Modelling spinal circuitry involved in locomotor pattern generation: insights from the effects of afferent stimulation

A computational model of the mammalian spinal cord circuitry incorporating a two-level central pattern generator (CPG) with separate half-centre rhythm generator (RG) and pattern formation (PF) networks has been developed from observations obtained during fictive locomotion in decerebrate cats. Sensory afferents have been incorporated in the model to study the effects of afferent stimulation on locomotor phase switching and step cycle period and on the firing patterns of flexor and extensor motoneurones. Here we show that this CPG structure can be integrated with reflex circuits to reproduce the reorganization of group I reflex pathways occurring during locomotion. During the extensor phase of fictive locomotion, activation of extensor muscle group I afferents increases extensor motoneurone activity and prolongs the extensor phase. This extensor phase prolongation may occur with or without a resetting of the locomotor cycle, which (according to the model) depends on the degree to which sensory input affects the RG and PF circuits, respectively. The same stimulation delivered during flexion produces a temporary resetting to extension without changing the timing of following locomotor cycles. The model reproduces this behaviour by suggesting that this sensory input influences the PF network without affecting the RG. The model also suggests that the different effects of flexor muscle nerve afferent stimulation observed experimentally (phase prolongation versus resetting) result from opposing influences of flexor group I and II afferents on the PF and RG circuits controlling the activity of flexor and extensor motoneurones. The results of modelling provide insights into proprioceptive control of locomotion.     

The activity of spinal commissural interneurons during fictive locomotion in the lamprey

Commissural interneurons in the lamprey coordinate activity of the hemisegmental oscillators to ensure proper left-right alternation during swimming. The activity of interneuronal axons at the ventral commissure was studied together with potential target motoneurons during fictive locomotion in the isolated lamprey spinal cord. To estimate the unperturbed activity of the interneurons, axonal recordings were chosen because soma recordings inevitably will affect the level of membrane depolarization and thereby spike initiation. Of 227 commissural axons recorded during locomotor activity, 14 produced inhibitory and 3 produced excitatory postsynaptic potentials (PSPs) in target motoneurons. The axons typically fired multiple spikes per locomotor cycle, with approximately 10 Hz sustained frequency. The average shortest spike interval in a burst corresponded to an instantaneous frequency of approximately 50 Hz for both the excitatory and inhibitory axons. The maximum number of spikes per locomotor cycle was inversely related to the locomotor frequency, in accordance with previous observations in the spinal hemicord preparation. In axons that fired multiple spikes per cycle, the mean interspike intervals were in the range in which the amplitude of the slow afterhyperpolarization (sAHP) is large, providing further support for the role of the sAHP in spike timing. One hundred ninety-five axons (86%) fired rhythmically during fictive locomotion, with preferred phase of firing distributed over either the segmental locomotor burst phase (40% of axons) or the transitional phase (between bursts; 60%). Thus in lamprey commissural interneurons, we found a broad distribution of firing rates and phases during fictive locomotion.     

Motoneuron input-resistance changes during fictive locomotion produced by stimulation of the mesencephalic locomotor region

Input-resistance changes during fictive locomotion were monitored in a variety of extensor and flexor hindlimb alpha-motoneurons in precollicular, postmammillary decerebrate cats induced to "walk" by electrical stimulation of the mesencephalic locomotor region (MLR). Using intracellular recording techniques and injected hyperpolarizing current pulses, the changes in the motoneuron input resistance recorded at the motoneuron soma were examined during nonlocomoting control periods as well as during the depolarized and hyperpolarized phases of the membrane potential oscillations (locomotor drive potentials, or LDPs) of fictive locomotion. In 28 of the 52 motoneurons examined, no change in the input resistance between the control and locomotor periods was observed. The remainder of the cells displayed a decrease (less than 20%) in input resistance when fictive stepping commenced. Over 80% of all the motoneurons depolarized (mean depolarization 4 mV), whereas only one LG motoneuron hyperpolarized (2 mV) with the onset of stimulation of the MLR. The remaining motoneurons did not display such changes. In 43 out of 52 motoneurons examined, no significant change in the input resistance could be observed between the depolarized and hyperpolarized phases of the step cycle. A decrease in the input resistance during the depolarized phase of the LDP was observed in four LG motoneurons, whereas five other motoneurons (2 LG, 1 TA, 1 PB, and 1 ST) displayed an increased input resistance during the depolarized phase compared with the hyperpolarized phase of locomotion. The data are consistent with the presence of an excitatory synaptic input alternating with an inhibitory input to the motoneuron during the fictive step cycle.(ABSTRACT TRUNCATED AT 250 WORDS)     

