Interactions of some partial agonists with high and low affinity binding sites in β-adrenoceptors

1987 ◽  
Vol 65 (1) ◽  
pp. 18-22 ◽  
Author(s):  
I. Takayanagi ◽  
K. Koike ◽  
A. Nakagoshi
Keyword(s):  
Guinea Pig ◽  
Binding Sites ◽  
Specific Binding ◽  
The Other ◽  
Affinity Binding ◽  
High Affinity ◽  
Partial Agonists ◽  
Significant Difference ◽  
High Affinity Site ◽  
Derivatives Of

Interactions of derivatives of befunolol (BFE-37, BFE-55, and BFE-61), carteolol, and pindolol with β-adrenoceptors were tested in guinea pig isolated taenia caecum. All the drugs used acted as partial agonists on the β-adrenoceptors when compared with isoprenaline, a full agonist. The pA2 values of BFE-61, carteolol, and pindolol were significantly larger than their pD2 values, while there was no significant difference between the pA2 and pD2 values for BFE-37 and BFE-55. The specific binding of [3H]befunolol to microsomal fractions from the guinea pig taenia caecum distinguished two binding sites, high affinity and low affinity sites. Both sites are considered to be bound by 50 nM of [3H]befunolol. Specific 3H binding was displaced by BFE-61, carteolol, and pindolol in a biphasic manner but in a monophasic manner by BFE-37 and BFE-55. Furthermore, [3H]befunolol binding was only partially displaced by BFE-55 but completely displaced by the other drugs used. These results, together with our previous findings, suggest that BFE-61, carteolol, and pindolol discriminate between the two affinity binding sites in the β-adrenoceptors, which are not discriminated between by BFE-37, and further that BFE-55 may bind with only the high affinity site.

scholarly journals Two Binding Sites for [3H]PBR28 in Human Brain: Implications for TSPO PET Imaging of Neuroinflammation

2010 ◽  
Vol 30 (9) ◽  
pp. 1608-1618 ◽  
Author(s):  
David R Owen ◽  
Owain W Howell ◽  
Sac-Pham Tang ◽  
Lisa A Wells ◽  
Idriss Bennacef ◽  
...  
Keyword(s):  
Binding Sites ◽  
Marked Reduction ◽  
Specific Binding ◽  
Translocator Protein ◽  
Affinity Binding ◽  
High Affinity Binding ◽  
Binding Characteristics ◽  
High Affinity ◽  
High Affinity Site ◽  
Site Binding

[11C]PBR28, a radioligand targeting the translocator protein (TSPO), does not produce a specific binding signal in approximately 14% of healthy volunteers. This phenomenon has not been reported for [11C]PK11195, another TSPO radioligand. We measured the specific binding signals with [3H]PK11195 and [3H]PBR28 in brain tissue from 22 donors. Overall, 23% of the samples did not generate a visually detectable specific autoradiographic signal with [3H]PBR28, although all samples showed [3H]PK11195 binding. There was a marked reduction in the affinity of [3H]PBR28 for TSPO in samples with no visible [3H]PBR28 autoradiographic signal ( K i=188±15.6 nmol/L), relative to those showing normal signal ( K i=3.4±0.5 nmol/L, P<0.001). Of this latter group, [3H]PBR28 bound with a two-site fit in 40% of cases, with affinities ( K i) of 4.0±2.4 nmol/L (high-affinity site) and 313±77 nmol/L (low-affinity site). There was no difference in Kd or Bmax for [3H]PK11195 in samples showing no [3H]PBR28 autoradiographic signal relative to those showing normal [3H]PBR28 autoradiographic signal. [3H]PK11195 bound with a single site for all samples. The existence of three different binding patterns with PBR28 (high-affinity binding (46%), low-affinity binding (23%), and two-site binding (31%)) suggests that a reduction in [11C]PBR28 binding may not be interpreted simply as a reduction in TSPO density. The functional significance of differences in binding characteristics warrants further investigation.

