A B S T R A C T Changes in [Mg [CPK], and ionic strength. Ca 2+-(or Sr2+-)activated steady-state tensions were recorded for three [Mg2+]'s: 5 X 10 -5 M, 1 X 10 -s M, and 2 X 10 -3 M; and these tensions were expressed as the percentages of maximum tension generation of the fibers for the same [Mg2+]. Maximum tension was not affected by [Mg 2+] within Ca2+-activating or Sr2+-activating sets of solutions; however, the submaximum Ca 2+-(or Sr~+-)activated tension is strongly affected in an inverse fashion by increasing [Mg2+]. Mg 2+ behaves as a competitive inhibitor of Ca 2+ and also affects the degree of cooperativity in the system. At [Mg 2+] -5 X 10 -~ M the shape of tension versus [Ca 2+] (or [Sr2+]) curve showed evidence of cooperativity of Ca 2+ (or Sr ~+) binding or activation of the contractile system. As [Mg 2+] increased, the apparent affinity for Ca 2+ or Sr 2+ and cooperativity of the contractile system declined. The effect on cooperativity suggests that as [Mg 2+] decreases a threshold for Ca 2+ activation appears.

The maximal calcium-activated isometric tension produced by a skinned frog single muscle fiber falls off as the ionic strength of the solution bathing this fiber is elevated declining to zero near 0.5 M as the ionic strength is varied using KC1. When other neutral salts are used, the tension always declines at high ionic strength, but there is some difference between the various neutral salts used. The anions and cations can be ordered in terms of their ability to inhibit the maximal calcium-activated tension. The order of increasing inhibition of tension (decreasing tension) at high ionic strength for anions is propionate--SO-4 < Cl-< Br-. The order of increasing inhibition of calciumactivated tension for cations is K + -Na +~ TMA+ < TEA + < TPrA + < TBuA+. The decline of maximal calcium-activated isometric tension with elevated salt concentration (ionic strength) can quantitatively explain the decline of isometric tetanic tension of a frog muscle fiber bathed in a hypertonic solution if one assumes that the internal ionic strength of a muscle fiber in normal Ringer's solution is 0.14-0.17 M. There is an increase in the base-line tension of a skinned muscle fiber bathed in a relaxing solution (no added calcium and 3 mM EGTA) of low ionic strength. This tension, which has no correlate in the intact fiber in hypotonic solutions, appears to be a noncalcium-activated tension and correlates more with a declining ionic strength than with small changes in [MgATP], [Mg], pH buffer, or [EGTA]. It is dependent upon the specific neutral salts used with cations being ordered in increasing inhibition of this noncalcium-activated tension (decreasing tension) as TPrA + < TMA+ < K + Na+. Measurements of potentials inside these skinned muscle fibers bathed in relaxing solutions produced occasional small positive values (<6 mV) which were not significantly different from zero.

1. Maximum and submaximum Ca-activated tension in mechanically disrupted rat ventricular fibres was examined in solutions containing 30 micron, 100 micron and 4 mM-MgATP and either 50 micron or 1 mM ionized Mg. 2. In the absence of added Ca, significant amounts of base-line tension (up to 50% of maximum) develop in solutions containing less than 30 micron-MgATP. This effect is Mg-dependent; more tension is produced with 50 micron-Mg than with 1 mM. 3. Increasing the MgATP concentration shifts the pCa-% maximum tension relationship in the direction of increasing Ca required for activation. At 50 micron-Mg the pCa which produces 50% maximum tension is 5-8, 5-3 and 5-5 for the 30 micron, 100 micron and 4 mM-MgATP solutions. The effect of MgATP on position is relatively independent of the Mg concentration. 4. The steepness of the pCa-% maximum tension curve increases as MgATP is elevated to the millimolar range. The Hill coefficients for the different MgATP curves at 50 micron-Mg are 1-1, 1-3 and 3-0. This change in steepness accounts for the slightly lower Ca concentration needed for half-maximum tension as the MgATP concentration is increased to millimolar levels. Raising the Mg concentration to 1 mM greatly diminishes the effect of MgATP on the slope of the pCa-tension relationship. 5. The maximum tnesion a fibre bundle can produce decreases as the amount of MgATP is raised from micromolar to millimolar levels. For 50 muM-Mg, maximum tension drops about 35% as MgATP is raised from 30 micronM to 4 mM. For any concentraiton of MgATP, maximum tension is higher at 1 mM-Mg than at 50 micron-Mg. 6. Existing theories of interaction between myosin heads and the thin filament are sufficient to account for the effects of MgATP on the position of the pCa-tension curves and on maximum tension. The effects on slope are less satisfactorily explained.

