Chemical stimuli are omnipresent in driving all kinds of animal behaviours. The comparative study of chemosensory systems has revealed a wide variety of organs, neuronal circuits, and above all of receptor proteins. Despite some conserved features, no clear picture has emerged yet about the origins of chemoreception, and new animal models are needed to understand its evolutionary history.
The marine worm Platynereis dumerilii (Nereididae, Polychaeta, Annelida, Lophotrochozoa) is a model system for evolutionary developmental biology, and recent achievements have revealed its potential for neurobiology. The head of marine annelids is generally equipped with abundant sensory appendages, chemosensory based on numerous morphological descriptions. Surprising as it may seem for this ecologically important phylum, there exists still no direct physiological proof of chemosensitivity in these prominent organs. Moreover, in the absence of appropriate assay systems, physiological experiments in Platynereis have so far concentrated on photic and mechanical stimuli.
The aim of this thesis was the description and physiological study of chemosensory systems in Platynereis, as well as the establishment of microfluidics-based methods to enable functional imaging and behavioural assays upon chemical stimulations. The 6-days-post-fertilisation larval stage (6dpf), at which most adult anatomical structures are already present, has emerged as a powerful stage for cross-species comparisons of cell types, thanks notably to a unique whole-body atlas of gene expression. It was thus chosen as a target stage for the study of chemosensation.
An anatomical investigation of Platynereis head appendages at various stages has allowed to better understand how the hemispheric head of larvae transforms into the complex, appendage-rich head of juveniles and adults. Neuroanatomical stainings have confirmed the presence already at 6dpf of different architectures, innervation patterns, and sensory cell types across appendages. A reference anatomical description has been established at 6dpf to characterise the position of nerves and sensory ganglia, which constitutes a useful basis for in vivo studies.
After having developed a microfluidic setup for confocal calcium imaging of the whole head upon chemical stimulations, I have tested the physiology of candidate chemosensory organs in 6dpf animals. These experiments have revealed that antennae, not nuchal organs as thought previously, are probably the main chemosensory organ in Platynereis, that nuchal organs and palps are endowed with chemosensitivity, and that so are tentacular cirri though to a lesser extent. Prominent fluctuating apical organ activity was seen, though not obviously related to chemosensation. Finally, new components of the chemosensory systems have been described based on their activity patterns, including sensory cells and probably interneurons. Based on these results, a first understanding of chemical stimulus detection has emerged. Partial evidence was given that Mushroom Bodies may play a role in these systems at 6dpf, which motivates the study of associative learning in relation with chemical cues.
To link chemical stimuli to larval behaviours, additional microfluidic devices have been developed in which freely-moving larvae can be exposed to controlled spatial and temporal patterns of chemical stimuli and their behaviour monitored. In the perspective of establishing an assay for chemosensory associative learning, aversive compounds such as quinine have been tested and found to produce stereotypical avoidance behavioural responses, thus they could be used as unconditioned stimuli in pavlovian assays. A neutral cue identified in functional imaging, 1-butanol, was shown to be a valid candidate as a conditioned stimulus. Thanks to these preliminary results, an experimental setup for quantitative studies of behavioural modifications is now available.
Overall, this work has laid a basis for the study of chemosensation in Platynereis, informed about sensory organ physiology in polychaetes, and shown the suitability of microfluidic setups for physiological and behavioural assays at larval stages. It suggests a possibly broad chemosensory repertoire in marine invertebrate larvae. Chemical stimuli in annelids are worth new attention for comparative studies of sensory systems, and in the search for associative learning abilities
