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S M Amir

Publications and source records attributed to S M Amir.

8 recordsLinked to original sources
PubMed1978

Interactions of bovine thyrotropin and human chorionic gonadotropin with adenylate cyclase in bovine thyroid membranes.

Previous studies have shown that crude preparations of human chorionic gonadotropin bind to bovine thyroid membranes, displace 125I-labeled bovine thyrotropin therefrom, and are weak agonists therein with respect to the activation of adenylate cyclase. The present studies reveal that concentrations of chorionic gonadotropin sufficient to elicit a maximal agonistic response of adenylate cyclase are strongly antagonistic to the stimulatory action of bovine thyrotropin in the thyroid membrane system. This effect is reminiscent of the inhibitory effects of crude human chorionic gonadotropin on other extragonadal tissues in vitro, and, like them, appears to be mediated by some factor(s) other than human chorionic gonadotropin itself, since highly purified human chorionic gonadotropin was without effect.

Adenylyl Cyclases↗
PubMed1978

Binding of bovine thyrotropin to receptors in rat testis and its interaction with gonadotropins.

Previously, we have shown that preparations of hCG bind to bovine thyroid membranes, as judged from their ability both to inhibit the binding of 125I-labeled bovine TSH (bTSH) and to activate adenylate cyclase (Amir, S.M., H. Uchimura, and S.H. Ingbar, J Clin Endocrinol Metab 45: 280, 1977). In the present studies, 125I-labeled, highly purified bTSH ([125I]bTSH) has been shown to bind specifically and saturably to receptors in a particulate fraction from rat testis. At 37 C, binding was rapid, reaching a maximum level in less than 15 min, but then declining markedly during the next several hours. At 22 C, binding reached a steady state after 2 h and remained unchanged for another 22 h. Binding of [125I]bTSH was greatest at pH 5.5, at which pH more than 50% of [125I]bTSH was bound in the presence of 330 microgram/ml particulate protein, the concentration of protein that yielded maximum binding. Nevertheless, the majority of experiments were conducted at lesser protein concentrations and at physiological pH (7.45), under which conditions total binding was only 25% of that measured at pH 5.5. Scatchard plots indicated the presence of a single binding site with a dissociation constant of 5.8 X 10(-8) M and a binding capacity of 0.22 nmol/mg protein on the basis of data obtained at 22 C and pH 7.45. Both crude and highly purified preparations of hCG inhibited the binding of [125I]bTSH to testis particulate fraction; crude hCG had 46 times the activity, and purified hCG had only one-tenth the activity of bTSH itself in this respect. This was true despite the fact that with respect to the displacement of [125I]hCG, crude and purified hCG were almost equally active. Bovine LH had one-third the activity of bTSH in displacing [125I]bTSH. Human FSH inhibited [125I]bTSH binding only slightly at the highest concentration tested, while glucagon, insulin, PRL, and GH were inactive. Purified bTSH inhibited the binding of [125I]hCG to testis particulate fraction but contained only about 2% of the activity of purified hCG. Lineweaver-Burk analysis suggested that inhibition of [125I]hCG binding by bTSH was competitive in nature. Purified bTSH stimulated cAMP production in Leydig cells, but with only about 0.1% of the activity of purified hCG. It is concluded that bTSH binds reversibly, saturably, and with relatively high affinity to receptors in rat testis that are either the same as receptors for hCG and LH or that interact therewith. bTSH, like hCG, is capable of stimulating the production of cAMP in rat Leydig cells, but is much less potent than hCG in this regard. Preparations of crude hCG contain a factor lacking hCG activity in bioassay, immunoassay, and receptor assay that is especially potent in displacing [125I]bTSH from receptors in testis, as has earlier been described for bTSH receptors in bovine thyroid membranes.

Animals↗
PubMed1976

The interaction of radioiodinated thyrotropin with plasma membranes. Evidence for high affinity binding sites in the thyroid.

The binding of biologically active [125I]thyrotropin to purified plasma membranes prepared from bovine thyroid glands was studied. At 4 degrees C, specific binding reached a maximum after 2 h of incubation and a plateau was maintained for up to 20 h. Degradation of [125I]thyrotropin was undetectable after 2 h of incubation and was only 10% of the total after 20 h. At pH 6.0, at which binding was maximal, a single class of binding sites, having a dissociation constant of approx. 25 nM, was evident. Dissociation studies revealed first order kinetics with a half-time of 2-3 min. At pH 7.5, binding curves were complex, suggesting two orders of binding sites with dissociation constants of approx. 200 nM and 80 pM. Further, at this pH, dissociation of the thyrotropin from its receptor was also complex, suggesting the presence of two first order reactions, one with a half-time similar to that seen at pH 6.0 and another with a half-time of 4 h. At both pH 6.0 and 7.5, insulin, glucagon, growth hormone, and prolactin were without effect on [125I]thyrotropin binding. Similar high affinity and low affinity binding sites were seen with porcine thyroid membranes, but only low affinity sites were seen with either rat liver membranes or human cultured lymphocytes.

Animals↗
PubMed1976

Properties of the interaction between bovine thyrotropin and bovine thyroid plasma membranes.

Studies have been conducted to characterize further the interaction between 125I-labeled bovine thyrotropin (TSH) and bovine thyroid plasma membranes. Sequential subcellular fractionation of thyroid homogenates yielded preparations of progressively greater specific binding activity, highest activity being found in fractions previously shown to contain predominately plasma membranes (Amir, S. M., Carraway, T.F., Kohn, L.D., and Winand, R.J. (1973) J. Biol. Chem. 248, 4092-4100). Although binding of 125I-TSH by plasma membranes was greatest at pH 6.0, studies were conducted at pH 7.45 as well as pH 6.0, and results obtained differed quantitatively, but not qualitatively. Binding was maximal at 0 degrees, 15 degrees, and 22 degrees and steady state values remained unchanged for at least 22 hours. At 37 degrees, binding was decreased by 40% at 1 hour; the loss was even greater (65%) at 50 degrees. A similar loss of binding was evident when membranes were preincubated without TSH at 37 degrees or higher and were then incubated with 125I-TSH at 0 degrees. Lineweaver-Burk analysis indicated that preincubation resulted in loss of receptor sites without change in affinity of residual receptors. Addition of Ca2+ (1 to 10 mM) to the preincubation medium prevented the effect of preincubation at 37 degrees by preserving the number of receptor sites without altering their affinity. Under similar conditions, Na+ and K+ were without protective effect. Membranes bound 45Ca2+ in a specific and saturable manner. Scatchard plots indicated a dissociatiion constant (Kd) of 9 X 10(-5) M and a capacity (n) of 54 nmol/mg of membrane protein. 45Ca2+ was also displaced from membranes by Mg2+ and Mn2+. Ca2+ had a biphasic effect on binding; low concentrations (1 to 10 muM) added to the incubation mixture stimulated binding, while higher concentrations (0.1 mM) caused inhibition. Mg2+ and Mn2+, at comparable concentrations, were also inhibitory, Na+ and K+ less so. In the case of Ca2+, both the stimulatory and inhibitory concentrations were lower than those required to achieve saturation of Ca2+-binding sites. Proteolytic enzymes (trypsin, alpha-chymotrypsin, and pronase) sharply reduced binding of 125I-TSH, owing to a decrease in receptor sites. Phospholipases A and C enhanced binding of TSH, while neuraminidase and beta-galactosidase were without measurable effect.

Animals↗