Phase response analyses support a relaxation oscillator model of locomotor rhythm generation in Caenorhabditis elegans

Neural circuits coordinate with muscles and sensory feedback to generate motor behaviors appropriate to an animal’s environment. In C. elegans, the mechanisms by which the motor circuit generates undulations and modulates them based on the environment are largely unclear. We quantitatively analyzed C. elegans locomotion during free movement and during transient optogenetic muscle inhibition. Undulatory movements were highly asymmetrical with respect to the duration of bending and unbending during each cycle. Phase response curves induced by brief optogenetic inhibition of head muscles showed gradual increases and rapid decreases as a function of phase at which the perturbation was applied. A relaxation oscillator model based on proprioceptive thresholds that switch the active muscle moment was developed and is shown to quantitatively agree with data from free movement, phase responses, and previous results for gait adaptation to mechanical loadings. Our results suggest a neuromuscular mechanism underlying C. elegans motor pattern generation within a compact circuit.

Recent Insights into the Rhythmogenic Core of the Locomotor CPG

In order for locomotion to occur, a complex pattern of muscle activation is required. For more than a century, it has been known that the timing and pattern of stepping movements in mammals are generated by neural networks known as central pattern generators (CPGs), which comprise multiple interneuron cell types located entirely within the spinal cord. A genetic approach has recently been successful in identifying several populations of spinal neurons that make up this neural network, as well as the specific role they play during stepping. In spite of this progress, the identity of the neurons responsible for generating the locomotor rhythm and the manner in which they are interconnected have yet to be deciphered. In this review, we summarize key features considered to be expressed by locomotor rhythm-generating neurons and describe the different genetically defined classes of interneurons which have been proposed to be involved.

Neural Interactions in Developing Rhythmogenic Spinal Networks: Insights From Computational Modeling

The mechanisms involved in generation of rhythmic locomotor activity in the mammalian spinal cord remain poorly understood. These mechanisms supposedly rely on both intrinsic properties of constituting neurons and interactions between them. A subset of Shox2 neurons was suggested to contribute to generation of spinal locomotor activity, but the possible cellular basis for rhythmic bursting in these neurons remains unknown. Ha and Dougherty (2018) recently revealed the presence of bidirectional electrical coupling between Shox2 neurons in neonatal spinal cords, which can be critically involved in neuronal synchronization and generation of populational bursting. Gap junctional connections found between functionally-related Shox2 interneurons decrease with age, possibly being replaced by increasing interactions through chemical synapses. Here, we developed a computational model of a heterogeneous population of neurons sparsely connected by electrical or/and chemical synapses and investigated the dependence of frequency of populational bursting on the type and strength of neuronal interconnections. The model proposes a mechanistic explanation that can account for the emergence of a synchronized rhythmic activity in the neuronal population and provides insights into the possible role of gap junctional coupling between Shox2 neurons in the spinal mechanisms for locomotor rhythm generation.

The M-current works in tandem with the persistent sodium current to set the speed of locomotion

The central pattern generator (CPG) for locomotion is a set of pacemaker neurons endowed with inherent bursting driven by the persistent sodium current (INaP). How they proceed to regulate the locomotor rhythm remained unknown. Here, in neonatal rodents, we identified a persistent potassium current critical in regulating pacemakers and locomotion speed. This current recapitulates features of the M-current (IM): a subthreshold noninactivating outward current blocked by 10,10-bis(4-pyridinylmethyl)-9(10H)-anthracenone dihydrochloride (XE991) and enhanced by N-(2-chloro-5-pyrimidinyl)-3,4-difluorobenzamide (ICA73). Immunostaining and mutant mice highlight an important role of Kv7.2-containing channels in mediating IM. Pharmacological modulation of IM regulates the emergence and the frequency regime of both pacemaker and CPG activities and controls the speed of locomotion. Computational models captured these results and showed how an interplay between IM and INaP endows the locomotor CPG with rhythmogenic properties. Overall, this study provides fundamental insights into how IM and INaP work in tandem to set the speed of locomotion.

The role of clockwork orange in the circadian clock of the cricket Gryllus bimaculatus

The circadian clock generates rhythms of approximately 24 h through periodic expression of the clock genes. In insects, the major clock genes period (per) and timeless (tim) are rhythmically expressed upon their transactivation by CLOCK/CYCLE, with peak levels in the early night. In Drosophila, clockwork orange (cwo) is known to inhibit the transcription of per and tim during the daytime to enhance the amplitude of the rhythm, but its function in other insects is largely unknown. In this study, we investigated the role of cwo in the clock mechanism of the cricket Gryllus bimaculatus. The results of quantitative RT-PCR showed that under a light/dark (LD) cycle, cwo is rhythmically expressed in the optic lobe (lamina-medulla complex) and peaks during the night. When cwo was knocked down via RNA interference (RNAi), some crickets lost their locomotor rhythm, while others maintained a rhythm but exhibited a longer free-running period under constant darkness (DD). In cwoRNAi crickets, all clock genes except for cryptochrome 2 (cry2) showed arrhythmic expression under DD; under LD, some of the clock genes showed higher mRNA levels, and tim showed rhythmic expression with a delayed phase. Based on these results, we propose that cwo plays an important role in the cricket circadian clock.

