Molecular Cloning and Pharmacological Characterization of the Canine B1 and B2 Bradykinin Receptors

2001 ◽  
Vol 382 (1) ◽  
pp. 123-129 ◽  
Author(s):  
J. Fred Hess ◽  
Patricia J. Hey ◽  
Tsing-Bau Chen ◽  
Julie OBrien ◽  
Stacey S. Omalley ◽  
...  
Keyword(s):  
Amino Acids ◽  
Cdna Sequence ◽  
Chinese Hamster ◽  
Physiological Role ◽  
Receptor Subtype ◽  
Receptor Subtypes ◽  
Pharmacological Properties ◽  
Bradykinin Receptors ◽  
B1 Receptor ◽  
Bradykinin Receptor

Abstract The dog is a valuable animal model in the study of the physiological role of both the B1 and B2 bradykinin receptors. To more thoroughly characterize the pharmacological properties of the canine kinin receptors we isolated the cDNA sequence encoding the B1 and B2 bradykinin receptor subtypes and overexpressed them in Chinese hamster ovary (CHO) cells. The cDNA sequence of the canine B1 bradykinin receptor encodes a protein comprised of 350 amino acids that is 76% identical to the human B1 bradykinin receptor. The cDNA sequence of the canine B2 bradykinin receptor encodes a protein of 392 amino acids that is 81% identical to the human B2 bradykinin receptor. The amino acid sequence of the canine B1 and B2 receptors are 35% identical. Pharmacological studies of the cloned receptors revealed that the agonist affinity of the dog B1 receptor is similar to the rodent B1 receptors, and differs from the human form in that there is no preference for the presence of the Nterminal Lys residue of [desArg10]Lysbradykinin. Significantly, the B1 receptor antagonist [desArg9,Leu8]BK behaves as partial agonist on the cloned dog B1 receptor. The dog B2 receptor exhibits the classical pharmacological properties of this receptor subtype.

scholarly journals Characterization of β-adrenergic receptors in bovine intramuscular and subcutaneous adipose tissue: comparison of lubabegron fumarate with β-adrenergic receptor agonists and antagonists

2021 ◽  
Vol 99 (8) ◽  
Author(s):  
Jinhee H Hwang ◽  
Michael E Spurlock ◽  
John C Kube ◽  
Xiang Z Li ◽  
Stephen B Smith
Keyword(s):  
Adipose Tissue ◽  
Adrenergic Receptor ◽  
Quantitative Polymerase Chain Reaction ◽  
Animal Health ◽  
Chinese Hamster ◽  
Ovary Cell ◽  
Receptor Subtype ◽  
Receptor Subtypes ◽  
Mrna Levels ◽  
Adipose Tissues

Abstract Chinese hamster ovary cell constructs expressing either the β 1-, β 2- or β 3-adrenergic receptor (AR) were used to determine whether a novel β-AR modulator, lubabegron fumarate (LUB; Experior, Elanco Animal Health) might exert greater potency for a specific β-AR subtype. EC50 values calculated based on cAMP accumulation in dose response curves indicate that LUB is highly selective for the β 3-AR subtype, with an EC50 of 6 × 10–9 M, with no detectible agonistic activity at the β 2-AR. We hypothesized that the accumulation of lipolytic markers would reflect the agonist activity at each of the β-receptor subtypes of the specific ligand; additionally, there would be differences in receptor subtype expression in subcutaneous (s.c.) and intrmuscular (i.m.) adipose tissues. Total RNA was extracted from adipose tissue samples and relative mRNA levels for β 1-, β2-, and β 3-AR were measured using real-time quantitative polymerase chain reaction. Fresh s.c. and i.m. adipose tissue explants were incubated with isoproterenol hydrochloride (ISO; β-AR pan-agonist), dobutamine hydrochloride (DOB; specific β 1-AA), salbutamol sulfate (SAL; specific β 2-AA), ractopamine hydrochloride (RAC), zilpaterol hydrochloride (ZIL), BRL-37344 (specific β 3-agonist), or LUB for 30 min following preincubation with theophylline (inhibitor of phosphodiesterase). Relative mRNA amounts for β 1-, β 2-, and β 3-AR were greater (P < 0.05) in s.c. than in i.m. adipose tissue. The most abundant β-AR mRNA in both adipose tissues was the β 2-AR (P < 0.05), with the β 1- and β 3-AR subtypes being minimally expressed in i.m. adipose tissue. ISO, RH, and ZH stimulated the release of glycerol and nonesterified fatty acid (NEFA) from s.c. adipose tissue, but these β-AR ligands did not alter concentrations of these lipolytic markers in i.m. adipose tissue. LUB did not affect glycerol or NEFA concentrations in s.c. or i.m. adipose tissue, but attenuated (P < 0.05) the accumulation of cAMP mediated by the β 1- and β 2-AR ligands DOB and SAL in s.c. adipose tissue. Collectively, these data indicate that bovine i.m. adipose tissue is less responsive than s.c. adipose tissue to β-adrenergic ligands, especially those that are agonists at the β 1- and β3-receptor subtypes. The minimal mRNA expression of the β 1- and β 3 subtypes in i.m. adipose tissue likely limits the response potential to agonists for these β-AR subtypes.

