following transfection. We selected a low cortisol dose (1 nM) for this study based on reports that this dose produced Type I GR activation, without Type II GR activation or TypeI:Type II dimerization, in the HT22 cell line 30. This is important because high cortisol doses (sufficient to activate Type II receptors) induce cell death in hippocampus 31, 32. Expression of DsRed was monitored 48h following treatment using an Olympus IX70 microscope (Olympus America, Melville, NY) interfaced with a Hamamatsu ORCA-ER digital camera (Hamamatsu, Tokyo, Japan). Region of interest analysis was performed using OpenLab® software. A CRH promoter expression value was obtained by dividing the DsRed intensity by that for GFP. Experiments were performed four times, with treatments in some instances performed in duplicate. Measurements were made by two observers and averaged. Data were analyzed by ANOVA, with treatment (No Treatment, Forskolin, Forskolin + Cort) and genotype (-2232 C and -2232 G) included as independent variables. Data were analyzed by ANOVA, with treatment (No Treatment, Forskolin, Forskolin + Cort) and genotype (-2232 C and -2232 G) included as independent variables.
