What single-cell stimulation has told us about neural coding

In recent years, single-cell stimulation experiments have resulted in substantial progress towards directly linking single-cell activity to movement and sensation. Recent advances in electrical recording and stimulation techniques have enabled control of single neuron spiking in vivo and have contributed to our understanding of neuronal coding schemes in the brain. Here, we review single neuron stimulation effects in different brain structures and how they vary with artificially inserted spike patterns. We briefly compare single neuron stimulation with other brain stimulation techniques. A key advantage of single neuron stimulation is the precise control of the evoked spiking patterns. Systematically varying spike patterns and measuring evoked movements and sensations enables ‘decoding’ of the single-cell spike patterns and provides insights into the readout mechanisms of sensory and motor cortical spikes.     

An image-free opto-mechanical system for creating virtual environments and imaging neuronal activity in freely moving Caenorhabditis elegans

Non-invasive recording in untethered animals is arguably the ultimate step in the analysis of neuronal function, but such recordings remain elusive. To address this problem, we devised a system that tracks neuron-sized fluorescent targets in real time. The system can be used to create virtual environments by optogenetic activation of sensory neurons, or to image activity in identified neurons at high magnification. By recording activity in neurons of freely moving C. elegans, we tested the long-standing hypothesis that forward and reverse locomotion are generated by distinct neuronal circuits. Surprisingly, we found motor neurons that are active during both types of locomotion, suggesting a new model of locomotion control in C. elegans. These results emphasize the importance of recording neuronal activity in freely moving animals and significantly expand the potential of imaging techniques by providing a mean to stabilize fluorescent targets.     

Direct and indirect assessment of gamma-motor firing patterns

The study of the patterns of gamma-motor activity which accompany natural contractions has been long and difficult, and has not as yet led to general agreement. In this review we have simplified matters by considering the case of locomotion in the cat only, and we have avoided discussion of the various hypotheses which have been advanced to provide general schemes of gamma control for a wide range of movements. The development of the subject is shown to depend very much on devising ingenious methods applicable to reduced and intact animals. Direct recording from gamma-motoneurones has only been possible in reduced preparations, whereas indirect assessment of gamma activity from spindle afferent recordings was used in these and in intact animals. At this point in time, we still have no direct recordings from gamma-motoneurones in normally behaving animals, but those obtained in decerebrate animals show distinct patterns of modulation for static and dynamic types with particular temporal relation to the stepping movements. The spindle recordings in intact animals potentially provide the most important information, and the problems of interpretation, which have previously caused difficulties, are beginning to be solved through the insights obtained from the reduced preparations.     

Gaze Strategy in the Free Flying Zebra Finch (Taeniopygia guttata)

Fast moving animals depend on cues derived from the optic flow on their retina. Optic flow from translational locomotion includes information about the three-dimensional composition of the environment, while optic flow experienced during a rotational self motion does not. Thus, a saccadic gaze strategy that segregates rotations from translational movements during locomotion will facilitate extraction of spatial information from the visual input. We analysed whether birds use such a strategy by highspeed video recording zebra finches from two directions during an obstacle avoidance task. Each frame of the recording was examined to derive position and orientation of the beak in three-dimensional space. The data show that in all flights the head orientation was shifted in a saccadic fashion and was kept straight between saccades. Therefore, birds use a gaze strategy that actively stabilizes their gaze during translation to simplify optic flow based navigation. This is the first evidence of birds actively optimizing optic flow during flight.     

Whole Body Mechanics of Stealthy Walking in Cats

The metabolic cost associated with locomotion represents a significant part of an animal''s metabolic energy budget. Therefore understanding the ways in which animals manage the energy required for locomotion by controlling muscular effort is critical to understanding limb design and the evolution of locomotor behavior. The assumption that energetic economy is the most important target of natural selection underlies many analyses of steady animal locomotion, leading to the prediction that animals will choose gaits and postures that maximize energetic efficiency. Many quadrupedal animals, particularly those that specialize in long distance steady locomotion, do in fact reduce the muscular contribution required for walking by adopting pendulum-like center of mass movements that facilitate exchange between kinetic energy (KE) and potential energy (PE) [1]–[4]. However, animals that are not specialized for long distance steady locomotion may face a more complex set of requirements, some of which may conflict with the efficient exchange of mechanical energy. For example, the “stealthy” walking style of cats may demand slow movements performed with the center of mass close to the ground. Force plate and video data show that domestic cats (Felis catus, Linnaeus, 1758) have lower mechanical energy recovery than mammals specialized for distance. A strong negative correlation was found between mechanical energy recovery and diagonality in the footfalls and there was also a negative correlation between limb compression and diagonality of footfalls such that more crouched postures tended to have greater diagonality. These data show a previously unrecognized mechanical relationship in which crouched postures are associated with changes in footfall pattern which are in turn related to reduced mechanical energy recovery. Low energy recovery was not associated with decreased vertical oscillations of the center of mass as theoretically predicted, but rather with posture and footfall pattern on the phase relationship between potential and kinetic energy. An important implication of these results is the possibility of a tradeoff between stealthy walking and economy of locomotion. This potential tradeoff highlights the complex and conflicting pressures that may govern the locomotor choices that animals make.     

