Behaviour and its consequences

In this thesis, I have examined the behaviour and some of its neural underpinnings of a ‘model’ animal, the tadpoles and froglets of Xenopus laevis, at different levels of description and detail. At a macroscopic level, I investigated the animals’ movements in a very simple space. Zooming in, I looked at locomotion in freely and fictively swimming animals as well as at some of the sensory and motor consequences of locomotion. For many of these projects, I tested not only one particular developmental stage but a range of stages, allowing me to test for changes in behaviour with development. Methodologically, I employed video tracking to quantify movements in space over a longer period of time, as well as at a higher temporal and spatial resolution for short periods to record head movements during swimming. Semi-intact in vitro preparations of tadpoles were used to examine fictive locomotion and its consequences using electrophysiological recordings of peripheral nerves. Movements in space remained fairly similar over development, from small tadpoles to froglets, with all animals following the walls in a square environment, although the strength of wall following (WF) increased with growth. Tentacles, which are putatively mechanosensory appendages that large tadpoles temporarily possess, did not play any role for the strength of WF. WF was passive at all developmental stages, meaning that the animals never actively turned at a convex curvature to follow the wall, but instead went straight and left the wall. This implies that WF is unlikely to serve a defensive or spatial function. Looking specifically at locomotion in tadpoles showed that these animals commonly swim at 20 - 40 mm/s forward speeds, and move their heads left to right at up to 2500°/s angular velocities. These velocities decrease with development, probably because swimming frequency also decreases, from about 8 to about 5 Hz. Developmentally appropriate swimming frequencies are also seen in fictive swimming when the animals are deprived of normal sensory feedback. The mechanisms behind the developmental decrease in swimming frequency remain to be elucidated; biomechanical factors might well play a role. The left- right head oscillations during swimming also represent vestibular self-stimulation, which reaches amplitudes that are much higher than any of the stimuli used in sensory vestibular experiments. Another consequence of locomotion was observed in large tadpoles with tentacles: These tentacles are retracted during swimming, via a locomotor corollary discharge from the spinal cord. What I have shown in this thesis is first, that navigational behaviour of X. laevis in a simple laboratory setting seems to be mainly driven and constrained by the environment. Second, I have quantified head movements during swimming and therefore vestibular reafference, and found a developmental decrease in the swimming frequency. Finally, I uncovered an unusual effect of locomotion, namely the retraction of the tentacles during swimming. Together, these studies deepen the understanding of behaviour and its consequences in X. laevis

Reproducing Five Motor Behaviors in a Salamander Robot With Virtual Muscles and a Distributed CPG Controller Regulated by Drive Signals and Proprioceptive Feedback

Diverse locomotor behaviors emerge from the interactions between the spinal central pattern generator (CPG), descending brain signals and sensory feedback. Salamander motor behaviors include swimming, struggling, forward underwater stepping, and forward and backward terrestrial stepping. Electromyographic and kinematic recordings of the trunk show that each of these five behaviors is characterized by specific patterns of muscle activation and body curvature. Electrophysiological recordings in isolated spinal cords show even more diverse patterns of activity. Using numerical modeling and robotics, we explored the mechanisms through which descending brain signals and proprioceptive feedback could take advantage of the flexibility of the spinal CPG to generate different motor patterns. Adapting a previous CPG model based on abstract oscillators, we propose a model that reproduces the features of spinal cord recordings: the diversity of motor patterns, the correlation between phase lags and cycle frequencies, and the spontaneous switches between slow and fast rhythms. The five salamander behaviors were reproduced by connecting the CPG model to a mechanical simulation of the salamander with virtual muscles and local proprioceptive feedback. The main results were validated on a robot. A distributed controller was used to obtain the fast control loops necessary for implementing the virtual muscles. The distributed control is demonstrated in an experiment where the robot splits into multiple functional parts. The five salamander behaviors were emulated by regulating the CPG with two descending drives. Reproducing the kinematics of backward stepping and struggling however required stronger muscle contractions. The passive oscillations observed in the salamander's tail during forward underwater stepping could be reproduced using a third descending drive of zero to the tail oscillators. This reduced the drag on the body in our hydrodynamic simulation. We explored the effect of local proprioceptive feedback during swimming and forward terrestrial stepping. We found that feedback could replace or reduce the need for different drives in both cases. It also reduced the variability of intersegmental phase lags toward values appropriate for locomotion. Our work suggests that different motor behaviors do not require different CPG circuits: a single circuit can produce various behaviors when modulated by descending drive and sensory feedback

