scholarly journals UTP binding and phosphoinositidase C activation in ampulla from frog semicircular canal

2000 ◽  
Vol 279 (3) ◽  
pp. R803-R812 ◽  
Author(s):  
Marie Teixeira ◽  
Evelyne Ferrary ◽  
Daniel Butlen
Keyword(s):  
Semicircular Canal ◽  
Binding Sites ◽  
Hill Coefficient ◽  
Disulfonic Acid ◽  
Scatchard Analysis ◽  
Affinity Binding ◽  
Reactive Blue 2 ◽  
High Affinity ◽  
Pyrimidine Nucleotide ◽  
Protein Dissociation

Pyrimidine nucleotide-sensitive phosphoinositidase C activity (PLC), previously identified in frog semicircular canal ampulla, was pharmacologically characterized. Binding of [3H]UTP and abilities of unlabeled nucleotide analogs to inhibit binding and to stimulate PLC in myo-[3H]inositol-loaded ampullas were determined. Specific [3H]UTP binding was competitively inhibited by UTP [apparent dissociation binding constant = 0.8 μM; Hill coefficient = 0.7]. Scatchard analysis revealed a minor class of high-affinity binding sites [45 fmol UTP bound/μg protein; dissociation constant ( K D1) = 0.4 μM] and a major class of moderate-affinity binding sites (365 fmol UTP bound/μg protein; K D2 = 10 μM). The stereospecificity pattern for UTP analog recognition was UMP > UDP ≥ ADP = UTP = dTTP > adenosine 5′- O-(3-thiotriphosphate) = ATP = CTP = 2′-and 3′- O-4-(benzoylbenzoyl)-ATP (Bz-ATP) ≥ AMP ≥ 2-methylthio-ATP = α,β-methylene-ATP > uridine = diadenosine tetraphosphate (Ap4A); cAMP and adenosine were inactive. Antagonist recognition pattern was DIDS = pyridoxal-phosphate-6-azophenyl-2′,4′-disulfonic acid (PPADS) = reactive blue 2 > suramin. The rank order of potencies for agonist-induced PLC activation was UDP ≥ UTP ≥ Ap4A ≥ UMP = Bz-ATP; uridine was inactive. UTP-stimulated PLC activity was inhibited by DIDS = reactive blue 2 = PPADS > suramin. These results suggest that the population of [3H]UTP-labeled binding sites is heterogeneous, with a low number of high-affinity UTP receptors whose function(s) need to be determined and a large number of moderate-affinity receptors triggering PLC activation.

Pharmacological characterization of ATP receptors in ampulla from frog semicircular canal

1998 ◽  
Vol 275 (1) ◽  
pp. R253-R261 ◽  
Author(s):  
Daniel Butlen ◽  
Christian Bernard ◽  
Evelyne Ferrary
Keyword(s):  
Semicircular Canal ◽  
Binding Sites ◽  
Minor Component ◽  
Hill Coefficient ◽  
Scatchard Analysis ◽  
Affinity Binding ◽  
Dissociation Kinetics ◽  
Pyrimidine Nucleotides ◽  
Reactive Blue 2 ◽  
A Minor

Phosphoinositidase C activities sensitive to purine and pyrimidine nucleotides have been identified earlier in ampulla from Rana ridibunda semicircular canal. The aim of this study was to characterize the pharmacological properties of other P2 receptors borne by this structure. A microassay was developed to measure the binding of [35S]adenosine 5′- O-(2-thiodiphosphate) ([35S]ADPβS) to a few ampullas microdissected from frog semicircular canals. When determined at 4°C in the absence of divalent cations, [35S]ADPβS binding was saturable with incubation time and reversible after elimination of free radioligand. The dissociation kinetics were biphasic and comprised a major component that was rapidly reversible and a minor component that dissociated slowly. [35S]ADPβS binding was competitively inhibited by unlabeled ADPβS with an apparent dissociation constant of 0.48 ± 0.09 μM and a Hill coefficient of 0.70 ± 0.06, and Scatchard analysis revealed a minor class of high-affinity binding sites (RT1 = 52 ± 11 fmol [35S]ADPβS bound/ampulla and K d1 = 0.15 ± 0.04 μM) and a major class of low-affinity binding sites (RT2 = 436 ± 79 fmol [35S]ADPβS bound/ampulla and K d2 = 2.0 ± 0.8 μM). The pattern of stereospecificity for recognition of unlabeled structural ATP analogs was ADPβS ≥ α,β-methyleneadenosine 5′-triphosphate = ADP = adenosine 5′- O-(3-thiotriphosphate) > ATP = diadenosine tetraphosphate = AMP > 2′- and 3′- O-(4-benzoylbenzoyl)-adenosine 5′-triphosphate ≥ 2-methylthioadenosine 5′-triphosphate > 2-desoxythymidine 5′-triphosphate = guanosine 5′-triphosphate = inosine-5′-triphosphate = xanthosine 5′-triphosphate = cytosine 5′-triphosphate = uridine 5′-triphosphate = uridine-5′-diphosphate, whereas cAMP and adenosine were devoid of activity. For antagonists, suramin revealed competitive inhibitor potencies, whereas reactive blue 2 and DIDS acted as pure noncompetitive inhibitors. Results suggest that the population of labeled receptors is heterogeneous and contains a low number of P2Y-like receptors and a large number of P2X-like receptors whose molecular subtypes and functions in endolymph homeostasis remain to be defined.