Visual Exploration during Locomotion Limited by Fear of Heights

Background

Visual exploration of the surroundings during locomotion at heights has not yet been investigated in subjects suffering from fear of heights.

Methods

Eye and head movements were recorded separately in 16 subjects susceptible to fear of heights and in 16 non-susceptible controls while walking on an emergency escape balcony 20 meters above ground level. Participants wore mobile infrared eye-tracking goggles with a head-fixed scene camera and integrated 6-degrees-of-freedom inertial sensors for recording head movements. Video recordings of the subjects were simultaneously made to correlate gaze and gait behavior.

Results

Susceptibles exhibited a limited visual exploration of the surroundings, particularly the depth. Head movements were significantly reduced in all three planes (yaw, pitch, and roll) with less vertical head oscillations, whereas total eye movements (saccade amplitudes, frequencies, fixation durations) did not differ from those of controls. However, there was an anisotropy, with a preference for the vertical as opposed to the horizontal direction of saccades. Comparison of eye and head movement histograms and the resulting gaze-in-space revealed a smaller total area of visual exploration, which was mainly directed straight ahead and covered vertically an area from the horizon to the ground in front of the feet. This gaze behavior was associated with a slow, cautious gait.

Conclusions

The visual exploration of the surroundings by susceptibles to fear of heights differs during locomotion at heights from the earlier investigated behavior of standing still and looking from a balcony. During locomotion, anisotropy of gaze-in-space shows a preference for the vertical as opposed to the horizontal direction during stance. Avoiding looking into the abyss may reduce anxiety in both conditions; exploration of the “vertical strip” in the heading direction is beneficial for visual control of balance and avoidance of obstacles during locomotion.     

Task-Level Strategies for Human Sagittal-Plane Running Maneuvers Are Consistent with Robotic Control Policies

The strategies that humans use to control unsteady locomotion are not well understood. A “spring-mass” template comprised of a point mass bouncing on a sprung leg can approximate both center of mass movements and ground reaction forces during running in humans and other animals. Legged robots that operate as bouncing, “spring-mass” systems can maintain stable motion using relatively simple, distributed feedback rules. We tested whether the changes to sagittal-plane movements during five running tasks involving active changes to running height, speed, and orientation were consistent with the rules used by bouncing robots to maintain stability. Changes to running height were associated with changes to leg force but not stance duration. To change speed, humans primarily used a “pogo stick” strategy, where speed changes were associated with adjustments to fore-aft foot placement, and not a “unicycle” strategy involving systematic changes to stance leg hip moment. However, hip moments were related to changes to body orientation and angular speed. Hip moments could be described with first order proportional-derivative relationship to trunk pitch. Overall, the task-level strategies used for body control in humans were consistent with the strategies employed by bouncing robots. Identification of these behavioral strategies could lead to a better understanding of the sensorimotor mechanisms that allow for effective unsteady locomotion.     

Pattern generation

Central pattern generators are neuronal ensembles capable of producing the basic spatiotemporal patterns underlying ‘automatic’ movements (e.g. locomotion, respiration, swallowing and defense reactions), in the absence of peripheral feedback. Different experimental approaches, from classical electrophysiological and pharmacological methods to molecular and genetic ones, have been used to understand the cellular and synaptic bases of central pattern generator organization and reconfiguration of generator operation in behaviorally relevant contexts. Recently, it has been shown that the high reliability and flexibility of central pattern generators is determined by their redundant organization. Everything that is crucial for generator operation is determined by a number of complementary mechanisms acting in concert; however, various mechanisms are weighted differently in determining different aspects of central pattern generator operation.     

