Theoretical and experimental investigations of intra- and inter-segmental control networks and their application to locomotion of insects and crustaceans

Movements of the walking legs in terrestrial animals have to be coordinated continuously in order to produce successful locomotion. Walking is a cyclic process: A single step consists of a stance phase and a swing phase. In the stance phase, the leg muscles provide propulsion of the animal’s body. During the swing phase, the leg is positioned to the starting position of the next stance phase. Sensory input, arising from sensory organs in the legs, modulates the rhythmic motoneuronal activity and therefore the rhythmic activity of the antagonistic muscles pairs in a leg. The coordination of leg joints, and thus of the respective muscle pairs, is called intra-segmental coordination. For coordinated walking not only the proper coordination of one leg is important, but also the coordination of contralateral and ipsilateral legs. The latter is called inter-segmental coordination and also strongly depends on sensory feedback. In this thesis I present three publications (Grabowska et al., 2012; Toth et al., 2013; Grabowska et al., in rev.) and results of an experimental study focusing on different aspects of intra- and inter-segmental coordination. Starting with experimental data on the stick insect Carausius morosus, a well studied model organism for locomotion, I analyzed inter-segmental coordination of legs during walking behavior of stick insects by video analysis. I also performed electrophysiological experiments that provide insight into the inter-segmental connections of different thoracic segments. Furthermore, experimental results were summarized in mathematical models in order to reproduce stick insect locomotion and to provide new hypotheses about so far unknown neuronal controlling processes. First, a study of the walking behavior of the stick insect is introduced (Grabowska et al., 2012). For this purpose, walking sequences of adult animals, walking straight on surfaces with increasing and decreasing slopes, were recorded. Depending on the slope, the animals used different coordination patterns. Subsequent, walking patterns of animals with amputated front, hind or middle legs were analyzed. It became evident that the resulting coordination patterns were regular or maladapted, depending on the amputated leg pairs. We therefore assumed that afferent information from walking front, middle, and hind legs contribute differently to coordination. The second part presents a neuromechanical model that describes starting and stopping of a stick insect leg during walking (Tóth et al., 2013). An existing model of the intra-segmental neuronal network of the stick insect leg was extended by a model of its musculo-skeletal system. The focus of the model was on the neuronal control of slow and fast muscle fiber activity of the three proximal leg muscle groups at start and stop of a leg within a stepping cycle. Using the effects of sensory signals that encode position and velocity of the leg joints like the temporal components of activated muscles during start and stop, observed in experiments, as well as the timing of starting and stopping processes within a step cycle, the simulation results were in good agreement with the observed data of the stick insect. Therefore, this model can be regarded as physiologically relevant and leads to hypotheses about the neuronal control of the musculo-skeletal system that can reveal details of stop and starting in the walking animals. In the third part of this thesis the above mentioned 3-CPG-MN network model, which has been developed based on stick insect data, was extended to serve as a basic module for eight-legged locomotion in walking crustaceans (Grabowska et al., in rev.). For this purpose, the existing 3-CPG-MN network model was extended by an additional segmental module. The basic properties of the 3-CPG-MN network modules remained unchanged. By testing two different network topologies of the new 4-CPG-MN network model, specific walking behavior (coordination patterns, stepping frequency, and transitions) of crustaceans could be replicated by only changing the timing of the inter-segmental excitatory sensory input on the influenced segment. Considering the topology of the 3-CPG-MN network model, namely a caudal-rostral inter-segmental connection connecting every second CPG, the 4-CPG-MN network model was able to reproduce all kinds of walking behavior of forward walking crabs and crayfish. This network stresses the importance of the timing of excitatory signals that are provided by inter-segmental pathways in animals with eight walking legs and four thoracic segments, and proposes possible inter-segmental sensory pathways. Finally, results of experimental data are introduced showing that the rhythm of protractor/retractor central pattern generating networks (thorax-coxa joint) in the prothoracic ganglion can be influenced by a stepping ipsilateral hind leg of the stick insect. This inter-segmental pathway was hypothesized in the 3-CPG-MN network model of Daun-Gruhn and Tóth (2011) for stick insect walking. The experiments showed that a pilocarpine-induced rhythm in the prothoracic protractor and retractor motoneurons could be entrained by an intact forward or backward walking hind leg. In stick insects, this is the evidence for a long range ipsilateral inter-segmental connection that mediates sensory information from a stepping hind leg to the prothoracic CPGs

Locomotion grows up: The neuromechanical control of interlimb coordinating mechanisms in crayfish

Locomotion requires many dynamic interactions between organism and environment at several levels. It is not known how the nervous system controls all of these relationships to ultimately produce and guide locomotor behavior. Furthermore, it is not known whether the nervous system needs to recognize and control all of the possible body-environment interactions. In this study the crayfish (Procambarus clarkii) is used as a model system to test how size influences locomotor behavior and how a single, simplified neuromechanical system can accommodate these changes.;A set of behavioral experiments was conducted to characterize kinematics of freely walking juvenile crayfish to compare with adults. The purpose of these studies was to determine how crayfish adapt to a great change in size during their ontogeny. Juvenile and adult crayfish show differences in limb function and coordination. Although crayfish are decapods, the juveniles predominantly use the posterior legs and behave more like four-legged walkers. The difference in locomotor behavior can best be explained by differences in chelae size. Allometric relationships between juveniles and adults show limb and body morphologies scale proportionately. Adult chelae, or claws, are twice as long and contribute ∼20% more to the total body mass in fully mature crayfish. This increase in chelae size shifts the location of the center of mass anterior as crayfish grow. The result is a change in relative load distribution that appears to affect individual limb behavior and interlimb coordination. Shifting the center of mass in adults by amputating the chelae resulted in limb behavior and interlimb coordination more similar to that observed in juveniles. Likewise, applying load to the rostrum of juveniles altered behavior and changed limb function in the posterior legs similar to adults with large chelae. The results of these experiments suggest that crayfish of all sizes adapt to changes in load distribution by adjusting behavior of individual legs.;To test whether developmental influences have an effect on walking behavior, juveniles were induced to walk on a treadmill at various speeds. The animals showed more consistent limb coordination as walking speed increased, similar to adults. Selected legs were then amputated to test how gait was affected. Amputating legs removes sensory feedback from the distal leg to the central nervous system. The behavior of the stump is therefore more representative of the endogenous rhythmicity of the central pattern generator (CPG). Juveniles showed no differences in coordination in individual legs. Coordination between adjacent ipsilateral legs was also the same as that observed in adults following amputation. Furthermore, intact legs acquired new interlimb coordination similar to adults. These results suggested that juvenile and adult crayfish have functionally similar nervous systems controlling walking.;Finally, a 3-D virtual crayfish was built to test whether differences in walking between juveniles and adults could be due to mechanical influences alone. The model crayfish lacked direct connections between legs. The model responded to shifts in the center of mass by showing more consistent limb coordination in those legs nearest the center of mass. This was achieved through indirect mechanical coupling of the legs through the environment and body of the crayfish. This mechanism also produced realistic adaptive behavior when limbs were amputated. This showed that differences between adult and juvenile walking are due solely to mechanical influences associated with the changing center of mass as the animals grow. These results suggest further that organisms do not need high levels of control to produce coordinated behavior. Locomotor behavior arises through interactions between body, limb, and environment that are a function of the spatio-temporal dynamics of body morphology. The results may be applicable to a large number of walking systems