scholarly journals Platelet membrane alterations induced by the local anesthetic dibucaine

Blood ◽  
1986 ◽  
Vol 68 (2) ◽  
pp. 463-471 ◽  
Author(s):  
EI Peerschke
Keyword(s):  
Monoclonal Antibody ◽  
Binding Sites ◽  
Periodic Acid ◽  
Platelet Membrane ◽  
Actin Binding ◽  
Scatchard Analysis ◽  
Affinity Binding ◽  
High Affinity ◽  
Fibrinogen Binding ◽  
Induced Aggregation

Abstract Tertiary amine local anesthetics modify a variety of platelet membrane- related functions. The present study explored dibucaine (DB)-induced inhibition of platelet cohesion by examining structural and functional alterations of the human platelet membrane glycoprotein IIb-IIIa complex (GPIIb-IIIa) and platelet Ca2+ homeostasis. Complete inhibition of ADP-induced aggregation was achieved five minutes after platelet exposure to 0.10 to 0.25 mmol/L of DB when fibrinogen binding was reduced by 50%. At higher concentrations of DB (approximately 1 mmol/L), ADP-induced fibrinogen binding was completely blocked. Scatchard analysis revealed loss of high-affinity binding sites in addition to reduction in Bmax. In contrast, chymotrypsin-treated platelets sustained 50% inhibition of fibrinogen binding when incubated with 0.4 to 0.5 mmol/L DB, and kinetic analysis showed that the high- affinity platelet-fibrinogen interactions were reduced but not absent. Fibrinogen binding to chymotrypsin-treated platelets could not be completely inhibited even at high DB concentrations (1 mmol/L). The inhibition of fibrinogen binding to chymotrypsin-treated platelets correlated with changes in binding of a monoclonal antibody (10E5) specific for an epitope on the GPIIb-IIIa complex. Sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) and radioelectroimmunoassay of DB-treated platelets, however, showed no evidence of a reduction or degradation of GP IIb or IIIa. Platelet incubation with DB (five minutes, 0.1 to 1.0 mmol/L) was also accompanied by: increased platelet membrane-associated Ca2+ involving low-affinity binding sites [Kd = 5 X 10(-5) mol/L-]; increased 45Ca2+ uptake which correlated with degradation of actin-binding protein (ABP) and digestion of GPIb as visualized on periodic-acid Schiff (PAS)- stained SDS gels and as inferred from decreased binding of a monoclonal antibody (6D1) directed against this glycoprotein; and enhanced Ca2+ exchange. Thus, exposure of platelets to DB results in membrane-related alterations that may contribute to inhibition of platelet cohesion: Decreased fibrinogen receptor exposure by traditional agonists and diminished accessibility of the GPIIb-IIIa complex to extracellular ligands correlate with DB-induced inhibition of platelet aggregation; and increased calcium uptake and exchange across the platelet membrane likely leads to activation of the calcium-dependent protease(s) which was previously shown to correlate with DB-induced inhibition of ristocetin-induced platelet agglutination.

