Little effect on creatine uptake rates has been observed with PCr, creatinine, ornithine, glycine, glutamic acid, histidine, alanine, arginine, leucine, methionine or cysteine (Fitch, 1988; Loike et al., 1988; Möller and Hamprecht, 1989). The sodium-dependent uptake of creatine in culture is sensitive to extracellular creatine concentration (Loike et al., 1988). Myoblasts maintained for 24 h in a medium containing 1 mM creatine exhibited one-third of the uptake activity of cells bathed for the same duration in a medium lacking creatine. The downregulation was slowed by inhibitors of protein synthesis. From this observation one can conclude that extracellular creatine regulates the induction of the expression of a protein that decreases the sodium-dependent uptake activity only after the creatine enters the cell.
No alteration in the efflux rate for creatine was observed in studies of the effects of changes in extracellular creatine concentrations (Loike et al., 1988). If creatine was leaving mainly by passive or facilitated diffusion, an alteration in efflux rates would be expected. Thus, it seems probable that the creatine is leaving mainly as creatinine, possibly with a small creatine component as they both move down their concentration gradients.
Adding thyroxine (T4) to mimic hyperthyroidism increases the uptake and efflux of creatine in the rat heart (Seppet et al., 1985). This group found a 20fold increase in Fmax and questioned the earlier findings which indicated that T3 induces inhibition of creatine uptake (Dinking et al., 1959, Fitch et al., 1960). Support for the findings of Seppet et al. (1985) comes from more recent studies in which the exposure of cultured mouse myoblasts to T3 produced increased creatine uptake (Odoom et al., 1993). This observation may result from the increase in the Na/K pump activity caused by T3 (Brodie and Sampson, 1988), an effect supported by the sensitivity of uptake to oubain (Möller and Hamprecht, 1989).
Creatine loading can be accomplished by incubating tissues with high concentrations of creatine. Total tissue creatine and PCr increase dramatically, probably because of creatine flooding across the membranes. This was performed in porcine carotid artery by Clark and Dillon (1995) to examine the flux through the CK system in the presence of increased metabolites. They found that despite increased PCr in the porcine carotid artery the CK reaction remained at equilibrium.
If porcine carotid arteries are incubated with 50 mM creatine substituted for sodium, in the absence of phosphate, the vessels tend to die. Figure 3.3 illustrates two 31P NMR spectra from two different porcine carotid arteries. Spectrum A is the control spectrum that was incubated for 12 h in a P, free Krebs buffer with glucose as the substrate in the absence of creatine. The PCr/ATP ratios are consistent with those reported by Clark and Dillon (1995) and show healthy carotid arteries. Spectrum B is from a porcine carotid artery that has been incubated with the same buffer but in the presence of 50 mM creatine. Interestingly, the PCr peak is greatly enlarged, while the ATP peaks are decreased. These results indicate that high levels of creatine in the presence of active CK will act as a phosphate sink, leading to the loss of ATP and possibly the nucleotide pool. Consistent with the equilibrium nature of CK, nucleotide levels appear to fall, but this occurs to the detriment of the cell.
Much of the research on creatine transport has been performed with a view towards therapeutic intervention in pathological conditions of the muscular system. Exercise combined with creatine ingestion enhances the increase in cell creatine (Harris et al., 1992). Thus, it may be possible to increase this effect by combining similar protocols with stimulatory hormones such as T3 or adrenaline (the latter because isoproterenol appears to enhance creatine uptake) (J.E. Odoom, unpublished data). As the hormones or other factors that affect transport become known, therapeutic interventions for particular disease states and individual patients can be devised.
The control of creatine transport may also have relevance for the physiology of adaptation to exercise. We know that there is a rapid influx of creatine compared with a much slower net efflux and this suggests that each bout of exercise may cause an increase in cell creatine content that persists and is cumulative
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