Dopamine1 receptors in rat kidneys identified with 125I-Sch 23982

1988 ◽  
Vol 255 (5) ◽  
pp. F970-F976 ◽  
Author(s):  
R. A. Felder ◽  
P. A. Jose
Keyword(s):  
Room Temperature ◽  
Specific Binding ◽  
Rat Kidney ◽  
Sch 23390 ◽  
Receptor Density ◽  
Single Class ◽  
Apparent Dissociation Constant ◽  
High Affinity ◽  
High Affinity Binding Site ◽  
Inhibition Constants

Dopamine1 receptors were studied in rat kidney using the selective dopamine1 antagonist 125I-labeled Sch 23982. The specific binding of 125I-Sch 23982 (defined by 5 microM Sch 23390) to renal cortical homogenates incubated at room temperature was rapid, saturable with time and ligand concentration, and reversible. Analysis of Rosenthal plots revealed a single class of receptors with an apparent dissociation constant of 12.2 +/- 1.9 nM and maximum receptor density of 1.03 +/- 0.15 pmol/mg protein (n = 6). However, competition experiments with the dopamine1 antagonist Sch 23390 revealed a low- and high-affinity binding site with inhibition constants of 1 x 10(-6) and 1 x 10(-8) M, respectively. The competition experiments were also indicative of dopamine1 receptors with stereoselectivity noted for dopamine1 but not for dopamine2 antagonists. The inhibition constants for dopamine1 antagonists and agonists were two orders of magnitude greater in renal cortical than striatal homogenates. Different buffers affected striatal but not renal cortical binding. Autoradiographic studies revealed 125I-Sch 23982 binding in renal cortical but not medullary tissue. These studies confirm the presence of dopamine1 receptors in the cortex of the rat kidney.

DA1 dopamine receptors in renal cortical collecting duct

1991 ◽  
Vol 261 (5) ◽  
pp. F890-F895 ◽  
Author(s):  
K. Ohbu ◽  
R. A. Felder
Keyword(s):  
Adenylate Cyclase ◽  
Specific Binding ◽  
Collecting Duct ◽  
Sch 23390 ◽  
Receptor Density ◽  
Cortical Collecting Duct ◽  
Tightly Coupled ◽  
Renal Dopamine ◽  
Dissociation Constant Kd ◽  
Stimulation Of

Renal dopamine DA1 receptors are linked to the regulation of sodium transport. We have previously reported the presence of DA1 receptors in the proximal convoluted tubule (PCT) but not in the distal convoluted tubule. However, the DA1 receptor in the collecting duct, the final determinant of electrolyte transport, has not been studied. DA1 receptors were studied in the microdissected cortical collecting duct (CCD) of rats by autoradiography with use of the selective DA1 radioligand 125I-Sch 23982 and by measurement of adenylate cyclase (AC) activity. Specific binding of 125I-Sch 23982 to CCD was saturable with radioligand concentration. The dissociation constant (Kd) was 0.46 +/- 0.08 nM (n = 5), and the maximum receptor density (Bmax) was 1.41 +/- 0.43 fmol/mg protein (n = 5). The DA1 antagonist Sch 23390 was more effective than the DA1 agonist fenoldopam in competing for specific 125I-Sch 23982 binding. Fenoldopam stimulated AC activity in CCD in a concentration-dependent (10(-9)-10(-6) M) manner. The ability of fenoldopam to stimulate AC activity was similar in CCD and PCT even though DA1 receptor density was 1,000 times greater in the CCD than in the PCT. In additional studies, fenoldopam stimulation of AC activity did not influence vasopressin-stimulated AC activity. We conclude that the DA1 receptor in rat CCD is tightly coupled to AC stimulation and that there is no interaction between DA1 agonist-stimulated and vasopressin-stimulated AC activity in the CCD.

Characterization of granulocyte-macrophage colony-stimulating factor receptor on the blast cells of acute myeloblastic leukemia

Blood ◽  
1990 ◽  
Vol 75 (1) ◽  
pp. 59-66 ◽  
Author(s):  
N Onetto-Pothier ◽  
N Aumont ◽  
A Haman ◽  
C Bigras ◽  
GG Wong ◽  
...  
Keyword(s):  
Binding Sites ◽  
Specific Binding ◽  
Receptor Occupancy ◽  
Gm Csf ◽  
High Affinity ◽  
High Affinity Binding Site ◽  
Macrophage Colony Stimulating Factor ◽  
Affinity Receptor ◽  
Acid Wash ◽  
Macrophage Colony

