Myofibrillar fatigue versus failure of activation during repetitive stimulation of frog muscle fibres.

1. Single fibres isolated from the anterior tibialis muscle of Rana temporaria (temperature, 2-5 degrees C; sarcomere length, 2.10 microns) were fatigued using two separate protocols that led to different degrees of depression of tetanic force. Under control conditions the fibre was stimulated to produce a 1 s fused isometric tetanus at 300 s intervals. A moderate degree of fatigue (tetanic force reduced to 70-80% of the control value) was produced by decreasing the intervals between tetani to 15 s ('fatiguing protocol 1'). A more pronounced depression of tetanic force (to 40-50% of the control value) was produced by evoking a single twitch at 1-2 s intervals ('fatiguing protocol 2'). 2. Fatiguing protocol 1 reduced the contracture response to submaximal and supramaximal concentrations of caffeine (3-15 mM) in proportion to the decrease in tetanic force. These results support the view that fatiguing stimulation according to protocol 1 leads to a true 'myofibrillar fatigue' with no failure of activation of the muscle fibre. 3. Fatiguing protocol 2 reduced the amplitudes of isometric twitch and tetanus to below 10 and 50% of the control values, respectively. By contrast, the maximal contracture response to caffeine (15 mM) was depressed by merely 2-3% of its prefatigue value. 4. Force and instantaneous fibre stiffness were recorded simultaneously during twitch and tetanus as fatigue was induced by protocol 2. During the initial part of fatigue (tetanic force reduced by 25% of control) stiffness was reduced by merely 9% in accordance with previous measurements during fatigue induced by protocol 1. However, with further depression of twitch and tetanus by protocol 2 there was a marked reduction of fibre stiffness. These results, together with the findings reported under point 3, strongly suggest that at an advanced state of fatigue induced by protocol 2 the decrease in active force is largely due to failure of activation of the contractile system. 5. Muscle fibres were quickly frozen for electron microscopical examination after shortening below slack length (to approximately 1.6 microns sarcomere spacing) during tetanic stimulation. In non-fatigued fibres, and in fibres fatigued according to protocol 1, the myofibrils exhibited a straight appearance throughout the preparation suggesting that the entire volume of the fibre was properly activated. In fibres fatigued by protocol 2, on the other hand, only the most peripheral layers of myofibrils remained straight after shortening, whereas the centre of the fibre showed marked waviness indicating failure of the inward spread of activation in this case.