Isolation of Axonal Varicosities (Autonomic Synaptosomes) From the Enteric Nervous System

A subcellular fraction enriched with varicosities of autonomic axons was obtained from homogenates of strips of guinea‐pig longitudinal muscle and adherent myenteric plexus using differential and sucrose‐or Ficoll‐density gradient ultracentrifugation. This fraction contained the marker, [G‐ 3 H]5‐hydroxytryptamine (5‐HT), taken up by serotonergic axon terminals present in the preparation during a period of incubation prior to homogenization. Electron microscopic examination showed that the isolated varicosities resembled CNS synaptosomes in containing vesicles and mitochondria but, as is characteristic of autonomic postganglionic terminals, they lacked synaptic membrane specializations. Quantitative electron microscopic radioautography revealed that [G‐ 3 H]5‐HT was confined to isolated varicosities. Isolated varicosities were capable of taking up and sequestering 5‐HT from the surrounding medium; this uptake was temperature‐sensitive and blocked by fluoxetine. The subcellular distribution of [G‐ 3 H] l ‐tryptophan was also studied by subfractionation of strips in an attempt to determine which structures were responsible for the high‐affinity uptake of that amino acid. Although most of the [G‐ 3 H] l ‐tryptophan was found in the high‐speed supernatant, particulate [G‐ 3 H] l ‐tryptophan was most concentrated in the subcellular fraction containing isolated axonal varicosities. These studies indicate that a fraction containing functional serotonergic varicosities can be isolated from the gut. These varicosities are probably one of the elements responsible for the high‐affinity uptake of l ‐tryptophan by the myenteric layer of the gut.