Effects of protein kinase A inhibition on rat diaphragm force generation

Although protein kinases are known to play a role in modulating a variety of intracellular functions, the direct effect of inhibition of these enzymes on skeletal muscle force production has not been studied. The purpose of the present study was to examine this issue by determining the effects produced on diaphragm force generation by two protein kinase inhibitors: (a) H7, an inhibitor of both cAMP-dependent protein kinase (PKA) and of protein kinase C, and (b) H89, a selective inhibitor of PKA. Experiments (n=15) were performed using isolated, arterially perfused, electrically stimulated rat diaphragms. Perfusate temperature was adjusted to maintain muscle temperature at 27°C and arterial pressure was kept at 150 Torr. Animals were divided into three groups: (a) a control group perfused with Krebs-Henselheit solution equilibrated with 95% O₂/5% CO₂, (b) a group in which H7 (2 μM) was added to the perfusate, and (c) a group perfused with solution containing H89 (4 μM). In all three groups, we assessed diaphragm twitch kinetics, force-frequency relationships and in vitro fatiguability. We found that both H7 and H89 administration slowed twitch relaxation, augmented force generation in response to low frequency stimulation, and increased the rate of development of fatigue. Specifically, for control, H7 and H89 groups, respectively, we found: (a) 1/2 relaxation time averaged 64±2 S.E.M., 87±6 and 90±2 ms, P<0.003, (b) force production during 10-Hz stimulation averaged 12.6±1.1, 20.1±2.3, and 20.3±2.1 N/cm², P<0.035, and (c) force fell to 14.3±2.0, 9.5±0.5 and 8.7±0.2% of its initial value after 20 min of fatiguing stimulation, P<0.035. These data show that it is possible to produce large increases in low frequency skeletal muscle force generation by directly inhibiting PKA. We speculate that it may be possible to pharmacologically augment respiratory muscle force and pressure generation in clinical medicine by administration of PKA inhibitors. Copyright (C) 2000 Elsevier Science B.V.