DOI: 10.5061/dryad.wh70rxwnz
title: Oxygen measurements in closed system respirometry using blubber explants from suckling and fasting grey seal pups

description: Understanding physiological responses of wildlife and domesticated animals
 to environmental challenges can be difficult or impossible to investigate
 at the whole animal level. Tissue culture approaches open up experimental
 possibilities, but are difficult to undertake in remote environments where
 facilities to assess tissue viability and minimise or monitor microbial
 contamination may not be available. Here we used planar optodes in closed
 system respirometry to measure oxygen use by blubber explants from grey
 seal (Halichoerus grypus) pups, obtained in a remote field environment in
 2017 and processed in a field laboratory with minimal equipment. These
 data show oxygen measurements at timed intervals and demonstrate that the
 tissues were respiring aerobically for the 24 h measurement period,
 compared to empty control vials. We also measured glucose, lactate and
 glycerol levels in culture media after 24 hours to allow correlation of
 glucose uptake, lactate accumulation and lipolytic rate, which are also
 useful markers of metabolic activity, with oxygen use data. Nutritional
 state of the pups (either suckling or fasting) and tissue depth (closer to
 muscle (inner) or closer to skin (outer)) were recorded to allow effects
 of tissue characteristics on metabolic parameters to be examined. We were
 also able to use the oxygen use profiles in explant-containing and control
 vials to identify those contaminated by microbes well ahead of any visible
 colour change in the media stocks. The use of planar optodes for oxygen
 measurement thus allows simultaneous physiological measurements and
 contamination monitoring to be undertaken in a minimally equipped field
 laboratory. Robinson et al. (2021) published in Methods in Ecology and
 Evolution provides the details of tissue collection and tissue culture.
 These data will facilitate comparison of metabolic activity of adipose
 tissue within and between species and assessment of efficacy of monitoring
 microbial contamination in tissue culture experiments, particularly in
 field laboratories where other methods may not be logistically possible.
description: This file contains data from 19 grey seal pups (Halichoerus
 grypus) captured on the Isle of May, Firth of Forth, Scotland (56°11′N,
 2°33′W) during the breeding season (Oct-Dec) of 2017. Individuals were
 sampled for blubber tissue, which was used to generate blubber explants of
 both inner (close to animal's core) and outer (close to the skin)
 tissue that were monitored for a 24 hour period for oxygen consumption in
 sealed vials. Oxygen sensitive planar optodes (OSPO) were used to measure
 oxygen use (see usage notes for more details). The culture media was
 collected after the 24 hour period ended for analysis of glucose, glycerol
 and lactate. In some vials (n=49 of 103) an experimental exposure of PCBs
 were added (see usage notes for details) however this treatment had no
 impact on blubber explants, with no difference in metabolite or oxygen use
 data from controls. Some animals were sampled during the suckling period
 (n = 9) and some were sampled their natural postweaning fast (n = 13). Of
 these, 3 individuals were sampled during both nutritional states.
 Information on capture, handling and samples taken for this dataset are
 given in the publication Robinson, K. J., Armstrong, H. A., Moss, S. E. W,
 Oller, L., Hall, A.J., & Bennett, K. A. (2021). Signs of life:
 Oxygen sensors confirm viability, measure oxygen consumption and provide
 rapid, effective contamination monitoring for field-based tissue culture.
 Methods in Ecology and Evolution (Robinson et al. 2021).
 Data were recorded in Microsoft Excel and was imported into R via
 R studio to allow data exploration and analysis. Oxygen sensors were
 supplied calibrated by the company that manufactures them, and do not need
 to be re-calibrated unless they exceed 10,000 reads, which did not occur
 in this study. Calibration to standardise oxygen reading for each optode
 at 35 degrees C is described in Robinson et al. 2021. Glucose, lactate and
 glycerol assays are performed in duplicate on all samples and are detailed
 in Robinson, K. J., Hall, A., Debier, C., Eppe, G., Thome, J-P., &
 Bennett, K. A. (2018). Persistent organic pollutant burden, experimental
 POP exposure and tissue properties affect metabolic profiles of blubber
 from grey seal pups. Environmental Science and Technology, 52(22),
 13523-13534. https://doi.org/10.1021/acs.est.8b04240 (Robinson et al.
 2018).