Single muscle fibers from rabbit soleus and adductor magnus and from semitendinosus muscles were peeled to remove the sarcolemma and then stimulated to release Ca2' by (a) caffeine application or (b) ionic depolarization accomplished via substitution of choline chloride for potassium propionate at constant [K+] X [Cl-] in the bathing solution . Each stimulus, ionic or caffeine, elicited an isometric tension transient that appeared to be due to Ca 21 released from the sarcoplasmic reticulum (SR) . The peak magnitude of the ionic (Cl--induced) tension transient increased with increasing Cl-concentration . The application of ouabain to fibers after peeling had no effect on either type of tension transient. However, soaking the fibers in a ouabain solution before peeling blocked the Cl --induced but not the caffeine-induced tension transient, which suggests that ouabain's site of action is extracellular, perhaps inside transverse tubules (TTs). Treating the peeled fibers with saponin, which should disrupt TTs to a greater extent than SR membrane, greatly reduced or eliminated the Cl --induced tension transient without significantly altering the caffeine-induced tension transient. These results suggest that the Cl --induced tension transient is elicited via stimulation of sealed, polarized TTs rather than via ionic depolarization of the SR .

Muscle atrophy is a significant consequence of chronic kidney disease (CKD) that increases a patient's risk of mortality and decreases their quality of life. The loss of lean body mass results, in part, from an increase in the rate of muscle protein degradation. In this review, the proteolytic systems that are activated during CKD and the key insulin signaling pathways that regulate the protein degradative processes are described.Skeletal muscle is a dynamic organ that provides a rich source of amino acids and carbon chains that can be mobilized during stress or chronic pathological conditions like chronic renal failure (CKD), sepsis or diabetes. Estimates of protein turnover rates in skeletal muscle of healthy individuals indicate that nearly 1.5 kg muscle mass turns over daily (1). Given this extraordinary level of protein metabolism in skeletal muscle, one can easily envision how even a minor change in the rate of protein degradation without a reciprocal change in synthesis would have profound physiological effects over time in conditions like CKD.There are numerous reports documenting the cachexia or loss of muscle mass that is frequently seen in patients with chronic kidney disease. Using experimental models of CKD, the causes of muscle atrophy have been extensively studied. May et al. found that CKD attenuated insulinstimulated protein synthesis and increased protein degradation in skeletal muscle (2). These studies were subsequently extended by identifying the proteolytic pathways in muscle that are augmented during CKD. The major pathway that is up-regulated during CKD, as well as most other chronic diseases, is the ubiquitin-proteasome pathway. This multi-enzyme system involves the targeting of muscle proteins by a series of enzymatic reactions and the subsequent degradation by a large protein complex called the proteasome (Figure 1). Proteins are targeted for proteasomal degradation by the covalent attachment of polymeric chains of a 7-kDa protein, ubiquitin, to the ε-amino group of lysine residues in the substrate proteins. The key substrate recognition component in this process is a group of enzymes called E3 ubiquitin ligases which are the largest known family of functionally-related proteins (>300) in mammals. Once the substrate protein is "polyubiquitinated", it is degraded by the proteasome, a large (2000 kDa)