Binding Sites for 125I-Labelled Endothelin-1 in the Kidneys: Differential Distribution in Rat, Pig and Man Demonstrated by Using Quantitative Autoradiography

1989 ◽  
Vol 77 (2) ◽  
pp. 129-131 ◽  
Author(s):  
Anthony P. Davenport ◽  
Derek J. Nunez ◽  
Morris J. Brown
Keyword(s):  
Binding Sites ◽  
Specific Binding ◽  
Receptor Autoradiography ◽  
Differential Distribution ◽  
Endothelin 1 ◽  
Specific Binding Sites ◽  
Vasa Recta ◽  
Von Willebrand ◽  
Willebrand Factor

1. Quantitative receptor autoradiography in vitro has been used to determine the distribution and density of specific binding sites for 125I-labelled endothelin-1 in human, porcine and rat kidneys. Immunocytochemistry was used to visualize von Willebrand factor-positive endothelial cells in adjacent sections to those used for autoradiography. 2. High levels of specific binding were detected in the vasa recta and papilla of all three species with lower levels in the medulla and cortex. 3. A major difference between the species was observed within the glomeruli, where high levels of binding were found in the rat but no detectable binding sites in pig or man.

Platelet-Binding of the von Willebrand Factor

1978 ◽  
Vol 39 (03) ◽  
pp. 689-694 ◽  
Author(s):  
David Green ◽  
Hendrik P Muller
Keyword(s):  
Binding Sites ◽  
Specific Binding ◽  
Platelet Membrane ◽  
Specific Binding Sites ◽  
Platelet Binding ◽  
Membrane Bound ◽  
Von Willebrand ◽  
Willebrand Factor ◽  
Aggregation Of Platelets ◽  
Partial Release

SummaryThe aggregation of platelets by the antibiotic, ristocetin, requires a plasma cofactor (VIII:vWF) and one or more specific binding sites on the platelet membrane. The interaction between VIII:vWF and the platelet was examined using VIII:vWF labelled with 125 I. In the presence of ristocetin (1.5 mg/ml), from 70 to 90% of the 125 I-VIII:vWF became platelet- bound. By contrast, only 21% was bound with thrombin (2.5 u/ml), and 2.2% with buffer alone. Fractionation of the platelets revealed that peak radioactivity was present in the membrane fraction. Treatment of ristocetin-reacted platelets with either chymotrypsin, 100 ug/ml, or trypsin, 75 ug/ml, resulted in the partial release of the membrane-bound radioactivity. It is concluded that VIII:vWF binds to the platelet membrane in the presence of ristocetin.

Atrial natriuretic factor binding sites in the jejunum

1989 ◽  
Vol 256 (2) ◽  
pp. G436-G441 ◽  
Author(s):  
C. Bianchi ◽  
G. Thibault ◽  
A. De Lean ◽  
J. Genest ◽  
M. Cantin
Keyword(s):  
Binding Sites ◽  
Atrial Natriuretic Factor ◽  
Specific Binding ◽  
Rat Tissues ◽  
Rat Jejunum ◽  
Specific Binding Sites ◽  
Natriuretic Factor ◽  
Atrial Natriuretic

We have studied the localization and the characterization of atrial natriuretic factor (ANF) binding sites by radioautographic techniques. Quantitative in vitro radioautography with a computerized microdensitometer demonstrated the presence of high-affinity, low-capacity 125I-ANF-(99-126) binding sites (Kd, 48 pM; Bmax, 63 fmol/mg protein) mainly in the villi of 20-microns slide-mounted transverse sections of the rat jejunum. Competition curves showed 50% inhibitory concentrations of 55 and 1,560 pM for ANF-(99-126) and ANF-(103-123), respectively. In vivo electron microscope radioautography showed that 80% of the silver grains were localized on the lamina propria fibroblast-like cells, 18% on mature enterocytes, and 2% on capillaries. Bradykinin and adrenocorticotropin did not compete with ANF binding. These results demonstrate that ANF binding sites in the rat jejunum possess the pharmacological characteristics of functional ANF receptors encountered in other rat tissues, and ultrastructural radioautographs show their cellular distribution. Taken together, these results demonstrate the presence and the localization of specific binding sites for ANF in the jejunal villi of the rat small intestine.

