No abstract

We examined the effects of spontaneous or evoked episodes of rhythmic activity on synaptic transmission in several spinal pathways of embryonic day 9-12 chick embryos. We compared the amplitude of synaptic potentials evoked by stimulation of the ventrolateral funiculus (VLF), the dorsal or ventral roots, before and after episodes of activity. With the exception of the short-latency responses evoked by dorsal root stimulation, the potentials were briefly potentiated and then reduced for several minutes after an episode of rhythmic activity. Their amplitude progressively recovered in the interval between successive episodes. The lack of post-episode depression in the shortlatency component of the dorsal root evoked responses is probably attributable to the absence of firing in cut muscle afferents during an episode of activity.The post-episode depression of VLF-evoked potentials was mimicked by prolonged stimulation of the VLF, subthreshold for an episode of activity. By contrast, antidromically induced motoneuron firing and the accompanying calcium entry did not depress VLF-evoked potentials recorded from the stimulated ventral root. In addition, post-episode depression of VLFevoked synaptic currents was observed in voltage-clamped spinal neurons. Collectively, these findings suggest that somatic postsynaptic activity and calcium entry are not required for the depression. We propose instead that the mechanism may involve a form of long-lasting activity-induced synaptic depression, possibly a combination of transmitter depletion and ligand-induced changes in the postsynaptic current accompanying transmitter release. This activity-dependent depression appears to be an important mechanism underlying the occurrence of spontaneous activity in developing spinal networks.

Video-rate imaging of spinal neurons loaded with calcium-sensitive dyes was used to investigate the calcium dynamics and cellular organization of spontaneously active rhythm-generating networks in the spinal cord of E9-E12 chick embryos. Spinal neurons were loaded with bath-applied fura-2am. Motoneurons were also loaded by retrograde labeling with dextran-conjugated, calcium-sensitive dyes. Dye-filled motoneurons exhibited large fluorescent changes during antidromic stimulation of motor nerves, and an increase in the 340/380 fura fluorescence ratio that is indicative of increased intracellular free calcium. Rhythmic fluorescence changes in phase with motoneuron electrical activity were recorded from motoneurons and interneurons during episodes of evoked or spontaneous rhythmic motor activity. Fluorescent responses were present in the cytosol and in the perinuclear region, during antidromic stimulation and network-driven rhythmic activity. Optically active cells were mapped during rhythmic activity, revealing a widespread distribution in the transverse and horizontal planes of the spinal cord with the highest proportion in the ventrolateral part of the cord. Fluorescent signals were synchronized in different regions of the cord and were similar in time course in the lateral motor column and in the intermediate region. In the dorsal region the rhythm was less pronounced and the signal decayed after a large initial transient. Video-rate fluorescent measurements from individual cells confirmed that fluorescent signals were synchronized in interneurons and in motoneurons although the time course of the signal could vary between cells. Some of the interneurons exhibited tonic elevations of fluorescence for the duration of the episode whereas others were rhythmically active in phase with motoneurons. At the onset of each cycle of rhythmic activity the earliest fluorescent change occurred ventrolaterally, in and around the lateral motor column, from which it spread to the rest of the cord. The results suggest that neurons in the ventrolateral part of the spinal cord are important for rhythmogenesis and that axons traveling in the ventrolateral white matter may be involved in the rhythmic excitation of motoneurons and interneurons. The widespread synchrony of the rhythmic calcium transients may reflect the existence of extensive excitatory interconnections between spinal neurons. The network-driven calcium elevations in the cytosol and the perinuclear region may be important in mediating activity-dependent effects on the development of spinal neurons and networks.

We have examined the regionalization and coordination of rhythm-generating networks in the isolated spinal cord of the chick embryo between embryonic days 9 and 13, by recording the pattern of rhythmic activity recorded from muscle nerves and ventral roots following a variety of lesions. We found that the capacity for rhythmic activity is distributed along the rostrocaudal axis of the cord but can be expressed in a single, isolated segment. Specializations within the lumbosacral cord were investigated by isolating particular regions and recording their motor output. The rostral part of the lumbosacral cord generates more cycles than the caudal part, and this difference becomes more pronounced with development. In the unlesioned cord, motoneuron activity is synchronized along the rostrocaudal axis. Lesion experiments revealed that the synchronization of motoneuron activity and the synaptic drive to caudal motoneurons is mediated in part by propriospinal pathways traveling in the ventrolateral white matter tracts and by synaptic interactions within the gray matter. The dorsal fiber tracts may also be involved but their effects appear to be weak. Lesions in dorsal-ventral and mediolateral planes were used to localize regions critical for rhythmogenesis and for the alternation of flexor and extensor motoneurons. Rhythmic activity with alternation persisted in spinal cords in which the dorsal and medial half had been removed. Severe medial or dorsal lesions, resulting in a thin strip of lateral or ventral gray matter, altered the phasing of motoneuron activity from alternating to synchronous without effects on cycle timing. These results suggest that the critical neural components for alternation are located close to and dorsomedial to the lateral motor column, and that the capacity for rhythmogenesis is distributed widely throughout the ventral gray matter and is not localized to specific nuclei.