Breathing maneuver protocol.
For the combined hyperventilation breath-hold (HVBH) maneuver, a single rest measurement was obtained in a short breath-hold (A). The animal was then manually hyperventilated for 60s (B) followed immediately by a long breath-hold (C) that was imaged throughout, with a repeating OS sequence. Hyperventilation analysis was always compared between rest and the start of the breath-hold (red arrow), while the breath-hold could be analyzed at multiple time points with comparison to data obtained at the beginning of the breath-hold (purple arrow). The long breath-hold (LBH) followed step C, starting after a normal ventilation pattern.