To explore the role of intrapulmonary neuropeptides in the development of airway hyperresponsiveness, we established an animal model of airway hyperresponsiveness (AHR) in rabbits by using ozone exposure. With the model, after test of the mechanics of respiration and bronchoalveolar lavage assay, the levels of vasoactive intestinal peptide (VIP) and calcitonin gene-related peptide (CGRP) in the lungs were determined by radioimmunoassay, and the expression of mRNA coding receptors of these two neuropeptides was evaluated by reverse transcriptional-polymerase chain reaction (RT-PCR). At the same time, the distribution of VIP receptor-1 (VIPR1) and CGRP receptor-1 (CGRPR1) in lung tissues and its time-course were examined by in situ hybridization. The results showed: (1) in ozone-stressing groups, airway resistance increased significantly and typical inflammatory pathological changes were observed in pulmonary tissue slides, including neutrophil and eosinophil infiltration, mucus exudation and bronchial epithelial cells (BECs) shedding; (2) with elongation of ozone exposure, the levels of VIP and CGRP in the lungs increased at first, reaching a peak on d 2 to 4, then decreased slowly, and CGRP peaked somewhat earlier than VIP; (3) mRNA expression of the two neuropeptide receptors in the lungs changed in a similar manner like VIP and CGRP, but the high level of mRNA expression of VIPR1 lasted longer than that of CGRPR1; and (4) in situ hybridization for neuropeptide receptors demonstrated that, in unstressed control, VIPR1 and CGRPR1 positive cells appeared in the airway epithelium, pulmonary interstitial and focal areas of airway and vascular smooth muscles. With the elongation of ozone exposure, hybridization stained deeper and the majority of positive cells were located around the vessels and bronchus except a few in the alveoli. At 8 d, only a small number of positive cells were seen in the lungs. From the results, it is concluded that ozone-stressing can induce the development of AHR, in which VIP and CGRP may play important roles. That implies, through binding to CGRPR1, CGRP stimulates an early inflammation response which contributes in cleaning up of irritants, while VIP exerts a later dampening of pulmonary inflammation response. These two neuropeptides may play sequential and complementary roles in the development of AHR.