Abstract Iodinated granulocyte-macrophage colony-stimulating factor (GM-CSF) was used to document the specific binding of GM-CSF to all acute myeloblastic leukemia (AML) samples examined in the present study. There was some heterogeneity in the number of GM-CSF binding sites per cell. To determine whether the low level of binding to some patient samples may be attributed to receptor occupancy by an endogenous source of GM-CSF, we devised an acid wash procedure that could remove surface- bound GM-CSF without affecting receptor properties. We thus document that GM-CSF specific binding to AML blasts before or after acid wash was the same, indicating that the observed heterogeneity in binding is not the result of receptor occupancy by an endogeneous source of GM- CSF. Saturation analyses are in favor of the presence of two classes of binding sites on AML blasts: a high-affinity receptor that binds GM-CSF with a dissociation constant (kd) of 3 to 73 pmol/L and a second class of low-affinity receptor that binds GM-CSF with a kd of 1 to 10 nmol/L. Binding studies with two established cell lines KG-1, and IRCM-8 also showed the presence of two classes of binding sites with high and low affinities. Analysis of GM-CSF titration curves in culture indicate that the median effective concentration required for stimulation of blast colony formation (EC50 = 5–36 pmol/L) were in the range of the kd of the high-affinity binding site, suggesting that this high-affinity binding site mediates the proliferative response.

Prostaglandin E2 binding sites in porcine oxyntic mucosa: effects of salicylates

1986 ◽  
Vol 64 (5) ◽  
pp. 515-520 ◽  
Author(s):  
B. L. Tepperman ◽  
B. D. Soper
Keyword(s):  
Binding Sites ◽  
Specific Binding ◽  
Prostaglandin E ◽  
Affinity Binding ◽  
Variable Degree ◽  
Single Class ◽  
Oxyntic Mucosa ◽  
High Affinity Binding ◽  
Mucosal Ulceration ◽  
High Affinity

These studies were designed to examine the changes in the characteristics of prostaglandin E2 (PGE2) binding to porcine oxyntic mucosa in the response to oral ingestion of salicylates. Either acetylsalicylic acid (ASA) or salicylic acid (SA) was administered to conscious pigs (100 mg/kg in 30 mL of an equimolar concentration of NaHCO3) once a day for 1, 3, 10, or 20 days. In control experiments a similar volume of 0.3 M NaHCO3 was administered for similar durations. Mucosal ulceration and the characteristics of the binding of [3H]PGE2 to a 30 000 × g membrane preparation of oxyntic mucosa were examined. Generation of mucosal PGE2 was measured by radioimmunoassay. ASA treatment resulted in an increase in the number and severity of mucosal ulcers and a decrease in PGE2 levels within the first treatment day. By day 20 the degree of ulceration had decreased in spite of a persistent reduction of mucosal PGE2 generation. A variable degree of ulceration was observed in SA-treated animals. In control animals only a single class of binding sites for [3H]PGE2 was evident. After 3 days of ASA treatment a second class of binding sites with a high affinity dissociation constant appeared. There was a decrease in the high affinity binding of [3H]PGE2 after 20 days of ASA ingestion. Low affinity binding was not altered. ASA treatment resulted in a significant increase in specific binding capacities for both families of binding sites. SA treatment did not consistently alter PGE2 binding characteristics from control at any time period studied. These data suggest that SA treatment results in a small degree of mucosal damage in the absence of a significant reduction in tissue generation of PGE2 or changes in PGE2 binding. Damage in response to ASA ingestion was associated with a reduction in both endogenous synthesis of PGE2 and an increase in the concentration of both low and high affinity binding sites for PGE2. The reduction in mucosal ulceration on day 20 in spite of depressed endogenous PGE2 coincides with an increase in PGE2 binding.

Specific binding of vasoactive intestinal peptide to human circulating mononuclear cells

1983 ◽  
Vol 61 (7) ◽  
pp. 664-671 ◽  
Author(s):  
C. A. Ottaway ◽  
C. Bernaerts ◽  
B. Chan ◽  
G. R. Greenberg
Keyword(s):  
Vasoactive Intestinal Peptide ◽  
Binding Sites ◽  
Mononuclear Cells ◽  
Specific Binding ◽  
Maximal Effect ◽  
Single Class ◽  
Apparent Dissociation Constant ◽  
Peptide Interactions ◽  
Tracer Dilution ◽  
Dose Dependent

