Posts Tagged ‘Rainbow Trout

A method that allows measurement of GH-R in individual testes at all stages of spermatogenesis was validated. Crude testicular preparations were chosen because they gave a less variable and a higher-recovery yield of receptors than protocols using enriched membrane preparations. In this latter protocol, dramatic changes in testis composition during the reproductive cycle influence the […]

A high level of low-affinity binding was found in a population consisting of a mixture of germ cells and various fibroblastic and somatic aggregated cells. Saturable binding with a high affinity (Ka = 3.8 X 109 M ‘) for 125I-rtGH was detected only in a fraction of testicular cells enriched in Sertoli cells after 4 […]

When the results were expressed in fmol/mg of protein, the binding tended to change similarly (significant decrease from the beginning of spermatogenesis [190 fmol/mg of protein] to stage VII [70 fmol/mg of protein], and a nonsignificant increase [120 fmol/mg of protein] at stage VIII). However, the decrease was less pronounced because 1 g of mature […]

Evolution of Testicular GH-R during Spermatogenesis Saturation experiments were conducted at the 8 stages of the spermatogenetic cycle (1-4 experiments per stage) on pools of gonads at the same histological stage (Table 1). Scatchard plots revealed that affinity constants were in the same order of magnitude (covariance analysis) during the entire spermatogenetic cycle, with a […]

The specificity of GH binding sites to crude testicular preparation was tested in competition experiments (Fig. 2). Unlabeled rtGH at concentrations of 0.1-250 ng/tube inhibited the specific binding of the tracer. Bovine GH and rtiGH competed with 125I-rtGH in a dose-dependent manner but appeared to be 75- and 350-fold less effective (calculated at 50% displacement), […]

Characterization of Testicular GH Binding Sites The specific binding of 125I-rtGH to increasing amounts of crude testicular preparations, obtained at two different stages of spermatogenesis (stages I and VI), was dependent on membrane concentration in the range of 5 to 40-50 mg of pellet per tube (~0.5 to 4-5 mg of protein per tube). In […]

Other Assays The protein concentration in the final (testis and liver) preparations was determined in duplicate by the bicinchon-inic acid method (BCA protein assay reagent; Pierce, Rockford, IL), with BSA as standard. The level of plasma GH was determined using a homologous RIA developed in our laboratory . The sensitivity of the assay (££>90) is […]