This study investigates the effect of increased ventilation on the expression of messenger ribonucleic acid (mRNA) levels of vascular endothelial growth factor (VEGF), basic fibroblast growth factor (bFGF) and transforming growth factor-beta1 (TGF-beta1) in the diaphragm of intact, awake, spontaneously breathing rats, compared with responses in paralysed, mechanically-ventilated animals at similar blood gas and ventilatory levels. Four groups of intact, rats were studied in a body box, each group breathing one of four gases: room air, 12% oxygen (O2), 5% carbon dioxide (CO2), or 12% O2+5% CO2 for 1 h. Another 4 groups of paralysed, mechanically-ventilated animals were matched for arterial blood gas and ventilatory level. The results showed that VEGF mRNA abundance was increased three-fold and that of bFGF 1.5-fold when 12% O2+5% CO2 were breathed, but TGF-beta1 did not change. A significant linear relationship of VEGF and bFGF mRNA to minute ventilation was observed in awake animals (r=0.98, p<0.02 and r=0.87, p<0.03, respectively). The paralysed, mechanically-ventilated animals showed no mRNA increases for any probe. Systemic hypoxia had no additional effect on VEGF or bFGF levels in the diaphragm. It was concluded that messenger ribonucleic acid for vascular endothelial growth factor and basic fibroblast growth factor in the diaphragm rises significantly as a result of active ventilation and not due to blood gas/pH changes or to passive muscle shortening per se.