ROLE OF GABA-A AND MITOCHONDRIAL DIAZEPAM-BINDING INHIBITOR RECEPTORSIN THE ANTI STRESS ACTIVITY OF NEUROSTEROIDS IN MICE

Neuroactive steroidal modulation of immobilization-stress and possible involvement of GABA-A and mitochondrial diazepam binding inhibitor (D BI) receptors (MDR) has been investigated in mice. Immobilization of m ice for 2 h induced intense antinociception, anxiety state, and associ ated with a fall in adrenal ascorbic acid levels. Pretreatment with hi gh dose of progesterone (10 mg/kg), a precursor of neurosteroids, sign ificantly decreased the stress-induced antinociception, anxiety and fa ll in adrenal ascorbic acid, while low doses (1 and 5 mg/kg) or hydroc ortisone (10 and 100 mg/kg) were ineffective. In contrast, progesteron e (1 mg/kg, for 9 days) produced a significant antistress effect, whic h was blocked by GABA-A antagonists picrotoxin (1 mg/kg) and bicuculli ne (1 mg/kg), but not by flumazenil (2 mg/kg), a specific benzodiazepi ne (BZD) antagonist. 4'-chlordiazepam (0.1 and 0.25 mg/kg), a specific high affinity MDR agonist, produced significant anti-stress effect in a flumazenil-insensitive manner, but was blocked by pretreatment with PK11195 (1.5 mg/kg), a selective partial agonist of MDR, and with bic uculline (1 mg/kg), a potent GABA-A receptor antagonist. At higher dos es, progesterone and 4'-chlordiazepam which are effective in immobiliz ation stress also reduced locomotion. However, lower doses of progeste rone (6.5 mg/kg) neither affected locomotion, nor produced any motor t oxicity on rota-rod test. At the lower doses, the MDR ligand 4'-chlord iazepam (50 mu g/kg) decreased locomotor activity, without altering mo tor toxicity on rota-rod test. Further the per se effects of these tre atments on unstressed mice were not significantly different from those of untreated controls, except for plus-maze test. The antistress prof ile of progesterone may be attributed to the in vivo production of neu rosteroid allopregnanolone, thus resembled that of BZDs. Furthermore, the antistress actions are flumazenil-resistant, reaffirming that ther e may be an increase in the levels of pregnane neurosteroids in vivo, which may act on a specific allosteric site on GABA-A receptors distin ct from BZD site. Because 4'-chlordiazepam binds to MDRs and stimulate mitochondrial neurosteroidogenesis, the anti-stress effects of 4'-chl ordiazepam may be imputed to its MDR-induced neurosteroids, which then act on GABA-A receptors. These data suggest a pivotal role for GABA-A and mitochondrial DBI receptors in the antistress actions of neuroste roids and reinforces their ameliorative effect in physiological stress .