Characteristics of Gait Ataxia in δ2 Glutamate Receptor Mutant Mice,ho15J

The cerebellum plays a fundamental, but as yet poorly understood, role in the control of locomotion. Recently, mice with gene mutations or knockouts have been used to investigate various aspects of cerebellar function with regard to locomotion. Although many of the mutant mice exhibit severe gait ataxia, kinematic analyses of limb movements have been performed in only a few cases. Here, we investigated locomotion in ho15J mice that have a mutation of the δ2 glutamate receptor. The cerebellum of ho15J mice shows a severe reduction in the number of parallel fiber-Purkinje synapses compared with wild-type mice. Analysis of hindlimb kinematics during treadmill locomotion showed abnormal hindlimb movements characterized by excessive toe elevation during the swing phase, and by severe hyperflexion of the ankles in ho15J mice. The great trochanter heights in ho15J mice were lower than in wild-type mice throughout the step cycle. However, there were no significant differences in various temporal parameters between ho15J and wild-type mice. We suggest that dysfunction of the cerebellar neuronal circuits underlies the observed characteristic kinematic abnormality of hindlimb movements during locomotion of ho15J mice.     

Symmetry-breaking bifurcation: A possible mechanism for 2:1 frequency-locking in animal locomotion

The generation and control of animal locomotion is believed to involve central pattern generators — networks of neurons which are capable of producing oscillatory behavior. In the present work, the quadrupedal locomotor central pattern generator is modelled as four distinct but symmetrically coupled nonlinear oscillators. We show that the typical patterns for two such networks of oscillators include 2:1 frequency-locked oscillations. These patterns, which arise through symmetry-breaking Hopf bifurcation, correspond in part to observed patterns of 2:1 frequency-locking of limb movements during electrically elicited locomotion of decerebrate and spinal quadrupeds. We briefly describe how our theoretical predictions could be tested experimentally.     

Measurement of Larval Activity in the Drosophila Activity Monitor

Drosophila larvae are used in many behavioral studies, yet a simple device for measuring basic parameters of larval activity has not been available. This protocol repurposes an instrument often used to measure adult activity, the TriKinetics Drosophila activity monitor (MB5 Multi-Beam Activity Monitor) to study larval activity. The instrument can monitor the movements of animals in 16 individual 8 cm glass assay tubes, using 17 infrared detection beams per tube. Logging software automatically saves data to a computer, recording parameters such as number of moves, times sensors were triggered, and animals’ positions within the tubes. The data can then be analyzed to represent overall locomotion and/or position preference as well as other measurements. All data are easily accessible and compatible with basic graphing and data manipulation software. This protocol will discuss how to use the apparatus, how to operate the software and how to run a larval activity assay from start to finish.     

The prisoner as model organism: malaria research at Stateville Penitentiary

In a military-sponsored research project begun during the Second World War, inmates of the Stateville Penitentiary in Illinois were infected with malaria and treated with experimental drugs that sometimes had vicious side effects. They were made into reservoirs for the disease and they provided a food supply for the mosquito cultures. They acted as secretaries and technicians, recording data on one another, administering malarious mosquito bites and experimental drugs to one another, and helping decide who was admitted to the project and who became eligible for early parole as a result of his participation. Thus, the prisoners were not simply research subjects; they were deeply constitutive of the research project. Because a prisoner’s time on the project was counted as part of his sentence, and because serving on the project could shorten one’s sentence, the project must be seen as simultaneously serving the functions of research and punishment. Michel Foucault wrote about such ‘mixed mechanisms’ in his Discipline and punish. His shining example of such a ‘transparent’ and subtle style of punishment was the panopticon, Jeremy Bentham’s architectural invention of prison cellblocks arrayed around a central guard tower. Stateville prison was designed on Bentham’s model; Foucault featured it in his own discussion. This paper, then, explores the power relations in this highly idiosyncratic experimental system, in which the various roles of model organism, reagent, and technician are all occupied by sentient beings who move among them fluidly. This, I argue, created an environment in the Stateville hospital wing more panoptic than that in the cellblocks. Research and punishment were completely interpenetrating, and mutually reinforcing.     

