Intro: Hypoxia leads to neonatal pulmonary vascular remodeling, and is associated with smooth muscle hypertrophy and hyperplasia.Hypothesis: We hypothesized that hypoxia increases neonatal pulmonary artery cell (PASMC) proliferation, and that this change is mediated either by induction of the transcription factors Hypoxia-Inducible Factor-1(HIF-1, known to be induced specifically by hypoxia), AP-1, and/or CREB, or by changes in activation of the Mitogen Activated Protein Kinase (MAPK) pathway.Methods: PASMC proliferation in response to varying oxygen tension was assessed by exposing near confluent PASMC in 24-well plates in MEM+10%FBS and in serum-free media to oxygen concentrations of 1-20% for 72 hours, followed by electronic cell counts (n=12 at each O2 level). For assay of the transcription factors HIF-1, AP-1, and CREB, confluent PASMC were exposed to 2, 7, or 20% O2 for four hours and nuclear extracts prepared. Gel shift analysis (n=9 at each O2 level) was done using 32P-labelled oligonucleotide probes. In addition to samples, specific (unlabelled similar oligo) and non-specific (unlabelled different oligo) inhibitors were used to confirm identity of the transcription factor. MAPK was estimated using a specific radioimmunoassay on cell lysates of confluent PASMC exposed for 20 minutes(Short-term) or 18 hours (Long-term) to 2, 7, or 20% O2. Results: O2 levels < 7% inhibited PASMC proliferation in serum-containing media. In serum-free media, cell counts were lower overall but O2 levels of 1% increased cell counts compared to 5% or 20%. No change was noted in HIF-1 levels, but AP-1 and CREB showed marked inter-sample variation (at each O2 level) and correlated with one another. MAPK was increased at 20 minutes in the 2% and 20% O2 samples compared to the 7% O2 sample (p<0.05), but MAPK levels were similar at 18 hours. Conclusions: Hypoxia inhibits PASMC proliferation in serum-containing media (but not in serum-free media), and this inhibition is not due to HIF-1 induction, or MAPK inhibition, but due to inhibition of serum-stimulated proliferation. Marked variations in AP-1 and CREB levels make it difficult to attribute changes of PASMC proliferation to these transcription factors. It is possible that other pathways important in serum response (SRE) or stress response (SAPK) may mediate hypoxia-induced changes in PASMC proliferation.