FUNCTIONAL CHANGES IN DIAPHRAGMATIC FATIGUE

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The purpose of the experiment proposed in this application is to provide a clearer understanding of cellular mechanisms responsible for the decreased force of contraction developed by the diaphragm exposed to acidosis. Recently, some clinical literature has suggested that patients with chronic obstructive pulmonary disease who are hypercapnic are unable to develop the same inspiratory pressures as equally obstructed eucapnic patients. Furthermore, a recent study in normal human subjects showed that artificially induced hypercapnia not only decreased the transdiaphragmatic pressure generated for a given electrical input, but also disposed the subjects to diaphragmatic fatigue more readily. We have demonstrated these same phenomena in the in vivo dog diaphragm and in the in vitro rat diaphragm. The overall hypothesis of this proposal is that acute and long term extracellular acidosis provokes mechanical dysfunction in the in vitro rat diaphragm by one or more of the following mechanisms: inhibiting the ability of cellular cation (Na-H) and anion (HCO3-C1) pumps to regulate intracellular pH; depleting high energy phosphates by inhibiting glycolytic or aerobic enzymes; modulating sarcolemmal or organellar calcium transport or altering intracellular calcium binding sites; provoking a depolarization of the resting membrane potential; changing the pattern of acetylcholine release at the neuromuscular junction of the diaphragm. Some of these mechanisms may be responsible acutely while others become so after a period of time. The proposal describes four sets of experiments which will test these mechanisms. Experiments using 31 phosphorus nuclear magnetic resonance will test hypotheses involving the effectos of acidosis on diaphragmatic intracellular pH and content of high energy phosphates. Experiments employing methods based on the use of 14C and 3H labelled glucoses will investigate the effects of acidosis on glycolytic and aerobic metabolism. Complete kinetic analysis of 45 calcium efflux data will permit quantitation of cellular calcium transport processes as well as changes in these processes caused by acidosis. Finally standard electrophysiological techniques will be used for the recording of resting membrane potentials and miniature end plate potentials.