Overexpression of the Steroidogenic Enzyme Cytochrome P450 Side Chain Cleavage in the Ventral Tegmental Area Increases 3 alpha,5 alpha-THP and Reduces Long-Term Operant Ethanol Self-Administration

Neuroactive steroids are endogenous neuromodulators capable of altering neuronal activity and behavior. In rodents, systemic administration of endogenous or synthetic neuroactive steroids reduces ethanol self-administration. We hypothesized this effect arises from actions within mesolimbic brain regions that we targeted by viral gene delivery. Cytochrome P450 side chain cleavage (P450scc) converts cholesterol to pregnenolone, the rate-limiting enzymatic reaction in neurosteroidogenesis. Therefore, we constructed a recombinant adeno-associated serotype 2 viral vector (rAAV2), which drives P450scc expression and neuroactive steroid synthesis. The P450scc-expressing vector (rAAV2-P450scc) or control GFP-expressing vector (rAAV2-GFP) were injected bilaterally into the ventral tegmental area (VTA) or nucleus accumbens (NAc) of alcohol preferring (P) rats trained to self-administer ethanol. P450scc overexpression in the VTA significantly reduced ethanol self-administration by 20% over the 3 week test period. P450scc overexpression in the NAc, however, did not alter ethanol self-administration. Locomotor activity was unaltered by vector administration to either region. P450scc overexpression produced a 36% increase in (3 alpha,5 alpha)-3-hydroxypregnan-20-one (3 alpha,5 alpha-THP, allopregnanolone)-positive cells in the VTA, but did not increase 3 alpha,5 alpha-THP immunoreactivity in NAc. These results suggest that P450scc overexpression and the resultant increase of 3 alpha,5 alpha-THP-positive cells in the VTA reduces ethanol reinforcement. 3 alpha,5 alpha-THP is localized to neurons in the VTA, including tyrosine hydroxylase neurons, but not astrocytes. Overall, the results demonstrate that using gene delivery to modulate neuroactive steroids shows promise for examining the neuronal mechanisms of moderate ethanol drinking, which could be extended to other behavioral paradigms and neuropsychiatric pathology.