Recovery of hindlimb locomotion after incomplete spinal cord injury in the cat involves spontaneous compensatory changes within the spinal locomotor circuitry

After incomplete spinal cord injury (SCI), compensatory changes occur throughout the whole neuraxis, including the spinal cord below the lesion, as suggested by previous experiments using a dual SCI paradigm. Indeed, cats submitted to a lateral spinal hemisection at T10-T11 and trained on a treadmill for 3-14 wk re-expressed bilateral hindlimb locomotion as soon as 24 h after spinalization, a process that normally takes 2-3 wk when a complete spinalization is performed without a prior hemisection. In this study, we wanted to ascertain whether similar effects could occur spontaneously without training between the two SCIs and within a short period of 3 wk in 11 cats. One day after the complete spinalization, 9 of the 11 cats were able to re-express hindlimb locomotion either bilaterally (n = 6) or unilaterally on the side of the previous hemisection (n = 3). In these 9 cats, the hindlimb on the side of the previous hemisection (left hindlimb) performed better than the right side in contrast to that observed during the hemispinal period itself. Cats re-expressing the best bilateral hindlimb locomotion after spinalization had the largest initial hemilesion and the most prominent locomotor deficits after this first SCI. These results provide evidence that 1) marked reorganization of the spinal locomotor circuitry can occur without specific locomotor training and within a short period of 3 wk; 2) the spinal cord can reorganize in a more or less symmetrical way; and 3) the ability to walk after spinalization depends on the degree of deficits and adaptation observed in the hemispinal period.     

Excitatory and inhibitory postsynaptic potentials in alpha-motoneurons produced during fictive locomotion by stimulation of the mesencephalic locomotor region

We tested the hypothesis that stimulation of the mesencephalic locomotor region (MLR) activates polysynaptic pathways that project to lumbar spinal motoneurons and are involved in the initiation of locomotion. Fictive locomotion was produced by MLR stimulation, and intracellular records of evoked postsynaptic potentials (PSPs) in alpha-motoneurons were computer analyzed. Stimulation of sites in the MLR that were maximally effective for the initiation of locomotion produced excitatory and inhibitory postsynaptic potentials (EPSPs and IPSPs) in all the motoneurons examined. The amplitudes of the PSPs increased as locomotion commenced. The EPSPs were largest during the depolarized phase of the step cycle, and in 17 of our 22 cells the EPSP was replaced by an IPSP of slightly longer latency during the hyperpolarized phase. The mean latency of the EPSPs measured from the stimulus artifact produced by stimulation of the MLR was 5.1 ms (3.0-7.0 ms). In all cases, the IPSP occurred 0.6 ms or more after the onset of the EPSP in the same cell. Later PSPs were sometimes observed as well. The effects of constant current injection on the membrane potential oscillations associated with fictive locomotion (locomotor drive potentials) were examined. The results showed that the amplitudes of the locomotor drive potentials (LDPs) could be affected by depolarizing and hyperpolarizing current injection. The data is consistent with the LDP having a predominant inhibitory component, which is more readily altered by current injection than is the excitatory component. The effect of constant current injections on the MLR-evoked PSPs was also examined, and it was observed that both EPSPs and IPSPs could be affected by the injected currents. The EPSPs increased in amplitude with constant hyperpolarizing current injection, and this fact rules out the possibility that the EPSP is actually a reversed IPSP. The IPSP was decreased in amplitude by hyperpolarizing current injection. Combined stimulation of the MLR and the ipsilateral high-threshold muscle or cutaneous afferents produced facilitation of both short- and long-latency MLR-evoked PSPs, suggesting that the two pathways share common interneurons. The possibility that the long-latency PSPs are produced by rapid oscillation in the locomotor central pattern generator is discussed. We concluded that MLR stimulation that evokes fictive locomotion produces both excitation and inhibition of spinal motoneurons. Spinal interneuronal systems are implicated and may be those involved in the initiation and control of locomotion. The probable relay sites for the descending pathway from the MLR to motoneurons are discussed.     