Axial dynamics during locomotion in vertebrates: lesson from the salamander

Much of what we know about the flexibility of the locomotor networks in vertebrates is derived from studies examining the adaptation of limb movements during stepping in various conditions. However, the body movements play important roles during locomotion: they produce the thrust during undulatory locomotion and they help to increase the stride length during legged locomotion. In this chapter, we review our current knowledge about the flexibility in the neuronal circuits controlling the body musculature during locomotion. We focus especially on salamander because, as an amphibian, this animal is able to display a rich repertoire of aquatic and terrestrial locomotor modes

In vitro-virtual-reality: an anatomically explicit musculoskeletal simulation powered by in vitro muscle using closed loop tissue-software interaction

Muscle force-length dynamics are governed by intrinsic contractile properties, motor stimulation and mechanical load. Although intrinsic properties are well-characterised, physiologists lack in vitro instrumentation accounting for combined effects of limb inertia, musculoskeletal architecture and contractile dynamics. We introduce in vitro virtual-reality (in vitro-VR) which enables in vitro muscle tissue to drive a musculoskeletal jumping simulation. In hardware, muscle force from a frog plantaris was transmitted to a software model where joint torques, inertia and ground reaction forces were computed to advance the simulation at 1 kHz. To close the loop, simulated muscle strain was returned to update in vitro length. We manipulated 1) stimulation timing and, 2) the virtual muscle's anatomical origin. This influenced interactions among muscular, inertial, gravitational and contact forces dictating limb kinematics and jump performance. We propose that in vitro-VR can be used to illustrate how neuromuscular control and musculoskeletal anatomy influence muscle dynamics and biomechanical performance

Initiation and maintenance of swimming in hatchling xenopus laevis tadpoles

Effective movement is central to survival and it is essential for all animals to react in response to changes around them. In many animals the rhythmic signals that drive locomotion are generated intrinsically by small networks of neurons in the nervous system which can be switched on and off. In this thesis I use a very simple animal, in which the behaviours and neuronal networks have been well characterised experimentally, to explore the salient features of such networks. Two days after hatching, tadpoles of the frog Xenopus laevis respond to a brief touch to the head by starting to swim. The swimming rhythm is driven by a small population of electrically coupled brainstem neurons (called dINs) on each side of the tadpole. These neurons also receive synaptic input following head skin stimulation. I build biophysical computational models of these neurons based on experimental data in order to address questions about the effects of electrical coupling, synaptic feedback excitation and initiation pathways. My aim is better understanding of how swimming activity is initiated and sustained in the tadpole. I find that the electrical coupling between the dINs causes their firing properties to be modulated. This allows two experimental observations to be reconciled: that a dIN only fires a single action potential in response to step current injections but the population fires like pacemakers during swimming. I build on this hypothesis and show that long-lasting, excitatory feedback within the population of dINs allows rhythmic pacemaker activity to be sustained in one side of the nervous system. This activity can be switched on and off at short latency in response to biologically realistic synaptic input. I further investigate models of synaptic input from a defined swim initiation pathway and show that electrical coupling causes a population of dINs to be recruited to fire either as a group or not at all. This allows the animal to convert continuously varying sensory stimuli into a discrete decision. Finally I find that it is difficult to reliably start swimming-like activity in the tadpole model using simple, short-latency, symmetrical initiation pathways but that by using more complex, asymmetrical, neuronal-pathways to each side of the body, consistent with experimental observations, the initiation of swimming is more robust. Throughout this work, I make testable predictions about the population of brainstem neurons and also describe where more experimental data is needed. In order to manage the parameters and simulations, I present prototype libraries to build and manage these biophysical model networks