The Binding Of 125I-Von Willebrand Factor To Human Platelets In The Presence Of Ristocetin

1981 ◽  
Author(s):  
P Silber ◽  
T H Finlay
Keyword(s):  
Von Willebrand Factor ◽  
Binding Sites ◽  
Binding Constant ◽  
Specific Binding ◽  
Human Platelets ◽  
Scatchard Analysis ◽  
Binding Data ◽  
Number Of Binding Sites ◽  
Von Willebrand ◽  
Willebrand Factor

The effect of ristocetin on the binding of 125I-porcine von Willebrand factor to human platelets was studied. Previously, we had shown that 125I-porcine von Willebrand factor binds to human platelets in the absence of ristocetin. The present work demonstrates that binding is stimulated by ristocetin and this stimulation is maximal at a ristocetin concentration of 2 mg/ml. At a ristocetin concentration of 0.5 mg/ml, Scatchard analysis indicates a binding constant of 5.18 × 10-9M and the presence of 105,000 binding sites. This compares with our previous finding, in the absence of ristocetin, of a binding constant of 2.92 × 10-7M and 4760 binding sites. These binding data assume the porcine von Willebrand factor to be a tetramer with a molecular weight of 9 × 105. This study indicates that ristocetin causes tighter binding and increases the number of binding sites on human platelets for porcine von Willebrand factor. Unlabelled porcine von Willebrand factor competitively inhibits the specific binding of the labelled protein and gives a binding constant of 0.17 × 10-9M. Similar results were obtained using human von Willebrand factor.

Specific binding sites for 125I-endothelin-1 in the porcine and human spinal cord

1992 ◽  
Vol 225 (4) ◽  
pp. 281-289 ◽  
Author(s):  
Masami Niwa ◽  
Tsutomu Kawaguchi ◽  
Akihiko Himeno ◽  
Meiko Fujimoto ◽  
Masaki Kurihara ◽  
...  
Keyword(s):  
Spinal Cord ◽  
Binding Sites ◽  
Specific Binding ◽  
Human Spinal Cord ◽  
Endothelin 1 ◽  
Specific Binding Sites

Autoradiographic detection of kinin receptors in the human medulla of control, hypertensive, and diabetic donors

2002 ◽  
Vol 80 (4) ◽  
pp. 249-257 ◽  
Author(s):  
Hudson de Sousa Buck ◽  
Brice Ongali ◽  
Gaétan Thibault ◽  
Charles J Lindsey ◽  
Réjean Couture
Keyword(s):  
Receptor Binding ◽  
Binding Sites ◽  
Specific Binding ◽  
Inferior Olivary Nucleus ◽  
Binding Studies ◽  
Specific Binding Sites ◽  
Kinin Receptors ◽  
Medullary Nuclei ◽  
Inferior Olivary

Kinins have been elected to the status of central neuromediators. Their effects are mediated through the activation of two G-protein-coupled receptors, denoted B1 and B2. Functional and binding studies suggested that B1 and B2 receptors are upregulated in the medulla and spinal cord of hypertensive and diabetic rats. The aim of this study was to localize and quantify kinin receptors in post-mortem human medulla obtained from normotensive, hypertensive, and diabetic subjects, using in vitro receptor autoradiography with the radioligands [125I]HPP-HOE140 (B2 receptor) and [125I]HPP[des-Arg10]-HOE140 (B1 receptor). Data showed specific binding sites for B2 receptor (0.4–1.5 fmol/mg tissue) in 11 medullary nuclei from 4 control specimens (paratrigeminal > ambiguus > cuneate, gelatinous layer of the caudal spinal trigeminal nucleus > caudal and interpolar spinal trigeminal, external cuneate, solitary tract > hypoglossal > gracile > inferior olivary nuclei). Increased density of B2 receptor binding sites was observed in seven medullary nuclei of four hypertensive specimens (paratrigeminal > external cuneate > interpolar and caudal spinal trigeminal, gracile, inferior olivary > hypoglossal nuclei). B2 receptor binding sites were seemingly increased in the same medullary nuclei of two diabetic specimens. Specific binding sites for B1 receptor (1.05 and 1.36 fmol/mg tissue) were seen only in the inferior olivary nucleus in two out of the ten studied specimens. The present results support a putative role for kinins in the regulation of autonomic, nociceptive, and motor functions at the level of the human medulla. Evidence is also provided that B2 receptors are upregulated in medullary cardiovascular centers of subjects afflicted of cardiovascular diseases.Key words: bradykinin, hypertension, diabetes, human brain.

