INTERACTION OF [125I]LH-RH AND OTHER OLIGOPEPTIDES WITH PLASMA MEMBRANES OF RAT ANTERIOR PITUITARIES

1979 ◽  
Vol 92 (2) ◽  
pp. 228-241 ◽  
Author(s):  
Rudolf Baumann ◽  
Herbert Kuhl
Keyword(s):  
Pituitary Gland ◽  
Binding Sites ◽  
Plasma Membranes ◽  
Binding Capacity ◽  
Specific Binding ◽  
Biological Activities ◽  
Single Type ◽  
Number Of Binding Sites ◽  
Specificity Of Binding ◽  
Lh Rh

ABSTRACT The specific binding of [125I]LH-RH to isolated plasma membranes of rat pituitaries was investigated. The binding process was found to be highly specific, temperature-dependent and saturable. The dissociation constant as calculated by three different methods was approximately 1.3 · 10−8 m, indicating a single type of binding sites. Maximal binding capacity was 1 · 10−12 moles/mg protein (=2 ng LH-RH/pituitary gland), and the number of binding sites was calculated to be 6 · 1011 per mg membrane protein (= 1 · 1010 binding sites/pituitary gland). When diluted with icecold buffer the dissociation of specifically bound LH-RH occurred very rapidly (half-life 3.17 min) with a rate constant of 0.219 min−1. The dissociation process followed first-order kinetics. Specificity of binding was demonstrated by dose-dependent competition of unlabelled LH-RH, the highly potent analogue D-glutamine-(cyclohexyl)6-LH-RH-nonapeptide-ethylamide and the fragment of an analogue (6-D-Ser(TBu))-LH-RH-(3-9)-heptapeptide-ethylamide with the binding [125I]LH-RH, while angiotensin I, II, oxytocin and bacitracin did not compete. The affinities of LH-RH and the analogue to the binding sites of the pituitary plasma membranes were not consistent with the respective biological activities.

Effects of hypophysectomy and GH administration on bovine and human GH binding to rat liver membranes

1985 ◽  
Vol 249 (1) ◽  
pp. E56-E62
Author(s):  
J. L. Messina ◽  
S. Eden ◽  
J. L. Kostyo
Keyword(s):  
Binding Sites ◽  
Plasma Membranes ◽  
Specific Binding ◽  
Human Growth ◽  
Male Rats ◽  
Normal Male ◽  
Number Of Binding Sites ◽  
Liver Membranes ◽  
Isolated Liver

Experiments were conducted to investigate the specific binding of highly purified bovine and human growth hormones (bGH and hGH) to purified liver plasma membranes of male rats at various times after hypophysectomy and after the acute intravenous administration of bGH. Liver membranes prepared from hypophysectomized male rats showed a two- to threefold increase in the specific binding of either [125I]iodo-bGH or [125I]iodo-hGH, when compared with membranes prepared from the livers of age-matched normal male rats. The increase in GH binding was apparent within 3 days after hypophysectomy and persisted for a number of weeks after the operation. The increase in GH binding produced by hypophysectomy appeared to be due to an increase in the number of binding sites present on the membranes. The intravenous injection of 200 micrograms of bGH into hypophysectomized male rats 5-60 min before they were killed markedly reduced the ability of liver membranes prepared from these animals to bind [125I]iodo-bGH specifically. This decrease in GH binding seen after the injection of bGH may have been due to the development of a slowly dissociating hormone-binding site complex, which thereby reduced the number of available binding sites. This conclusion is supported by the finding that bGH, which is bound in vitro to isolated liver membranes, dissociates slowly and incompletely in the presence of an excess of unlabeled hormone. Moreover, the degree to which the bound hormone can dissociate appears to depend on the length of time that association is allowed to occur.(ABSTRACT TRUNCATED AT 250 WORDS)

scholarly journals Characterization of a thyroid-hormone-binding site on nuclear envelopes and nuclear matrices of the male-rat liver

1984 ◽  
Vol 219 (3) ◽  
pp. 1001-1007 ◽  
Author(s):  
Y A Lefebvre ◽  
J T Venkatraman
Keyword(s):  
Rat Liver ◽  
Binding Sites ◽  
Intact Animal ◽  
Binding Capacity ◽  
Specific Binding ◽  
Male Rat ◽  
Scatchard Analysis ◽  
Specific Binding Sites ◽  
Number Of Binding Sites