Design for an Increasingly Protean Machine

Data-driven, rather than hypothesis-driven, approaches to robot design are becoming increasingly widespread, but they remain narrowly focused on tuning the parameters of control software (neural network synaptic weights) inside an overwhelmingly static and presupposed body. Meanwhile, an efflorescence of new actuators and metamaterials continue to broaden the ways in which machines are free to move and morph, but they have yet to be adopted by useful robots because the design and control of metamorphosing body plans is extremely non-intuitive. This thesis unites these converging yet previously segregated technologies by automating the design of robots with physically malleable hardware, which we will refer to as protean machines, named after Proteus of Greek mythology. This thesis begins by proposing an ontology of embodied agents, their physical features, and their potential ability to purposefully change each one in space and time. A series of experiments are then documented in which increasingly more of these features (structure, shape, and material properties) were allowed to vary across increasingly more timescales (evolution, development, and physiology), and collectively optimized to facilitate adaptive behavior in a simulated physical environment. The utility of increasingly protean machines is demonstrated by a concomitant increase in both the performance and robustness of the final, optimized system. This holds true even if its ability to change is temporarily removed by fabricating the system in reality, or by “canalization”: the tendency for plasticity to be supplanted by good static traits (an inductive bias) for the current environment. Further, if physical flexibility is retained rather than canalized, it is shown how protean machines can, under certain conditions, achieve a form of hyper-robustness: the ability to self-edit their own anatomy to “undo” large deviations from the environments in which their control policy was originally optimized. Some of the designs that evolved in simulation were manufactured in reality using hundreds of highly deformable silicone building blocks, yielding shapeshifting robots. Others were built entirely out of biological tissues, derived from pluripotent Xenopus laevis stem cells, yielding computer-designed organisms (dubbed “xenobots”). Overall, the results shed unique light on questions about the evolution of development, simulation-to-reality transfer of physical artifacts, and the capacity for bioengineering new organisms with useful functions

