The formation of chemical synapses between cell-cultured neuronal somata

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Abstract
The study of the development and plasticity of chemical synaptic connections is frequently restricted by the lack of access to the synaptic terminals. This can, in part, be overcome by plating neurons into cell culture where all regions of a neuron are made experimentally accessible. However, the small size of synaptic terminals still makes direct experimental manipulation difficult. In this study we have found in the absence of neurite extension, directly contacting cell somata (diameter 50–100 mumol) will form chemical synapses. Identified neurons B5 and B19 of Helisoma were plated into culture under conditions that promote adhesion between cell pairs. Under these conditions, neurite outgrowth was absent, but action potentials in B5 evoked inhibitory postsynaptic potentials in B19 that were reversed in sign by the injection of chloride ions and were blocked by tubocurare (10(-5) M), reduced extracellular Ca2+, and Cd2+ ions. Such synapses exhibited classical properties of chemical synapses, including the spontaneous release of neurotransmitter. Since somatic synapses represent an appropriate model of synaptic transmission, this system was utilized to study the role of mutual neuronal contact in the development of transmitter release capabilities. Future pre- and postsynaptic somata were cultured separately for 3 d, the period required for the development of synaptic transmission under conditions of maintained contact. Then, neurons were made to contact and intracellular recordings taken within 0 to 4 hours. Postsynaptic potentials were detected as early as 10 sec following contact. Thus, qualitatively the development of transmitter release capabilities does not require maintained contact.