Does sex matter in neurons’ response to hypoxic stress?

Background: Stroke exhibits significant sex differences in incidence, response to treatment, and outcome. Preclinical studies suggest that hormones, particularly estrogens, are key to differential sensitivity, as female neurons demonstrate enhanced resilience compared to males in both in vivo and in vitro models. This study investigates whether these sex-specific differences in neuronal vulnerability extend to the ischemic penumbra and explores the effects of estrogens under such conditions.

Methods: Primary cortical neuronal networks were generated from male and female newborn Wistar rats and cultured on micro-electrode arrays or glass coverslips. Male and female networks were subjected to hypoxic conditions, followed by a recovery phase, with or without exogenous estrogen treatment. Electrophysiological activity, including spikes and bursts, was monitored and analyzed. Apoptosis was assessed through immunocytochemistry, focusing on caspase-dependent and apoptosis-inducing factor (AIF)-dependent pathways.

Results: Under hypoxia, male and female networks showed similar reductions in firing and burst rates with longer burst durations. Exogenous estrogen altered these dynamics, leading to increased burst rates and shorter burst durations for both sexes. During recovery, two-way ANOVA suggested higher burst rates in estrogen-treated networks and sex differences in burst duration at 24h, but these effects were not confirmed by non-parametric analysis. Immunocytochemistry revealed that estrogen significantly reduced caspase-dependent apoptosis, but not AIF-dependent apoptosis. Mean firing rates and overall network viability did not differ between groups, indicating no clear long-term survival benefit.

Conclusion: In our model of the ischemic penumbra, exogenous estrogen modulated neuronal network activity, with sex-dependent differences evident under normoxic but not hypoxic or recovery conditions. These effects reflect context-dependent responsiveness rather than intrinsic sex differences, and provide no evidence for enhanced neuronal survival after hypoxia.