Caffeine- and ryanodine-sensitive Ca2+ stores of canine cerebrum and cerebellum neurons

1991 ◽  
Vol 261 (6) ◽  
pp. C1048-C1054 ◽  
Author(s):  
L. G. Meszaros ◽  
P. Volpe
Keyword(s):  
Binding Sites ◽  
Striated Muscle ◽  
Binding Capacity ◽  
Ruthenium Red ◽  
Hill Coefficient ◽  
Stopped Flow ◽  
High Affinity ◽  
Protein Dissociation ◽  
Dissociation Constant Kd ◽  
Slight Inhibition

[3H]ryanodine binding to and Ca2+ release from microsomal fractions derived from canine cerebrum (CBR) and cerebellum (CBL) were investigated. High-affinity ryanodine binding sites were detected in both cerebrum and cerebellum microsomes [CBR: maximal binding capacity (Bmax) = 446 fmol/mg protein, dissociation constant (Kd) = 9 nM, Hill coefficient (n) = 0.95; CBL: Bmax = 650, Kd = 12, n = 1.8]. Ryanodine binding in both fractions was increased by millimolar concentrations of ATP [or its nonhydrolyzable analogue beta, gamma-methyleneadenosine 5'-triphosphate (AMP-PCP)] and micromolar concentrations of Ca2+ but was decreased by micromolar concentrations of ruthenium red, similar to that found in sarcoplasmic reticulum (SR) of striated muscle. The addition of caffeine or the sudden elevation of extravesicular Ca2+ induced a rapid La(3+)-sensitive Ca2+ release from both CBR and CBL microsomal fractions with rate constants of approximately 100 s-1, as determined by stopped-flow photometry of the Ca2+ indicator arsenazo III. The release of Ca2+ was activated by either millimolar ATP or AMP-PCP, blocked by micromolar concentrations of La3+, and significantly inhibited by 50 microM ryanodine. Mg2+ and ruthenium red in millimolar and micromolar concentrations, respectively, caused only a slight inhibition of Ca2+ release. These results indicate that rapid Ca2+ release occurs from caffeine-, Ca2+- and ryanodine-sensitive Ca2+ stores in both CBR and CBL microsomal fractions.

scholarly journals Platelet membrane alterations induced by the local anesthetic dibucaine

Blood ◽  
1986 ◽  
Vol 68 (2) ◽  
pp. 463-471
Author(s):  
EI Peerschke
Keyword(s):  
Monoclonal Antibody ◽  
Binding Sites ◽  
Periodic Acid ◽  
Platelet Membrane ◽  
Actin Binding ◽  
Scatchard Analysis ◽  
Affinity Binding ◽  
High Affinity ◽  
Fibrinogen Binding ◽  
Induced Aggregation

Tertiary amine local anesthetics modify a variety of platelet membrane- related functions. The present study explored dibucaine (DB)-induced inhibition of platelet cohesion by examining structural and functional alterations of the human platelet membrane glycoprotein IIb-IIIa complex (GPIIb-IIIa) and platelet Ca2+ homeostasis. Complete inhibition of ADP-induced aggregation was achieved five minutes after platelet exposure to 0.10 to 0.25 mmol/L of DB when fibrinogen binding was reduced by 50%. At higher concentrations of DB (approximately 1 mmol/L), ADP-induced fibrinogen binding was completely blocked. Scatchard analysis revealed loss of high-affinity binding sites in addition to reduction in Bmax. In contrast, chymotrypsin-treated platelets sustained 50% inhibition of fibrinogen binding when incubated with 0.4 to 0.5 mmol/L DB, and kinetic analysis showed that the high- affinity platelet-fibrinogen interactions were reduced but not absent. Fibrinogen binding to chymotrypsin-treated platelets could not be completely inhibited even at high DB concentrations (1 mmol/L). The inhibition of fibrinogen binding to chymotrypsin-treated platelets correlated with changes in binding of a monoclonal antibody (10E5) specific for an epitope on the GPIIb-IIIa complex. Sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) and radioelectroimmunoassay of DB-treated platelets, however, showed no evidence of a reduction or degradation of GP IIb or IIIa. Platelet incubation with DB (five minutes, 0.1 to 1.0 mmol/L) was also accompanied by: increased platelet membrane-associated Ca2+ involving low-affinity binding sites [Kd = 5 X 10(-5) mol/L-]; increased 45Ca2+ uptake which correlated with degradation of actin-binding protein (ABP) and digestion of GPIb as visualized on periodic-acid Schiff (PAS)- stained SDS gels and as inferred from decreased binding of a monoclonal antibody (6D1) directed against this glycoprotein; and enhanced Ca2+ exchange. Thus, exposure of platelets to DB results in membrane-related alterations that may contribute to inhibition of platelet cohesion: Decreased fibrinogen receptor exposure by traditional agonists and diminished accessibility of the GPIIb-IIIa complex to extracellular ligands correlate with DB-induced inhibition of platelet aggregation; and increased calcium uptake and exchange across the platelet membrane likely leads to activation of the calcium-dependent protease(s) which was previously shown to correlate with DB-induced inhibition of ristocetin-induced platelet agglutination.