Nuclear envelopes and nuclear matrices were isolated from the male-rat liver. Incubation of 125I-labelled 3,3′,5-tri-iodothyronine (T3) with the nuclear-envelope fraction resulted in specific binding of T3 to the membranes. Maximum specific binding occurred at 30 degrees C after 2h incubation. Storage for 1 week at -80 degrees C resulted in no loss of binding. Scatchard analysis revealed a class of binding sites with KD 86 nM. 3,3′,5′-Tri-iodothyronine was as effective a competitor of [125I]T3 binding to nuclear envelopes as was L-T3 itself, and tri-iodothyroacetic acid was 70% as potent as T3. L- and D-thyronine did not compete for [125I]T3 binding. Incubation of nuclear envelopes with 0.6 M-NaCl before addition of T3 resulted in the complete loss of specific binding sites, whereas exposure of the membranes to 2.0 M-NaCl after incubation with T3 did not extract binding sites. Nuclear matrices, after incubation with [125I]T3 under the same conditions, were shown to possess a class of binding sites with a similar KD but with approx. 30% of the maximum binding capacity. Nuclear envelopes from hypothyroid animals may possess slightly lower numbers of binding sites compared with nuclear envelopes from the intact animal, whereas nuclear matrices from hypothyroid animals have the same number of binding sites as do nuclear envelopes from the intact animal. In conclusion, nuclear envelopes and nuclear matrices have a class of binding sites with relatively high affinity for T3. It is distinct from nuclear and cytosolic binding sites.

Insulin binding to liver plasma membranes in salmonids with modified plasma insulin levels

1991 ◽  
Vol 69 (11) ◽  
pp. 2745-2750 ◽  
Author(s):  
Joaquim Gutiérrez ◽  
Erika M. Plisetskaya
Keyword(s):  
Binding Affinity ◽  
Plasma Insulin ◽  
Plasma Membranes ◽  
Coho Salmon ◽  
Binding Capacity ◽  
Specific Binding ◽  
Insulin Binding ◽  
Term Treatment ◽  
Liver Plasma Membranes ◽  
Number Of Binding Sites

Insulin binding to the liver plasma membranes was assessed in coho salmon (Oncorhynchus kisutch) injected intraperitoneally with 6.6 μmol arginine/g body weight, and in rainbow trout (Oncorhynchus mykiss) fasted for 40 days and then refed for 15 days. Corresponding control groups of fish were injected with saline (coho salmon) or fed regularly (trout). Arginine injection was followed by a substantial elevation of plasma insulin titres from 12.5 to 76.8 ng/mL, an increase in the binding capacity and, consequently, an increase in specific binding of insulin to the liver plasma membranes from 3.4 to 5.5%. The binding affinity remained unchanged. Food deprivation lowered plasma insulin titres from 13.2 to 3.0 ng/mL, increased the binding capacity, but decreased the binding affinity, so the specific binding remained essentially unchanged (5.6% in fed versus 5.1% in fasted fish). Refeeding of fasted fish resulted in restoration of insulin levels and an increase in binding affinity relative to both control and fasted groups of fish. This led to a substantial elevation of the specific binding of insulin up to 7.1%. The binding of insulin to liver plasma membranes in salmonids depends both on the number of binding sites (binding capactity) and on the binding affinity. Long-term treatment, such as food deprivation, resulted in altered affinity and capacity of binding, whereas short-term treatment, such as arginine injection, affected mostly the binding capacity. Modulation of the number of binding sites in liver plasma membrane according to the insulin level was observed only in experiments on trout conducted at a higher (15 °C) water temperature.

scholarly journals Contribution of gangliosides to thyroxin binding with plasma membranes

1995 ◽  
Vol 41 (2) ◽  
pp. 28-30
Author(s):  
T. S. Saatov ◽  
F. Ya. Gulyamova ◽  
G. U. Usmanova
Keyword(s):  
Binding Sites ◽  
Plasma Membranes ◽  
Binding Capacity ◽  
Specific Binding ◽  
Brain Cell ◽  
Cell Recognition ◽  
High Affinity ◽  
Specific Binding Sites

Besides intracellular receptors of thyroid hormones, specific binding sites for T3 and T4 were detected on plasma membranes (PM) of some cells and a relationship between membrane reception .and lipid composition of membranes shown. The parameters of 125I-T4 binding to highly purified PM of hepatic and cerebral cells of rats were studied. The hepatic and cerebral cellular membranes were found to contain two sites of hormone binding each, one of these sites being characterized by a high affinity and low capacity, and the other by low affinity and a higher binding capacity. The association constant of highly affine site of hepatocyte membranes was found to be higher than that of brain cell membranes. T4 membranous receptors may be significant in the process of cell “recognition" by the hormone. In vivo and in vitro experiments with 125I-T4 and 14C-labeled thyroxin in ganglioside fractions showed appreciable binding of the hormone to Gm3 fraction, this evidently pointing to participation of this, ganglioside in T4 interaction with membrane receptor. It is possible that gangliosides situated on membranous surface are components of or function as receptors.