Motoneuronal and muscle synergies involved in cat hindlimb control during fictive and real locomotion: a comparison study

We compared the activity profiles and synergies of spinal motoneurons recorded during fictive locomotion evoked in immobilized decerebrate cat preparations by midbrain stimulation to the activity profiles and synergies of the corresponding hindlimb muscles obtained during forward level walking in cats. The fictive locomotion data were collected in the Spinal Cord Research Centre, University of Manitoba, and provided by Dr. David McCrea; the real locomotion data were obtained in the laboratories of M. A. Lemay and B. I. Prilutsky. Scatterplot representation and minimum spanning tree clustering algorithm were used to identify the possible motoneuronal and muscle synergies operating during both fictive and real locomotion. We found a close similarity between the activity profiles and synergies of motoneurons innervating one-joint muscles during fictive locomotion and the profiles and synergies of the corresponding muscles during real locomotion. However, the activity patterns of proximal nerves controlling two-joint muscles, such as posterior biceps and semitendinosus (PBSt) and rectus femoris (RF), were not uniform in fictive locomotion preparations and differed from the activity profiles of the corresponding two-joint muscles recorded during forward level walking. Moreover, the activity profiles of these nerves and the corresponding muscles were unique and could not be included in the synergies identified in fictive and real locomotion. We suggest that afferent feedback is involved in the regulation of locomotion via motoneuronal synergies controlled by the spinal central pattern generator (CPG) but may also directly affect the activity of motoneuronal pools serving two-joint muscles (e.g., PBSt and RF). These findings provide important insights into the organization of the spinal CPG in mammals, the motoneuronal and muscle synergies engaged during locomotion, and their afferent control.     

Stumbling corrective reaction during fictive locomotion in the cat

An obstacle contacting the dorsal surface of a cat's hind foot during the swing phase of locomotion evokes a reflex (the stumbling corrective reaction) that lifts the foot and extends the ankle to avoid falling. We show that the same sequence of ipsilateral hindlimb motoneuron activity can be evoked in decerebrate cats during fictive locomotion. As recorded in the peripheral nerves, twice threshold intensity stimulation of the cutaneous superficial peroneal (SP) nerve during the flexion phase produced a very brief excitation of ankle flexors (e.g., tibialis anterior and peroneus longus) that was followed by an inhibition for the duration of the stimulus train (10-25 shocks, 200 Hz). Extensor digitorum longus was always, and hip flexor (sartorius) activity was sometimes, inhibited during SP stimulation. At the same time, knee flexor and the normally quiescent ankle extensor motoneurons were recruited (mean latencies 4 and 16 ms) with SP stimulation during fictive stumbling correction. After the stimulus train, ankle extensor activity fell silent, and there was an excitation of hip, knee, and ankle flexors. The ongoing flexion phase was often prolonged. Hip extensors were also recruited in some fictive stumbling trials. Only the SP nerve was effective in evoking stumbling correction. Delivered during extension, SP stimulus trains increased ongoing extensor motoneuron activity as well as increasing ipsilateral hip, knee, and ankle hindlimb flexor activity in the subsequent step cycle. The fictive stumbling corrective reflex seems functionally similar to that evoked in intact, awake animals and involves a fixed pattern of short-latency reflexes as well as actions evoked through the lumbar circuitry responsible for the generation of rhythmic alternating locomotion.     

