cloned human and rat Y(1) receptors with K(i) values of 0.29 and 0.54 nM, respectively, and inhibited the NPY (10 nM)-induced increase in intracellular calcium levels (IC(50) = 3.2 nM) in cells expressing human Y(1) receptors. In contrast, J-104870 showed low affinities for human Y(2) (K(i) > 10 microM), Y(4) (K(i) > 10 microM), and Y(5) receptors (K(i) = 6 microM). In rat hypothalamic membranes, J-104870 also completely displaced the binding of [(125)I]1229U91, which is known to bind to the typical Y(1) receptor, with a high affinity (K(i) = 2.0 nM). Intracerebroventricular (ICV) injection of J-104870 (200 microg) significantly suppressed NPY (5 microg)-induced feeding in satiated Sprague-Dawley rats by 74%. Furthermore, ICV and oral administration of J-104870 (200 microg and 100 mg/kg, respectively) significantly suppressed spontaneous food intake in Zucker fatty rats. These findings suggested that J-104870 is a selective and potent nonpeptide Y(1) antagonist with oral bioavailability and brain penetrability. In addition, the anorexigenic effect of J-104870 clearly revealed the participation of the Y(1) receptor in NPY-mediated feeding regulation. The potent and orally active Y(1) antagonist J-104970 is a useful tool for elucidating the physiological roles of NPY in obesity.     

GPC receptors and not ligands decide the binding mode in neuropeptide Y multireceptor/multiligand system

Many G protein-coupled receptors belong to families of different receptor subtypes, which are recognized by a variety of distinct ligands. To study such a multireceptor/multiligand system, we investigated the Y-receptor family. This family consists of four G protein-coupled Y receptors in humans (hY 1R, hY 2R, hY 4R, and hY 5R) and is activated by the so-called NPY hormone family, which itself consists of three native peptide ligands named neuropeptide Y (NPY), pancreatic polypeptide (PP), and peptide YY (PYY). The hY 5R shows high affinity for all ligands, although for PP binding, the affinity is slightly decreased. As a rational explanation, we suggest that Tyr (27) is lost as a contact point between PP and the hY 5R in contrast to NPY or PYY. Furthermore, several important residues for ligand binding were identified by the first extensive mutagenesis study of the hY 5R. Using a complementary mutagenesis approach, we were able to discover a novel interaction point between hY 5R and NPY. The interaction between NPY(Arg (25)) and hY 5R(Asp (2.68)) as well as between NPY(Arg (33)) and hY 5R(Asp (6.59)) is maintained in the binding of PYY and PP to hY 5R but different to the PP-hY 4R and NPY-hY 1R contact points. Therefore, we provide evidence that the receptor subtype and not the pre-orientated conformation of the ligand at the membrane decides the binding mode. Furthermore, the first hY 5R model was set up on the basis of the crystal structure of bovine rhodopsin. We can show that most of the residues identified to be critical for ligand binding are located within the now postulated binding pocket.     

A Y2 receptor mimetic aptamer directed against neuropeptide Y.

Neuropeptide Y (NPY) is a 36-amino acid neuropeptide that exerts its activity by at least five different receptor subtypes that belong to the family of G-protein-coupled receptors. We isolated an aptamer directed against NPY from a nuclease-resistant RNA library. Mapping experiments with N-terminally, C-terminally, and centrally truncated analogues of NPY revealed that the aptamer recognizes the C terminus of NPY. Individual replacement of the four arginine residues at positions 19, 25, 33, and 35 by l-alanine showed that arginine 33 is essential for binding. The aptamer does not recognize pancreatic polypeptide, a highly homologous Y4 receptor-specific peptide of the gut. Furthermore, the affinity of the aptamer to the Y5 receptor-selective agonist [Ala(31),Aib(32)]NPY and the Y1/Y5 receptor-binding peptide [Leu(31),Pro(34)]NPY was considerably reduced, whereas Y2 receptor-specific NPY mutants were bound well by the aptamer. Accordingly, the NPY epitope was recognized by the Y2 receptor, and the aptamer was highly similar. This Y2 receptor mimicking effect was further confirmed by competition binding studies. Whereas the aptamer competed with the Y2 receptor for binding of [(3)H]NPY with high affinity, a low affinity displacement of [(3)H]NPY was observed at the Y1 and the Y5 receptors. Consequently, competition at the Y2 receptor occurred with a considerably lower K(i) value compared with the Y1 and Y5 receptors. These results indicate that the aptamer mimics the binding of NPY to the Y2 receptor more closely than to the Y1 and Y5 receptors.     

