Using µCT in live larvae of a large wood-boring beetle to study tracheal oxygen supply during development

Philipp Lehmann,Marion Javal,Anton Du Plessis,John S Terblanche

doi:10.1016/j.jinsphys.2021.104199

Philipp Lehmann, Marion Javal + Show 2 more

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https://doi.org/10.1016/j.jinsphys.2021.104199

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Abstract

How respiratory structures vary with, or are constrained by, an animal’s environment is of central importance to diverse evolutionary and comparative physiology hypotheses. To date, quantifying insect respiratory structures and their variation has remained challenging due to their microscopic size, hence only a handful of species have been examined. Several methods for imaging insect respiratory systems are available, in many cases however, the analytical process is lethal, destructive, time consuming and labour intensive. Here, we explore and test a different approach to measuring tracheal volume using X-ray micro-tomography (µCT) scanning (at 15 µm resolution) on living, sedated larvae of the cerambycid beetle Cacosceles newmannii across a range of body sizes at two points in development. We provide novel data on resistance of the larvae to the radiation dose absorbed during µCT scanning, repeatability of imaging analyses both within and between time-points and, structural tracheal trait differences provided by different image segmentation methods. By comparing how tracheal dimension (reflecting metabolic supply) and basal metabolic rate (reflecting metabolic demand) increase with mass, we show that tracheal oxygen supply capacity increases during development at a comparable, or even higher rate than metabolic demand. Given that abundant gas delivery capacity in the insect respiratory system may be costly (due to e.g. oxygen toxicity or space restrictions), there are probably balancing factors requiring such a capacity that are not linked to direct tissue oxygen demand and that have not been thoroughly elucidated to date, including CO2 efflux. Our study provides methodological insights and novel biological data on key issues in rapidly quantifying insect respiratory anatomy on live insects.

Highlights

What determines the metabolic scope of animals? One important aspect is the delivery capacity of the oxygen supply system
Matched growth of structures linked to oxygen demand and supply was proposed within the framework of symmorphosis (Weibel et al, 1991)
We were suc cessfully able to sedate larvae (MacMillan et al, 2017), scan and reconstruct tracheal structures, and can show that exposure to the anesthetic and μCT radiation dose does not lead to increased mortality or a systematic difference in growth

Summary

Introduction

What determines the metabolic scope of animals? One important aspect is the delivery capacity of the oxygen supply system. Oxygen is supplied through air-filled tubes called tracheae (Chapman, 2012; Harrison et al, 2012). The smallest tracheae, with a diameter smaller than ~2 μm, are called tra cheoles and the major site of oxygen exchange (Schmitz and Perry, 1999; Wigglesworth, 1983). As insects grow, their body mass increases exponentially and as tis sue is being added, it is expected that the cost of maintaining these tissues, reflected in the resting metabolic rate (rmr), increases as well (Greenlee and Harrison, 2005; Kivelaet al., 2016). The theory suggests that excess functional capacity of tissues in a supply-demand network should be selected against over evolutionary timescales, as tissues, especially neural, muscle and gut, are energetically costly (Niven and Laughlin, 2008)

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