DOI: 10.5061/dryad.hdr7sqvmw
title: Metabolic rate increases with thermal acclimation and is associated with mitochondrial function in some tissues of threespine stickleback

description: The metabolic rate (ṀO2) of eurythermal fishes changes in response to
 temperature, yet it is unclear how changes in mitochondrial function
 contribute to changes in ṀO2. We hypothesized that ṀO2 would increase with
 acclimation temperature in the threespine stickleback (Gasterosteus
 aculeatus) in parallel with metabolic remodeling at the cellular level but
 that changes in metabolism in some tissues and organs, such as liver,
 would contribute more to changes in ṀO2 than others. Threespine
 stickleback were acclimated to 5, 12 and 20°C for 21 weeks. At each
 temperature, standard and maximum metabolic rate (SMR and MMR,
 respectively), and aerobic scope (AS) were quantified, along with
 mitochondrial respiration rates in liver, oxidative skeletal, and cardiac
 muscles, and the maximal activity of citrate synthase (CS) and lactate
 dehydrogenase (LDH) in liver, and oxidative and glycolytic skeletal
 muscles. SMR, MMR and AS increased with acclimation temperature, along
 with rates of mitochondrial phosphorylating respiration in all tissues.
 Low SMR and MMR at 5°C were associated with low or undetectable rates of
 mitochondrial complex II activity and a greater reliance on complex I
 activity in liver, oxidative skeletal muscle, and heart. SMR was
 positively correlated with cytochrome c oxidase (CCO) activity in liver
 and oxidative muscle but not mitochondrial proton leak, while MMR was
 positively correlated with CCO in liver. Overall, the results suggest that
 changes in ṀO2 in response to temperature are driven by changes in some
 aspects of mitochondrial function in some, but not all tissues of
 threespine stickleback.
description: Threespine stickleback were
 collected using minnow traps. Animals were transported to the University
 of Alaska Fairbanks where they were maintained between 7 to 21 weeks at
 three different acclimation temperatures (5, 12 or 20°C). Standard and
 maximum metabolic rates (SMR and MMR, respectively) were measured using a
 170 mL glass Loligo swim tunnel respirometer (Loligo Systems, Viborg,
 Denmark). Mitochondrial respiration rates were measured in liver,
 oxidative muscle and heart at the acclimation temperature of the animal
 (5, 12 or 20°C) using an Oroboros O2k high resolution respirometer
 (Oroboros Instruments, Innsbruck, Austria) following a
 substrate-uncoupler-inhibitor titration (SUIT) protocol. Values were
 normalized per g tissue. Maximal activity of citrate synthase and lactate
 dehydrogenase were measured following a modified method by Srere et al.
 (1963) and by Hansen and Sidell (1983), respectively, in liver, oxidative
 and glycolytic muscle in fish acclimated to the three temperature
 groups. Metabolic rates data were collected using
 AutoResp version 2.3.0 (Loligo Systems, Viborg, Denmark) and processed
 using RStudio version 1.3.1093 (R Studio Team 2016). Code was modified
 from the “FishResp” package version 1.0.3 (Morozov et al., 2019) to
 correct raw data for background ṀO2. To obtain SMR values, dissolved
 oxygen (DO) values were smoothed by removing cycles with r2 &lt; 0.95.
 The final SMR estimate for each animal was calculated using the mean of
 the lowest normal distribution (MLND) when the coefficient of variation of
 MLND (CVMLND) ≥ 5.4, and the 20% quantile method when CVMLND ≤ 5.4. To
 obtain MMR values, we either 1) fitted DO values to a linear model to
 estimate the slope of the line. In this case, the entire (5 min)
 measurement period that yielded the highest estimate was chosen as MMR
 (hereafter referred to as “traditional MMR”). And 2) employed a second
 approach (hereafter referred to as “truncated MMR”), in which each
 measurement period was edited to retain only the steepest declines in DO
 as long as there was a minimum of 100 s (33% of the measurement period) of
 data remaining (mean ± s.d. = 225 ± 78 s, range = 100 – 300 s) and
 processed as described above for SMR. Finally, Absolute aerobic scope
 (AAS) was calculated as MMR – SMR.
 The temperature coefficient
 (Q10) were calculated for each mitochondrial function and for metabolic
 rate between 5 and 12°C and between 12 and 20°C using the following
 formula: Q10 = (R2/R1)^(10/(T2-T1)), where R1 and R2 are the mean values
 after removing outliers at temperature T1 or T2, respectively.
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