Aims: The coordination of neurons to execute brain functions requires plenty of oxygen. Thus, it is not surprising that the chronic hypoxia resulting from chronic obstructive pulmonary diseases (COPD) can cause neuronal damage. Injury in the cortex can give rise to anxiety and cognitive dysfunction. This study investigated what causes hypoxia-induced neuronal injury and what strategies might be used to protect neurons against such damage. Results: This study found that hypoxia in primary cortical neurons caused neurite retraction, a caspase-6-dependent process. The hypoxic stress activated caspase-6 within the neurite, leading to microtubule disassembly and neurite retraction. The effect of hypoxia on caspase-6 activation, microtubule disassembly, and neurite retraction was alleviated by nitrite treatment. The protective role of nitrite was further supported by the observation that the active-site Cys146 of caspase-6 was S-nitrosylated in hypoxic neuro-2a cells treated with nitrite. We further validated the beneficial effect of nitrite on neuronal function against hypoxic stress in vivo. Using the wild-type or Apo E-/- mice exposed to chronic hypoxia as a model, we demonstrated that supplementing drinking water with nitrite suppressed active caspase-6 in the cortex of the brain, concomitant with the prevention of hypoxia-induced anxiety in the animals. Innovation: These results are the first evidence of a new pathway for the activation of caspase-6 and the first to indicate that nitrite can protect neurons against chronic hypoxic insult. Conclusion: Our findings suggest that nitrite holds great potential for the treatment of diseases such as COPD associated with hypoxia-induced neuronal injury.