Neurite growth patterns leading to functional synapses in an identified embryonic neuron

Abstract

We explored the relationship between neurite outgrowth and the onset of synaptic activity in the central neuropil of the leech embryo in vivo. To follow changes in early morphology and the onset of synaptic activity in the same identified neuron, we obtained whole-cell patch-clamp recordings and fluorescent dye fills from dorsal pressure-sensitive (P) cells, the first neurons that could be reliably identified in the early embryo. We followed the development of the P cell from the first extension of neurites to the elaboration of an adult-like arbor. After the growth of primary neurites, we observed a profuse outgrowth of transient neurites within the neuropil. Retraction of the transient neurites left the primary branches studded with spurs. After a dormant period, stable secondary branches grew apparently from the spurs and became tipped with terminals. At this time, neurites of the Retzius (R) cell, a known presynaptic partner in the adult, were observed to apparently contact the terminals. Although voltage-dependent currents were seen in the P cell at the earliest stage, spontaneous synaptic activity was only observed when terminals had formed. Spontaneous release was observed before evoked release could be detected from the R cell. Our results suggest that transient neurites are formed during an exploratory phase of development, whereas the more precisely timed outgrowth of stable neurites from the spurs signals functional differentiation during synaptogenesis. Because spurs have also been observed in neurons of the mammalian brain, they may constitute a primordial synaptic organizer.