DOI: 10.5061/dryad.pg4f4qrzr
title: Modulation of gill surface area does not correlate with oxygen loss in Chitala ornata

description: Air-breathing fish risk losing aerially sourced oxygen to ambient hypoxic
 water since oxygenated blood from the air-breathing organ returns through
 the heart to the branchial basket before distribution. This loss is
 thought to have helped drive the evolutionary reduction in gill size with
 the advent of air-breathing. In many teleost fish, gill size is known to
 be highly plastic by modulation of their anatomic diffusion factor (ADF)
 with interlamellar cell mass (ILCM). In the anoxia-tolerant crucian carp,
 ILCM recedes with hypoxia but regrows in anoxia. The air-breathing
 teleost, Chitala ornata has been shown to increase gill ADF from normoxic
 to mildly hypoxic water by reducing ILCM. Here, we test the hypothesis
 that ADF is modulated to minimize oxygen loss in severe aquatic hypoxia by
 measuring ADF and gas exchange, and by using CT scans to reveal possible
 trans-branchial shunt vessels. Contrary to our hypothesis, ADF does not
 modulate to prevent oxygen loss and despite no evident trans-branchial
 shunting, C. ornata loses only 3% of its aerially-sourced O2 while still
 excreting 79% of its CO2 production to the severely hypoxic water. We
 propose this is achieved by ventilatory control and by compensating the
 minor oxygen loss by extra aerial O2 uptake.
description: # Modulation of gill surface area does not correlate with oxygen loss in
 *Chitala ornata*
 [https://doi.org/10.5061/dryad.pg4f4qrzr](https://doi.org/10.5061/dryad.pg4f4qrzr) ## Description of the data and file structure The data was collected using intermittent closed respirometry and stereology techniques.  ### Files and variables #### File: Respirometry\_and\_stereology\_data.xlsx * Sheet 1: The Respirometry Excel sheet contains the following columns. * Sheet 2: The Stereology Excel sheet contains the following columns. * Sheet 3: The harmonic mean of blood-water diffusion distance. Variables: ##### Sheet 1: Respirometry Excel sheet contains the following columns: A) An ID column showing which fish is measured (F for fish and number for date) B) A column describing the number of measurements, each number is one measurement. C) A treatment column showing if water was normoxic or hypoxic. D-F) Columns show O2 uptake from air, water, and total in addition to branchial loss in severe hypoxia. Units are μmol/min/kg. G-I) columns show CO2 excretion to air, water, and total. Units are μmol/min/kg. J-L) columns show the respiratory exchange ratio (RER) from air, water, and total (unit less).      M-P) Partitioning of O2 and CO2 (%) from water and air. Q) Loop, describes the ID of each measurement. It shows what raw data measurement corresponds to the data across all the methods. R) The branchial O2 loss of total O2 uptake in %. S) Time since the start. Hypoxia is always set to start at measurement 34 at 19.83 hours. (unit = hours). T-U) A 9-row table, with extra information (mass of replicates and their name).  ##### Sheet 2: Stereology Excel sheet contains the following columns: Columns A through Q shows data used for volume estimation. – for raw counts see column AK-AP. Numbers are unitless unless a unit is described in the header (P and pi describe a count). SSF = section sampling fraction, ASF = area sampling fraction, TSF = tissue sampling fraction. L/P = length per point, A/P = area per point. ILCM = interlamellar cell mass. ILCM count = number of counts on lamellae covered by ILCM. NO ILCM count = number of counts on lamellae without ILCM.   Column S-AE shows the data used for the surface area of exposed and covered lamellae estimations together with the amount of ILCM present. For raw data counts see column AU-BA. Units a denoted in the header where appropriate.  Fish ID in each replicate for volume, surface area, harmonic mean diffusion distance, and ILCM: Fish ID: 1, 4, 7, 10, 13, 16 used in Normoxia treatment Fish ID: 3, 6, 9, 12, 15, 18 used in Mild hypoxia treatment Fish ID: 2, 5, 8, 11, 14, 17 used in Severe hypoxia treatment ##### Sheet 3: harmonic mean of blood-water diffusion distance: Column A-CK shows every raw measurement in μm between blood vessels and water. The data is presented with every fish replicate followed by the measurements for that fish on gill arches 1 to 4. All numbers are in μm.   Please note that some columns and rows have been left blank for organizational purposes. #### Ct-scan (added in the 22-aug-2024 version) See the paper methods description for details. The CT-scan-based pictures on the highest quality replicate have been uploaded as three zip files: **A high-resolution scan (160µm) of the entire body of a *Chitala ornata* called Chitala-ornata_1_FullBody.** CT-scan settings: Al-filter1-5mm, 80kV, 95W, 160um, 25ms, 9avg. **A medium-resolution scan (55µm) of the entire air-breathing organ (the most important set for the anatomical model in the paper) called Chitala-ornata_1_ABO.** CT-scan settings: Al-filter 1-5mm, 80kV, 55W, 55um, 90ms, 4avg. **A high-resolution scan (25µm) of the anterior part of the air-breathing organ where a majority of the vascular branching happens in the zip called Chitala-ornata_1_ABO-anterior.** Ct-scan settings: Al-filter1-5, 80kV, 25W, 25um, 165ms, 9avg. **Supplementary Video V1 shows a** Virtual axial slice video going through the images of one replicate. ## Code/software ImageJ - free software that can be used to load the ct-scan image stacks.