Nerve growth factor enhances cholinergic innervation and contractile response to electric field stimulation in a murine in vitro model of chronic asthma

Background Asthma is a chronic inflammatory disease characterized by airway hyper-responsiveness. Alterations in the neurogenic control are believed to contribute to the pathogenesis. Yet, the long-term interaction between nerves and inflammatory mediators, such as the neurotrophin nerve growth factor (NGF), are not fully understood much due to the absence of appropriate experimental assays. Objective To develop an ex vivo mouse organ culture assay and to investigate the effects of NGF on nerve-mediated airway contractions. Method Mouse tracheal segments were cultured in periods of up to 16 days. Their contractile responses to electric field stimulation (EFS) were investigated. In addition, the effect of 4 days of NGF treatment was analysed using EFS and immunohistochemistry. Results EFS (0.2-25.6 Hz) induced reproducible and frequency-dependent cholinergic contractions of both fresh and cultured tracheal segments. The main part of the EFS response was blocked by tetrodotoxin or atropine. After 4 days in culture, regional differences appeared, with stronger EFS responses in distal than in proximal segments. More nerve fibres were seen in distal segments than in proximal segments. Treatment with NGF during 4 days of culture increased the innervation of the proximal segments, at the same time as the cholinergic contractile responses to EFS were enhanced dose-dependently. Conclusion Culture of tracheal segments appears to be a suitable assay for the examination of long-term effects induced by inflammatory mediators on neurally mediated airway contractions. NGF treatment enhanced the cholinergic, nerve-dependent contractions and increased the amount of nerve fibres seen in the murine tracheal segments, suggesting a role for NGF in the development of airway hyper-responsiveness.