UBC Theses and Dissertations
Metabolic regulation in skeletal muscle during exercise : a fish-mammal comparison Dobson, Geoffrey P.
The aim of the present investigation was to examine the control of anaerobic glycogenolysis in working and fatigued skeletal muscle. The two animals chosen for the study were a teleost fish and the laboratory rat. The rationale behind using a comparative approach to investigate fundamental questions on metabolic control resides in the different abilities of each animal to perform exercise, and to their markedly different myofibrillar organization. In the process of defining the hierarchical recruitment of fuel and pathway selection in rainbow trout fast-twitch white skeletal muscle, it was clear that the near-maximal myosin ATPase activity was supported solely by PCr hydrolysis. It was not until the rate and force of contraction decreased that the relative contribution of anaerobic glycogenolysis became increasingly important. Despite glycogenolysis possessing a lower maximal ATP generating potential than PCr hydrolysis, it has the advantage of being less constrained by time, and is recruited to extend muscle performance, but at submaximal workloads. Demonstration of the same temporal pattern of activation was not attempted for rat skeletal muscle because of complex fiber heterogeneity. The etiology of fatigue after 10 and 30 minutes of burst swimming in trout was due to the near depletion of glycogen in white muscle. Inhibition of anaerobic glycogenolysis was not correlated with limitations to either the availability of ADP or NAD⁺ or inhibition ofphosphofructokinase (PFK-1). Similarly, the onset of fatigue and inhibition of glycogenolysis in three different skeletal muscles (gastrocnemius, plantaris and soleus) of the rat after 30 min of endurance treadmill running (25 meters/min), was not related to ADP availability, but associated with the near-depletion of muscle glycogen. As the endogenous stores of glycogen became limiting, hexokinase (Hk) appeared to be activated in trout white muscle after 10 min, and in the three rat skeletal muscles after 30 min, Indicating an increase in uptake and phosphorylation of blood-borne glucose. In rats running at a high speed for 2 minutes, glycogenosis was maintained through the coordination of glycogen phosphorylase and PFK-1. Muscle performance in these rats was maintained despite large percentage swings in cytosolic redox, the ATP/ADP ratio and phosphorylation potential. A common belief in the literature is that inhibition of glycogenolysis during short-term strenuous exercise is brought about by the pH dependent ATP inhibition of PFK-catalysis. Evidence was provided indicating PFK-1 is operational in skeletal muscle at about pH 6.6 for both the fish and the rat. Fish partially solved the problem of PFK-1 inhibition by lowering ATP, whereas the rat appeared to rely on the synergistic action of a number of positive modulators. A detailed kinetic analysis of purified rabbit muscle PFK-1 revealed that any modulator that increases the ratio of unprotonated to protonated form of the enzyme, could supply the muscle cell with a means of maintaining glycogenolytic flux despite falling pH. A number of striking differences were apparent between the regulation of glycogenolysis in fish and rat skeletal muscle. The first major difference was the direction of change in cytosolic redox or the NAD+/NADH ratio. In fish white muscle the ratio increased, and the cytosol became more oxidized with exercise, whereas the opposite occurred in rat fast-twitch skeletal muscle. The difference was a consequence of lactate retention in fish white muscle. On the basis of crossover analysis, pyruvate kinase (PK) appeared to be activated in trout white muscle at both fatigue states. However, this was misleading, and also considered a consequence of rising pyruvate and due to lactate retention via the mass action effect at the LDH equilibrium. Obviously, the change in redox and the apparent crossover at PK are linked, and a literature survey revealed that during short-term maximal work, a mammalian skeletal muscle may indeed behave as fish white muscle. The other contrasting feature of this comparative analysis was the demonstration that ATP in trout white muscle can fall by 80% at exhaustion. No such large percentage reductions in ATP occurred in either of the rat fast-twitch skeletal muscles, or indeed have been reported in skeletal muscle of any other exercising animal. In all cases the total nucleotide pool remained constant. A general conclusion to be drawn from this study is that muscle fatigue should be viewed as a multi-component process in response to limiting glycogen, and not leveled at any one particular step of the glycogenolytic pathway.