Permissive role of brain allopregnanolone content in the regulation of pentobarbital-induced righting reflex loss.

Allopregnanolone [3alpha-hydroxy-5alpha-pregnan-20-one] (ALLO), a potent neurosteroid that positively modulates gamma-aminobutyric acid (GABA) action at various GABA(A) receptor subtypes is synthesized in nanomolar concentrations and stored non uniformly in various brain structures of mammals. We have measured brain ALLO content and its precursors by negative ion chemical ionization-mass-spectrometry after purification and separation of the different steroids with HPLC and gas chromatography. Our procedure measures steroids in the femtomolar range with structural information and unsurpassed selectivity. We were able to establish an association between the decrease in content of ALLO in mouse brain cortex elicited by either long-lasting social isolation or by the administration of 17beta-17 [bis (1-methylethyl) amino carbonyl] androstane-3,5-dilene-3-carboxylic acid (SKF 105111). an inhibitor of Types I and II 5alpha reductases, and the shortening of the righting reflex loss elicited by pentobarbital (PBT). SKF 105111 added to cortical brain slices in concentrations up to 10(-5) M failed per se to alter GABAergic currents or their potentiation by PTB recorded from pyramidal neurons. Fluoxetine (1.45 or 2.9 micromol/kg i.p.) doses that fail to change the PTB-induced loss of righting reflex and the level of brain ALLO in group-housed mice normalized both parameters in socially-isolated mice. In addition, we could detect both fluoxetine actions in socially isolated mice pretreated with doses of p-chlorophenylalanine (1.2 mmol/kg i.p. at 72, 48, and 24 h) that substantially inhibit brain serotonin 5HT synthesis as shown by an 80% drop of brain 5HT content. These studies for the first time have provided evidence suggesting that the endogenous cortical stores of ALLO physiologically upregulate GABAergic tone and by such a mechanism play a permissive or facilitatory role on the PTB-induced loss of the righting reflex. In the absence of such a permissive physiological influence by endogenous ALLO, the righting reflex inhibition by PTB is down regulated.