scholarly journals Cloning and functional expression of the canine anaphylatoxin C5a receptor. Evidence for high interspecies variability

1992 ◽  
Vol 288 (3) ◽  
pp. 911-917 ◽  
Author(s):  
J J Perret ◽  
E Raspe ◽  
G Vassart ◽  
M Parmentier
Keyword(s):  
Chinese Hamster Ovary Cells ◽  
Single Gene ◽  
Mammalian Species ◽  
Chinese Hamster ◽  
Orphan Receptor ◽  
Free Calcium ◽  
Receptor Subtype ◽  
Receptor Subtypes ◽  
Binding Studies ◽  
C5a Receptor

A cDNA clone, DTJP03, encoding an orphan receptor, was isolated from a canine thyroid library, and found to exhibit 68.6% amino-acid identity with the recently described human C5a receptor. This relatively low similarity first suggested that DTJP03 encoded either a C5a receptor subtype, or the presumably related C3a receptor. Binding studies performed on membranes from COS-7 cells expressing the recombinant receptor demonstrated that DTJP03 encoded a high-affinity C5a receptor, with a Kd of 1.2 nM. C3a was unable to compete for C5a binding. Intracellular free calcium concentrations were measured by Quin-2 fluorescence assays in Chinese hamster ovary cells stably transfected with the canine C5a receptor. C5a addition elicited an increase in the intracellular calcium concentration. Extracellular EGTA partially prevented this response, suggesting that activation of the C5a receptor promotes both the release of calcium from intracellular stores, and the influx of extracellular calcium. Genes encoding C5a-receptor subtypes were subsequently searched for by PCR in genomic DNA from human, canine, rat and bovine sources. The result was the amplification of a single gene fragment from each species, with about 70% identity between any two of them. The canine C5a receptor has therefore to be considered as orthologous to the human C5a receptor described previously. The low similarity between C5a receptors from different mammalian species is quite unusual for a G-protein-coupled receptor.

Different actions of secretin and Gly-extended secretin predict secretin receptor subtypes

2001 ◽  
Vol 280 (1) ◽  
pp. G88-G94 ◽  
Author(s):  
Travis E. Solomon ◽  
Gabor Varga ◽  
Ning Zeng ◽  
S. Vincent Wu ◽  
John H. Walsh ◽  
...  
Keyword(s):  
Acid Secretion ◽  
Gastric Acid Secretion ◽  
Gastric Acid ◽  
Chinese Hamster Ovary Cells ◽  
Chinese Hamster ◽  
Receptor Subtype ◽  
Receptor Subtypes ◽  
Secretin Receptor ◽  
D Cells