Characterization of intestinal smooth muscle responses and binding sites for endothelin

1992 ◽  
Vol 70 (3) ◽  
pp. 377-384 ◽  
Author(s):  
Gordon T. Bolger ◽  
Francine Liard ◽  
Michel Garneau ◽  
Jorge Jaramillo
Keyword(s):  
Smooth Muscle ◽  
Binding Sites ◽  
Contractile Activity ◽  
Benzodiazepine Receptor ◽  
Intestinal Smooth Muscle ◽  
Scatchard Analysis ◽  
Affinity Binding ◽  
Receptor Antagonists ◽  
High Affinity Binding ◽  
High Affinity

The contractile activity of and binding sites for endothelin-1 (ET-1) were investigated in isolated guinea-pig ileal longitudinal smooth muscle (GPILM). ET-1 produced concentration-dependent contractions of GPILM that either slowly subsided in the continued presence of ET-1 or rapidly subsided following washing of the tissue. The ED50 value for ET-1 contractions was 4.2 ± 1.3 × 10−9 M. The removal of extracellular calcium or pretreatment with nifedipine produced a complete inhibition of the contractions to ET-1. The IC50 value of nifedipine for inhibition of ET-1 mediated contractions was 3.0 ± 0.8 × 10−8 M. ET-1 produced a marked prolonged homologous desensitization of its contractile response but did not affect the responses mediated by carbachol, histamine, serotonin, substance P, and PLA2. High-affinity binding sites for 125I-labelled ET-1 were identified on microsomal membranes prepared from GPILM with Kd and Bmax values obtained by Scatchard analysis of 3.5 ± 0.6 × 10−10 M and 2138 ± 159 fmol/mg protein, respectively. The binding of 125I-labelled ET-1 to GPILM microsomes was characterized by a rapid association (kob value of 0.077 min−1 at a radioligand concentration of 0.45 nM and an extremely slow dissociation (kl value of 0.011 min−1; t1/2 value of 793 min). The binding was unaffected by the calcium channel antagonists nifedipine, verapamil, and diltiazem (10−6 M); the receptor antagonists phenoxybenzamine, atropine, and naloxone (10−6 M) and propranolol; and the peripheral benzodiazepine receptor antagonists Ro 5-4864 and PK 11195 and psychotomimetic drug phencyclidine (10−5 M). Incubation of GPILM with ET-1 (2 × 10−8 M) for 10 min followed by washing of the tissue for 1 h resulted in a significant (p < 0.05 unpaired Student's t-test) reduction (33%) of 125I-labelled ET-1 binding that partially recovered following 2 h of washing the tissue. These results demonstrate that ET-1 is an intestinal smooth muscle spasmogen that produces its pharmacologic effects by a mechanism(s) that is not shared by other major intestinal neurotransmitters. Furthermore, intestinal smooth muscle contains specific high-affinity binding sites that likely mediate the contractile responses to ET-1.Key words: intestine, smooth muscle, endothelin, calcium channels, contraction.

Purine and pyrimidine nucleotide-sensitive phosphoinositidase C in ampulla from frog semicircular canal

1997 ◽  
Vol 272 (1) ◽  
pp. R51-R58 ◽  
Author(s):  
D. Butlen ◽  
C. Bernard ◽  
A. Ammar ◽  
E. Ferrary
Keyword(s):  
Semicircular Canal ◽  
Competitive Inhibition ◽  
Rank Order ◽  
Semicircular Canals ◽  
Disulfonic Acid ◽  
Sensory Cells ◽  
Reactive Blue 2 ◽  
Pyrimidine Nucleotide ◽  
Noncompetitive Inhibitor ◽  
Dose Dependent

