Phosphorus NMR spectroscopy of cat biceps and soleus muscles

It is clear that the combination of 31P NMR studies of muscle and of other organs with biochemical and physiological measurements is an extremely powerful method for studying cellular energetics. We have presented examples of our on-going experiments. Well defined and stable in vitro preparations of mammalian fast-twitch (cat biceps) and slow-twitch (cat soleus) muscles are developed. Erythrocyte-containing perfusate is needed for adequate oxygenation during stimulation. It is not clear why fluorocarbon suspensions are inadequate. Inorganic phosphate levels in the fast-twitch biceps as determined from NRM spectra are close to lmM, that is very much lower than the value (6 mM) derived from chemical measurements of muscle extracts. A similar discrepancy in the slow-twitch soleus cannot yet be established; if it exists, the discrepancy is small. From the metabolite contents, the free energy available from ATP splitting is similar in both muscle types: -63 kJ/mole in biceps and -57 kJ/mole in soleus. Kinetic NMR measurements indicate the creatine phosphokinase reaction is at equilibrium, and in biceps the unidirectional fluxes are 5 mumole per gram per sec. Transient alkalinization of intracellular pH during stimulation periods and acidification during recovery periods are clearly seen. In the biceps the alkalinization is much smaller than in soleus, and suggests a greater buffer capacity in fast-twitch muscles. Both muscles recovery from prolonged ischemic periods upon reperfusion, although the recovery rate is very much slower than that from 10-20 min periods of stimulation in which similar decreases of PCr and increases of Pi occur. The pattern of recovery is however quite similar. Perfused muscles may be a useful model for studying ischemia, anoxia and hypoxia and are useful preparations for a wide variety of metabolic and energetic studies of fast-twitch and slow-twitch mammalian skeletal muscles.