Age-related differences in the adaptive potential of type I skeletal muscle fibers.

This study was designed to investigate whether the relationship of fiber size and force is maintained with aging and inactivity. We hypothesized that fiber size and fiber force-generating capacity would decrease in parallel, thus resulting in no change in specific force with either age or inactivity. Thirty male Fischer 344/Brown Norway F1 hybrid rats, 6-23-month old (young adult), 24-33-month old (middle-aged) and 34-40 month old (old) were hindlimb unweighted for 14 days. Single permeabilized type I fibers from the soleus and gastrocnemius muscles were evaluated for size and contractile function. The diameter of the fibers from the soleus muscle declined with age and unweighting. In contrast, the fibers from the gastrocnemius showed no age-related atrophy. In both the soleus and gastrocnemius, there was a significant decrease in the force generation with age and unweighting. When comparing size and peak absolute force in the type I fibers from control young and middle-aged animals a positive relationship was observed whereas no significant relationship between size and peak absolute force was observed in the old animals. Following unweighting, fibers from young and middle aged rats showed a significant relationship between force and size. Fibers from old animals did not exhibit a relationship between size and force following unweighting. These results suggest aged skeletal muscle has an attenuated ability to adapt to inactivity by altering its size in response to inactivity.

Altered proteasome structure, function, and oxidation in aged muscle.

The proteasome is the main protease for degrading oxidized proteins. We asked whether altered proteasome function contributes to the accumulation of oxidized muscle proteins with aging. Proteasome structure, function, and oxidation state were compared in young and aged F344BN rat fast-twitch skeletal muscle. In proteasome-enriched homogenates from aged muscle, we observed a two- to threefold increase in content of the 20S proteasome that was due to a corresponding increase in immunoproteasome. Content of the regulatory proteins, PA700 and PA28, relative to the 20S were reduced 75% with aging. Upon addition of exogenous PA700, there was a twofold increase in peptide hydrolysis in aged muscle, suggesting the endogenous content of PA700 is inadequate for complete activation of the 20S. Measures of catalytic activity showed a 50% reduction in specific activity for proteasome-enriched homogenates with aging. With purification of the 20S, proteasome specific activity was equivalent between ages, indicating that endogenous regulators inhibit proteasome in aged muscle. Significantly less degradation of oxidized calmodulin by the 20S from aged muscle was observed. Partial rescue of activity for aged 20S by DTT implies oxidation of functionally significant cysteines. These results demonstrate significant age-related changes in proteasome structure, function, and oxidation state that could inhibit removal of oxidized proteins.

Myofibrillar myosin ATPase activity in hindlimb muscles from young and aged rats.

We tested the hypothesis that Ca(2+)-activated myosin ATPase activity is lower in muscles of aged rats relative to muscles of young rats, independent of changes in myosin isoform expression. Myofibrils were prepared from permeabilized fibers of soleus, plantaris, and semimembranosus muscles of young (8-12 months) and aged (32-38 months) F344 x BN rats and assayed for resting myosin ATPase, Ca(2+)-activated myosin ATPase, and myosin heavy chain (MHC) and myosin light chain (MLC) isoform compositions. Resting myosin ATPases were not affected by age in any muscle (P > or = 0.42). Ca(2+)-activated myosin ATPases of soleus and plantaris myofibrils were not affected by age (P > or = 0.31) but were 16% lower in semimembranosus myofibrils from aged rats (0.448 +/- 0.019 micromol P(i)/min/mg) compared to young rats (0.533 +/- 0.031 micromol P(i)/min/mg; P = 0.03). Correspondingly, maximal unloaded shortening velocity of single semimembranosus fibers from aged rats was slow (4.6 +/- 0.2 fiber lengths/s) compared with fibers from young rats (5.8 +/- 0.3 fiber lengths/s; P < 0.01). No age-related changes in MHC or regulatory MLC isoforms were detected in any muscle (P > or = 0.08) but changes in the essential MLC occurred in plantaris and semimembranosus muscles. The data indicate that Ca(2+)-activated myosin ATPase activity is reduced with age in semimembranosus muscle, independent of age-related changes in MHC isoform expression, and is one mechanism contributing to age-related slowing of contraction in that muscle.

Altered proteasome function and subunit composition in aged muscle.

Myofibrillar protein degradation is mediated through the ubiquitin-proteasome pathway. To investigate if altered proteasome activity plays a role in age-related muscle atrophy, we examined muscle size and proteasome function in young and aged F344BN rats. Significant age-related muscle atrophy was confirmed by the 38% decrease in cross-sectional area of type 1 fibers in soleus muscle. Determination of proteasome function showed hydrolysis of fluorogenic peptides was equivalent between ages. However, when accounting for the 3-fold increase in content of the 20S catalytic core in aged muscle, the lower specific activity suggests a functional loss in individual proteins with aging. Comparing the composition of the catalytic beta-subunits showed an age-related 4-fold increase in the cytokine-inducible subunits, LMP2 and LMP7. Additionally, the content of the activating complexes, PA28 and PA700, relative to the 20S proteasome was reduced 50%. These results suggest significant alterations in the intrinsic activity, the percentage of immunoproteasome, and the regulation of the 20S proteasome by PA28 and PA700 in aged muscle.