Studies on Corticosterone–Receptor Complexes from Mouse Placenta

1974 ◽  
Vol 52 (3) ◽  
pp. 190-195 ◽  
Author(s):  
Ming D. Wong ◽  
A. F. Burton
Keyword(s):  
Binding Sites ◽  
Time Course ◽  
Specific Binding ◽  
Receptor Site ◽  
Binding Properties ◽  
Receptor Complexes ◽  
Specific Binding Sites ◽  
Mouse Placenta ◽  
Vitro Binding

The in vitro binding of radioactive steroids to components of mouse placental nuclei and cytoplasm was investigated using Sephadex or charcoal to remove unbound steroid. Specificity was indicated in competition experiments using excess unlabelled competing steroids. Only the active glucocorticoids formed complexes that could be isolated from the nucleus. The binding properties of the cytoplasmic steroid–receptor complex were studied. From the time course of binding the complex was shown to be more stable at 0° than at 37°, and the distribution of receptors in the cytosol appeared to be homogeneous. The complex was labile to heat and to proteolytic digestion but did not appear to be affected by nucleases or sulfhydryl reagents. Kinetic analysis revealed the presence of high affinity specific binding sites with a dissociation constant of 17.5 nM and a receptor site concentration of 0.26 pmol/mg protein. The corticosterone isolated from nuclear complexes and dexamethasone from cytoplasmic complexes were identified by chromatography and by cocrystallization as the unchanged steroid in each case.

scholarly journals A quantitative binding model for the Apl protein, the dual purpose recombination-directionality factor and lysis-lysogeny regulator of bacteriophage 186

2020 ◽  
Vol 48 (16) ◽  
pp. 8914-8926
Author(s):  
Erin E Cutts ◽  
J Barry Egan ◽  
Ian B Dodd ◽  
Keith E Shearwin
Keyword(s):  
Binding Sites ◽  
Specific Binding ◽  
Attachment Site ◽  
Binding Energies ◽  
Sequence Specificity ◽  
Binding Studies ◽  
Binding Model ◽  
Specific Binding Sites

Abstract The Apl protein of bacteriophage 186 functions both as an excisionase and as a transcriptional regulator; binding to the phage attachment site (att), and also between the major early phage promoters (pR-pL). Like other recombination directionality factors (RDFs), Apl binding sites are direct repeats spaced one DNA helix turn apart. Here, we use in vitro binding studies with purified Apl and pR-pL DNA to show that Apl binds to multiple sites with high cooperativity, bends the DNA and spreads from specific binding sites into adjacent non-specific DNA; features that are shared with other RDFs. By analysing Apl's repression of pR and pL, and the effect of operator mutants in vivo with a simple mathematical model, we were able to extract estimates of binding energies for single specific and non-specific sites and for Apl cooperativity, revealing that Apl monomers bind to DNA with low sequence specificity but with strong cooperativity between immediate neighbours. This model fit was then independently validated with in vitro data. The model we employed here is a simple but powerful tool that enabled better understanding of the balance between binding affinity and cooperativity required for RDF function. A modelling approach such as this is broadly applicable to other systems.

ENDOTHELIN STIMULATES TESTOSTERONE SECRETION BY RAT LEYDIG CELLS

1993 ◽  
Vol 136 (2) ◽  
pp. R1-R4 ◽  
Author(s):  
D. Conte ◽  
P. Questino ◽  
S. Fillo ◽  
M. Nordio ◽  
A. Isidori ◽  
...  
Keyword(s):  
Binding Sites ◽  
Leydig Cells ◽  
Channel Blocker ◽  
Specific Binding ◽  
Human Chorionic Gonadotrophin ◽  
Paracrine Factor ◽  
Testosterone Secretion ◽  
Specific Binding Sites ◽  
Testicular Steroidogenesis

ABSTRACT The present study was designed to evaluate the effects of endothelin (ET) on rat testicular steroidogenesis in vitro and the involvement of prostaglandins (PG) and extracellular calcium in its mechanism of action. To this purpose we examined the effects of ET-1 and ET-3 on basal testosterone secretion, the influence of ET-1 on PGE2, release, the interaction of ET-1 and ET-3 with human chorionic gonadotrophin (hCG) and the interference of indomethacin (an inhibitor of cycloxygenase) and nifedipine (a calcium-channel blocker) in purified rat Leydig cells. The data indicate that ET-1 and ET-3 stimulate basal and hCG-induced testosterone production although the effects of ET-3 were less marked. In addition, a concomitant release of PGE2, was observed after exposure to ET-1. A sinergistic interaction between ET-1 and hCG in stimulating testicular steroidogenesis was revealed. Indomethacin was ineffective in modifying ET-1 evoked testosterone output, while in the presence of nifedipine the stimulatory effect of ET-1 was completely abolished. Since it has been shown by others that ET-1 is produced by rat Sertoli cells and specific binding sites are present in Leydig cells, the results of our study indicate that such a peptide may be regarded as a new paracrine factor able to influence steroidogenesis in Leydig cells. The action of ET-1 requires the activity of voltage-operated Ca2+ channels, while PGE2 activation is not essential for its steroidogenic effect.

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