Location of U.V.-absorbing substance in isolated skeletal muscle fibres. The effect of stimulation.

1. In an attempt to study the intracellular location of ATP in skeletal muscle the distribution of substances which absorb light at 260 nm wave-length has been studied in isolated muscle fibres with the aid of a modified U.V.-microscope.2. U.V.-absorption in resting frog fibres was found to be higher in the I band than in the A band which confirms earlier findings. In stretched fibres (sarcomere length 2.9-3.6 mum) an absorbing substance could be seen to be concentrated in a pair of narrow lines, centred at the Z-line. The separation of the lines increased with increasing sarcomere length.3. Snake fibres, with sparse triads located at the A-I junction, displayed an absorption pattern very similar to that of frog fibres. It is concluded that it is unlikely that the absorbing substance is associated with the sarcotubular system.4. The absorption pattern of frog fibres remained unchanged during a tetanus. No clear changes could be detected after a period of stimulation, neither after single twitches nor after repeated tetani.5. In further attempts to cause exhaustion, metabolically poisoned fibres were stimulated repetitively until they went into rigor. The absorption pattern was essentially unchanged also when rigor tension started to develop.6. The characteristic absorption pattern was observed also in glycerol-extracted fibres. It was confirmed by spectrophotometry that glycerol-extraction led to the disappearance of a large amount of a substance with the spectral characteristics of ATP.7. The higher U.V.-absorption in the I band does not prove that the major part of the ATP in the fibre is concentrated here; the absorption could either be due to a minor fraction of the ATP or to RNA.