Y1 and Y2 receptors for neuropeptide Y

By using monoiodinated radioligands of both intact neuropeptide Y (NPY) and of a long C-terminal fragment, NPY13-36, two subtypes of binding sites, which differ in affinity and specificity, have been characterized. The Y1 type of binding site, characterized on a human neuroblastoma cell line, MC-IXC, and a rat pheochromocytoma cell line, PC-12, binds NPY with a dissociation constant (Kd) of a few nanomolar but does not bind NPY13-36. The Y2 type of binding site, characterized on porcine hippocampal membranes and on another human neuroblastoma cell line, SMS-MSN, is of higher affinity and binds both NPY and NPY13-36. None of the binding sites distinguish between NPY and the homologous peptide YY (PYY). It is concluded that NPY/PYY-binding sites occur in two subtypes which may represent two types of physiological receptors.     

Functional CCK-A and Y2 receptors in guinea pig esophagus

Effects of cholecystokinin octapeptide (CCK-8), peptide YY (PPY), neuropeptide Y (NPY) and their analogs on muscle contractions of esophageal strips were investigated. CCK-8 induced a tetrodotoxin and atropine-sensitive contraction. The relative potencies for CCK related peptides to induce contractions were CCK-8 > desulfated CCK-8 > gastrin-17-I. The CCK-A receptor antagonist L-364,718 was 300-fold more potent than the CCK-B receptor antagonist L-365,260 at inhibiting CCK-8-induced contraction. These indicate that neural CCK-A receptors mediate this contraction. PYY or NPY did not cause muscle contraction or inhibit muscle contraction induced by carbachol, endothelin-1 or KCl. However, both PYY and NPY concentration-dependently inhibited contraction induced by CCK-8. This inhibition was not affected by nitric oxide (NO) synthase inhibitors L-NMMA or L-NAME. The relative potencies of PYY related peptides to inhibit CCK-8 induced contraction were PYY > NPY > NPY13-36 > [Leu(31), Pro(34)]NPY > pancreatic polypeptide (PP). We conclude that CCK interacts with neural CCK-A receptors to cause esophageal muscle contraction. PYY and NPY interact with Y2 receptors to inhibit this CCK-induced muscle contraction by an effect not related to NO.     

Cloning,pharmacology, and distribution of the neuropeptide Y-receptor Yb in rainbow trout

This work describes the isolation and pharmacological characterization of a neuropeptide Y (NPY) receptor from rainbow trout (Oncorhynchus mykiss). The receptor exhibits approximately 45% amino acid sequence identity to mammalian Y1-subfamily receptors, Y1, Y4 and y6, a similar degree of identity as these subtypes display to one another. Because it displays highest sequence identity to zebrafish Yb (75%), we named it the trout Yb receptor. The receptor exhibits high binding affinity for zebrafish and human NPY and peptide YY (PYY) but not truncated forms of the peptides. Human pancreatic polypeptide (PP) also binds with high affinity. Y1 selective antagonists exhibit poor binding as is the case for Y2 and Y5 selective ligands. This binding profile supports membership in the Y1 subfamily. Sequence data also support this relationship suggesting that Yb is a fourth and separate member of the Y1 subfamily. NPY has a number of important physiological functions such as regulating food intake and reproduction. The expression of the receptor in the hypothalamus and telencephalon suggests a possible role in these processes. This and other receptors from this species have potential for improving aquaculture.     

Blockade of pancreatic polypeptide-sensitive neuropeptide Y (NPY) receptors by agonist peptides is prevented by modulators of sodium transport. Implications for receptor signaling and regulation

