Neurosteroid inhibition of cell death

Diverse γ-aminobutyric acid (GABA(A)) receptor modulators exhibited novel cytoprotective effects and mechanisms of action in rabbit renal proximal tubules subjected to mitochondrial inhibition (antimycin A) or hypoxia. Cytoprotective potencies (50% effective concentration, EC₅₀) were 0.3 nM allopregnanolone (AP) > 0.4 nM 17α-OH-allopregnanolone (17α-OH-AP) > 30 nM dehydroepiandrosterone sulfate (DHEAS) = 30 nM pregnenolone sulfate (PS) > 500 nM pregnenolone (PREG) > 30 μM museimol > 10 mM GABA following antimycin A exposure. Maximal protection with AP and 17α-OH-AP was 70%, whereas DHEAS, PS, PREG, and muscimol produced 100% cytoprotection. Experiments with AP, PS, and muscimol revealed the return of mitochondrial function and active Na⁺ transport following hypoxia/reoxygenation. Muscimol inhibited the antimycin A-induced influx of both extracellular Ca²⁺ and Cl^- that occurs during the late phase of cell injury, whereas the neurosteroids only inhibited influx of Cl^-. Radioligand binding studies with AP and PS did not reveal a specific binding site; however, structural requirements were observed for cytoprotective potency and efficacy. In conclusion, we suggest that the GABA(A) receptor modulators muscimol and neurosteroids are cytoprotective at different cellular sites in the late phase of cell injury; muscimol inhibits Ca²⁺ and subsequent Cl^- influx, whereas the neurosteroids inhibit Cl^- influx.