Mode-dependent effects of chemical, intermittent, and hypobaric hypoxia on HIF-1α, HIF-2α, HIF-3α, PACAP, and PAI mRNA expression in the single prolonged stress model of PTSD.

INTRODUCTION: Although hypoxic interventions are increasingly explored as non-pharmacological modulators of stress-related pathology, their ability to benefit PTSD remains poorly understood. We compared behavioural and molecular outcomes of three hypoxic regimes - intermittent normobaric hypoxia (IHT), hypobaric hypoxia (Bar), and chemical hypoxia (CH) via cobalt chloride (CoCl₂) - in rats with PTSD induced by single prolonged stress (SPS). METHODS: Male albino rats (n = 105; eight groups) received one of the three treatments after control or SPS conditioning. Anxiety-like behaviour was assessed by Elevated Plus Maze (EPM), Open Field Test (OFT), and Dark-Light Box (DLB); HIF-1α, HIF-2α, HIF-3α, PACAP, and PAI-1 mRNA were quantified in the medial prefrontal cortex (mPFC) and hippocampus by real-time qPCR. RESULTS: Mild IHT was the only regimen to produce robust PTSD-specific anxiolysis: PTSD+IHT animals showed a four-fold reduction in EPM freezing relative to Control+IHT (5.4 ± 4.9 vs 21.6 ± 14.9 s; p < 0.01), a near four-fold reduction in OFT freezing (30.3 ± 8.6 vs 119.9 ± 46.3 s; p < 0.001), and greater open-arm engagement. CH and Bar produced limited or no behavioural benefit. All three regimes substantially reduced HIF-1α mRNA and strongly reduced PAI-1 (>20-fold in most groups; p ≤ 0.001) irrespective of trauma status. Only HIF-1α was significantly elevated in the hippocampus of untreated PTSD animals relative to controls (2.78 ± 1.97 vs 1.10 ± 0.50; q = 0.039); HIF-2α and HIF-3α showed a similar, non-significant upward trend (both ns after FDR correction, reflecting wide between-animal variability). Group means for all three HIF subunits converged toward the control range under all three interventions. HIF-3α responses to IHT or Bar were consistently blunted in PTSD animals across both regions, while PACAP was uniformly suppressed with stronger trauma-dependent attenuation under Bar. DISCUSSION: As protein-level and canonical HIF target-gene readouts were not assessed, the reported transcript changes should be interpreted as a downstream feedback signature rather than a direct index of functional HIF activity. Hypoxic modality, not hypoxia per se, determines behavioural and molecular responses in PTSD; mild IHT uniquely normalised both in traumatised animals, supporting its development as a state-dependent non-pharmacological intervention for trauma-related conditions.