Young and middle‐aged mouse breathing behavior during the light and dark cycles

Unrestrained barometric plethysmography is a common method used for characterizing breathing patterns in small animals. One source of variation between unrestrained barometric plethysmography studies is the segment of baseline. Baseline may be analyzed as a predetermined time‐point, or using tailored segments when each animal is visually calm. We compared a quiet, minimally active (no sniffing/grooming) breathing segment to a predetermined time‐point at 1 h for baseline measurements in young and middle‐aged mice during the dark and light cycles. Additionally, we evaluated the magnitude of change for gas challenges based on these two baseline segments. C57BL/6JEiJ x C3Sn.BliA‐Pde6b (+)/DnJ male mice underwent unrestrained barometric plethysmography with the following baselines used to determine breathing frequency, tidal volume (VT) and minute ventilation (VE): (1) 30‐sec of quiet breathing and (2) a 10‐min period from 50 to 60 min. Animals were also exposed to 10 min of hypoxic (10% O(2), balanced N(2)), hypercapnic (5% CO (2), balanced air) and hypoxic hypercapnic (10% O(2), 5% CO (2), balanced N(2)) gas. Both frequency and VE were higher during the predetermined 10‐min baseline versus the 30‐sec baseline, while VT was lower (P < 0.05). However, VE/V(O) (2) was similar between the baseline time segments (P > 0.05) in an analysis of one cohort. During baseline, dark cycle testing had increased VT values versus those in the light (P < 0.05). For gas challenges, both frequency and VE showed higher percent change from the 30‐sec baseline compared to the predetermined 10‐min baseline (P < 0.05), while VT showed a greater change from the 10‐min baseline (P < 0.05). Dark cycle hypoxic exposure resulted in larger percent change in breathing frequency versus the light cycle (P < 0.05). Overall, light and dark cycle pattern of breathing differences emerged along with differences between the 30‐sec behavior observational method versus a predetermined time segment for baseline.