A microassay was developed to screen the abilities of ATP analogues to stimulate phosphoinositidase C in single ventral regions (including dark cells and sensory cells) of ampullas microdissected from posterior vertical semicircular canals of Rana ridibundo and labeled with myo-[3H]inositol. ATP induced a dose-dependent and saturable increase of total [3H]linositol phosphate production accompanied by an equivalent decrease in the [3H]phosphoinositide pool. The rank order of analogues revealing agonistic potencies for phosphoinositidase C activation was as follows: uridine 5'-triphosphate > or = adenosine 5'-O-[3-thiophosphate] tetralithium > adenosine 5'-O-[2-thiodiphosphate] trilithium > or = ATP > or = ADP = inosine 5'-triphosphate > or = guanosine 5'-triphosphate > or = 2-methylthio-adenosine 5'-triphosphate tetrasodium > or = 2'-desoxy-thymidine 5'-triphosphate > or = cytidine 5'-triphosphate = (alpha, beta)-methyl ATP > AMP, whereas adenosine 3',5'-cyclic monophosphate and adenosine were almost devoid of activity. For antagonists, 1,4'-diisothiocyanostilbene-2, 2'-disulfonic acid was far more active than suramin for competitive inhibition of ATP-induced enzyme stimulation, whereas reactive blue 2 acted as a noncompetitive inhibitor. Results indicate that the putative P2 receptors triggering phosphoinositidase C activation in ventral ampullary epithelium from frog semicircular canal exhibit mainly the functional properties of P2Y and P2U receptors.

Vagus nerve modulates secretin binding sites in the rat forestomach

1999 ◽  
Vol 276 (4) ◽  
pp. G1052-G1058 ◽  
Author(s):  
Hyeok Y. Kwon ◽  
Ta-Min Chang ◽  
Kae Y. Lee ◽  
William Y. Chey
Keyword(s):  
Gastric Motility ◽  
Binding Sites ◽  
Intravenous Infusion ◽  
Intraperitoneal Injection ◽  
Specific Binding ◽  
Scatchard Analysis ◽  
Affinity Binding ◽  
High Affinity Binding ◽  
High Affinity ◽  
Secretin Receptor

Secretin is well known for its inhibitory action on gastric motility. It has been reported that secretin in a physiological dose inhibits gastric motility through mediation by the vagal afferent pathway. Secretin also elicited relaxation of carbachol-stimulated rat forestomach muscle strips by binding to its receptors, suggesting a direct action on this peripheral tissue. We hypothesized that vagal input may affect the action of secretin by modulating the level of secretin receptor in the forestomach. Several treatments, including vagal ligation, vagotomy, perivagal application of capsaicin or colchicine, intravenous infusion of tetrodotoxin, and intraperitoneal injection of atropine, were performed to investigate their effects on secretin receptor binding to forestomach membranes. Specific binding of125I-labeled secretin to forestomach membranes was significantly decreased (45%) by vagal ligation, vagotomy (50%), or perivagal colchicine treatment (40%). On the contrary, specific binding of125I-secretin was not affected by perivagal capsaicin treatment, intravenous infusion of tetrodotoxin, or intraperitoneal injection of atropine. By Scatchard analysis of the binding data, the capacity of the high-affinity binding sites in forestomach membranes was found to decrease significantly after vagal ligation compared with membranes from the sham-operated group. However, the affinity at the high-affinity binding sites, the binding parameters of the low-affinity binding sites, and binding specificity were not changed. Vagal ligation but not perivagal capsaicin treatment reduced the inhibitory effect of secretin on bethanechol-stimulated contraction of isolated forestomach muscle strips, causing a right shift in the dose-response curve. These results suggest that vagal input through axonal transport plays a significant role on secretin action by modulating the capacity of secretin binding sites (but not affinity or specificity), at least in rat forestomach.

PGE2 binding, synthesis, and distribution in hen oviduct

1985 ◽  
Vol 248 (1) ◽  
pp. E80-E88
Author(s):  
G. Asboth ◽  
H. Todd ◽  
M. Toth ◽  
F. Hertelendy
Keyword(s):  
Binding Sites ◽  
Scatchard Analysis ◽  
Affinity Binding ◽  
Shell Gland ◽  
Temperature Dependent ◽  
Single Class ◽  
High Affinity Binding ◽  
High Affinity ◽  
Prostaglandin F2 Alpha ◽  
20 Nm

