Rhythm generation in neuronal networks relies on synaptic interactions and pacemaker properties. Little is known about the contribution of the latter mechanisms to the integrated network activity underlying locomotion in mammals. We tested the hypothesis that the persistent sodium current (I NaP ) is critical in generating locomotion in neonatal rodents using both slice and isolated spinal cord preparations. After removing extracellular calcium, 75% of interneurons in the area of the central pattern generator (CPG) for locomotion exhibited bursting properties and I NaP was concomitantly upregulated. Putative CPG interneurons such as commissural and Hb9 interneurons also expressed I NaP -dependent (riluzole-sensitive) bursting properties. Most bursting cells exhibited a pacemaker-like behavior (i.e., burst frequency increased with depolarizing currents). Veratridine upregulated I NaP , induced riluzole-sensitive bursting properties, and slowed down the locomotor rhythm. This study provides evidence that I NaP generates pacemaker activities in CPG interneurons and contributes to the regulation of the locomotor activity.

SUMMARY Changes in the extracellular ionic concentrations occur as a natural consequence of firing activity in large populations of neurons. The extent to which these changes alter the properties of individual neurons and the operation of neuronal networks remains unknown. Here, we show that the locomotor-like activity in the isolated neonatal rodent spinal cord reduces the extracellular calcium ([Ca2+]o) to 0.9 mM and increases the extracellular potassium ([K+]o) to 6 mM. Such changes in [Ca2+]o and [K+]o trigger pacemaker activities in interneurons considered to be part of the locomotor network. Experimental data and a modeling study show that the emergence of pacemaker properties critically involves a [Ca2+]o-dependent activation of the persistent sodium current (INaP). These results support a concept for locomotor rhythm generation in which INaP-dependent pacemaker properties in spinal interneurons are switched on and tuned by activity-dependent changes in [Ca2+]o and [K+]o.

The persistent sodium current (I(NaP)) is known to play a role in rhythm generation in different systems. Here, we investigated its contribution to locomotor pattern generation in the neonatal rat spinal cord. The locomotor network is mainly located in the ventromedial gray matter of upper lumbar segments. By means of whole cell recordings in slices, we characterized membrane and I(NaP) biophysical properties of interneurons located in this area. Compared with motoneurons, interneurons were more excitable, because of higher input resistance and membrane time constant, and displayed lower firing frequency arising from broader spikes and longer AHPs. Ramp voltage-clamp protocols revealed a riluzole- or TTX-sensitive inward current, presumably I(NaP), three times smaller in interneurons than in motoneurons. However, in contrast to motoneurons, I(NaP) mediated a prolonged plateau potential in interneurons after reducing K(+) and Ca(2+) currents. We further used in vitro isolated spinal cord preparations to investigate the contribution of I(NaP) to locomotor pattern. Application of riluzole (10 muM) to the whole spinal cord or to the upper lumbar segments disturbed fictive locomotion, whereas application of riluzole over the caudal lumbar segments had no effect. The effects of riluzole appeared to arise from a specific blockade of I(NaP) because action potential waveform, dorsal root-evoked potentials, and miniature excitatory postsynaptic currents were not affected. This study provides new functional features of ventromedial interneurons, with the first description of I(NaP)-mediated plateau potentials, and new insights into the operation of the locomotor network with a critical implication of I(NaP) in stabilizing the locomotor pattern.

The structural organization of excitatory inputs supporting spike-timing-dependent plasticity (STDP) remains unknown. We performed a spine STDP protocol using two-photon (2P) glutamate uncaging (pre) paired with postsynaptic spikes (post) in layer 5 pyramidal neurons from juvenile mice. Here we report that pre-post pairings that trigger timing-dependent LTP (t-LTP) produce shrinkage of the activated spine neck and increase in synaptic strength; and post-pre pairings that trigger timing-dependent LTD (t-LTD) decrease synaptic strength without affecting spine shape. Furthermore, the induction of t-LTP with 2P glutamate uncaging in clustered spines (<5 μm apart) enhances LTP through a NMDA receptormediated spine calcium accumulation and actin polymerization-dependent neck shrinkage, whereas t-LTD was dependent on NMDA receptors and disrupted by the activation of clustered spines but recovered when separated by >40 μm. These results indicate that synaptic cooperativity disrupts t-LTD and extends the temporal window for the induction of t-LTP, leading to STDP only encompassing LTP.

Locomotor disorders profoundly impact quality of life of patients with spinal cord injury. Understanding the neuronal networks responsible for locomotion remains a major challenge for neuroscientists and a fundamental prerequisite to overcome motor deficits. Although neuronal circuitry governing swimming activities in lower vertebrates has been studied in great details, determinants of walking activities in mammals remain elusive. The manuscript reviews some of the principles relevant to the functional organization of the mammalian locomotor network and mainly focuses on mechanisms involved in rhythmogenesis. Based on recent publications supplemented with new experimental data, the authors will specifically discuss a new working hypothesis in which pacemakers, cells characterized by inherent oscillatory properties, might be functionally integrated in the locomotor network in mammals.

The development and the ionic nature of bistable behavior in lumbar motoneurons were investigated in rats. One week after birth, almost all (ϳ80%) ankle extensor motoneurons recorded in whole-cell configuration displayed self-sustained spiking in response to a brief depolarization that emerged when the temperature was raised Ͼ30°C. The effect of L-type Ca 2ϩ channel blockers on self-sustained spiking was variable, whereas blockade of the persistent sodium current (I NaP ) abolished them. When hyperpolarized, bistable motoneurons displayed a characteristic slow afterdepolarization (sADP). The sADPs generated by repeated depolarizing pulses summed to promote a plateau potential. The sADP was tightly associated with the emergence of Ca 2ϩ spikes. Substitution of extracellular Na ϩ or chelation of intracellular Ca 2ϩ abolished both sADP and the plateau potential without affecting Ca 2ϩ spikes. These data suggest a key role of a Ca

Maturation of inhibitory postsynaptic transmission onto motoneurons in the rat occurs during the perinatal period, a time window during which pathways arising from the brainstem reach the lumbar enlargement of the spinal cord. There is a developmental switch in miniature IPSCs (mIPSCs) from predominantly long-duration GABAergic to short-duration glycinergic events. We investigated the effects of a complete neonatal [postnatal day 0 (P0)] spinal cord transection (SCT) on the expression of Glycine and GABA A receptor subunits (GlyR and GABA A R subunits) in lumbar motoneurons. In control rats, the density of GlyR increased from P1 to P7 to reach a plateau, whereas that of GABA A R subunits dropped during the same period. In P7 animals with neonatal SCT (SCT-P7), the GlyR densities were unchanged compared with controls of the same age, while the developmental downregulation of GABA A R was prevented. Whole-cell patch-clamp recordings of mIPSCs performed in lumbar motoneurons at P7 revealed that the decay time constant of miniature IPSCs and the proportion of GABAergic events significantly increased after SCT. After daily injections of the 5-HT 2 R agonist DOI, GABA A R immunolabeling on SCT-P7 motoneurons dropped down to values reported in control-P7, while GlyR labeling remained stable. A SCT made at P5 significantly upregulated the expression of GABA A R 1 week later with little, if any, influence on GlyR. We conclude that the plasticity of GlyR is independent of supraspinal influences whereas that of GABA A R is markedly influenced by descending pathways, in particular serotoninergic projections.

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