description: Each row contains all the data for one vial containing 1.5ml of
 cell culture media and an oxygen spot sensor that measures oxygen
 concentrations in the media across a 24 hour period. Some vials contain
 100mg explants of adipose tissue from the seal pups sampled (n=70) and
 some vials contain no tissue to act as controls for this validation study
 (n=33). Variable List: Pup_ID: one
 of 2 letters followed by 4 digits. Unique identifier for each pup in the
 study based on PHATS_letter for each year and the year in which it was
 born. This can be cross referenced with
 File_1_ile_1_Bennett_Grey_Seal_Pups_2015_to_2017_POPS_mass_TH<br>
 PHATS_letter: one or two letters. Pups are assigned a letter
 alphabetically within each year in the order in which they enter the
 study, which helps with keeping track of the sampling during the field
 season.<br> Vial_ID: Text string containing the pup ID plus an X
 where the animal was fasted and a unique number for the individual vial or
 C_ followed by a number for the controls that did not contain
 explants<br> Empty_filled: Single word, 'filled' indicates
 vials that contain explants, 'empty' indicates vials that do not
 contain explants.<br> Nutritional_state: Single word,
 'feed' indicates vials contain explants from animals that were
 suckling at the time of sampling, 'fast' indicates vials contain
 explants from animals that had weaned and were in a fasting state at the
 time of sampling<br> Tissue_type: Single word, 'inner'
 indicates vials contain explants consisting of tissue from the inner
 portion of the biopsy sample which contains cells close to the body core,
 'outer' indicates vials contain explants consisting of tissue
 from the outer portion of the biopsy sample which contains cells close to
 the skin<br> Microbe_contamination: Single word, 'clean'
 indicates vials that were not suspected to have been contaminated while in
 the experiment, 'infected' indicates vials that are suspected to
 have been contaminated while in the experiment<br> Treatment: Single
 word, 'control' indicates vials containing cell culture medium
 with no pollutants added, 'PCB' indicates vials containing cell
 culture media that has been spiked with 150 ng/mL POP mix (PCB Aroclor
 standard mix 1 (Sigma-Aldrich), see Robinson, K. J., Hall, A. J., Debier,
 C., Eppe, G., Thomé, J. P., & Bennett, K. A. (2018) Persistent
 organic pollutant burden, experimental POP exposure, and tissue properties
 affect metabolic profiles of blubber from gray seal pups Environmental
 science & technology 52, no. 22 (2018): 13523-13534 (Robinson et
 al. 2018))<br> GLU: number to 6 dp. Glucose removal rate in vials in
 µmol per 100 mg tissue per hour. Internal quality control measurements lay
 within &lt; +/− 15%. Intra and inter assay variability for sample
 analysis was &lt;15% and &lt;10% respectively. For assay details
 and QC see Robinson et al. 2018 and Bennett, K.A., Robinson, K.J.,
 Armstrong, H.C., Moss, S.E., Scholl, G., Tranganida, A., Eppe, G., Thomé,
 J.P., Debier, C. and Hall, A.J., (2021). Predicting consequences of
 POP-induced disruption of blubber glucose uptake, mass gain rate and
 thyroid hormone levels for weaning mass in grey seal pups.
 <i>Environment international</i>, <i>152</i>,
 p.106506 (Bennett et al. 2021).<br> LAC: number to 6 dp. Lactate
 accumulation rate in vials in µmol per 100 mg tissue per hour. Internal
 quality control measurements lay within &lt; +/− 15%. Intra and inter
 assay variability for sample analysis was &lt;15% and &lt;10%
 respectively. See Robinson et al. 2018 and Bennett et al. 2021 for assay
 details and QC<br> GLY: number to 6 dp. Glycerol accumulation rate
 in vials in µmol per 100 mg tissue per hour. Internal quality control
 measurements lay within &lt; +/− 15%. Intra and inter assay
 variability for sample analysis was &lt;15% and &lt;10%
 respectively. See Robinson et al. 2018 and Bennett et al. 2021 for assay
 details and QC<br> O2_0Hr: number to 6 dp. Dissolved oxygen
 concentration in mg/L in the cell culture media in the vials measured by
 oxygen spot sensor. This column contains pre-explant addition readings,
 which were all taken at 17 hours after vial creation and 10 hours before
 explant addition. See Robinson et al. 2021 for further methods
 details.<br> O2_1Hr: number to 6 dp. Dissolved oxygen concentration
 in mg/L in the cell culture media in the vials measured by oxygen spot
 sensor one hour after explants had been added to begin an experiment. See
 Robinson et al. 2021 for further methods details.<br> O2_5Hr: number
 to 6 dp. Dissolved oxygen concentration in mg/L in the cell culture media
 in the vials measured by oxygen spot sensor five hours after explants had
 been added to begin an experiment. See Robinson et al. 2021 for further
 methods details.<br> O2_9Hr: number to 6 dp. Dissolved oxygen
 concentration in mg/L in the cell culture media in the vials measured by
 oxygen spot sensor nine hours after explants had been added to begin an
 experiment. See Robinson et al. 2021 for further methods
 details.<br> O2_24Hr: number to 6 dp. Dissolved oxygen concentration
 in mg/L in the cell culture media in the vials measured by oxygen spot
 sensor twenty-four hours after explants had been added to begin an
 experiment, and just prior to the experiment ending and all cells and
 culture media being collected for analysis. See Robinson et al. 2021 for
 further methods details.<br> Sensor_ID: Text string containing S and
 a unique number for the individual sensor ID in the vial used to obtain
 oxygen concentration. Sensors are oxygen sensitive planar optodes (OSPO),
 type SP-PSt3-NAU made by Presens (Precision Sensing GmbH). See Robinson et
 al. 2021 for further methods details. Missing data
 codes:<br> null = not collected for that vial. Only present for
 glucose, glycerol and lactate values for control vials containing no
 explants, or vials that were suspected to be contaminated.<br> NA =
 Not applicable for that vial type.