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Subjects always commenced with the single breath of room air, but the order of presentation http://www.selleckchem.com/products/Methazolastone.html of 100% N2 or 10% CO2 in N2 was randomized. For the maximal inspiratory-capacity apnoeas, subjects were instructed to perform a maximal inspiration and hold their breath at maximal volume (against a closed glottis) for 40?s. In four subjects each manoeuvre was repeated once, while in six subjects the manoeuvre was repeated twice; in these cases the mean responses were calculated for each subject. In one subject each manoeuvre was performed only once. Analysis of MSNA burst parameters was carried out using Chart software (version?5.1; ADInstruments) after shifting the neurogram back in time by 1.15�C1.35?s so as to remove peripheral conduction delays (Fagius & Wallin, 1980) and align the peak of an individual burst to the R?wave of the ECG responsible for its inhibition. The period of inhibition of MSNA at the asphyxic break point was measured with cursors from the start of the inspiratory airflow to the first burst of MSNA. Systolic and diastolic pressures were calculated for each of the R?waves during baseline and during the apnoea, immediately prior to the single breath of 100% N2 or 10% CO2 in 90% N2. An additional 10 R?waves were measured immediately following the single breath of 100% N2. Only five R?waves were measured following Moroxydine the single breath of 10% Vismodegib ic50 CO2 in 90% N2, because the breath hold could only be maintained for a short period owing to the higher chemical drive. Systolic and diastolic pressures were also calculated for 10 R?waves during the initial recovery period. Average pressures were calculated over 10?s periods prior to the apnoea, immediately before and after the single breath and during the recovery period, with the exception of the post-10% CO2 in 90% N2 breath, for which a 5?s period was used. All data are presented as means?��?SEM. Analysis of variance and Student's paired t?tests were used for statistical analysis (Prism?5; GraphPad Software, San Diego, CA, USA). Differences were considered significant at P?