Neural interactions in developing rhythmogenic spinal networks: Insights from computational modeling

The mechanisms involved in generation of rhythmic locomotor activity in the mammalian spinal cord remain poorly understood. These mechanisms supposedly rely on both intrinsic properties of constituting neurons and interactions between them. A subset of Shox2 neurons was found to contribute to generation of spinal locomotor activity, but the possible cellular basis for rhythmic bursting in these neurons remains unknown. Ha and Dougherty (2018) recently revealed the presence of bidirectional electrical coupling between Shox2 neurons in neonatal spinal cords, which can be critically involved in neuronal synchronization and generation of populational bursting. Gap junctional connections found between functionally-related Shox2 interneurons decrease with age, possibly being replaced by increasing interactions through chemical synapses. Here, we developed a computational model of a heterogeneous population of neurons sparsely connected by electrical or/and chemical synapses and investigated the dependence of frequency of populational bursting on the type and strength of neuronal interconnections. The model proposes a mechanistic explanation for emergence of a synchronized rhythmic activity in the neuronal population and provides insights into the mechanisms of the locomotor rhythm generation.

Long ascending propriospinal neurons provide flexible, context-specific control of interlimb coordination

Within the cervical and lumbar spinal enlargements, central pattern generator (CPG) circuitry produces the rhythmic output necessary for limb coordination during locomotion. Long propriospinal neurons that inter-connect these CPGs are thought to secure hindlimb-forelimb coordination, ensuring that diagonal limb pairs move synchronously while the ipsilateral limb pairs move out-of-phase during stepping. Here, we show that silencing long ascending propriospinal neurons (LAPNs) that inter-connect the lumbar and cervical CPGs disrupts left-right limb coupling of each limb pair in the adult rat during overground locomotion on a high-friction surface. These perturbations occurred independent of the locomotor rhythm, intralimb coordination, and speed-dependent (or any other) principal features of locomotion. Strikingly, the functional consequences of silencing LAPNs are highly context-dependent; the phenotype was not expressed during swimming, treadmill stepping, exploratory locomotion, or walking on an uncoated, slick surface. These data reveal surprising flexibility and context-dependence in the control of interlimb coordination during locomotion.

Phase response analyses support a relaxation oscillator model of locomotor rhythm generation in Caenorhabditis elegans

Neural circuits work together with muscles and sensory feedback to generate motor behaviors appropriate to an animal’s environment. In C. elegans, forward locomotion consists of dorsoventral undulations that propagate from anterior to posterior. How the worm’s motor circuit generates these undulations and modulates them based on external loading is largely unclear. To address this question, we performed quantitative behavioral analysis of C. elegans during free movement and during transient optogenetic muscle inhibition. Undulatory movements in the head were found to be highly asymmetric, with bending toward the ventral or dorsal directions occurring slower than straightening toward a straight posture during the locomotory cycle. Phase shifts induced by brief optogenetic inhibition of head muscles showed a sawtooth-shaped dependence on phase of inhibition. We developed a computational model based on proprioceptive postural thresholds that switch the active moment of body wall muscles. We show that our model, a type of relaxation oscillator, is in quantitative agreement with data from free movement, phase responses, and previous results for frequency and amplitude dependence on the viscosity of the external medium. Our results suggest a neuromuscular mechanism that enables C. elegans to coordinate rhythmic motor patterns within a compact circuit.

It takes two to tango: M-current swings with the persistent sodium current to set the speed of locomotion

SummaryThe central pattern generator (CPG) for locomotion is set of pacemaker neurons endowed with inherent bursting driven by the persistent sodium current (INaP). How they proceed to regulate the locomotor rhythm remained unknown. Here, in neonatal rodents, we identified a persistent potassium current, critical in regulating pacemakers and locomotion speed. This current recapitulates features of the M-current (IM); a subthreshold non-inactivating outward current blocked by XE991 and enhanced by ICA73. Immunostaining and mutant mice highlight an important role of axonal Kv7.2 channels in mediating IM. Pharmacological modulation of IM regulates the emergence and the frequency regime of both pacemaker and CPG activities, and controls the speed of locomotion. Computational models captured these results and show howed an interplay between IM and INaP that endows the locomotor CPG with rhythmogenic properties. Overall, this study provides fundamental insights into how IM and INaP work in tandem to set the speed of locomotion.