Respiratory pattern and expression of muscarinic receptors in AIRmin and AIRmax mice. Respiratory pattern was determined in awake, unrestrained mice by noninvasive whole-body barometric plethysmography. (a) Penh values were used as an index of bronchoconstriction induced after sequential delivery of increasing concentrations of MCh (3, 6, 12, and 25 mg/mL) and (b) provocative concentration of aerosol MCh at a 200% increase (PC200) over baseline values. Gene (c and e) or protein (d and f) expression of M2 and M3 muscarinic receptors was evaluated by real-time PCR or Western blot analysis in lungs from AIRmin and AIRmax mice. Real-time PCR was carried out using  β -actin gene expression as internal control for normalization of M3R (c) and M2R (e) mRNA transcription levels. All PCR reactions were quantitative reactions made by real-time PCR. In Western blot analysis the density of M3R (d) and M2R (f) protein expression was nor aerosol at a malized to actin expression in each sample. Western blots were quantified by densitometry using the ImageJ software (NIH). Real-time PCR and Western-blot analyses were performed using pooled lungs from 5 mice. Data are expressed as mean ± SEM of five mice per group and are representative of three experiments; * P