Functional and non-functional contacts between ciliary neurones and muscle grown in vitro

1. In cultures of chick embryo skeletal muscle and ciliary ganglia, muscle fibres near a ganglion were contacted by many individual nerve processes. Experiments were performed to determine if these muscle fibres were multiply innervated, and if any of the nerve-muscle contacts were non-synaptic. 2. Synaptic potentials evoked by electrical stimulation of a ganglion were graded with stimulus strength. When two ganglia were plated near each other, synaptic potentials could be evoked in some muscle fibres by stimulation of either ganglion. These observations suggest that muscle fibres were multiply innervated. 3. Spontaneous synaptic potentials recorded from single muscle fibres with two widely spaced micropipettes varied in a manner which suggested that the synapses were distributed at different points on the surface of the muscle fibres. 4. Stimulation of some nerve processes failed to evoke synaptic potentials in muscle fibres contacted by those processes. Such nerve-muscle contacts were not strongly adhesive, and the nerves were peeled easily from the surface of the muscle with a micropipette. On the other hand, nerve processes which formed synaptic contacts with muscle fibres seemed to be tightly adherent to the muscle. 5. Electron microscopic observations of nerve-muscle contacts revealed that the vast majority of such contacts lacked morphological specializations characteristic of mature neuromuscular synapses.