Glucocorticoid and mineralocorticoid receptors in rat neocortical and hippocampal brain cells in culture: characterization and regulatory studies

Glucocorticoid and mineralocorticoid binding sites were characterized in cell cultures derived from neocortical and hippocampal brain tissue from fetal (E18) rats. Specific and saturable binding was detected in living cells with a sensitive whole cell binding method using [3H]dexamethasone ([3H]DEX) and [3H]aldosterone ([3H]ALDO) (in the presence of RU 28362, a selective glucocorticoid receptor (GR) agonist) as ligands for the measurement of glucocorticoid and mineralocorticoid receptors (MRs), respectively. Specific corticosteroid binding was demonstrated as early as day 4 in culture in neocortical cells, with a time-dependent increase in binding sites during further culturing time. At 7-9 days in vitro, Scatchard analysis of [3H]DEX binding revealed a maximum binding capacity (Bmax) of 83.4 +/- 5.0 fmol/mg protein and a binding affinity (Kd) of 3.6 +/- 0.4 nM in neocortical brain cells. Competition binding studies with [3H]DEX demonstrated a glucocorticoid specificity of receptor sites (relative binding affinity: RU 28362 = DEX > PROG > ALDO). Similar binding characteristics were demonstrated for GRs in cultures derived from fetal hippocampal tissue (Bmax 49.1 +/- 5.8 fmol/mg protein, Kd 3.5 +/- 0.2 nM). Analysis of MRs with [3H]ALDO (+RU 28362) revealed specific and saturable binding in hippocampal cultures, with a Bmax of 8.0 +/- 0.5 fmol/mg protein and a Kd of 0.2 +/- 0.1 nM. Competition studies with [3H]ALDO showed a mineralocorticoid-like pattern of receptor binding (relative binding affinity: CORT = ALDO > PROG > DEX). In addition, small numbers of MRs were detectable in cortex-derived cultures (Bmax: 3.7 +/- 0.8 fmol/mg protein, Kd: 0.3 +/- 0.2 nM).(ABSTRACT TRUNCATED AT 250 WORDS)