Testosterone is Sufficient to Impart Susceptibility to Isoflurane Neurotoxicity in Female Neonatal Rats

Volatile anesthetic exposure during development leads to long-term cognitive deficits in rats which are dependent on age and sex. Female rats are protected relative to male rats for the same exposure on postnatal day 7. Here we test our hypothesis that androgens can modulate chloride cotransporter expression to alter the susceptibility to neurotoxicity from GABAergic drugs using female rats with exogenous testosterone exposure.

The expression of chloride cotransporters, NKCC1 and KCC2, is altered by testosterone in female rats and corresponds to a cognitive deficit after isoflurane exposure. This confirms the role of androgens in perinatal anesthetic neurotoxicity and supports our hypothesis that the developing GABAergic system plays a critical role in the underlying mechanism.

Female rats were injected with testosterone (100 μg/animal) or vehicle on postnatal days 1 to 6. On postnatal day 7, the animals were randomized to either isoflurane exposure or sham. Spatial memory was assessed with the Barnes maze starting on postnatal day 41. Western blots were run from testosterone treated postnatal day 7 animals to measure levels of chloride cotransporters sodium-potassium-chloride symporter (NKCC1) and chloride-potassium symporter 5 (KCC2).

Exogenous testosterone modulated isoflurane anesthetic neurotoxicity in female rats based on poor performance in the probe trial of the Barnes Maze. By contrast, females with vehicle and isoflurane exposure were able to differentiate the goal position. These behavioral differences corresponded to differences in the protein levels of NKCC1 and KCC2 after exogenous testosterone exposure, with NKCC1 increasing ( P <0.001) and KCC2 decreasing ( P =0.003) relative to female controls. Conclusions: The expression of chloride cotransporters, NKCC1 and KCC2, is altered by testosterone in female rats and corresponds to a cognitive deficit after isoflurane exposure. This confirms the role of androgens in perinatal anesthetic neurotoxicity and supports our hypothesis that the developing GABAergic system plays a critical role in the underlying mechanism.