EFFECT OF Ca2+ and Mg2+ ON PLATELET ACTIVATING FACTOR (PAF) INDUCED AGGREGATION AND SPECIFIC BINDING TO HUMAN PLATELETS

1987 ◽  
Author(s):  
C M Chesney ◽  
D D Pifer
Keyword(s):  
Platelet Aggregation ◽  
Calcium Channel ◽  
Binding Sites ◽  
Competitive Inhibition ◽  
Specific Binding ◽  
Human Platelets ◽  
High Affinity ◽  
Specific Binding Sites ◽  
Significant Difference ◽  
Induced Aggregation

Gel filtered human platelets (GFP) collected in Tyrode's buffer containing 0.5 mM Ca+2, ImM Mg+2, and 0.35% albumin exhibit high affinity binding of 3H-PAF with a Kd of 0.109 α 0.029 nM (mean α SD; n=13) and 267 α 70 sites per platelet. When fibrinogen (1.67 mg/ml final concentration) is added to these GFP preparations biphasic aggregation is observed with PAF (4 nM). Normal aggregation is also observed with other platelet agonists including ADP, epinephrine, collagen, arachidonic acid, A23187 and thrombin. If GFP is prepared without added Ca+2 or Mg+2 in the presence of 3mM EDTA, platelets do not aggregate in response to PAF. However the number of specific binding sites remains unchanged (387 per platelet) with some decrease in affinity of binding (Kd = 0.2l4nM). In the presence of ImM Mg+2 there is no significant difference in binding kinetics over a range of Ca+2 concentrations (0-2mM). On the other hand the calcium channel blocker verapamil (5-10uM) exhibits competitive inhibition of 3H-PAF as analyzed by Lineweaver-Burk plots. Specific binding of 3H-PAF to GFP in the presence of ImM Mg+2 and ImM EGTA shows Kd of 0.l66nM but with increase in specific binding sites to 665. Despite increase in number of sites and no change in binding affinity, GFP under these conditions does not exhibit platelet aggregation with PAF in doses up to 80 nM.From these data it appears that external Ca+2 is not necessary for specific binding of 3H-PAF to its high affinity receptor. However, calcium does appear to be necessary for second wave aggregation with PAF. While Mg+2 appears to enhance 3H-PAF binding to platelets Mg+2 cannot substitute for Ca+2 in PAF induced platelet aggregation. Although verapamil appears to competitively inhibit binding of PAF to GFP it is not clear whether the inhibition is due to competition at or near the actual PAF receptor or at a site involving the calcium channel.

Demonstration of High-Affinity Binding Sites for C3a Anaphylatoxin on Guinea-Pig Platelets

1978 ◽  
Vol 8 (6) ◽  
pp. 551-555 ◽  
Author(s):  
S. BECKER ◽  
U. HADDING ◽  
H. U. SCHORLEMMER ◽  
D. BITTER-SUERMANN
Keyword(s):  
Guinea Pig ◽  
Binding Sites ◽  
Affinity Binding ◽  
High Affinity Binding ◽  
High Affinity

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.

Uptake of 3,5,3′-l-tri-iodothyronine in human erythrocytes

1989 ◽  
Vol 121 (3) ◽  
pp. 585-591 ◽  
Author(s):  
K. Yamauchi ◽  
R. Horiuchi ◽  
H. Takikawa
Keyword(s):  
Binding Sites ◽  
Binding Capacity ◽  
Human Erythrocytes ◽  
The Other ◽  
Transport Pathway ◽  
High Affinity ◽  
High Affinity Site ◽  
Maximum Binding Capacity ◽  
Energy Dependent ◽  
Dependent Pathway

