Growth cone "collapse" in vivo: are inhibitory interactions mediated by gap junctions?

Abstract

In the leech embryo, oppositely directed axons of homologous anterior pagoda (AP) neurons overlap with each other extensively within the nerves that link adjacent ganglia, and inhibit each other's further growth (Gao and Macagno, 1987b). During this 5-8 d period of inhibition, the axons begin to grow thin, and eventually they retract completely. However, deletion of one overlapping AP cell results in the renewed growth of the remaining AP cell's axon, which then innervates territory vacated by the killed cell. Thus, each neuron can detect the presence of the other, and adjust its branching pattern accordingly. To begin to explore how these neurons detect and inhibit each other, we tested for direct communication between them. Dye fills with fluorescent chromophores suggested direct contact between their axons at the light level, and this was confirmed by serial-section electron microscopic analysis. Morphological features resembling aspects of gap junctions were observed where the projections were closely apposed, and subsequent electrophysiological recordings demonstrated electrical coupling between the mutually inhibited axons. Confirmation that these projections communicate via gap junctions was obtained using intracellular injection of 5-HT as a tracer, followed by anti-5-HT immunohistochemistry. The tracer passed selectively between AP neurons. We propose that the gap junctions formed between the transient projections of the developing AP neurons may mediate the exchange of the signals that permit homologs to recognize each other and to inhibit the further forward progress of these projections.