EXPRESSION OF MINERALOCORTICOID TYPE-I AND GLUCOCORTICOID TYPE-II RECEPTORS IN ASTROCYTE GLIA AS A FUNCTION OF TIME IN CULTURE

In the present study we have examined the expression of mineralocorticoid Type I and glucocorticoid Type Il receptors in astrocyte glia maintained in culture for different periods of time. Cytosolic mineralocorticoid Type I receptors were labeled with [H-3]aldosterone (ALDO) in the presence of a 500-fold molar excess of the potent Type II receptor ligand RU 28362. [H-3]Dexamethasone (DEX) was used to label cytosolic Type 11 receptors. Both Type I and Type II receptor binding was saturable in astrocyte glia that had been maintained in culture for 20 and 30 days following final plating (i.e. 20- and 30-day-old cultures). Scatchard analysis of [H-3]ALDO binding revealed a single class of Type I receptors, with dissociation constants (K(d)) of 0.45 +/- 0.13 nM and 0.53 +/- 0.07 nM, respectively, in 20- and 30-day-old cultures. The number of Type I receptors in 30-day-old cultures was nearly half that found in 20-day-old cultures (22.06 vs 42.64 fmol/mg protein). Linear Scatchard plots were also obtained for [H-3]DEX binding to cytosol prepared from 20- and 30-day-old cultures. There were no significant differences in the K(d) or B(max) values for [H-3]DEX binding in 20- or 30-day-old cultures, i.e. 2.06 +/- 0.15 nM and 247.36 +/- 18.16 fmol/mg protein for 20-day-old cells and 2.3 +/- 0.74 nM and 261.02 +/- 3.08 fmol/mg protein for 30-day-old cells. These B(max) values are more than double the B(max) value for [H-3]DEX binding observed in our previous studies in 10-day-old astrocyte glial cultures. Switching cultured astrocyte glial from serum-supplemented to serum-free medium had no significant effects on the K(d) values of Type I or Type Il receptors in all the cultures tested. However, treatment with serum-free medium increased the number of Type I receptors in 30-day-old cultures to a level similar to that found in 20-day-old cultures. Taken together, these binding data suggest that Type I and Type II receptors are expressed differently in astrocyte glia as a function of time in culture.