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Differences in the characteristics and distribution of rat luteal receptors for native and deglycosylated human choriogonadotropin

Differences in the characteristics and distribution of rat luteal receptors for native and deglycosylated human choriogonadotropin

Biology of Reproduction 47(1): 97-104

Previous work has suggested that rat luteal cells have two populations of LH/hCG receptors that are located in different parts of the cell membrane. The possibility that these two receptor pools may have functional differences has been investigated through examination of the binding and action of native and deglycosylated hCG to different membrane fractions. Ovaries from eCG/hCG-primed immature female rats were separated into 1000 .times. g (heavy) and 20,000 .times. g (light) particulate fractions. Increasing concentrations of NaCl had a biphasic effect on the binding of native and deglycosylated hCG to both membrane fractions, causing an increase in binding at low concentrations and a decrease in binding at higher concentrations. The binding of deglycosylated hCG to both membrane preparations and the binding of native hCG to light-membrane preparations was maximal at approximately the same NaCl concentration (50-65 mM). This was higher than the concentration of NaCl necessary for maximal binding of native nCG to the heavy-membrane preparation. In addition, maximal native hCG binding to this preparation occurred over a broader NaCl concentration range (15-65 mM). Equilibrium binding experiments showed differences in hCG binding to both fractions. In light membranes there were significantly more receptor sites for deglycosylated hCG (11.2 .+-. 4.8 fmol/mg ovary) than for native hCG (4.8 .+-. 0.7 fmol/mg ovary), with no significant different in affinity. In contrast, in heavy membranes the affinity for deglycosylated hCG (6.30 .+-. 0.19 .cntdot. 109 M-1) was significantly higher than that for native nCG (2.60 .+-. 0.13 .cntdot. 109 M-1), with no significant differences in receptor number. Displacement binding assays showed that deglycosylated hCG was as good as, or slightly better than, native hCG in displacing labeled native hCG in either membrane fraction; however, native hCG was only one-fourth to one-fifth as effective as deglycoslyated hCG in displacing labeled deglycosylated hCG in either membrane fraction. Fractionation of the crude preparations on sucrose density gradients revealed differences in the distribution of native and deglycosylated hormone binding and adenylyl cyclase activity. In the light preparation, binding activities for the two ligands were distributed similarly, and no significant adenylyl cyclase activity was found. In the heavy preparation, deglycosylated hCG binding was found only in a less dense portion of the gradient, while native nCG binding was found in the same region and also in a more dense region that also contained adenylyl cyclase activity. Deglycosylated hCG binding was not modulated by guanine nucleotides under conditions that allowed for modulation of native hCG. The results suggest that in the rat ovary there are multiple receptor sites for hCG that have different subcellular localizations and perhaps different functions.

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Accession: 002072359

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PMID: 1637954

DOI: 10.1095/biolreprod47.1.97

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