ABSTRACT The mechanisms of 3,5,3′-l-tri-iodothyronine (T3) uptake into human erythrocytes were examined. Purified membranes of human erythrocytes were shown to have two classes of T3-binding sites with one being a high-affinity site (dissociation constant, 59·2±17·8 nmol/l; maximum binding capacity, 344·3 ± 95·5 fmol/μg protein). Furthermore, it was shown that there were two pathways for T3 uptake in human erythrocytes; one was saturable, stereospecific (T3»thyroxine > 3,5,3′-d-tri-iodothyronine), energydependent and dominant at 15 °C; the other was not displaced by unlabelled T3 and was energyindependent but did not occur by passive diffusion. The former pathway which, it is suggested, is a receptor-mediated transport pathway, was inhibited by monodansylcadaverine, phloretin or oligomycin at 15 or 37 °C, but the latter pathway was not inhibited by these inhibitors. Our results strongly suggest that uptake of T3 by the energy-independent pathway became predominant over the energy-dependent pathway at 37 °C and accounted for 83% of total T3 uptake of human erythrocytes. Journal of Endocrinology (1989) 121, 585–591

scholarly journals Relations between high-affinity binding sites for l-tryptophan, diazepam, salicylate and Phenol Red on human serum albumin

1983 ◽  
Vol 209 (1) ◽  
pp. 135-142 ◽  
Author(s):  
U Kragh-Hansen
Keyword(s):  
Human Serum Albumin ◽  
Serum Albumin ◽  
Binding Sites ◽  
Association Constants ◽  
The Other ◽  
Affinity Binding ◽  
Phenol Red ◽  
High Affinity Binding ◽  
High Affinity ◽  
High Affinity Binding Site

Binding of L-tryptophan, diazepam, salicylate and Phenol Red to defatted human serum albumin was studied by ultrafiltration at pH 7.0. All ligands bind to one high-affinity binding site with association constants of the order of 10(4)-10(5)M-1. The number of secondary binding sites was found to vary from zero to five, with association constants about 10(3)M-1. Competitive binding studies with different pairs of the ligands were performed. Binding of both ligands was determined simultaneously. L-Tryptophan and diazepam were found to compete for a common high-affinity binding site on albumin. The following combinations of ligands do not bind competitively to albumin: L-tryptophan-Phenol Red, L-tryptophan-salicylate and Phenol Red-salicylate. On the other hand, high-affinity bindings of the three ligands do not take place independently but in such a way that binding of one of the ligands results in a decrease in binding of the other ligands. The decreases in binding are reciprocal and can be accounted for by introducing a coupling constant. The magnitude of the constant is dependent on the ligands being bound. In the present study, the mutual decrease in binding was more pronounced with L-tryptophan-salicylate and Phenol Red-salicylate than with L-tryptophan-Phenol Red.

Comparison of interactions of R-(+)- and S-(−)-isomers of β-adrenergic partial agonists, befunolol and carteolol, with high affinity site of β-adrenoceptors in isolated rabbit ciliary body and guinea-pig taenia caeci

1991 ◽  
Vol 69 (7) ◽  
pp. 951-957 ◽  
Author(s):  
Katsuo Koike ◽  
Hisashi Hagiwara ◽  
Issei Takayanagi
Keyword(s):  
Cyclic Amp ◽  
Guinea Pig ◽  
Binding Site ◽  
Ciliary Body ◽  
Partial Agonist ◽  
Affinity Binding ◽  
High Affinity Binding ◽  
High Affinity ◽  
Partial Agonists ◽  
High Affinity Binding Site

The stereoselectivities of β-adrenergic partial agonists for the high affinity binding site of β-adrenoceptors in the rabbit ciliary body and the guinea-pig taenia caeci were studied. The pA2 values of the S-(−)-isomers of befunolol and carteolol against S-(−)-isoprenaline, which were calculated from the shift of each concentration–response curve in increasing cyclic AMP levels, were significantly larger than those of the R-(+)-isomers in the guinea-pig taenia caeci, while the pA2 values of the S-(−)-isomers were not significantly larger than those of the R-(+)-isomers in the rabbit ciliary body. The pKi values determined from the binding experiments were in good agreement with the pA2 values from the increases in cyclic AMP levels. These results suggest that the high affinity binding site of β-adrenoceptors in the guinea-pig taenia caeci may be able to discriminate stereoselectively between the R-(+)- and S-(−)-isomers, while in the rabbit ciliary body there is no stereoselectivity between the two enantiomers.Key words: stereoselectivity, β-adrenoceptor, partial agonist, rabbit ciliary body, guinea-pig taenia caeci.