Specific binding sites for ovine prolactin in three amphibian cell lines

1985 ◽  
Vol 248 (1) ◽  
pp. C80-C87 ◽  
Author(s):  
M. Dunand ◽  
M. L. Aubert ◽  
J. P. Kraehenbuhl ◽  
B. C. Rossier
Keyword(s):  
Growth Hormone ◽  
Cell Lines ◽  
Binding Sites ◽  
Binding Capacity ◽  
Specific Binding ◽  
Human Growth ◽  
Functional Differentiation ◽  
Water Metabolism ◽  
Ovine Prolactin ◽  
Follicle Stimulating Hormones

Established cell lines (TB-6c and TB-M) obtained by continuous culture of epithelial cells from toad Bufo marinus urinary bladder, which, in culture, maintained a high degree of functional differentiation, exhibited a significant number of high-affinity (KA = 1-2 X 10(10) M-1) binding sites detected both with radioiodinated (125I) ovine prolactin (oPRL) and human growth hormone (hGH). Binding capacity was higher in the case of TB-6c cells (7,573 +/- 581 sites/cell) than with the TB-M cells (1,160 +/- 87). Similarly, binding sites for oPRL were characterized on Xenopus laevis kidney-derived cell line A6. With oPRL used both as tracer and standard, significant cross-reaction was observed with hGH, less with human or rat prolactin (PRL), and none with human chorionic somatomammotropin, bovine growth hormone, and rat luteinizing hormone or follicle-stimulating hormones. B. marinus pituitary extracts completely displaced the binding of 125I-oPRL to toad bladder binding sites. This finding of specific sites for PRL on amphibian bladder and kidney cells confirms that PRL exerts specific biological actions for the control of electrolyte and water metabolism in the amphibians.

Distribution of opioid receptors in canine small intestine: implications for function

1989 ◽  
Vol 256 (6) ◽  
pp. G966-G974 ◽  
Author(s):  
H. D. Allescher ◽  
S. Ahmad ◽  
P. Kostka ◽  
C. Y. Kwan ◽  
E. E. Daniel
Keyword(s):  
Small Intestine ◽  
Binding Sites ◽  
Myenteric Plexus ◽  
Plasma Membranes ◽  
Binding Capacity ◽  
Opiate Receptors ◽  
Circular Muscle ◽  
Maximum Binding Capacity ◽  
Opiate Binding ◽  
Subtype Selective

Distribution of the binding sites for [3H]diprenorphine, a non-selective opiate ligand, was studied in membrane fractions from longitudinal muscle/myenteric plexus and circular muscle containing deep muscular plexus. [3H]saxitoxin was used as a marker for neuronal plasma membranes and 5'-nucleotidase as a marker for smooth muscle plasma membranes. Saxitoxin binding correlated strongly with diprenorphine binding, but 5'-nucleotidase correlated poorly with diprenorphine or saxitoxin binding in these fractions. Opiate binding sites in membranes of myenteric and deep muscular plexus were of high affinity (Kd = 0.12 and 0.18 nM, respectively) with maximum binding capacity of 400 and 500 fmol/mg protein, respectively. Competition experiments using subtype-selective opiate ligands indicated that all three subtypes of opiate receptors were present in the same ratio of 40-45% mu-subtypes, 40-45% delta-subtypes, and 10-15% kappa-subtypes on both plexuses. Opiate receptors of canine small intestine, therefore, are located primarily or exclusively on nerves with similar distributions in nerve membranes containing only axonal varicosities (deep muscular plexus) as in those containing neurons, dendrites, and varicosities (myenteric plexus).

Endothelin ETA and ETB receptors in postnatal intestine

2001 ◽  
Vol 280 (4) ◽  
pp. G555-G562 ◽  
Author(s):  
Craig A. Nankervis ◽  
David J. Dunaway ◽  
Charles E. Miller
Keyword(s):  
Binding Sites ◽  
Specific Binding ◽  
Ischemia Reperfusion ◽  
Age Groups ◽  
Scatchard Analysis ◽  
Binding Assays ◽  
Site Model ◽  
Number Of Binding Sites ◽  
Competitive Binding Assays

We aimed to characterize endothelin (ET) receptors in the swine intestinal vasculature and to determine ischemia-reperfusion (I/R) effects on these receptors. Saturation and competitive binding assays were performed on mesenteric artery protein membranes from 1- and 40-day-old animals, both control and those subjected to 1 h of partial ischemia followed by 6 h of reperfusion in vivo. Scatchard analysis of saturation binding with 125I-labeled ET-1 in membranes from endothelium-denuded (E−) vessels revealed that the maximum number of binding sites was greater in younger animals. Competitive125I-ET-1 binding was significant for a one-site model with ET-1, ET-3, and sarafotoxin S6c (S6c) in membranes from endothelium-intact (E+) and E− vessels in both age groups. The maximum number of ET-1 binding sites was significantly greater in younger animals. In the presence of the ETAreceptor antagonist BQ-123, competitive 125I-ET-1 binding was significant for a one-site model with ET-1 and S6c in membranes from E+ vessels in both age groups. The maximum number of ET-1 binding sites was significantly greater in younger animals. After I/R, the maximum number of ET-1 binding sites was unchanged. In the presence of BQ-123, specific binding by ET-1 and S6c was eliminated in both age groups after I/R. These results suggest that both ET receptor populations are expressed to a greater degree in younger animals and I/R significantly affects the ETB receptor.

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