An outflow of acetylcholine from normal and regenerating ventral roots of the cat.

1. An outflow of acetylcholine (ACh) has been measured from cat ventral roots in situ and in vitro, using standard bio-assay techniques. The ACh was collected in chambers, filled with physiological solution containing anticholinesterase, which surrounded the root.2. The outflow fell rapidly at first and then progressively more slowly, taking several hours to fall to a level near the assay threshold. The outflow of ACh resulted from cutting a root or from such conventional manipulations as stretching the root or teasing apart root filaments. The outflow could occur either from the end of a root (following cutting) or from along the length of the root (following stretching or teasing). The outflow was only detected in the presence of an anticholinesterase.3. ACh was resynthesized by roots at a rate which was usually rapid enough to maintain their ACh content at its original level in spite of a considerable ACh outflow. In roots subjected to a minimum of interference their ACh content built up to about twice the initial content after 5-7 hr soaking in eserinized Krebs solution although the total ACh outflow was equivalent to the original ACh content.Roots which were cut or stretched several times within a few hours put out a large amount of ACh and showed a fall in their ACh content below the initial level.4. The time course of sealing of the damaged ends of axons following crushing was assessed by measuring the injury potential for 1-2 hr after crushing at room temperature; in normal roots the half-time for decline of the injury potential was 91 +/- 10 min. This was slower than the rate of decline of the ACh outflow.5. An explanation for the observed outflow of ACh is discussed in terms of diffusion of ACh and sealing of axon ends. It is concluded that much of the ACh in normal ventral root axons is present in a readily diffusible form.6. Similar experiments on roots regenerating 9-10 days after crushing showed that the ACh outflow from regenerating roots was much smaller than from normal axons, although their ACh content was similar. This could have been due to more rapid sealing of the cut regenerating axons since the half-time for the decline of the injury potential following crushing of the regenerating axons was less than half that in control roots. The alternative possibility is considered that a high proportion of the ACh might be much less diffusible than in normal axons.7. The subcellular distribution of ACh in normal and regenerating peripheral axons is discussed.