Adaptation of a perfused rat hemidiaphragm preparation to the study of intermediary metabolism

Abstract

Glucose catabolism of a vascular perfused rat hemidiaphragm was determined at rest and during stimulation of the phrenic nerve with trains of either 5 (T5) or 15 (T15) pulses (20 msec intervals) per second. Tissues were perfused and bathed in HEPES-buffered medium containing 11 mM d-[U- 14C, 5- 3H]glucose, equilibrated with 100% O 2. Resting glucose catabolism via the Emden-Meyerhof pathway was indicated by a 3H 2O production rate per hemidiaphragm of 1.42 ± 0.07 μmol/h, of which 47% was recovered as [ 14C]lactate with the remainder asssumed to be metabolised by mitochondria. During the first 30 min of T5 and T15 stimulation, peak isometric tension declined from an initial value of 105 ± 8 g by 54% and 79%, respectively. The resulting peak tension of 48 and 22 g remained constant for the next 60 min. These tensions were associated with linear rates of 3H 2O production of 2.93 ± 0.41 and 2.84 ± 0.25 μmol/h. Stimulation by T5 and T15 increased mitochondrial metabolism of glucose by 64% adn 95%, respectively, with no significant alterations in lactate formation from either exogenous or endogenous sources. The results suggest that the initial decline in tension is due to fatigue of the fast anaerobic myofibers; whereas, the sustained force beyond 30 min is attributable to the mitochondrial-rich slow myofibers.