Ligand binding to rodent pancreatic polypeptide-responding neuropeptide Y (NPY) receptors (here termed PP/NPY receptors), or to cloned Y4 or Y5 receptors, is selectively inhibited by amiloride, peptide or alkylating modulators of sodium transport. The PP/NPY and Y4 receptors are also selectively blocked by human or rat pancreatic polypeptide (PP) and the blocking peptides are not dissociated by high concentrations of alkali chlorides (which restore most of the binding of subtype-selective agonists to Y1 and Y2 sites). The PP/NPY receptors could also be blocked by NPY and related full-length peptides, including Y1-selective agonists (IC50 300-400 pM). The cloned Y(4) receptors from three species are much less sensitive to NPY or PYY. The sensitivity of both the PP/NPY sites and the Y(4) sites to Y2-selective peptides is quite low. The ligand attachment to PP/NPY sites is also very sensitive to peptidic Y1 antagonist ((Cys31,NVal34NPY27-36))2, which however blocks these sites at much higher molarities. Blockade of PP/NPY and Y4 sites by agonist peptides can be largely prevented by N5-substituted amiloride modulators of Na+ transport, and by RFamide NRNFLRF.NH2, but not by Ca2+ channel blockers, or by inhibitors of K+ transport. Protection of both PP/NPY and Y4 sites against blockade by human or rat pancreatic polypeptide is also afforded by short N-terminally truncated NPY-related peptides. The above results are consistent with a stringent and selective activity regulation for rabbit PP/NPY receptor(s) that may serve to differentiate agonists and constrain signaling, and could involve transporter-like interactants.     

Y-receptor affinity modulation by the design of pancreatic polypeptide/neuropeptide Y chimera led to Y(5)-receptor ligands with picomolar affinity

Neuropeptide Y (NPY) and pancreatic polypeptide (PP) bind to the Y-receptors with very different affinities: NPY has high affinity for the receptors Y(1), Y(2) and Y(5), while PP binds only to Y(4)-receptor with picomolar affinity. By exchanging of specific amino acid positions between the two peptides, we developed 38 full-length PP/NPY chimeras with binding properties that are completely different from those of the two native ligands. Pig NPY (pNPY) analogs containing the segment 19-23 from human PP (hPP) bound to the Y-receptors with much lower affinity than NPY itself. The affinity of the hPP analog containing the pNPY segments 1-7 and 19-23 was comparable to that of pNPY at the Y(1)- and Y(5)-receptor subtypes, and to that of hPP at the Y(4)-receptor. Furthermore, the presence of the segments 1-7 from chicken PP (cPP) and 19-23 from pNPY within the hPP sequence led to a ligand with IC(50) of 40 pM at the Y(5)-receptor. This is the most potent Y(5)-receptor ligand known so far, with 15-fold higher affinity than NPY.     

Feeding after fourth ventricular administration of neuropeptide Y receptor agonists in rats

Neuropeptide Y (NPY) and peptide YY (PYY) stimulate food intake after injection into the fourth cerebral ventricle, suggesting that NPY receptors in the hindbrain are targets for the stimulatory effect of these peptides on food intake. However, the NPY/PYY receptor subtype mediating the feeding response in the hindbrain is not known. To approach to this question we compared dose-effect of several NPY receptor agonists to stimulate food intake in freely-feeding rats 60- and 120-min after injection into the fourth cerebral ventricle. At the 120-min time point, PYY was 2- to 10-times as potent as NPY over the dose-response range and stimulated twice the total intake at the maximally effective dose (2-fold greater efficacy). NPY was 2-times as potent as the Y1, Y5 receptor agonist, [Leu(31)Pro(34)]NPY but acted with comparable efficacy. The Y5-, Y2-differentiating receptor agonist, NPY 2-36, was comparable in potency to PYY at low doses but equal in efficacy NPY and [Leu(31)Pro(34)]NPY. The Y2 receptor agonist, NPY 13-36, produced only a marginal effect on total food intake. The profile of agonist potency after fourth cerebral ventricle administration is similar to the profile obtained when these or related agonists are injected in the region of the hypothalamus. Agonists at both Y1 and Y5 receptors stimulated food intake with a rank order of potency that does not conclusively favor the exclusive involvement of a single known NPY receptor subtype. Thus it is possible that the ingestive effects of NPY and PYY are mediated by multiple or novel receptor subtypes in the hindbrain. And the relatively greater potency and efficacy of PYY raises the possibility that a novel PYY-preferring receptor in the hindbrain is involved in the stimulation of food intake.     