The interaction of the neuropeptide vasoactive intestinal peptide with human circulating mononuclear cells has been studied. Mononuclear cells were able to bind radiolabelled vasoactive intestinal peptide; the binding was rapid, reversible, saturable, and specific for vasoactive intestinal peptide. A fragment of vasoactive intestinal peptide (10–28) was 25-fold less effective than intact peptide as a competitor for the binding of the tracer. Secretin was 100-fold less effective as a competitor and glucagon competed poorly even at concentrations 10 000 times greater than that of the tracer molecule. In tracer dilution studies, the binding suggested a single class of binding sites with an apparent dissociation constant (Kd) of 2.4 × 10−10 M and a capacity of 2 000 sites per cell. In the presence of vasoactive intestinal peptide, there was a dose-dependent augmentation of cyclic AMP in the mononuclear cells. The concentration of vasoactive intestinal peptide which produced a half-maximal effect was the same as the Kd for the peptide binding. We conclude that mononuclear cells have specific high-affinity binding sites for vasoactive intestinal peptide. Interactions between mononuclear cells and vasoactive intestinal peptide may be an important mechanism modulating local immune responses within tissues innervated by vasoactive intestinal peptide containing neurons.

Alpha-adrenergic receptors on rat ventricular myocytes: characteristics and linkage to cAMP metabolism

1986 ◽  
Vol 251 (2) ◽  
pp. H307-H313 ◽  
Author(s):  
I. L. Buxton ◽  
L. L. Brunton
Keyword(s):  
Dissociation Constant ◽  
Ventricular Myocytes ◽  
Specific Binding ◽  
Single Class ◽  
Adult Rat ◽  
Alpha Adrenergic Receptors ◽  
High Affinity ◽  
Adrenergic Antagonist ◽  
Dependent Protein ◽  
Camp Metabolism

When incubated with purified cardiomyocytes from adult rat ventricle, the alpha 1-antagonist [3H]prazosin binds to a single class of sites (8 X 10(4) per cell) with high affinity [dissociation constant (KD) = 82 pM]. Competition for [3H]prazosin binding by the alpha 2-selective antagonist yohimbine [inhibitor dissociation constant (KI) = 714 nM] and the nonselective alpha-antagonist phentolamine (KI = 168 nM) demonstrates that these receptors are of the alpha 1-subtype. In addition, incubation of myocyte membranes with [3H]yohimbine results in no measurable specific binding. Agonist competition for [3H]prazosin binding to membranes prepared from purified myocytes demonstrates the presence of two components of binding: 28% of alpha 1-receptors interact with norepinephrine with high affinity (KD = 36 nM), whereas the majority of receptors (72%) have a low affinity for agonist (KD = 2.2 microM). After addition of 10 microM GTP, norepinephrine competes for [3H]prazosin binding to a single class of sites with lower affinity (KD = 2.2 microM). Incubation of intact myocytes for 2 min with 1 microM norepinephrine leads to significantly less cyclic AMP (cAMP) accumulation (13.6 pmol/mg) than stimulation with either norepinephrine plus prazosin or isoproterenol (18 pmol/mg). Likewise, incubation of intact myocytes with 10(-6) M norepinephrine leads to significantly less activation of cAMP-dependent protein kinase (71 +/- 4%) than when myocytes are stimulated by both norepinephrine and the alpha 1-adrenergic antagonist, prazosin (95 +/- 1%), or the beta-adrenergic agonist, isoproterenol (100%).(ABSTRACT TRUNCATED AT 250 WORDS)

Oxytocin receptors in rat kidney during development

1990 ◽  
Vol 259 (6) ◽  
pp. F872-F881 ◽  
Author(s):  
A. Schmidt ◽  
S. Jard ◽  
J. J. Dreifuss ◽  
E. Tribollet
Keyword(s):  
Binding Sites ◽  
Specific Binding ◽  
Rat Kidney ◽  
Distal Tubule ◽  
Juxtaglomerular Apparatus ◽  
Binding Studies ◽  
Single Class ◽  
Ligand Selectivity ◽  
Oxytocin Receptors ◽  
Two Stages

The development and characteristics of oxytocin (OT) receptors in the rat kidney were studied by light-microscopic autoradiography and on membrane preparations using the iodinated OT antagonist 125I-d(CH2)5[Tyr(Me)2,Thr4,Orn8,Tyr(NH2)9]OT. Specific binding was first detected by autoradiography at embryonic day 17 (E17) in both the cortex and the medulla. Cortical labeling was found thereafter at all ages examined including in the adult (postnatal day 90, PN90). It was localized on the distal tubule at the level of the juxtaglomerular apparatus. Medullary binding was detected only transiently during two stages of development, first, before PN6, and second, at the approximate time of weaning (PN20-PN30). Binding studies on crude membranes prepared from whole kidneys of animals aged between PN1 and PN15 showed a single class of high-affinity binding sites, with a dissociation constant of 0.13 +/- 0.08 nM. Thus transient OT binding sites expressed in the medulla do not differ from cortical OT binding sites; moreover, the ligand selectivity of kidney OT receptors regardless of location and age appears similar to that of previously characterized OT receptors. Our results suggest that OT may play a role in both renal development and renal function.

