Accurate quantification of lung disease severity is essential for optimal clinical management. Measurement of neural respiratory drive (NRD) using the electromyogram of the parasternal intercostal muscles (EMGpara), obligate, extra-diaphragmatic inspiratory muscles, provides a non-invasive index of overall respiratory system load-capacity balance. Previous studies in patients with obstructive lung diseases have shown strong relationships between EMGpara and measures of both airflow obstruction and hyperinflation at a single time point, but have not examined the relative contribution of each to overall respiratory load, either at a single time point or when such parameters are changing dynamically. The aim of this study was to examine the incremental changes in EMGpara induced by chemical airway challenge testing, and to determine the relative influences of bronchoconstriction and increased end-expiratory lung volume on EMGpara activity. We hypothesised that EMGpara would increase in relation to the degree of bronchoconstriction, and that subjects demonstrating hyperinflation would show greater increases in EMGpara. Twenty adult humans with a self-reported history of atopy, asthma or bronchial hyper-responsiveness underwent incremental methacholine challenge testing (maximum dose 32mg.ml-1) using a six-breath dosimeter technique with diluent (0.9% saline) step until a decrease in FEV1 of ≥20% was observed. FEV1 and inspiratory capacity (IC) were measured immediately after each dose, followed by three minutes of tidal breathing during which EMGpara was recorded using surface electrodes over the second intercostal space 3cm lateral to the sternal edge. EMGpara was converted to root-mean-square and expressed as mean peak RMS EMGpara per breath over the final minute of each recording. Significant hyperinflation was defined as a decreased in IC of ≥300ml. Linear mixed model (LMM) analysis was used to analyse dose-wise changes in both FEV1 and EMGpara, and to determine the influence of changes in FEV1 and IC on change in EMGpara. Wilcoxon’s matched pairs testing demonstrated significant decreases in median (IQR) FEV1%predicted (96.9 (86.3-106.9)% to 67.5 (59.4-75.5)%, p<0.0001) and IC%predicted (91.9 (75.6-120.6)% to 84.9 (73.1-103.3)%, p=0.0037) from baseline to end-of-test, and a significant increase in EMGpara (5.4 (4.2-6.1)µV to 6.6 (5.7-8.9)µV, p=0.0012). LMM analysis showed a significant interaction between methacholine dose and induced change in EMGpara (slope (95% confidence interval) 0.27 (0.12-0.42), p=0.001). The degree of change in EMGpara was significantly associated with the extent of decline in FEV1 (mean (95% CI) increase in slope of 0.003 (0.005-0.001) per %predicted fall in FEV1, p=0.011), but not with change in IC. These data suggest that bronchoconstriction exerts a more potent influence on NRD than changes in end expiratory lung volume during methacholine challenge.