Caffeine inhibits hypoxia-induced nuclear accumulation in HIF-1α and promotes neonatal neuronal survival

Apnea of prematurity (AOP) defined as cessation of breathing for 15-20 s, is commonly seen in preterm infants. Caffeine is widely used to treat AOP due to its safety and effectiveness. Caffeine releases respiratory arrest by competing with adenosine for binding to adenosine A₁ and A_2A receptors (A₁R and A_2AR). Long before its use in treating AOP, caffeine has been used as a psychostimulant in adult brains. However, the effect of caffeine on developing brains remains unclear. We found that A₁R proteins for caffeine binding were expressed in the brains of neonatal rodents and preterm infants (26-27 weeks). Neonatal A₁R proteins colocalized with PSD-95, suggesting its synaptic localization. In contrast, our finding on A₂R expression in neonatal neurons was restricted to the mRNA level as detected by single cell RT/PCR due to the lack of specific A_2AR antibody. Furthermore, caffeine (200 μM) at a dose twice higher than the clinically relevant dose (36-130 μM) had minor or no effects on several basic neuronal functions, such as neurite outgrowth, synapse formation, expression of A₁R and transcription of CREB-1 and c-Fos, further supporting the safety of caffeine for clinical use. We found that treatment with CoCl₂ (125 μM), a hypoxia mimetic agent, for 24 h triggered neuronal death and nuclear accumulation of HIF-1α in primary neuronal cultures. Subsequent treatment with caffeine at a concentration of 100 μM alleviated CoCl₂-induced cell death and prevented nuclear accumulation of HIF-1α. Consistently, caffeine treatment in early postnatal life of neonatal mice (P4-P7) also prevented subsequent hypoxia-induced nuclear increase of HIF-1α. Together, our data support the utility of caffeine in alleviating hypoxia-induced damages in developing neurons.