Trained vs. Untrained: Lower Extremity Pressure Changes (TULiP)

Purpose and hypothesis

The purpose of the current study is to determine the impact of endurance training status and biological sex on pulmonary diffusing capacity for carbon monoxide (DLCO) and its components (pulmonary capillary blood volume [Vc] and pulmonary membrane diffusing capacity [Dm]) during acute changes in central blood volume using varying levels of lower body positive and negative pressure (LBPP and LBNP) and head down and head up tilt (HDT and HUT).

The investigators hypothesize that for a given reduction (i.e., LBNP, HUT) or increase (i.e., LBPP, HDT) in central blood volume, endurance-trained individuals will demonstrate greater changes in DLCO compared to untrained individuals. As a second exploratory aim, the investigators hypothesize that females will have larger changes in DLCO compared to males.

Research Method/Procedures

All visits will happen on separate days with at least 24 hours between visits. All visits will take place within 4 months.

Visit 1: Screening

On the screening day, the participant will complete the informed consent procedure, fill out a medical history form, and be screened for exercise using the modified medical research council (mMRC) dyspnea scale, PAR-Q+ exercise readiness questionnaire and the Physical Activity and Sedentary Behaviour Questionnaire (PASB-Q). The investigators will then perform pulmonary function tests on each participant. Finally, the participant will complete a graded exercise test on an upright cycle ergometer to volitional exhaustion to determine maximal oxygen consumption (V̇O2max). Evaluation of dyspnea including intensity and unpleasantness will be performed at each workload. Further, reasons for test termination, dyspnea, and qualitative descriptors of dyspnea will be evaluated at peak exercise. Hemoglobin concentration will be measured via finger prick prior to and following completion of exercise testing.

Visit 2: Central Blood Volume Manipulation and DLCO

The participant will perform the DLCO test at various levels of LBNP/LBPP ranging from -80 mmHg to +40 mmHg, or HUT/HDT ranging between 90° and -12°. Head tilt and lower body pressure changes provide a similar intervention that can be used interchangeably based on equipment availability. Changes in central blood volume during LBNP/HUT are equivalent to standing upright (-80 mmHg LBNP or 90° HUT), and changes in central blood volume during LBPP/HDT are equivalent to someone lying down with their feet above their head (+40 mmHg LBPP or -12° HDT). For the LBNP/LBPP trials, participants will lay supine with their lower body (from the iliac crest to their toes) inside the pressure chamber. The participant's lower body will be sealed within the differential pressure chamber using a neoprene skirt to prevent any air leakage from the insides of the chamber. Negative and positive pressures are induced by a vacuum pump that either reduces or increases pressure inside the chamber in a controlled and precise manner. The vacuum pump is controlled by a trained member of the research team, and chamber pressure is continuously monitored via a digital manometer. For the HUT/HDT trials, the participant will lie supine on a tilt table. The table will be electronically adjusted to the desired angle at a rate of 1°/second.

Once participants are secured in the lower body pressure chamber or on the tilt table, a certified technician will insert an intravenous (IV) catheter into an antecubital vein which will allow peripheral venous pressure measurements. Participants will also be equipped with a continuous blood pressure monitor, a brachial blood pressure cuff, an electrocardiogram, and a thoracic impedance device. Each stage of LBPP/LBNP or HUT/HDT will begin with two minutes of stabilization followed by two DLCO breath-hold maneuvers. At each stage participants will also breathe on a mouthpiece for two minutes to collect metabolic data: ventilation (V̇E), oxygen consumption (V̇O2), and carbon dioxide production (V̇CO2). Cardiovascular parameters (blood pressure, peripheral venous pressure, heart rate, thoracic impedance) will be measured continuously. Hemoglobin will be measured at each stage via a small (< 1 mL) blood sample from the IV. At the end of the trial, a certified technician will obtain a ~5 mL blood sample from the IV for sex hormone analyses.

Participants will not complete more than 15 DLCO breath holds during each experimental visit. As per recommendations, participants will perform standard DLCO breath-hold maneuvers for 6 seconds, and maneuvers will be performed twice at each stage. A washout period of at least 4 minutes between each breath hold will be completed to ensure adequate recovery and blood gas equilibration.

Visit 3: DLCO with Exercise

Exercise will be used as a physiological stressor to increase central blood volume beyond that of LBPP or HDT. The participant will perform the DLCO maneuvers during exercise at varying exercise intensities (~20%-100% of an individual's V̇O2max). Participants will not complete more than 15 breath holds during the experimental visit. As per recommendations, participants will perform standard DLCO breath-hold maneuvers for 6 seconds, and maneuvers will be performed twice at each stage. At each exercise intensity, participants will breathe on a mouthpiece for two minutes to collect metabolic data; heart rate and peripheral oxygen saturation will be measured continuously for the same two minutes at each intensity. Blood pressure will be measured once per stage with an automatic sphygmomanometer. Hemoglobin via finger prick will also be measured at each intensity. A washout period of at least 2 minutes between each breath hold during exercise will be completed to ensure adequate recovery and blood gas equilibration. At the end of the trial, a certified technician will obtain a ~5 mL blood sample from an antecubital vein for sex hormone analyses.

Visit 4: Ultrasound with LBNP/HUT, LBPP/ HDT, and Exercise

Participants will complete the same interventions as visit 2 (LBNP/LBPP -80 to +40 mmHg, or HUT/HDT 90° to -12°) and visit 3 (exercise at ~20%-100% of an individual's V̇O2max). At each stage of LBNP/LBPP or HUT/HDT, and exercise, a trained sonographer will obtain images of the heart via echocardiography. Participants will then undergo a carbon monoxide rebreathe protocol to estimate blood volume. A certified technician will insert an IV catheter into an antecubital vein to collect serial blood samples. While lying supine, participants will inhale a bolus of carbon monoxide (0.7-1.0 mL/kg bodyweight) mixed with 100% oxygen on a rebreathing circuit for two minutes and will then continue to rest for eight additional minutes. Venous blood samples measuring carboxyhemoglobin before the rebreathe and eight minutes after the rebreathe will allow the calculation of blood volume.

Visit 5: Computed Tomography

Participants will report to a Medical Imaging Centre for the computed tomography (CT) scan. Following a briefing by the Medical Imaging Centre staff, participants will lay supine on the CT scanner and the region of interest in the chest will be obtained. The participant will remain as still as possible and inhale to total lung capacity and hold their breath. During the breath hold, images will be acquired. The participant will then empty their lungs as far as they can and imaging will be repeated.

CT can noninvasively detect features of pulmonary vascular dysfunction. CT outcomes-such as pulmonary artery diameter, vessel pruning, perfusion heterogeneity, and right ventricular morphology-correlate with hemodynamic abnormalities and impaired pulmonary vascular reserve. These imaging biomarkers enable early identification and monitoring of pulmonary vascular disease, providing clinically relevant information beyond conventional structural assessment. In healthy individuals, CT can provide valuable information characterizing pulmonary structure not available using other non-invasive techniques. For example, CT can provide quantitative information on pulmonary vascular and airway branching to show potential structural differences inherent to highly fit individuals. CT is therefore a tool to characterize differences in lung structure which may also potentially explain differences in lung function elucidated by the other measures in this study.