Methodology:

Pteropod adult (>0.5 mm, with fully developed wings/parapodia) were captured for experiments using a Reeve net with 333-m mesh and a large cod end that was deployed at slow speeds, for a short duration (< 1 h) with the aim of gently sampling live, undamaged specimens on various cruises. Tows were conducted at multiple stations in the Gulf of Maine (C42 22.1’ - 42 0.0’ N and 69 42.6’ - 70 15.4’ W). Individuals for respiration experiments were returned via coolers to a walk-in cold-room facility at the Woods Hole Oceanographic Institution within 8 h of collection. The cold-room was set at a constant temperature of 8C for all experiments. During each experimental period, water had been collected from an offshore site concurrent to pteropod sampling and had been returned to the lab in advance for filtration (0.2-m) and thermal equilibration. It had been bubbled with ambient air, which ranged from a calculated 380 to 440 atm over the seasonal cycle, for ~12 h prior to the arrival of the pteropods. Upon arrival, pteropods were placed in 1-L beakers of water containing the filtered in situ water (15 ind L-1) for 8-12 h to allow for further temperature acclimatization and gut clearance. After this period, healthy looking individuals (actively swimming or with parapodia extended) were placed into glass respiration chambers (containing optode spots; OXFOIL: PyroScience, Aachen Germany) with 2-3 mL fresh 0.2-m filtered water. A control chamber, without a pteropod, was set up for every fourth chamber to determine background bacterial respiration. The oxygen concentration was measured non-invasively at the start of the experiment and 24 h later using a FireStingO2 optical oxygen meter (PyroScience, Aachen Germany). At the conclusion of the experiment, each organism was visually inspected to confirm survival, briefly rinsed with DI water, placed in a pre-weighed aluminum dish, and weighed on a Cahn microbalance (C-33; 1 g precision) for wet mass. They were then dried for 3-7 days in a drying oven at 60C and weighed again to obtain dry mass. Final oxygen consumption rates were calculated based on the change in oxygen between the final and initial oxygen measurements, corrected for the bacterial respiration from the control chambers (mol O2 h-1).

Sampling and analytical procedures:

Temperature in the experiments was continually monitored using the Pyrosciences temperature logger that comes with the Firesting meter and are reported for only the ~24 duration of the respiration experiments.

Carbonate chemistry parameters are calculated as the average (over the course of the whole 14 day experiment) of measured pH, DIC and TA values using CO2sys. To monitor the carbonate chemistry during the experiments, subsamples of water (50 mL) from each carboy were taken every 2–3 days and measured spectrophotometrically for pH as described in White et al. (2013). To more fully characterize the carbonate chemistry of the experimental treatments, discrete samples for analysis of dissolved inorganic carbon (DIC) and total alkalinity (TA) were also collected from the pre-equilibrated water at weekly intervals, prior to its transfer to the carboys (day 0 and 7) and from the water in each carboy right before it was replaced during water transfers (day 7 and 14). This provided both the starting and ending DIC and TA of each batch of water for the first two weeks, and the starting conditions for week 3. DIC was measured using an Apollo SciTech DIC auto-analyzer, while TA was measured using an Apollo SciTech alkalinity auto-titrator, a Ross combination pH electrode, and a pH meter (ORION 3 Star) based on a modified Gran titration method (detailed in Wang et al., 2017). pH is reported on the seawater scale (pHsw). Salinity was measured with a Hannah refractometer.