Cortical tubular and glomerular dopamine receptors in the rat kidney

1984 ◽  
Vol 246 (5) ◽  
pp. F557-F568 ◽  
Author(s):  
R. A. Felder ◽  
M. Blecher ◽  
G. M. Eisner ◽  
P. A. Jose
Keyword(s):  
Adenylate Cyclase ◽  
Dopamine Receptors ◽  
Binding Sites ◽  
Radioligand Binding ◽  
Rat Kidney ◽  
D2 Dopamine Receptors ◽  
Single Class ◽  
Apparent Dissociation Constant ◽  
Tubular Membranes ◽  
Dissociation Constant Kd

Dopamine receptors in glomeruli and renal cortical tubules were characterized using radioligand binding and adenylate cyclase studies. The binding of [3H]haloperidol to glomeruli and tubules was rapid, saturable with time and ligand concentration, reversible, of high affinity, and demonstrated stereoselectivity and antagonist and agonist rank potency for binding to dopamine receptors. Analysis of kinetic data and Rosenthal plots in glomeruli revealed a single class of [3H]haloperidol binding sites with an apparent dissociation constant (Kd) of 6 nM and maximum receptor density (Bmax) of 0.42 pmol/mg protein. In tubules, at least two binding sites were noted, one with an apparent Kd of 38 nM and Bmax of 1.90 pmol/mg protein and another with an apparent Kd of 183 nM and Bmax of 3.50 pmol/mg protein. Dopamine and apomorphine increased adenylate cyclase in tubular membranes while no increases were noted in glomeruli. These studies suggest that glomeruli have D2 dopamine receptors, while renal cortical tubules contain the D1 dopamine receptor.

Somatotrophic receptors in hepatic tissue of the developing male pig

1989 ◽  
Vol 123 (1) ◽  
pp. 25-31 ◽  
Author(s):  
B. H. Breier ◽  
P. D. Gluckman ◽  
H. T. Blair ◽  
S. N. McCutcheon
Keyword(s):  
Binding Site ◽  
Specific Binding ◽  
Plasma Concentrations ◽  
Age Groups ◽  
Affinity Binding ◽  
Lower Affinity ◽  
High Affinity ◽  
Igf I ◽  
High Affinity Binding Site ◽  
Scatchard Plots

ABSTRACT The development of hepatic somatotrophic receptors and plasma concentrations of insulin-like growth factor-I (IGF-I) were investigated at five different ages (2, 20, 35, 105 and 165 days) in four male pigs per group. The specific binding of 125I-labelled porcine GH (pGH) to hepatic somatotrophic membranes was very low at 2 days of age (0·53±0·12%), and increased progressively (P <0·01) with advancing age to 3·60 ± 0·95% at 165 days of age. Specific binding of 125I-labelled bovine GH (bGH) to the same membrane preparations was markedly higher than binding of 125I-labelled pGH; it also showed a distinct developmental increase (P <0·01) with age from 4·4 ± 0·55% at 2 days of age to 24·0±1·90% at 165 days of age. Plasma concentrations of IGF-I increased significantly (P <0·01) from 79 ± 14·0 μg/l at 2 days of age to 610 ± 64·0 μg/l at 165 days of age. Non-linear regression analysis of the competitive binding data using bGH as labelled and unlabelled ligands showed linear Scatchard plots in the three youngest age groups, with an association constant (Ka) of approximately 3·5 litres/nmol. Curvilinear Scatchard plots were observed in the two oldest age groups. The Ka for the higher affinity binding site (approximately 5·0 litres/nmol) was very similar to that for the sole site observed in the younger animals. The Ka of the lower affinity binding site was approximately 0·35 litres/nmol. There was a significant (P <0·01) developmental increase in the capacity of the higher affinity binding site from 12 ± 4·6 pmol/100 mg liver at 2 days of age to 91 ± 23·0 pmol/100 mg liver at 165 days of age. These studies demonstrate heterogeneity of somatotrophic hepatic membranes in the pig and show that low concentrations of a high affinity binding site are already present in newborn pigs. A considerable developmental increase was observed in the capacity of this binding site which correlated significantly (r = 0·82, P <0·01, n = 20) with plasma concentrations of IGF-I. The role of the lower affinity binding site which was observed in addition to the high affinity binding site in older pigs is less clear. The data from the present study support the hypothesis that the postnatal rise in plasma concentrations of IGF-I is associated with the developmental increase of the capacity of the high affinity somatotrophic receptors. Journal of Endocrinology (1989) 123, 25–31

Sign in / Sign up

Export Citation Format

Share Document