Serotonergic Retzius (R) neurons of the leech Hirudo medicinalis in culture reform inhibitory synapses with pressure sensitive (P) neurons while selectively reducing an extrasynaptic, depolarizing response to serotonin (5-HT) in the P neuron. We have examined if the selection of 5-HT responses is restricted to sites of contact between processes and growth cones of these cells. As measured by intracellular recording at the soma, focal application of 5-HT depolarized uncontacted P cell bodies, neurites and growth cones but not processes contacted by R cells. In patch clamp recordings of the depolarizing channels, application of 5-HT modulated channel activity in uncontacted but not in contacted growth cones. The selection of transmitter responses during synaptogenesis is therefore localized to discrete sites of contact specifically between synaptic partners. / Prior experiments have shown that tyrosine kinases play a crucial part in the selection of responses to 5-HT that occurs in the P cell (Catarsi and Drapeau, 1993). To further examine the mechanism responsible for this change in transmitter responses, we have utilized a monoclonal antibody against phosphotyrosine to determine if tyrosine phosphorylation could be detected in P and R cell pairs placed in contact. Our results revealed bright, punctate cytoplasmic staining in P cells paired with R cells. / Embryonic leeches were used to examine how R to P synaptogenesis proceeds in vivo. By filling the R and P neurons with different fluorescent dyes (Lucifer Yellow and Rhodamine-Dextran), confocal microscopy established that putative contact between neuropilar processes were made as early as 11 days of development. Spontaneous, chloride-dependent synaptic potentials in embryonic P cells similar to those seen in adult P cells were observed as early as day 10 of development.

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