An optogenetic assay of Drosophila larval motor neuron performance in vivo

Abstract

Background

Over fifty million people worldwide currently live with neurodegenerative diseases, many of which are the result of pathogenic gene variants. Genetically malleable model organisms provide an avenue for research into the genetic bases of these diseases, and the large motor neurons of fruit fly larvae provide a test bed for these investigations. However, it is challenging to collect information from these neurons under physiological conditions as they terminate on rhythmically contracting muscle fibers.

New Method

As a test of in vivo neuronal performance, we expressed light-activated opsins in motor-neurons of unrestrained intact Drosophila larva and used light pulses to drive cyclical body-wall contractions that were captured on camera and analyzed offline.

Results

We describe the assembly of an apparatus to systematically activate motor-neurons in Drosophila larvae and an image acquisition system to capture the resulting body-wall contractions. To test the utility of the assay we performed a motor-neuron specific knock-down of Miro, an adaptor [protein, MIRO] for mitochondrial transport into motor-neuron terminals.

Comparison with Existing Methods

This in vivo assay allows for a test of sustained neuronal performance while sidestepping the shortcomings of electrophysiological assays of neurotransmission in situ where recordings are mechanically disrupted at endogenous firing rates. Secondly, unlike adult climbing assays and larval locomotion assays, performance is assayed independently of the organism’s motivation to perform or ability to detect stimuli.

Conclusions

Here we demonstrated an optogenetic assay for quantifying motor neuron output of intact Drosophila 3rd instar larvae. Our data established the robustness of the assay and its capacity to discriminate impaired motor neuron performance.