Data-driven analysis of motor activity implicates 5-HT2A neurons in backward locomotion of larval Drosophila

Abstract

Rhythmic animal behaviors are regulated in part by neural circuits called the central pattern generators (CPGs). Classifying neural population activities correlated with body movements and identifying the associated component neurons are critical steps in understanding CPGs. Previous methods that classify neural dynamics obtained by dimension reduction algorithms often require manual optimization which could be laborious and preparation-specific. Here, we present a simpler and more flexible method that is based on the pre-trained convolutional neural network model VGG-16 and unsupervised learning, and successfully classifies the fictive motor patterns in Drosophila larvae under various imaging conditions. We also used voxel-wise correlation mapping to identify neurons associated with motor patterns. By applying these methods to neurons targeted by 5-HT2A-GAL4, which we generated by the CRISPR/Cas9-system, we identified two classes of interneurons, termed Seta and Leta, which are specifically active during backward but not forward fictive locomotion. Optogenetic activation of Seta and Leta neurons increased backward locomotion. Conversely, thermogenetic inhibition of 5-HT2A-GAL4 neurons or application of a 5-HT2 antagonist decreased backward locomotion induced by noxious light stimuli. This study establishes an accelerated pipeline for activity profiling and cell identification in larval Drosophila and implicates the serotonergic system in the modulation of backward locomotion.

A pair of ascending neurons in the subesophageal zone mediates aversive sensory inputs-evoked backward locomotion in Drosophila larvae

“…R73F04-LexA by cloning R73F04 from the genome of w 1118 using the primers described in FlyLight ( http://flweb.janelia.org/ ) into pENTR 1A (Thermo Fisher Scientific, A10462) and subsequently to pBPnlsLexA :: p65Uw (Addgene, #26230) according to the previously described method [ 45 ]. We generated Gad1-2A-GAL80 using CRISPR/Cas9 system as described in the previous studies [ 46 , 47 ]. We inserted a transgene encoding the 2A peptide and GAL80 gene in front of the stop codon of the Gad1 gene with the following 20-bp guide RNA (gRNA) sequence: 5′-GCCTGGGCGACGACTTGTAA-3′.…”

Regulation of coordinated muscular relaxation in Drosophila larvae by a pattern-regulating intersegmental circuit

“…1c, d , Supplementary Fig. 1d, e , and Supplementary Movie 2 ) 28 , 29 . Canon neurons were activated after aCC MNs in the same neuromere and at a similar timing to aCC MNs in neuromeres 2–3 segments more anterior suggesting possible roles in the termination of segmental motor activity (Fig.…”

Regulation of coordinated muscular relaxation by a pattern-generating intersegmental circuit

“…1c, d, Supplementary Fig. 1d, e, and Supplementary Movie 2) 27,28 . Canon neurons were activated after aCC MNs in the same neuromere and at a similar timing to aCC MNs in neuromeres 2–3 segments more anterior suggesting possible roles in the termination of segmental motor activity (Fig.…”