FIGURE 3.
Candidate gene screening for GnRH and CTX responsiveness and Gαs and Gαq/11 protein dependence. A, effect of GnRH or CTX stimulation on the expression of 19 candidate genes in LβT2 cells. Cells were serum-starved overnight and stimulated with either 1 nm GnRH, 5 μg/ml CTX, or vehicle for 10 h. B, effect of Gαs or Gαq/11 silencing on the expression of 19 candidate genes in LβT2 cells. Cells were serum-starved overnight and transfected with either scrambled (control), Gαs, or Gαq/11 siRNA. Transfections were carried out in the Nucleofector 96-well Shuttle as described under” Experimental Procedures.” Cells were harvested 3 days post-transfection. C, effect of GnRH or CTX stimulation on the expression of 19 candidate genes in purified primary gonadotropes. Cells were stimulated with either 1 nm GnRH, 5 μg/ml CTX, or vehicle for 10 h. mRNA expression levels of the indicated genes were determined by qPCR. Data shown are the mean from one experiment performed in triplicate and are representative of three independent experiments. Error bars represent S.E. ***, p < 0.001; **, p < 0.01; *, p < 0.05, one-way ANOVA.