Neural and non-neural acetylcholine in the rat diaphragm

The compartmentation of acetylcholine (ACh) and of choline acetyltransferase in the rat diaphragm was analysed by measuring their contents in muscle segments containing endplates (e.p.) and endplate-free segments (non-e.p.) at different times following section of the phrenic nerve. In addition ACh release was determined before and after denervation. Freshly dissected hemidiaphragms contained about 125 pmol of ACh; more than 90% of this was localized in the e.p. portion. Between 10 and 18 h after denervation the ACh content of the e.p. portion decreased by 80% and its ACh concentration became approximately equal to that in the non-e.p. region, whose ACh content did not change. Spontaneous release of ACh was reduced by denervation and ACh release evoked by 50 mM KC1 was practically abolished. Choline acetyltransferase activity in freshly dissected preparations was about 30 nmol of ACh per gram per hour, Km 0.5 mM. About 65% of the enzyme disappeared in the first 24 h and the remaining 35% between 24 and 50 h after denervation. A different enzyme capable of ACh synthesis was found in the muscle fibres; its activity did not decrease after denervation. It is concluded that about 70% of the ACh in the diaphragm is contained in the motor nerve terminals, about 10% in the intramuscular nerve fibres and the remainder in the muscle fibres, and that about 65% of choline acetyltransferase is in the motor terminals and 35% in the nerve fibres.