Protecting the cellular energy state during contractions: role of AMP deaminase.

AMP deaminase activity (AMP->IMP+NH3) is the entry reaction to the purine nucleotide cycle. In skeletal muscle, excessive energy demands during contractions leads to a net production of ADP, because ATP hydrolysis exceeds ADP rephosphorylation. Elevations in ADP increase AMP, via the myokinase reaction. This accumulation of ATP hydrolysis products should lead to a catastrophic reduction in the energy state of the myocyte. The removal of AMP to IMP in times of excessively high energy demands have been hypothesized as essential to protect the energy state of the cell. While AMP deamination leads to a net loss of adenine nucleotides (principally, as ATP), the viability of the myocyte is preserved. Following these demanding contraction conditions, the concentration of IMP of fast-twitch muscle is rapidly reduced, typically with the return of the muscle adenine nucleotide content (ATP + ADP + AMP) to pre-contraction levels. While these observations are generally observed for fast-twitch skeletal muscle and consistent with the hypothesis, there has been no direct experimental evaluation. In the AK1 (-/-) mouse, there is a markedly reduced accumulation of AMP, during conditions of excessive contractile activity. Rather, there is a high ADP concentration, approaching 1.5 mM, that remains unbound 'free' within the muscle. This contributes to an inordinate reduction in the ATP/ADP ratio. At the same time, PCr hydrolysis is nearly complete leading to a large increase in orthophosphate. In combination, this leads to an exceptional decline in the free energy of ATP hydrolysis. This is projected to impair Ca(2+) handling by the sarcoplasmic reticulum and slow cross-bridge cycling rate. The outcome should be slowed contraction characteristics and possible contracture. While some contractile changes were observed, there was a remarkable ability of the muscle to function under these challenging energetic conditions. Thus, it is not essential that the AMP deaminase reaction be operating during intense contraction conditions. This helps explain why patients deficient in AMP deaminase do not always exhibit an impaired muscle function.

Influence of ribose on adenine salvage after intense muscle contractions.

The influence of ribose supplementation on skeletal muscle adenine salvage rates during recovery from intense contractions and subsequent muscle performance was evaluated using an adult rat perfused hindquarter preparation. Three minutes of tetanic contractions (60 tetani/min) decreased ATP content in the calf muscles by approximately 50% and produced an equimolar increase in IMP. Effective recovery of muscle ATP 1 h after contractions was due to reamination of IMP via the purine nucleotide cycle and was complete in the red gastrocnemius but incomplete in the white gastrocnemius muscle section. Adenine salvage rates in recovering muscle averaged 45 +/- 4, 49 +/- 5, and 30 +/- 3 nmol. h(-1). g(-1) for plantaris, red gastrocnemius, and white gastrocnemius muscle, respectively, which were not different from values in corresponding nonstimulated muscle sections. Adenine salvage rates increased five- to sevenfold by perfusion with approximately 4 mM ribose (212 +/- 17, 192 +/- 9, and 215 +/- 14 nmol. h(-1). g(-1) in resting muscle sections, respectively). These high rates were sustained in recovering muscle, except for a small (approximately 20%) but significant (P < 0.001) decrease in the white gastrocnemius muscle. Ribose supplementation did not affect subsequent muscle force production after 60 min of recovery. These data indicate that adenine salvage rates were essentially unaltered during recovery from intense contractions.

Purine salvage to adenine nucleotides in different skeletal muscle fiber types.

Rates of purine salvage of adenine and hypoxanthine into the adenine nucleotide (AdN) pool of the different skeletal muscle phenotype sections of the rat were measured using an isolated perfused hindlimb preparation. Tissue adenine and hypoxanthine concentrations and specific activities were controlled over a broad range of purine concentrations, ranging from 3 to 100 times normal, by employing an isolated rat hindlimb preparation perfused at a high flow rate. Incorporation of [(3)H]adenine or [(3)H]hypoxanthine into the AdN pool was not meaningfully influenced by tissue purine concentration over the range evaluated (approximately 0.10-1.6 micromol/g). Purine salvage rates were greater (P < 0.05) for adenine than for hypoxanthine (35-55 and 20-30 nmol x h(-1) x g(-1), respectively) and moderately different (P < 0.05) among fiber types. The low-oxidative fast-twitch white muscle section exhibited relatively low rates of purine salvage that were approximately 65% of rates in the high-oxidative fast-twitch red section of the gastrocnemius. The soleus muscle, characterized by slow-twitch red fibers, exhibited a high rate of adenine salvage but a low rate of hypoxanthine salvage. Addition of ribose to the perfusion medium increased salvage of adenine (up to 3- to 6-fold, P < 0.001) and hypoxanthine (up to 6- to 8-fold, P < 0.001), depending on fiber type, over a range of concentrations up to 10 mM. This is consistent with tissue 5-phosphoribosyl-1-pyrophosphate being rate limiting for purine salvage. Purine salvage is favored over de novo synthesis, inasmuch as delivery of adenine to the muscle decreased (P < 0.005) de novo synthesis of AdN. Providing ribose did not alter this preference of purine salvage pathway over de novo synthesis of AdN. In the absence of ribose supplementation, purine salvage rates are relatively low, especially compared with the AdN pool size in skeletal muscle.