Acetylcholine storage, release and leakage at the neuromuscular junction of mature adult and aged rats.

Abstract

Acetylcholine (ACh) content, release during stimulation and spontaneous leakage were studied at the diaphragm neuromuscular junction in rats aged 10 and 28 months. In addition, quantal release statistics were determined. The results were related to the number of nerve terminals per end-plate and to the age of the animals. Nerve terminal ACh content was lower in the 28-month-old rats; in nonstimulated tissue the average (+/- S.E.) ACh levels were 29.3 (+/- 4.1) and 17.2 (+/- 2.4) fmol per end-plate in the 10- and 28-month-old rats, respectively. During nerve stimulation, greater amounts of ACh were released by the older rats. After 1000 impulses, the average (+/- S.E.) ACh release was 3.2 (+/- 0.2) and 3.9 (+/- 0.2) fmol per end-plate in the 10- and the 28-month-old animals, respectively; however, the amounts released per nerve terminal were not significantly different. The fraction of the resting ACh content released per action potential was greater in the aged rats; this may be the cause of enhanced synaptic depression in the older animals. Leakage of ACh from cytoplasmic sources was assessed by measuring the hyperpolarization following application of d-tubocurarine (dTC). The hyperpolarizing responses were of similar magnitude in both age groups; however, the older rats were less sensitive to ACh, indicating that cytoplasmic leakage was greater in these animals. Miniature end-plate potentials (m.e.p.p.s) occurred twice as frequently in the aged rats; this increase could be attributed to an age-related increase in the number of nerve terminals per end-plate. After correcting for variations in membrane input resistance and resting potential, m.e.p.p. amplitudes were compared and found to be significantly smaller in the aged rats. Quantal release during nerve stimulation was measured and found to obey binomial statistics. Estimated values of m and n were significantly larger in the older rats; there were no age-related differences in p. The increases in m and n could be attributed to the larger number of nerve terminals per end-plate in the aged animals. Acetylcholinesterase (AChE) activity was assayed. In innervated tissue there were no significant differences with age; however, in non-innervated tissue AChE activity was significantly higher in the 28-month-old rats.