Only one secretin receptor has been cloned and its properties characterized in native and transfected cells. To test the hypothesis that stimulatory and inhibitory effects of secretin are mediated by different secretin receptor subtypes, pancreatic and gastric secretory responses to secretin and secretin-Gly were determined in rats. Pancreatic fluid secretion was increased equipotently by secretin and secretin-Gly, but secretin was markedly more potent for inhibition of basal and gastrin-induced acid secretion. In Chinese hamster ovary cells stably transfected with the rat secretin receptor, secretin and secretin-Gly equipotently displaced125I-labeled secretin (IC50 values 5.3 ± 0.5 and 6.4 ± 0.6 nM, respectively). Secretin, but not secretin-Gly, caused release of somatostatin from rat gastric mucosal D cells. Thus the equipotent actions of secretin and secretin-Gly on pancreatic secretion appear to result from equal binding and activation of the pancreatic secretin receptor. Conversely, secretin more potently inhibited gastric acid secretion in vivo, and only secretin released somatostatin from D cells in vitro. These results support the existence of a secretin receptor subtype mediating inhibition of gastric acid secretion that is distinct from the previously characterized pancreatic secretin receptor.

Bradykinin regulation of airway submucosal gland secretion: role of bradykinin receptor subtype

1996 ◽  
Vol 270 (6) ◽  
pp. L907-L913 ◽  
Author(s):  
M. Nagaki ◽  
S. Shimura ◽  
T. Irokawa ◽  
T. Sasaki ◽  
T. Oshiro ◽  
...  
Keyword(s):  
Receptor Antagonist ◽  
Acinar Cells ◽  
Gland Secretion ◽  
Receptor Subtypes ◽  
Dependent Manner ◽  
Submucosal Gland ◽  
Patch Clamp Technique ◽  
B1 Receptor ◽  
Bradykinin Receptor

To clarify the role of bradykinin receptor subtypes, we examined the effect of bradykinin on feline tracheal and human airway submucosal gland secretion using an isolated gland preparation. Bradykinin induced a significant increase in [3H]glycoconjugate secretion in a dose-dependent manner from isolated glands, which was significantly inhibited by D-Arg-(Hyp3, Thi5,8, D-Phe7)-bradykinin (the B2-receptor antagonist), whereas Des-Arg9-(Leu8)-bradykinin (B1-receptor antagonist) or indomethacin did not significantly alter it. Nitric oxide synthase inhibitor (nitro-L-arginine methyl ester) caused a significant inhibition of bradykinin-induced glycoconjugate secretion, which was reversed by the addition of L-arginine. Bradykinin evoked bidirectional current responses, and an initial inward current (Cl- current) was followed by an outward current (K+ current) of the acinar cells in a whole cell configuration by patch-clamp technique. Bradykinin induced an immediate increase in intracellular calcium concentration ([Ca2+]i) of the acinar cells followed by a prolonged plateau, and Ca2+ removal resulted in an initial increase alone. [Ca2+]i rise was significantly inhibited by the B2-receptor antagonist, whereas the B1-receptor antagonist did not significantly alter it. These findings suggest that B2-receptor stimulation and the resultant [Ca2+]i rise induced both mucus glycoprotein and electrolyte secretions, involving NO formation in airway submucosal gland cells.

Demonstration of a common DPhe7 to DNal(2')7 peptide ligand antagonist switch for the melanocortin-3 and melanocortin-4 receptors identifies systematic mischaracterization of the pharmacological properties of melanocortin peptides

2022 ◽  
Author(s):  
Luis E. Gimenez ◽  
Terry A. Noblin ◽  
Savannah Y Williams ◽  
Satarupa Mullick Bagchi ◽  
Ren-Lei Ji ◽  
...  
Keyword(s):  
Cell Lines ◽  
Melanocortin Receptor ◽  
Hek293 Cells ◽  
Peptide Ligand ◽  
Receptor Subtype ◽  
Receptor Subtypes ◽  
Pharmacological Properties ◽  
Pharmacological Characterization ◽  
Melanocortin Peptides ◽  
The University

