Ytterbium-doped fibre femtosecond laser offers robust operation with deep and precise microsurgery of C. elegans neurons

M. B. Harreguy,G. Haspel,S. H. Chung,C. V. Gabel,V. Marfil,N. W. F. Grooms

doi:10.1038/s41598-020-61479-0

M. B. Harreguy, G. Haspel + Show 4 more

Open Access

https://doi.org/10.1038/s41598-020-61479-0

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Abstract

Laser microsurgery is a powerful tool for neurobiology, used to ablate cells and sever neurites in-vivo. We compare a relatively new laser source to two well-established designs. Rare-earth-doped mode-locked fibre lasers that produce high power pulses recently gained popularity for industrial uses. Such systems are manufactured to high standards of robustness and low maintenance requirements typical of solid-state lasers. We demonstrate that an Ytterbium-doped fibre femtosecond laser is comparable in precision to a Ti:Sapphire femtosecond laser (1–2 micrometres), but with added operational reliability. Due to the lower pulse energy required to ablate, it is more precise than a solid-state nanosecond laser. Due to reduced scattering of near infrared light, it can lesion deeper (more than 100 micrometres) in tissue. These advantages are not specific to the model system ablated for our demonstration, namely neurites in the nematode C. elegans, but are applicable to other systems and transparent tissue where a precise micron-resolution dissection is required.

Highlights

Laser microsurgery is a powerful tool for neurobiology, used to ablate cells and sever neurites in-vivo
Axotomy using nanosecond and femtosecond pulses at high or low repetition rate varies in the extent of damage to surrounding tissues and in the size of the gap induced in a severed axon, but axon regeneration seems to occur at comparable extents and rates after axotomy[12]
The basic design is similar to Ti:Sapphire lasers that are very commonly used for multiphoton imaging and axotomy[15,16]

Summary

Introduction

Laser microsurgery is a powerful tool for neurobiology, used to ablate cells and sever neurites in-vivo. Microsurgery of neurites (termed axotomy) was pioneered in C. elegans[7] where a laser-pumped titanium-sapphire laser was used to cut commissure neurites of motoneurons These Ti:Sapphire lasers are typically configured to produce near-infrared (NIR) pulses with energies up to 50 nJ, a centre wavelength of approximately 800 nm, pulse duration of 100–200 femtoseconds, and repetition rates of 80 MHz. Ablation at 80 MHz does not allow complete dissipation of pulse energy[8] and in many cases an external electro-optic pulse picking device is added to reduce the repetition rate and ablate at 1 to 10 kHz. The longer intervals between pulses at these lower repetition rates allow the deposited energy to dissipate completely[8,9,10], improving the surgical resolution. For any energy level there is a minimal number of pulses that will initiate ablation but adding pulses does not substantially increase the region damaged[3,13]

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