![]() We then measured the animals' paths in reference to these origins, revealing robust, highly replicable modules termed excursions, which are performed from the origin into the environment and back to the origin. In previous phenotyping studies of mouse and rat exploratory behavior we developed a computational exploratory data analysis methodology including videotaping, tracking, preparatory methods for customized data analysis, a methodology for improving the replicability of results across laboratories, and algorithmic design for exposing the natural reference places (origins) used by animals during exploration. Thus, the IMBA is an easy-to-use toolbox allowing an unprecedentedly rich view of the behaviour and behavioural variability of individual Drosophila larvae, with utility in multiple biomedical research contexts. This is shown for the transient backward locomotion induced by brief optogenetic activation of the brain-descending 'mooncrawler' neurons, and the variability in this behaviour. ![]() Strikingly, IMBA can also be used to analyse 'silly walks', that is patterns of locomotion it was not originally designed to investigate. The IMBA further allows us to determine, at the level of individual animals, the modulation of locomotion across repeated activations of dopamine neurons. ![]() We then report the discovery of a novel, complex locomotion phenotype of a mutant lacking an adhesion-type GPCR. We take advantage of the IMBA first to systematically describe the inter- and intra-individual variability in free, unconstrained locomotion in wild-type animals. It does not require specific hardware and can therefore be used in non-expert labs. Using a combination of computational modelling and statistical approaches, the IMBA reliably resolves individual identity across collisions. Here we present the IMBA (Individual Maggot Behaviour Analyser) for tracking and analysing the behaviour of individual larvae within groups. In principle or in practice, this in particular rules out grasping the inter- and intra-individual variability in locomotion and its genetic and neuronal determinants. The alternative is to measure animals one at a time, an extravagance for larger-scale analyses. However, although the faculty of locomotion clearly pertains to the individual animal, present studies of locomotion in larval Drosophila mostly use group assays and measurements aggregated across individual animals. Due to its numerically simple brain and neuromuscular system and its genetic accessibility, the larva of the fruit fly Drosophila melanogaster is an established model to study these processes at tractable levels of complexity. Neuronally orchestrated muscular movement and locomotion are defining faculties of multicellular animals. Dots represent the switching rate change for every fly. (F) Overall the difference of the switching differences after immobility and before it is always positive (p < 0.00195, sign test) and has a median of. Dots represent the score for individual flies and lines connect the results for the same individual. (E) Switching rate, calculated as the number of transitions per second, decreases in the interval preceding immobility (left panel, p < 0.039, Sign test) and tends to increase in the interval following immobility (right panel, not significant). To examine the change in switching across the session, the period leading to immobility is partitioned into the segments preceding and following maximal rotation (same for the period leading out of immobility). ![]() (D) Global changes in body orientation obtained from the absolute time derivative of the curve in (A). As shown, there is an overall drift in one direction, concurrent with a decrease in the number of transitions as a function of time. Zero crossings in the rotational speed mark switching, corresponding to the vertical bars in (B). (C) Time derivative of the fly's momentary body orientation. Each transition is marked by a small black dot on the curve and a corresponding vertical bar on the transitions raster plot below. As illustrated, while the fly globally rotates in one particular direction, locally it keeps switching between clockwise and counterclockwise rotations. (B) Zoomed in time segment in (A) of cumulative body rotation. ![]() Note that during transition into immobility the fly performs in the order of 50 full body rotations in 5 minutes, and during transition out of immobility, as many as 100 full body rotations in 5 minutes. As shown, the general tendency of this fly is to rotate clockwise both before and after immobility. The blue shaded area marks a zoomed-in time interval presented in (B,C). The gray shaded area marks the period of immobility. (A) Cumulative body orientation as a function of time for a single fly across the session. Switching between clockwise and counterclockwise rotation decreases into immobility and increases out of immobility. ![]()
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