Melanocortin peptides containing a D-naphthylalanine residue in position 7 (DNal(2')7), reported as melanocortin-3 receptor (MC3R) subtype-specific agonists in two separate publications, were found to lack significant MC3R agonist activity. The cell lines used at the University of Arizona for pharmacological characterization of these peptides, consisting of HEK293 cells stably transfected with human melanocortin receptor subtypes MC1R, MC3R, MC4R, or MC5R, were then obtained and characterized by quantitative PCR. While the MC1R cell line correctly expressed only the hMCR1, the three other cell lines were mischaracterized with regard to receptor subtype expression. Demonstration that a D-naphthylalanine residue in position 7, irrespective of the melanocortin peptide template, results primarily in antagonism of the MC3R and MC4R, then allowed us to search the published literature for additional errors. The erroneously characterized DNal(2')7-containing peptides date back to 2003; thus, our analysis suggests that systematic mischaracterization of the pharmacological properties of melanocortin peptides occurred.

scholarly journals GRKs as Modulators of Neurotransmitter Receptors

Cells ◽  
2020 ◽  
Vol 10 (1) ◽  
pp. 52
Author(s):  
Eugenia V. Gurevich ◽  
Vsevolod V. Gurevich
Keyword(s):  
G Protein ◽  
General Model ◽  
G Protein Coupled Receptors ◽  
Receptor Internalization ◽  
Neurotransmitter Receptors ◽  
Receptor Subtype ◽  
Receptor Subtypes ◽  
Receptor Kinase ◽  
G Protein Coupled ◽  
Homologous Desensitization

Many receptors for neurotransmitters, such as dopamine, norepinephrine, acetylcholine, and neuropeptides, belong to the superfamily of G protein-coupled receptors (GPCRs). A general model posits that GPCRs undergo two-step homologous desensitization: the active receptor is phosphorylated by kinases of the G protein-coupled receptor kinase (GRK) family, whereupon arrestin proteins specifically bind active phosphorylated receptors, shutting down G protein-mediated signaling, facilitating receptor internalization, and initiating distinct signaling pathways via arrestin-based scaffolding. Here, we review the mechanisms of GRK-dependent regulation of neurotransmitter receptors, focusing on the diverse modes of GRK-mediated phosphorylation of receptor subtypes. The immediate signaling consequences of GRK-mediated receptor phosphorylation, such as arrestin recruitment, desensitization, and internalization/resensitization, are equally diverse, depending not only on the receptor subtype but also on phosphorylation by GRKs of select receptor residues. We discuss the signaling outcome as well as the biological and behavioral consequences of the GRK-dependent phosphorylation of neurotransmitter receptors where known.

scholarly journals A new 68Ga-labeled somatostatin analog containing two iodo-amino acids for dual somatostatin receptor subtype 2 and 5 targeting

2020 ◽  
Vol 10 (1) ◽  
Author(s):  
Rosalba Mansi ◽  
Karim Abid ◽  
Guillaume P. Nicolas ◽  
Luigi Del Pozzo ◽  
Eric Grouzmann ◽  
...  
Keyword(s):  
Amino Acids ◽  
Somatostatin Receptor ◽  
Somatostatin Analog ◽  
Receptor Subtype ◽  
Somatostatin Receptor Subtype 2

scholarly journals Transmitter and ion channel profiles of neurons in the primate abducens and trochlear nuclei

2021 ◽  
Author(s):  
Ümit Suat Mayadali ◽  
Jérome Fleuriet ◽  
Michael Mustari ◽  
Hans Straka ◽  
Anja Kerstin Ellen Horn
Keyword(s):  
Eye Movements ◽  
Ion Channel ◽  
Membrane Properties ◽  
Receptor Subtype ◽  
Receptor Subtypes ◽  
Voltage Gated ◽  
Calcium Ion Channels ◽  
Cell Type Specific ◽  
Transmitter Receptors ◽  
Intrinsic Membrane Properties

