Chronic or prolonged low-intensity loading of the inspiratory muscles has recently been shown to produce diaphragm injury. The present study was designed to examine whether an acute episode of inspiratory resistive loading (IRL) could produce secondary diaphragm inflammation and injury. On Day 1, three groups of anesthetized and intubated New Zealand White rabbits were subjected to moderate IRL (Pao of approximately 30 cm H2O), high IRL (Pao of approximately 45 cm H2O), or no load for 1.5 h. On Day 3, costal and crural diaphragms, parasternals, and gastrocnemius muscles were taken to assess injury by point counting. Normal muscle, abnormal and inflamed muscle, and connective tissue on hematoxylin and eosin-stained cross-sections were expressed as percentage of the total points for that cross-section. For the costal diaphragm, both the abnormal muscle (7.3 +/- 0.6% versus 1.1 +/- 0.2%; p < 0.001) and connective tissue (8.0 +/- 0.6% versus 5.7 +/- 0.2%; p < 0.01) in the high IRL group were higher than control, whereas in the moderate IRL group they were not significantly different from control. Total calpain-like activity was increased in the moderate IRL group but not in the high IRL group. Injury was observed in the parasternal muscles but to a lesser extent. No injury was observed in the gastrocnemius muscle. We conclude that secondary diaphragm injury occurs after acute IRL but only when the IRL exceeds the fatigue threshold.

The present study was designed to examine the effect of delayed diaphragm injury produced by inspiratory resistive loading (IRL) on diaphragm force production. On Day 1, three groups of anesthetized and intubated NZW rabbits (n = 7 in each group) were subjected to moderate IRL (Pao approximately 30 cm H2O), high IRL (Pao approximately 45 cm H2O), or no load for 1.5 h. On Day 3, the baseline twitch transdiaphragmatic pressure (Pdi) and Pdi at 10 to 80 Hz were measured during bilateral phrenic stimulation and these measurements were repeated after another IRL (high level) in all three groups. Diaphragm injury was assessed by the point-counting technique. Marked diaphragm injury was observed in the high-IRL group (p < 0.01), but no significant diaphragm injury was observed in the moderate-IRL or control groups. The baseline twitch Pdi was maintained in both IRL groups, whereas the baseline Pdi-frequency values in the high-IRL group were significantly reduced at most frequencies (p < 0.05). The decreases in twitch and Pdi at different frequencies were more pronounced after the IRL on Day 3 in the high-IRL group compared with controls. Moderate IRL did not decrease diaphragm force either before or after the high IRL on Day 3. We conclude that the diaphragm injury induced by high IRL has a significant impact on diaphragm force production and the attendant force loss produced by IRL is dependent on the intensity of inspiratory loading.

We developed a new technique of diaphragmatic stimulation by apposing plate electrodes directly against the diaphragm (DPS) in adult Golden Syrian hamsters. The electrophysiological and the mechanical responses to DPS were compared with those with phrenic nerve stimulation. In four animals, evaluation of the electromyogram before and after curare demonstrated that plate electrode stimulation occurred via the phrenic nerve filaments. In four animals, similar transdiaphragmatic pressure was produced at maximal current with DPS and phrenic nerve stimulation. Using DPS increasing current beyond a certain level resulted in recruitment of muscles besides the diaphragm. In six animals, an external abdominal pressure of 15 cmH2O produced maximal transdiaphragmatic pressure, suggesting that the diaphragm was contracting near optimal position with this external abdominal pressure. In another four animals the twitch and pressure-frequency characteristics with the use of DPS were found to be reproducible over a 2-h period. We conclude that DPS is an effective method of diaphragmatic stimulation and should prove to be a valuable technique to study the diaphragm in long-term studies of small rodents.

The purpose of this study was to determine the diurnal fluctuation of glycogen stores for the whole hemidiaphragm and within a specific myofibrillar ATPase (M-ATPase) fibre type and diaphragmatic region. Fifty-six golden Syrian hamsters were randomly divided into six groups according to the time of sampling biopsies from the diaphragm: 03:00, 07:00, 11:00, 15:00, 19:00, and 23:00. The right hemidiaphragm was quick frozen and biochemically assayed for glycogen levels. Biopsies from the left hemidiaphragm of the same animal were cut from the anterior costal and crural regions, and stained with periodic acid--Schiff (PAS) and for M-ATPase. Optical density measures of PAS-stained fibres were determined to quantitate glycogen in different M-ATPase fibre types and diaphragmatic regions. Biochemical assay of the entire hemidiaphragm showed slightly greater glycogen content of biopsies taken at 11:00 and 15:00 than at 03:00, 19:00, and 23:00 (range of differences: 6.4-10.0%). However, glycogen levels within a specific M-ATPase fibre type and diaphragm region were not different in biopsies sampled at different times. Because the hamster has a small diurnal variation of glycogen in the diaphragm, which is similar to the small diurnal variation of glycogen in human skeletal muscle, this species may be a good animal model for metabolic studies of the diaphragm that could be affected by diurnal glycogen variability.