Interlimb coordination during fictive locomotion in the thalamic cat

Summary Efferent discharges in muscle nerves of the four limbs were recorded simultaneously during spontaneous fictive locomotion in thalamic cats with the goal of understanding how the central nervous system controls interlimb coordination during stepping. The onset of the bursts of activity in the nerve of a selected flexor muscle in each limb allowed the temporal and the phase relationships between the fictive step cycle of a pair of limbs to be determined. Our main results are the following: 1) the fictive step cycles of the two forelimbs are always strictly alternated whereas the phasing of the step cycles of either the two hindlimbs or pairs of homolateral or diagonal limbs is more variable; 2) the time interval between the onsets of the flexor bursts of one of the two pairs of diagonal limbs is independent of the step cycle duration; 3) distinct patterns of interlimb coordination exist during fictive locomotion; a small number of patterns of coordination involving all four limbs, which correspond to the walking and the trotting gaits in the intact cat, occur very frequently. The results demonstrate that the central nervous system deprived of phasic afferent inputs from the periphery has the capacity to generate most of the patterns of interlimb coordination which occur during real locomotion. They further support the view that the central pattern of interlimb coordination essentially results from diagonal interactions between a forelimb generator for locomotion and a hindlimb one.Abbreviations CD step cycle duration - LF left forelimb - LH left hindlimb - m slope of correlation curve - N number of step cycles - r correlation coefficient - RF right forelimb - RH right hindlimb - Ti time interval     

Intracellular QX-314 causes depression of membrane potential oscillations in lamprey spinal neurons during fictive locomotion

Spinal neurons undergo large cyclic membrane potential oscillations during fictive locomotion in lamprey. It was investigated whether these oscillations were due only to synaptically driven excitatory and inhibitory potentials or if voltage-dependent inward conductances also contribute to the depolarizing phase by using N-(2,6-dimethylphenyl carbamoylmethyl)triethylammonium bromide (QX-314) administered intracellularly during fictive locomotion. QX-314 intracellularly blocks inactivating and persistent Na+ channels, and in some neurons, effects on certain other types of channels have been reported. To detail the effects of QX-314 on Na+ and Ca2+ channels, we used dissociated lamprey neurons recorded under whole cell voltage clamp. At low intracellular concentrations of QX-314 (0.2 mM), inactivating Na+ channels were blocked and no effects were exerted on Ca2+ channels (also at 0.5 mM). At 10 mM QX-314, there was, however a marked reduction of I(Ca). In the isolated spinal cord of the lamprey, fictive locomotion was induced by superfusing the spinal cord with Ringer's solution containing N-methyl-D-aspartate (NMDA), while recording the locomotor activity from the ventral roots. Simultaneously, identified spinal neurons were recorded intracellularly, while infusing QX-314 from the microelectrode. Patch electrodes cannot be used in the intact spinal cord, and therefore "sharp" electrodes were used. The amplitude of the oscillations was consistently reduced by 20-25% in motoneurons (P < 0.05) and unidentified spinal neurons (P < 0.005). The onset of the effect started a few minutes after impalement and reached a stable level within 30 min. These effects thus show that QX-314 causes a reduction in the amplitude of membrane potential oscillations during fictive locomotion. We also investigated whether QX-314 could affect glutamate currents by applying short pulses of glutamate from an extracellular pipette. No changes were observed. We also found no evidence for a persistent Na+ current in dissociated neurons, but these cells have a much-reduced dendritic tree. The results indicate that there is an inward conductance, which is sensitive to QX-314, during membrane potential oscillations that "boosts" the synaptic drive during fictive locomotion. Taken together, the results suggest that inactivating Na+ channels contribute to this inward conductance although persistent Na+ channels, if present on dendrites, could possibly also contribute to shaping the membrane potential oscillations.     

Reversal of the late phase of spike frequency adaptation in cat spinal motoneurons during fictive locomotion

In spinal motoneurons, late spike frequency adaptation (SFA) is defined as the slowing of the firing rate over tens of seconds and can be seen during sustained or intermittent current injection. Although the function of late SFA is not known, it may result in a decrease in force production over time, or muscle fatigue. Because locomotion can persist for long periods of time without fatigue, late SFA was studied using intracellular recordings from adult cat motoneurons during fictive locomotion. Of eight lumbar motoneurons studied, all showed late adaptation during control conditions, but none demonstrated late adaptation during locomotor activity. The most consistent properties that correlated with the presence or absence of late SFA were those related to availability of fast, inactivating sodium channels, particularly action potential rate of rise. Evidence of the reversal of late SFA during locomotion was present for several minutes following locomotor trials, consistent with the suggestion that SFA is modulated through slow metabotropic pathways. The abolition of late adaptation in spinal motoneurons during fictive locomotion is an example of a state-dependent change in the "intrinsic" properties of mammalian motoneurons. This change contributes to increased excitability of motoneurons during locomotion and results in robust firing during sustained locomotion.     