Transmission des voies olfactives aux cellules réticulospinales de la lamproie

Les informations olfactives sont connues pour leur capacité à induire des comportements moteurs spécifiques. En dépit de nombreuses observations comportementales chez les vertébrés, on ne connaît toujours pas les mécanismes et les voies nerveuses qui sous-tendent ces phénomènes de transformation olfacto-locomotrices. Chez la lamproie, des travaux récents ont permis de décrire cette voie, et les mécanismes responsables de la transformation des entrées olfactives en activité locomotrice (Derjean et al., 2010). Cette voie prend origine dans la partie médiane du bulbe olfactif, et envoie des projections vers le tubercule postérieur, une région qui se trouve dans le diencéphale. De là, les neurones projettent directement vers la Région Locomotrice Mésencéphalique, connue pour envoyer des connexions vers les neurones réticulospinaux, et activer la locomotion. L’objectif de cette étude était d’établir si l’ensemble des neurones réticulospinaux répond aux stimulations olfactives. Pour ce faire, nous avons utilisé sur une préparation de cerveau isolé de lamproie des techniques d’électrophysiologie et d’imagerie calcique. La stimulation électrique des nerfs olfactifs, de la région médiane du bulbe olfactif ou du tubercule postérieur a provoqué une activation de toutes les cellules réticulospinales qui se retrouvent dans les quatre noyaux réticulaires (ARRN : Noyau Réticulaire Rhombencéphalique Antérieur; MRN : Noyau Réticulaire Mésencéphalique; MRRN : Noyau Réticulaire Rhombencéphalique Moyen; PRRN : Noyau Réticulaire Rhombencéphalique Postérieur). Seule la partie médiane du bulbe olfactif est impliquée dans le passage de l’information olfactive vers les neurones réticulospinaux. Nous avons aussi découvert que le blocage des récepteurs GABAergiques dans la partie médiane du bulbe olfactif augmentait les réponses olfactives de façon considérable dans les cellules réticulospinales. Nous avons montré ainsi qu’il existe un tonus inhibiteur impliqué dans la dépression modulatrice de la voie olfacto-locomotrice. Ce travail a permis de montrer que la stimulation des afférences sensorielles olfactives active simultanément l’ensemble des populations de neurones réticulospinaux qui commandent la locomotion. De plus, il existerait un tonus inhibiteur GABAergique, au niveau de la partie médiane du bulbe olfactif, responsable d’une dépression modulatrice dans la voie olfacto-locomotrice.Olfactory inputs are known for their ability to induce specific motor behaviors. Despite numerous behavioral observations in vertebrates, the mechanisms and the neural pathways underlying the olfactory-locomotor transformation are still unknown. In lamprey, recent studies have described this pathway and the mechanism underlying the transformation of olfactory input into a locomotor activity (Derjean et al., 2010). This pathway originates in the medial part of the olfactory bulb, sends projections to the posterior tuberculum, a diencephalic region. From there, the neurons project directly to the mesencephalic locomotor region that is known to send projections to the reticulospinal neurons to activate locomotion. Using lamprey brain preparation, electrophysiology and calcium imaging, the aim of this study was to establish whether all reticulospinal neurons respond to olfactory stimuli. Electrical stimulation of the olfactory nerves, the medial part of the olfactory bulb or the posterior tuberculum activates all reticulospinal cells in the four reticular nuclei (ARRN: Anterior rhombencephalic reticular nucleus; MRN: middle mesencephalic reticular nucleus; MRRN: middle rhombencephalic reticular nucleus; PRRN: posterior rhombencephalic reticular nucleus). The medial part of the olfactory bulb is the only region that is implicated in transmitting the olfactory information to reticulospinal neurons. We also discovered that when blocking the GABAergic receptors in the medial part of the olfactory bulb, the reticulospinal neurons have a stronger response to olfactory stimulation. Thus we showed that a tonic inhibition is involved in the modulating depression of the olfacto-locomotor pathway. Altogether, this work shows that stimulation of the olfactory sensory inputs activates simultaneously the entire population of reticulospinal neurons that control locomotion. In addition, there is a GABAergic tonic inhibition at the level of the medial part of the olfactory bulb that causes a modulating depression in the olfacto-locomotor pathway

Seasonal proteome variation in intertidal shrimps under a natural setting: connecting molecular networks with environmental fluctuations

The ability of intertidal organisms to maintain their performance via molecular and physiological adjustments under low tide, seasonal fluctuations and extreme events ultimately determines population viability. Analyzing this capacity in the wild is extremely relevant since intertidal communities are under increased climate variability owing to global changes. We addressed the seasonal proteome signatures of a key intertidal species, the shrimp Palaemon elegans, in a natural setting. Shrimps were collected during spring and summer seasons at low tides and were euthanized in situ. Environmental variability was also assessed using hand-held devices and data loggers. Muscle samples were taken for 2D gel electrophoresis and protein identification through mass spectrometry. Proteome data revealed that 55 proteins (10.6% of the proteome) significantly changed between spring and summer collected shrimps, 24 of which were identified. These proteins were mostly involved in cytoskeleton remodelling, energy metabolism and transcription regulation. Overall, shrimps modulate gene expression leading to metabolic and structural adjustments related to seasonal differences in the wild (i.e. abiotic variation and possibly intrinsic cycles of reproduction and growth). This potentially promotes performance and fitness as suggested by the higher condition index in summer-collected shrimps. However, inter-individual variation (% coefficient of variation) in protein levels was quite low (min-max ranges were 0.6-8.3% in spring and 1.2-4.8% in summer), possibly suggesting reduced genetic diversity or physiological canalization. Protein plasticity is relevant to cope with present and upcoming environmental variation related to anthropogenic forcing (e.g. global change, pollution) but low inter-individual variation may limit evolutionary potential of shrimp populations.publishe