Mechanisms that initiate spontaneous network activity in the developing chick spinal cord

Many developing networks exhibit a transient period of spontaneous activity that is believed to be important developmentally. Here we investigate the initiation of spontaneous episodes of rhythmic activity in the embryonic ch ick spinal cord. These episodes recur regularly and are separated by quiesc ent intervals of many minutes. We examined the role of motoneurons and thei r intraspinal synaptic targets (R-interneurons) in the initiation of these episodes. During the latter part of the inter-episode interval, we recorded spontaneous, transient ventral root depolarizations that were accompanied by small, spatially diffuse fluorescent signals from interneurons retrograd ely labeled with a calcium-sensitive dye. A transient often could be resolv ed at episode onset and was accompanied by an intense pre-episode (similar to 500 ms) motoneuronal discharge (particularly in adductor and sartorius) but not by interneuronal discharge monitored from the ventrolateral funicul us (VLF). An important role for this pre-episode motoneuron discharge was s uggested by the finding that electrical stimulation of motor axons, suffici ent to activate R-interneurons, could trigger episodes prematurely. This ef fect was mediated through activation of R-interneurons because it was preve nted by pharmacological blockade of either the cholinergic motoneuronal inp uts to R-interneurons or the GABAergic outputs from R-interneurons to other interneurons. Whole-cell recording from R-interneurons and imaging of calc ium dye-labeled interneurons established that R-interneuron cell bodies wer e located dorsomedial to the lateral motor column (R-interneuron region). T his region became active before other labeled interneurons when an episode was triggered by motor axon stimulation. At the beginning of a spontaneous episode, whole-cell recordings revealed that R-interneurons fired a high-fr equency burst of spikes and optical recordings demonstrated that the R-inte rneuron region became active before other labeled interneurons. In the pres ence of cholinergic blockade, however, episode initiation slowed and the in ter-episode interval lengthened. In addition, optical activity recorded fro m the R-interneuron region no longer led that of other labeled interneurons . Instead the initial activity occurred bilaterally in the region medial to the motor column and encompassing the central canal. These findings are co nsistent with the hypothesis that transient depolarizations and firing in m otoneurons, originating from random fluctuations of interneuronal synaptic activity, activate R-interneurons, which then trigger the recruitment of th e rest of the spinal interneuronal network. This unusual function for R-int erneurons is likely to arise because the output of these interneurons is fu nctionally excitatory during development.