Fig 1.
Schematic of the workflow for swim-up of bull sperm and subsequent metabolic measurements.
Fig 2.
Basal sperm metabolic rates over time in three individuals belonging to different breeds.
Oxygen consumption rates (OCR) (top) and extracellular acidification rates (bottom) of bull sperm before swim-up (red symbols), upper (yellow symbols) and lower fractions (blue symbols) after swim-up procedure of three bulls of different cattle breeds. Each data point is obtained from a mean value from 3 wells and the data were normalized to the number of cells in each well. Error bars are standard deviation from 15 reads and 3 wells.
Fig 3.
Oxygen consumption rate (OCR) in low viscosity medium (Assay Medium, indicated as light green) and high viscosity medium (Assay medium supplemented with 1% MC, indicated with darker green) of bull sperm after undergoing the swim-up procedure in low viscosity sperm medium SP-TALP. a) Schematic of the sample distribution in the 8 well plate. This triple-well measurement was performed on each fraction (upper, lower and before swim-up) and two control wells with the two levels of viscosity were used as background level. b) Oxygen consumption rate (OCR) and c) extracellular acidification rate (ECAR) for the before swim-up sperm, the upper and lower swim-up fractions of 12 bulls of three different breeds (Holstein, Fleckvieh, Angus) in two levels viscosity (labelled with light and dark green background). The horizontal lines across the boxes display the median values. The upper fraction shows significantly higher OCR (OCR, p = 0.02) and ECAR (p = 0.00019) compared to the lower fraction and before swim-up. Viscosity does not change the OCR or ECAR values significantly.
Fig 4.
ATP production by oxidative phosphorylation of different swim-up fractions.
a) ATP production from oxphos is calculated as the difference between basal OCR rate and OCR rate after oligomycin injection. b) Average ATP production due to oxphos of unselected sperm (before-swim-up), upper and lower swim-up fractions. Each box contains data from 12 bulls. Horizontal lines through each box are the median values, lower and upper limits are 25th and 75th percentile, respectively.
Fig 5.
Total ATP content per sperm cell of before swim-up fractions, upper and lower swim-up fractions from 12 bulls measured with a luminescence-based ATP kit.
Each condition was measured three times.
Fig 6.
Sperm flagella length of swim-up fractions from 9 bulls of different breeds.
100 sperm cells were measured for each bar. Error bars are standard error of the mean. Upper fractions contain consistently longer sperm tails (estimate = 1.05, p = 0.04) compared to the initial sperm sample. The lower fractions contain consistently shorter tails (estimate = -0.98, p = 0.05). BS = before swim-up. UF = upper fraction. LF = lower fraction. H0-3 = Holstein bulls, FV0-3 = Fleckvieh bulls, A0-2 = Angus bulls.