Effects of acute and chronic denervation on release of acetylcholinesterase and its molecular forms in rat diaphragms

Hemidiaphragms were removed from rats at various times after intrathoracic transection of the left phrenic nerve and were incubated in organ baths containing 1.5 ml of oxygenated, buffered physiologic saline solution, with added glucose and bovine serum albumin. After incubation, the acetylcholinesterase (AChE: EC 3.1.1.7) activities of the bath fluid and of the muscle were determined. Innervated left hemidiaphragms were found to release 107 units of AChE over a 3-h period, corresponding to 1.9% of their total AChE activity. Denervation led to a rapid loss of AChE from the muscle coincident with a transient increase in the outpouring of enzyme activity into the bath fluid. Thus, 1 day after nerve transection the left hemidiaphragm contained only 68% of the control amounts of AChE activity, but released 140% as much as control. After 3 or 4 days of denervation, the AChE activity of the diaphragm stabilized at 35% of the control value. Release also fell below control by this time, but not as far. One week after denervation the release, 69 units per 3 hr, correspond to 3.3% of the reduced content of AChE activity in the muscle, indicating that denervation caused an increase in the proportion of AChE released. Sucrose density gradient ultracentrifugation showed that 10S AChE accounted for more than 80% of the released enzyme activity at all times. The results did not rule out the possibility, however, that the released enzyme originally stemmed from 4S or 16S AChE in the diaphragms.