Receptor subtype-specific docking of Asp6.59 with C-terminal arginine residues in Y receptor ligands

Y receptors (YRs) are G protein-coupled receptors whose Y(1)R, Y(2)R, and Y(5)R subtypes preferentially bind neuropeptide Y (NPY) and peptide YY, whereas mammalian Y(4)Rs show a higher affinity for pancreatic polypeptide (PP). Comparison of YR orthologs and paralogs revealed Asp(6.59) to be fully conserved throughout all of the YRs reported so far. By replacing this conserved aspartic acid residue with alanine, asparagine, glutamate, and arginine, we now show that this residue plays a crucial role in binding and signal transduction of NPY/PP at all YRs. Sensitivity to distinct replacements is, however, receptor subtype-specific. Next, we performed a complementary mutagenesis approach to identify the contact site of the ligand. Surprisingly, this conserved residue interacts with two different ligand arginine residues by ionic interactions; although in Y(2)R and Y(5)R, Arg(33) is the binding partner of Asp(6.59), in Y(1)R and Y(4)R, Arg(35) of human PP and NPY interacts with Asp(6.59). Furthermore, Arg(25) of PP and NPY is involved in ligand binding only at Y(2)R and Y(5)R. This suggests significant differences in the docking of YR ligands between Y(1/4)R and Y(2/5)R and provides new insights into the molecular binding mode of peptide agonists at GPCRs. Furthermore, the proposed model of a subtype-specific binding mode is in agreement with the evolution of YRs.     

Neuropeptide Y induced modulation of dopamine synthesis in the striatum

The purpose of the present study was to determine whether the activation of NPY receptors alters catecholamines (CA) synthesis in the central nervous system and, if so, to identify the NPY receptor subtype(s) mediating this effect. Tyrosine hydroxylation, the rate-limiting step in CA synthesis, was assessed by measuring the accumulation of 3,4-dihydroxyphenyalanine (DOPA) by high pressure liquid chromatography coupled to electrochemical detection (HPLC-EC) in rat striatal dices following incubation of the tissue with the aromatic L-amino acid decarboxylase inhibitor m-hydroxybenzyl hydrazine (NSD 1015). Treatment with NSD 1015 resulted in an increase in DOPA accumulation that was increased even further following depolarization with a high potassium (KCl) buffer. PYY13-36 and NPY13-36 both produced a significant enhancement of the KCl-induced increase in DOPA accumulation. The effect of PYY13-36 was completely attenuated by the selective Y2 antagonist BIIE0246 suggesting that activation of Y2 receptors enhanced the synthesis of dopamine. In contrast to the effects of NPY13-36 and PYY13-36; NPY, PYY and PYY3-36 all produced a significant attenuation of the KCl-induced increase in DOPA accumulation. The Y1 antagonist BIBO3304 and the Y5-antagonist CGP71683A, both prevented the inhibitory effect of NPY converting it to a stimulatory effect. The enhancement of the NPY induced increase in DOPA accumulation observed by BIBO3304 was attenuated when examined in the presence of the Y2 antagonist BIIE0246. These results suggest that activation of NPY receptors can modulate the synthesis of CA in the rat striatum. The Y1 and Y5 receptor appear to be involved in attenuation, while Y2 receptors are involved in the stimulation of synthesis.     

Multiple Y receptors mediate pancreatic polypeptide responses in mouse colon mucosa

A functional study has been performed to characterise the Y receptors responsible for NPY, PYY and PP-stimulated responses in mouse colonic mucosal preparations. Electrogenic ion secretion was stimulated with VIP following which NPY, PYY and PP analogues were, to varying degrees, inhibitory. PYY(3-36), hPP, Gln(23)hPP and rPP were effective but less potent than full length PYY, NPY or their Pro(34)-substituted analogues, while the Y(5) agonist Ala(31), Aib(32)hNPY was the least active peptide tested. The Y(1) antagonists, BIBP3226 and BIBO3304 virtually abolished Pro(34)PYY and PYY responses while PYY(3-36) responses were selectively inhibited by the Y(2) antagonist, BIIE0246. A combination of BIBO3304 and BIIE0246 also partially attenuated hPP responses, leaving residual effects that were most probably Y(4)-mediated. Thus we conclude that Y(1), Y(2) and Y(4) receptors attenuate ion secretion in mouse colon.     

PYY preference is a common characteristic of neuropeptide Y receptors expressed in human, rat, and mouse gastrointestinal epithelia

This investigation describes the relative potencies of four peptide agonists, namely, peptide YY (PYY), [Leu3l,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 Y1 receptor (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 Y1 receptors) 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 Y1 receptor 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.