Rhythms of locomotion expressed by Limulus polyphemus, the American horseshoe crab: II. Relationship to circadian rhythms of visual sensitivity

In the laboratory, horseshoe crabs express a circadian rhythm of visual sensitivity as well as daily and circatidal rhythms of locomotion. The major goal of this investigation was to determine whether the circadian clock underlying changes in visual sensitivity also modulates locomotion. To address this question, we developed a method for simultaneously recording changes in visual sensitivity and locomotion. Although every animal (24) expressed consistent circadian rhythms of visual sensitivity, rhythms of locomotion were more variable: 44% expressed a tidal rhythm, 28% were most active at night, and the rest lacked statistically significant rhythms. When exposed to artificial tides, 8 of 16 animals expressed circatidal rhythms of locomotion that continued after tidal cycles were stopped. However, rhythms of visual sensitivity remained stable and showed no tendency to be influenced by the imposed tides or locomotor activity. These results indicate that horseshoe crabs possess at least two biological clocks: one circadian clock primarily used for modulating visual sensitivity, and one or more clocks that control patterns of locomotion. This arrangement allows horseshoe crabs to see quite well while mating during both daytime and nighttime high tides.     

Elegant‐crested Tinamous Eudromia elegans do not synchronize head and leg movements during head‐bobbing

Head‐bobbing is the fore–aft movement of the head relative to the body during terrestrial locomotion in birds. It is considered to be a behaviour that helps to stabilize images on the retina during locomotion, yet some studies have suggested biomechanical links between the movements of the head and legs. This study analysed terrestrial locomotion and head‐bobbing in the Elegant‐crested Tinamou Eudromia elegans at a range of speeds by synchronously recording high‐speed video and ground reaction forces in a laboratory setting. The results indicate that the timing of head and leg movements are dissociated from one another. Nonetheless, head and neck movements do affect stance duration, ground reaction forces and body pitch and, as a result, the movement of the centre of mass in head‐bobbing birds. This study does not support the hypothesis that head‐bobbing is itself constrained by terrestrial locomotion. Instead, it suggests that visual cues are the primary trigger for head‐bobbing in birds, and locomotion is, in turn, constrained by a need for image stabilization and depth perception.     

Multidimensional recording (MDR) and data sharing: an ecological open research and educational platform for neuroscience

Primate neurophysiology has revealed various neural mechanisms at the single-cell level and population level. However, because recording techniques have not been updated for several decades, the types of experimental design that can be applied in the emerging field of social neuroscience are limited, in particular those involving interactions within a realistic social environment. To address these limitations and allow more freedom in experimental design to understand dynamic adaptive neural functions, multidimensional recording (MDR) was developed. MDR obtains behavioral, neural, eye position, and other biological data simultaneously by using integrated multiple recording systems. MDR gives a wide degree of freedom in experimental design because the level of behavioral restraint is adjustable depending on the experimental requirements while still maintaining the signal quality. The biggest advantage of MDR is that it can provide a stable neural signal at higher temporal resolution at the network level from multiple subjects for months, which no other method can provide. Conventional event-related analysis of MDR data shows results consistent with previous findings, whereas new methods of analysis can reveal network mechanisms that could not have been investigated previously. MDR data are now shared in the public server Neurotycho.org. These recording and sharing methods support an ecological system that is open to everyone and will be a valuable and powerful research/educational platform for understanding the dynamic mechanisms of neural networks.     

Imaging morphogenesis, in Xenopus with Quantum Dot nanocrystals

Mesoderm migration is a well studied morphogenetic movement that takes place during Xenopus gastrulation. The study of mesoderm migration and other morphogenetic movements has been primarily based on in vitro assays due to the inability to image deep tissue movements in the opaque embryo. We are the first to report the use of Near Infra Red Quantum Dots (NIR QD’s) to image mesoderm migration in vivo with single cell resolution and provide quantitative in vivo data regarding migration rates. In addition we use QD’s to address the function of the focal adhesion kinase (FAK) in this movement. Inhibition of FAK blocks mesoderm spreading and migration both in vitro and in vivo without affecting convergent extension highlighting the molecular differences between the two movements. These results provide new insights about the role of FAK and of focal adhesions during gastrulation and provide a new tool for the study of morphogenesis in vivo.     

Coordinated network functioning in the spinal cord: An evolutionary perspective

The successful achievement of harmonious locomotor movement results from the integrated operation of all body segments. Here, we will review current knowledge on the functional organization of spinal networks involved in mammalian locomotion. Attention will not simply be restricted to hindlimb muscle control, but by also considering the necessarily coordinated activation of trunk and forelimb muscles, we will try to demonstrate that while there has been a progressive increase in locomotor system complexity during evolution, many basic organizational features have been preserved across the spectrum from lower vertebrates through to humans. Concerning the organization of axial neuronal networks that control trunk muscles, it has been found across the vertebrate range that during locomotor movement a motor wave travels longitudinally in the spinal cord via the coupling of rhythmic segmental networks. For hindlimb activation it has been found in all species studied that the rostral lumbar segments contain the key elements for pattern generation. We also showed that rhythmic arm movements are under the control of cervical forelimb generators in quadrupeds as well as in human. Finally, it is highlighted that the coordination of quadrupedal movements during locomotion derives principally from an asymmetrical coordinating influence occurring in the caudo-rostral direction from the lumbar hindlimb networks.     