scholarly journals Species differences in nucleoside transport. A study of uridine transport and nitrobenzylthioinosine binding by mammalian erythrocytes

1982 ◽  
Vol 208 (1) ◽  
pp. 83-88 ◽  
Author(s):  
S M Jarvis ◽  
J R Hammond ◽  
A R P Paterson ◽  
A S Clanachan
Keyword(s):  
Guinea Pig ◽  
Binding Sites ◽  
Species Differences ◽  
Cell Types ◽  
Nucleoside Transport ◽  
Kinetic Properties ◽  
Affinity Binding ◽  
Transport Capacity ◽  
High Affinity Binding ◽  
High Affinity

A kinetic study of the inward transport of uridine in erythrocytes of rabbit, human, mouse, rat and guinea-pig demonstrated that the apparent Km of this process was similar (about 0.2mM) in these cell types, but Vmax. values differed markedly. In this array of cell types, Vmax. values were proportional to the number of transport-inhibitory, high-affinity binding sites present per cell of each type. Transport of uridine or adenosine was not detected in dog erythrocytes, nor was saturable, high-affinity binding of nitrobenzylthioinosine demonstrable. These findings demonstrate that species differences in nucleoside transport capacity are attributable to differences in the cell-surface content of functional nucleoside transport sites, rather than to differences in the kinetic properties of these sites.

Evaluation of the Binding to Fixed Platelets of Agonists and Antagonists of ADP-Induced Aggregation

1989 ◽  
Vol 62 (04) ◽  
pp. 1103-1106 ◽  
Author(s):  
Ashok K Agarwal ◽  
Narendra N Tandon ◽  
Nicholas J Greco ◽  
Noel J Cusack ◽  
G A Jamieson
Keyword(s):  
Binding Sites ◽  
Binding Assay ◽  
Relative Magnitude ◽  
Affinity Binding ◽  
Inhibitory Potency ◽  
Competitive Binding Assay ◽  
High Affinity ◽  
High Affinity Site ◽  
Purine Ring ◽  
Induced Aggregation

SummarySteady state binding of eleven different ADP analogues to formaldehyde-fixed platelets has been determined in a competitive binding assay using 3H-ADP. The compounds tested were the inactive analogues L-ADP and L-ATP; the agonists 2-chloroadenosine 5’-diphosphate, adenosine 5’-O-(2-thiodiphosphate)and the diastereoisomeric pair Sp-adenosine 5’-(1-thiodiphosphate) (Sp-ADP-α-S) and Rp-adenosine 5’-(1-thiodiphosphate) (Rp-ADP-α-S); and the antagonists adenosine 5’-O-thiomonophosphate, 2-chloroadenosine 5’-O-thiomonophosphate, 2-chloroadenosine 5’-triphoshate, and the diastereoisomeric pair 5’-(1-thiotriphosphate) (Sp-ATP-α-S) and Rp-adenosine 5’-(1-thiotriphosphate) (Rp-ATP-α-S). All compounds tested competed at the high affinity binding sites for ADP previously identified (Blood 1988; 71: 110-6) but in some cases competition could not be demonstrated at the low affinity sites because of the high nucleotide concentrations required. As a group, C2-substituted analogues bound less strongly (Ki >2 μM) than did the analogues without substituents in the purine ring (Ki <0.7 μM). With the pair of diastereoisomeric agonists Sp-ADP-α-S and Rp-ADP-α-S the Ki values at the high affinity site (210 nM and 560 nM) were of the same relative magnitude and in the same direction as their reported potencies as agonists (Ki 4 μM and 20 μM). With the diastereoisomeric antagonists Sp-ATP-α-S and Rp-ATP-α-S a similar relationship was seen between affinity (17 nM and 156 nM) and inhibitory potency (Ki 4 μM and 20 μM). These results may help to differentiate possible mechanisms in the interaction of ADP with its receptors.

Sign in / Sign up

Export Citation Format

Share Document