Different extrinsic trophic factors regulate neurite outgrowth and synapseformation between identified Lymnaea neurons

The requirement for trophic factors in neurite outgrowth is well establishe d, though their role in synapse formation is yet to be determined. Moreover , the issue of whether the trophic factors mediating neurite outgrowth are also responsible for synapse specification has not yet been resolved. To te st whether trophic factors mediating neurite outgrowth and synapse formatio n between identified neurons are conserved in two molluscan species and whe ther these developmental processes are differentially regulated by differen t trophic factors, we used soma-soma and neurite-neurite synapses between i dentified Lymnaea neurons. We demonstrate here that the trophic factors pre sent in Aplysia hemolymph, although sufficient to induce neurite outgrowth from Lymnaea neurons, do not promote specific synapse formation between exc itatory partners. Specifically, the identified presynaptic neuron visceral dorsal 4 (VD4) and postsynaptic neuron left pedal dorsal 1 (LPeD1) were eit her paired in a soma-soma configuration or plated individually to allow neu ritic contacts. Cells were cultured in either Lymnaea brain-conditioned med ium (CM) or on poly-L-lysine dishes that were pretreated with Aplysia hemol ymph (ApHM), but contained only Lymnaea defined medium (DM; does not promot e neurite outgrowth). In ApHM-coated dishes containing DM, Lymnaea neurons exhibited extensive neurite outgrowth, but appropriate excitatory synapses failed to develop between the cells. Instead, inappropriate reciprocal inhi bitory synapses formed between VD4 and LPeD1, Similar inappropriate inhibit ory synapses were observed in Aplysia hemolymph-pretreated dishes that cont ained dialyzed Aplysia hemolymph, These inhibitory synapses were novel and inappropriate, because they do not exist in vivo. A receptor tyrosine kinas e inhibitor (Lavendustin A) blocked neurite outgrowth induced by both Lymna ea Chi and ApHM. However, it did not affect inappropriate inhibitory synaps e formation between the neurons. These data demonstrate that neurite outgro wth but not inappropriate inhibitory synapse formation involves receptor ty rosine kinases. Together, our data provide direct evidence that trophic fac tors required for neurite outgrowth are conserved among two different mollu scan species, and that neurite extension and synapse specification between excitatory partners are likely mediated by different trophic factors. (C) 2 000 John Wiley & Sons, Inc.