AbstractExtraocular motoneurons initiate dynamically different eye movements, including saccades, smooth pursuit and vestibulo-ocular reflexes. These motoneurons subdivide into two main types based on the structure of the neuro-muscular interface: motoneurons of singly-innervated (SIF), and motoneurons of multiply-innervated muscle fibers (MIF). SIF motoneurons are thought to provoke strong and brief/fast muscle contractions, whereas MIF motoneurons initiate prolonged, slow contractions. While relevant for adequate functionality, transmitter and ion channel profiles associated with the morpho-physiological differences between these motoneuron types, have not been elucidated so far. This prompted us to investigate the expression of voltage-gated potassium, sodium and calcium ion channels (Kv1.1, Kv3.1b, Nav1.6, Cav3.1–3.3, KCC2), the transmitter profiles of their presynaptic terminals (vGlut1 and 2, GlyT2 and GAD) and transmitter receptors (GluR2/3, NMDAR1, GlyR1α) using immunohistochemical analyses of abducens and trochlear motoneurons and of abducens internuclear neurons (INTs) in macaque monkeys. The main findings were: (1) MIF and SIF motoneurons express unique voltage-gated ion channel profiles, respectively, likely accounting for differences in intrinsic membrane properties. (2) Presynaptic glutamatergic synapses utilize vGlut2, but not vGlut1. (3) Trochlear motoneurons receive GABAergic inputs, abducens neurons receive both GABAergic and glycinergic inputs. (4) Synaptic densities differ between MIF and SIF motoneurons, with MIF motoneurons receiving fewer terminals. (5) Glutamatergic receptor subtypes differ between MIF and SIF motoneurons. While NMDAR1 is intensely expressed in INTs, MIF motoneurons lack this receptor subtype entirely. The obtained cell-type-specific transmitter and conductance profiles illuminate the structural substrates responsible for differential contributions of neurons in the abducens and trochlear nuclei to eye movements.

Reduced responses of retinal vessels of the newborn pig to prostaglandins but not to thromboxane

1994 ◽  
Vol 72 (2) ◽  
pp. 168-173 ◽  
Author(s):  
Daniel Abran ◽  
Daya R. Varma ◽  
Ding-You Li ◽  
Sylvain Chemtob
Keyword(s):  
Blood Flow ◽  
Receptor Agonist ◽  
Perfusion Pressure ◽  
Retinal Blood Flow ◽  
Receptor Subtype ◽  
Retinal Vasculature ◽  
Receptor Subtypes ◽  
Retinal Vessels ◽  
Limit Blood Flow ◽  
Arteries And Veins

The upper blood pressure limit of retinal blood flow autoregulation is lower in the newborn than in the adult; this suggests an insufficient vasoconstrictor response in the newborn when perfusion pressure is increased. Because prostaglandins (PGs) have an important role in autoregulation of retinal blood flow, we compared the effects of PGE2, PGF2α, carbacyclin (PGI2 analogue), and U46619 (thromboxane analogue), as well as that of agonists for the three different PGE2 receptor subtypes, 17-phenyl trinor PGE2 (EP1), butaprost (EP2), and M&B 28,767 (EP3), on the retinal vasculature of newborn and adult pigs, using isolated eyecup preparations. PGF2α and PGE2 caused a markedly greater constriction of retinal arteries and veins of the adult than of the newborn animals. Further analysis of the response to PGE2, using receptor subtype agonists, revealed that the EP1 receptor agonist, 17-phenyl trinor PGE2, and the EP3 receptor agonist, M&B 28,767, caused a significant constriction of adult arteries and veins but produced minimal effects on newborn vessels; the EP2 receptor agonist, butaprost, caused a small and comparable dilation of newborn and adult arteries and veins. The PGI2 analogue, carbacyclin, caused a greater dilation of the adult than of the newborn arteries, but produced comparable dilation of veins from both newborn and adult animals. In contrast to the effects of PGF2α and PGE2, the thromboxane analogue, U46619, as well as the α1-adrenoceptor agonist, phenylephrine, significantly constricted newborn arteries and veins, and this effect was comparable with that observed on retinal vessels of the adult. Our findings indicate that the retinal vasculature of the newborn responds minimally to prostaglandins, primarily PGF2α and PGE2, compared with the adult, but constricts effectively to thromboxane. Since prostaglandins play an important role in the autoregulation of retinal blood flow, our observations provide an explanation for the inability of the newborn to limit blood flow when perfusion pressure is raised.Key words: retinal vascular responses, prostaglandins, thromboxane, PGE2 receptor subtypes.

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