Cutaneous fiber groups involved in the inhibition of fictive locomotion in the rabbit

Summary Decorticate, paralyzed unanaesthetized rabbit preparations can display motoneuron discharges (spontaneous or elicited through various somatic stimulations) which are related to locomotion. These activities are suppressed when manual pressure is exerted on the dorso-lumbar skin. The present study indicates that: (1) locomotor discharges can also be inhibited through repetitive electrical stimulation of one of the skin nerves belonging to dorsal dermatomes (TH5 to L5); (2) other skin nerves, especially those of the limbs, do not present the same inhibitory properties; (3) among the various fiber groups composing these dorsal skin nerves, only A delta are inhibitory; other groups, on the contrary, display moderate (A alpha and A beta) or strong (unmyelinated C fibers) excitatory actions; (4) the inhibitory action of A delta fibers counteracts the excitatory action of C fibers, when both groups are stimulated together. The relationship between this form of motor inhibition and the well known phenomenon of reflex immobility is briefly considered.     

Gamma-motoneurone discharge patterns during fictive locomotion in the decerebrate cat

Triceps surae gamma-motoneurones were recorded during fictive locomotion in the paralysed high decerebrate cat. Two distinctive patterns of discharge were observed which were similar to those reported for static and dynamic gamma-motoneurones during locomotion in the same preparation, but without paralysis (Murphy, Stein & Taylor, 1984). These results suggest that movement-related afferent feedback is not essential for the generation of the basic patterns of static and dynamic gamma-motoneurone activity during locomotion. The results are discussed in relation to the generation of alpha and gamma locomotor rhythms.     

Activity of interneurons within the L4 spinal segment of the cat during brainstem-evoked fictive locomotion

Summary Extracellular recordings from interneurons located in the L4 spinal segment were made during fictive locomotion produced by electrical stimulation of the mesencephalic locomotor region (MLR) in the paralysed decerebrate cat. Only interneurons within the L4 segment which received group II input from quadriceps, sartorius or the pretibial flexor muscle afferents and which had axonal projections to motor nuclei in L7 were selected for analysis. During the fictive step cycle two thirds of these interneurons fired action potentials during the time of activity in the ipsilateral hindlimb flexor neurograms. These cells were also less responsive to peripheral input during the extension phase of the fictive locomotion cycle. The remaining one third of the interneurons examined were not rhythmically active during locomotion. The possible contributions of the midlumbar interneurons to motoneuron activity during locomotion are discussed.     

Quantitative investigation of calcium signals for locomotor pattern generation in the lamprey spinal cord

Locomotor pattern generation requires the network coordination of spinal ventral horn neurons acting in concert with the oscillatory properties of individual neurons. In the spinal cord, N-methyl-d-aspartate (NMDA) activates neuronal oscillators that are believed to rely on Ca(2+) entry to the cytosol through voltage-operated Ca(2+) channels and synaptically activated NMDA receptors. Ca(2+) signaling in lamprey ventral horn neurons thus plays a determinant role in the regulation of the intrinsic membrane properties and network synaptic interaction generating spinal locomotor neural pattern activity. We have characterized aspects of this signaling quantitatively for the first time. Resting Ca(2+) concentrations were between 87 and 120 nM. Ca(2+) concentration measured during fictive locomotion increased from soma to distal dendrites [from 208 +/- 27 (SE) nM in the soma to 335 +/- 41 nM in the proximal dendrites to 457 +/- 68 nM in the distal dendrites]. We sought to determine the temporal and spatial properties of Ca(2+) oscillations, imaged with Ca(2+)-sensitive dyes and correlated with fluctuations in membrane potential, during lamprey fictive locomotion. The Ca(2+) signals recorded in the dendrites showed a great deal of spatial heterogeneity. Rapid changes in Ca(2+)-induced fluorescence coincided with action potentials, which initiated significant Ca(2+) transients distributed throughout the neurons. Ca(2+) entry to the cytosol coincided with the depolarizing phase of the locomotor rhythm. During fictive locomotion, larger Ca(2+) oscillations were recorded in dendrites compared with somata in motoneurons and premotor interneurons. Ca(2+) fluctuations were barely detected with dyes of lower affinity providing alternative empirical evidence that Ca(2+) responses are limited to hundreds of nanomolars during fictive locomotion.