Emotional reactions to learning in cattle

It has been suggested that during instrumental learning, animals are likely to react emotionally to the reinforcer. They may in addition react emotionally to their own achievements. These reactions are of interest with regard to the animals’ capacity for self-awareness. Therefore, we devised a yoked control experiment involving the acquisition of an operant task. We aimed to identify the emotional reactions of young cattle to their own learning and to separate these from reactions to a food reward. Twelve Holstein–Friesian heifers aged 7–12 months were divided into two groups. Heifers in the experimental group were conditioned over a 14-day period to press a panel in order to open a gate for access to a food reward. For heifers in the control group, the gate opened after a delay equal to their matched partner’s latency to open it. To allow for observation of the heifers’ movements during locomotion after the gate had opened, there was a 15 m distance in the form of a race from the gate to the food trough. The heart rate of the heifers, and their behaviour when moving along the race towards the food reward were measured. When experimental heifers made clear improvements in learning, they were more likely than on other occasions to have higher heart rates and tended to move more vigorously along the race in comparison with their controls. This experiment found some, albeit inconclusive, indication that cattle may react emotionally to their own learning improvement.     

Limb movement,intensity of integral afferent impulses from limb receptors,and level of primary afferent terminal polarization during locomotion in cats decerebrated at high level

The relationship between parameters of electrical muscle activity, changes at hindlimb joint angles, intensity of integral afferent flow, and dorsal root potential during real-life locomotion was investigated in cats decerebrated at high level. Characteristics of rear limb movements before and after deafferentation were described. It was found that afferent activity during locomotion motion consists, of two components — a tonic and a periodic phasic stage. Three main waves may be distinguished in the latter, each of which gives rise to associated changes in the level of primary afferent terminal polarization. These changes in turn are summated with the effects produced by the central generator. Correlations, between the parameters of these processes were investigated and the mechanisms underlying afferent control of locomotion generator function discussed.A. A. Bogomolets Institute of Physiology, Academy of Sciences of the Ukrainian SSR, Kiev. Translated from Neirofiziologiya, Vol. 20, No. 1, pp. 119–127, January–February, 1988.     

In vivo recordings of neuroendocrine cells (caudo-dorsal cells) in the pond snail

Summary Ovulation and egg-laying behavior in the pond snailLymnaea stagnalis are controlled by the neuroendocrine caudodorsal cells (CDCs), constituting two clusters — one in each cerebral ganglion — totaling about 100 cells. In vitro studies have shown that the CDCs release their products, including the ovulation hormone, during a burst of spiking activity lasting for about 30 min (CDC discharge). This burst can be initiated by repeated intracellular stimulation with depolarizing current pulses, in which case the firing pattern is termed afterdischarge.Using cuff electrodes we recorded extracellularly from the intercerebral commissure, (the neurohaemal area of the CDCs) to study the activity of these cells during spontaneous egg-laying of freely behaving snails.The cuff-implanted snails showed long-lasting spiking activity prior to every bout of egg-laying. These spontaneous long-lasting discharges had several features in common with the intracellularly recorded activity of the CDCs in vitro: the time courses of spike broadening and of firing rates in the cuff-implanted animals were very similar to the characteristic patterns found in the isolated brain. Firing rates were higher and durations were longer in the cuff-implanted animals, however. In vitro, the duration of the discharge could be prolonged appreciably by recording in blood instead of normal saline, indicating that the bathing fluid normally used causes shortening of the CDC discharge. The way in which CDC discharges are triggered is discussed as a possible explanation for the differences in firing rates.The pattern of locomotion during spontaneous egg-laying was largely similar in cuff-implanted and unoperated animals. The level of locomotion was lower in the experimental animals. In addition, the rate of locomotion only partially returned to pre-oviposition levels. This is ascribed to the effect of the operation.It is concluded that the afterdischarge is the natural firing pattern of the caudodorsal cells ofLymnaea, and that this firing pattern constitutes the central event in the egg-laying behavior of this animal.Abbreviations CDC caudodorsal cells - CDCH caudodorsal cell hormone - CDCA caudodorsal cell autotransmitter