Radical new technique lets researchers see inside live insects in 3D for the first time

Imaging live insects is challenging because of the radiation doses required
In addition, the movement of the insects compromises image quality
But in a new study, researchers used carbon dioxide gas to anesthetize the insects to prevent them from moving during 3D visualization

Published: 21:12, 1 August 2017 | Updated: 21:32, 1 August 2017

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Researchers have imaged whole live insects using a technique called X-ray micro-computed tomography (micro-CT).

Imaging whole live insects is challenging because of the radiation doses required to obtain the images and the movement of the insects, compromising image quality - so most research involving involving invertebrates imaging requires animal sacrifice.

But in a new study, researchers used carbon dioxide gas to anesthetize the insects to prevent them from moving during 3D visualization, and according to the researchers, the method had very little impact on the insects' longevity.

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3D-CT reconstruction (20 micrometer resolution) showing the internal structures of a live anesthetized male true armyworm at 1 (sexually immature) and 4 (sexually mature) days old. Top: 3D-volume of the male (♂) body exoskeleton. Middle: 3D-MPR (multi-planar reformatted) views of the whole body showing the internal structures; and Bottom: The tracheal system

In order to image live insects, the researchers say it's crucial to fully immobilize them, deliver a low-enough radiation dose to allow for repeated scans of the same individuals for time-relevant studies, and provide adequate image resolution and quality to distinguish internal structures.

The researchers, based at the University of Western Ontario, scanned Colorado potato beetles (Leptinotarsa decemlineata) and true armyworms (Pseudaletia unipuncta).

HOW THEY DID THE STUDY

The researchers, based at the University of Western Ontario, scanned Colorado potato beetles (Leptinotarsa decemlineata) and true armyworms (Pseudaletia unipuncta).

The study relied on giving carbon dioxide gas to insects to induce hypoxia - oxygen deficient conditions.

The insects were placed in a custom designed tube for micro-CT scanning, and a syringe filter was attached to the tube's open end to deliver the carbon dioxide gas.

Schematic of the new experimental set-up summarizing the incorporation of carbon dipxides gas as anesthetic during X-ray micro-computed tomography (micro-CT) of live insect imaging. The insects were placed in a tube (pictured) for the micro-CT scanning

The experiment relied on the fact that these insects in particular have remarkable abilities ti survive prolonged hypoxia and ionising radiation doses - at least 100 times more than the lethal dose for humans.

This approach allowed the researchers to keep the live insects still for a prolonged period of time - up to 7 hours - while repeatedly scanning the same insect using X-ray micro-computed tomography (micro-CT).

According to Harvard University's Center for Nanoscale Systems, X-ray micro-computed tomography is an X-ray imaging method capable of generating 3-D, cross-sectional and internal structure images.

It uses micro-focused X-rays to create a non-destructive shadow image of an an object based on the X-ray absorption of the material.

Then, a computer processed images from all directions and reconstructs 3-D structures of the object.

The study relied on giving carbon dioxide gas to insects to induce hypoxia - oxygen deficient conditions.

The insects were placed in a custom designed tube for micro-CT scanning, and a syringe filter was attached to the tube's open end to deliver the carbon dioxide gas.

According to an article written by the researcher for BioMed Central, this approach allowed the researchers to keep the live insects still for a prolonged period of time - up to 7 hours - while repeatedly scanning the same insect using X-ray micro-computed tomography (micro-CT).

3D-CT reconstruction of a live anesthetized Colorado potato beetle adult female depicting the body and the tracheal system

According to Harvard University's Center for Nanoscale Systems, X-ray micro-computed tomography is an X-ray imaging method capable of generating 3-D, cross-sectional and internal structure images.

It uses micro-focused X-rays to create a non-destructive shadow image of an an object based on the X-ray absorption of the material

Then, a computer processed images from all directions and reconstructs 3-D structures of the object.

The researchers say that when the correct scan protocol was used, insects recovered with minimal impact on their behavior and reproduction.

'We We have demonstrated that using CO2 gas during micro-CT imaging effectively induces safe, repeatable, whole-body, and temporary immobilization of live insects for 3D visualizations without motion artefacts,' the researchers concluded in their study.

'Moreover, we have shown that exposed TAW (true armyworm) individuals made a full recovery with very little impact on subsequent longevity, and mating success post hypoxia.

'This method is applicable to other imaging modalities and could be used for routine exploratory and time-course studies, for repeated scanning of live and intact individuals,' they researchers wrote.

The method of using carbon dioxide gas to temporarily paralyze insects has been successfully tested for other insect life stages. Pictured are 3D-CT reconstruction images of a live (i) true armyworm pupa, and (ii) a hornworm caterpillar