Prostaglandin E2 (PGE2) bound specifically to particulate fractions prepared from the vagina and uterus (shell gland) portions of the hen oviduct in a time and temperature dependent fashion. Scatchard analysis indicated a single class of high-affinity binding sites in the vagina (Kd congruent to 1 nM), whereas the myometrium exhibited two kinds of binding site populations (Kd1 congruent to 1 nM, Kd2 congruent to 20 nM). It is suggested that these binding sites represent specific PGE2 receptors mediating the effects of PGE2 in oviductal smooth muscle. Vaginal particulate fractions produced approximately four times more prostanoids from [3H]-arachidonate than did uterine preparations. In the presence of epinephrine both tissues synthesized mainly thromboxane (TxB2), PGE2, and significantly less prostaglandin F2 alpha (PGF2 alpha). Addition of glutathione (GSH) or cytosol prepared from the oviduct markedly increased the yield of PGE2 at the expense of TxB2. Of the five morphologically discrete regions of the oviduct the vagina, infundibulum, and uterus contained the highest amounts of PGE and PGF, whereas the magnum and isthmus portions contain the least. TxB2 and 6-keto PGF1 alpha could not be detected in significant quantities in either region. These studies support the notion that PGE2 play a key role in the physiology of oviposition.

Flunitrazepam binding sites in rat diaphragm. Receptors for direct neuromuscular effects of benzodiazepines?

1982 ◽  
Vol 60 (7) ◽  
pp. 1003-1005 ◽  
Author(s):  
M. Wilkinson ◽  
Dale Grovestine ◽  
J. T. Hamilton
Keyword(s):  
Binding Site ◽  
Muscle Relaxant ◽  
Binding Sites ◽  
Scatchard Analysis ◽  
Affinity Binding ◽  
Rat Diaphragm ◽  
High Affinity Binding ◽  
High Affinity ◽  
Site Density ◽  
Peripheral Type

The evidence for direct muscle relaxant effects of benzodiazepines is controversial. We now show that a crude membrane preparation of rat diaphragm possesses binding sites for [3H]flunitrazepam (FNZ). Scatchard analysis gave a binding site density of 1689 ± 143 fmol/mg protein (Kd = 25.6 ± 2.6 nM). These sites are of the "peripheral" type since clonazepam fails to displace [3H]FNZ as effectively as R05-4864 (IC50 values: 7.5 × 10 6 M and 8 × 10−9 M, respectively). Diazepam is almost as effective as R05-4864 and potently displaces [3H]FNZ binding (IC50 = 3 × 10−8 M). We propose that the previously described effects of diazepam on rat diaphragm are mediated through high-affinity binding sites.

Chronic treatment with bovine growth hormone upregulates high-affinity hepatic somatotropic receptors in sheep

1991 ◽  
Vol 124 (3) ◽  
pp. 307-313 ◽  
Author(s):  
Helga Sauerwein ◽  
Bernhard H. Breier ◽  
John J. Bass ◽  
Peter D. Gluckman
Keyword(s):  
Growth Hormone ◽  
Binding Sites ◽  
Dissociation Constants ◽  
Specific Binding ◽  
Hormone Treatment ◽  
Bovine Growth Hormone ◽  
Scatchard Analysis ◽  
Affinity Binding ◽  
Gh Therapy ◽  
High Affinity

Abstract. We evaluated the effect of chronic bovine growth hormone treatment on the hepatic somatotropic receptor. Growing lambs were treated with bGH at 0, 0.05, 0.15, 0.25 or 0.55 mg · kg−1 · day−1 daily (N=5/group) for 56 days. The binding of ovine GH to hepatic membranes washed with 4 mol/l MgCl2 and prepared in the presence of aprotinin was examined. The specific binding of oGH was increased (p<0.01) from 7.1±1.2% in saline-treated controls to 17.4±1.5% in the 0.55 mg · kg−1 · day−1 group. Scatchard analysis showed curvilinear plots that best fitted a two-site model in 22/25 livers. The two sites had estimated dissociation constants (Kd) of 3 to 13 nmol/l for the low-affinity site and a Kd ranging from 0.17 to 0.31 nmol/l for the high-affinity site. Treatment with bGH had no consistent effect on the affinity of either binding site. However, bGH therapy was associated with a dose-dependent increase (p<0.01) in the number of high-affinity somatotropic receptors. There was no effect of bGH therapy on the concentration of low-affinity binding sites. The concentration of high-affinity receptors correlated with weight gain (r=0.54, p<0.01), fat content (r=−0.54, p<0.01), protein content (r=0.40, p<0.05), and plasma IGF-I (r=0.57, p<0.005). The concentrations of low-affinity binding sites showed no such correlations. These observations demonstrate that an important effect of chronic GH therapy in animals with an intact somatotropic axis is to increase significantly the concentration of high-affinity somatotropic receptors. It is suggested that enhancement of the number of high-affinity somatotropic receptors is central to the determination of the efficacy of GH therapy.

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