The site and mechanism of action of fatigue was investigated in the isolated rat phrenic nerve diaphragm preparation, with indirect and direct stimulation at 20 Hz and recording of tension and EMG. An equal decay of the subtetanic tension during indirect and direct stimulation, and a parallel decay of tension and EMG, suggested a mechanism of fatigue localized to structures that were 'seen' by the EMG electrodes. A comparison of the responses to sub- and supra-maximum direct stimulation did not show increased fatigue at sub-maximum stimulation. Therefore, the fatigue was probably not caused by an increased threshold of the excitability of the sarcolemma. However, prolongation of the stimulus pulse during direct stimulation from 0.5 ms to 5 ms in the fatigued preparation caused a two-phasic recovery of tension. The initial phase, but not the slow phase, was inhibited by tetrodotoxin (TTX). Thus a recovery of sarcolemma action potentials could explain the initial phase. The slow phase was probably caused by a mechanism localized at more distal potential-dependent sites, probably in the T tubules.
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