Skeletal muscle relaxation rate after fasting or hypocaloric feeding.

The effect of malnutrition on skeletal muscle relaxation is not entirely clear; some studies indicate no change and others a slowing of the relaxation rate. We investigated whether these different results were due to type of malnutrition, muscle fiber type composition, or the index used to express relaxation rate. The effect of a 2-day fast (16% body wt loss) or 1 wk of hypocaloric feeding (22.6% wt loss) on relaxation rates of soleus and extensor digitorum longus (EDL) muscles was studied in situ with the use of anesthetized adult Wistar rats. Relaxation rates were assessed for twitch contractions using half-relaxation times and exponential phase half-times and for tetanic contractions using exponential phase half-times. The rate of relaxation was unaffected by fasting, whereas hypocaloric feeding reduced relaxation rates after twitch and tetanic contractions in both soleus and EDL muscles. We conclude that slowing of skeletal muscle relaxation rate occurs after 1 wk of hypocaloric feeding but not after 2 days of fasting. The slowing is independent of muscle fiber composition, type of contraction, or the index used to express relaxation rate.

Effects of hypothyroidism and aortic construction on mitochondria during cardiac hypertrophy.

We evaluated mitochondrial adaptations in the hearts of euthyroid and hypothyroid rats subject to aortic constriction for 2, 4, 7, 14, 21, and 28 d to induce a pressure-overload (PO), compared to sham-operated (SH) controls. PO animals attained higher arterial pressures than SH animals, by 55% in the euthyroid group, but only 14% in hypothyroid rats after 28 d. The left ventricle/body weight ratio was increased 44% by PO in the euthyroid group, and 26% in the hypothyroid group. PO attenuated the decline in cardiac growth in the hypothyroid group. Thus, hypothyroidism reduces the magnitude of the PO, but not the potential for hypertrophy in response to PO. Cytochrome c oxidase activity (CYTOX) was unchanged by PO in the euthyroid animals, indicating that the synthesis of mitochondria paralleled adaptive growth. However, CYTOX activity decreased up to 20% in the hypothyroid groups (P < 0.05) and was unaltered by PO. Thus, PO prevented the decline in growth, but not the decline in mitochondrial enzymes due to hypothyroidism. The lack of effect of PO on mitochondria was partly due to pretranslational changes since CYTOX subunit VIc mRNA was reduced by PO in the hypothyroid animals, but not in the euthyroid group. Levels of the chaperones HSP60 and GRP75, as well as HSP60 mRNA were unaffected by hypothyroidism, but paralleled adaptive growth induced by PO. Hypothyroidism changes the pattern of gene expression within the heart leading to altered mitochondrial composition. This cannot be compensated for by conditions of increased physiological demand.

Comparison of six methods for force normalization in muscles from malnourished rats.

We have investigated six methods for normalizing isometric force in skeletal muscles from malnourished rats. Adult male Wistar rats were anesthetized, and the maximum isometric force production of soleus and extensor digitorum longus (EDL) muscles was measured after a 2-d fast or 1 wk of hypocaloric feeding. The force was expressed per unit muscle wet weight, dry weight, cross-sectional area, total creatine, total protein, and myofibrillar protein. We found that the absolute force of the muscles (N) and the force expressed in relation to total creatine were similar in muscles from fasted, hypocaloric, and control rats. Regardless of the method of normalization, forces produced by soleus muscles from fasted rats were the same as control values. Force expressed in relation to wet weight, dry weight, cross-sectional area, and total protein was greater in muscles from malnourished animals except for the soleus muscles from fasted rats. Force per mg myofibrillar protein was greater in the EDL muscles from fasted and hypocaloric rats than in controls, but the values for the soleus muscles were similar in the three groups. The data indicate that different methods of force normalization can lead to different conclusions concerning the ability of muscles to generate force after nutritional intervention.

Effect of malnutrition on aerobic and anaerobic performance of fast- and slow-twitch muscles of rats.

The effect of malnutrition on the functional properties of fast- and slow-twitch muscles from rats was studied using aerobic and anaerobic preparations. A 2-day fast and hypocaloric feeding to a weight loss of 25% were used as models of malnutrition. Soleus (slow-twitch) and extensor digitorum longus (EDL) (fast-twitch) muscles were studied using an in situ preparation with the blood supply intact and an in vitro preparation to which cyanide had been added to render the muscles anaerobic. We found that a 2-day fast had little effect on the function of muscles stimulated in situ, whereas anaerobic stimulation produced a decrease in force per gram of muscle weight in the soleus, but not in the EDL, compared with control values. Hypocaloric feeding resulted in a slowed relaxation rate, an increased Fs/Fmax ratio, and an upward shift of the force-frequency curve relative to controls when studied in situ. Under anaerobic conditions, soleus muscles from hypocaloric rats continued to show a slow relaxation rate and demonstrated a loss of force per gram of muscle weight compared with controls, particularly at low stimulation frequencies. EDL muscles from hypocaloric rats had an increased relaxation rate and were able to maintain force with anaerobic stimulation. Soleus and EDL muscles from the fasted and hypocaloric groups had lower activities of phosphofructokinase. We conclude that slow-twitch muscles from malnourished rats are at a disadvantage when required to function under anaerobic conditions. These findings suggest that muscle performance may be impaired in malnourished patients subjected to hypoxia.

Effect of hypothyroidism on the expression of cytochrome c and cytochrome c oxidase in heart and muscle during development.

The effect of thyroid hormone on the expression of mitochondrial proteins was evaluated during development by measuring cytochrome c oxidase (CYTOX) activity and cytochrome c protein and mRNA levels in heart and skeletal muscle of control and hypothyroid rats. Animals were killed at the late fetal, early, and late postnatal stages up to 56 days of age. In heart, CYTOX activity increased 2.3-fold above the fetal level throughout development, most of which occurred prior to 2 days of age. No increase was observed in muscle. CYTOX activity was reduced in hypothyroid animals throughout development in heart compared to controls (by 50% at 56 days), but in muscle no effect of hypothyroidism was observed. In muscle and heart 4- and 1.5-fold increases in cytochrome c above the fetal level were evident by 1 day of age, with further increases to 8.5- and 2.7-fold by 56 days, respectively. The increase in cytochrome c differed from the increase in CYTOX, indicating changes in mitochondrial composition. Hypothyroidism reduced cytochrome c in muscle by 30-35% at 56 days, but had no effect in heart, indicating a muscle type-specific effect of thyroid hormone on cytochrome c protein expression. Cytochrome c mRNA increased rapidly to 4-5 fold above the fetal level in both heart and muscle by 6 h post-partum. Between 7 and 56 days of age, further increases to 6- and 25-fold were observed in muscle and heart, respectively. In muscle, the 6-fold developmental increase in mRNA paralleled that of the protein, suggesting transcriptional regulation.(ABSTRACT TRUNCATED AT 250 WORDS)

Mitochondrial adaptations to chronic muscle use: effect of iron deficiency.

1. The effects of chronic muscle use on mitochondrial structure, enzymes and gene expression is reviewed. The role of iron deficiency in modulating this adaptation is discussed. 2. Chronic muscle use and disuse alter mitochondrial composition and affect mitochondrial subpopulations differentially. This has implications for an understanding of organelle assembly. 3. Iron deficiency decreases mitochondrial functional mass within muscle by reducing the level of heme and non-heme iron-containing components. This alters the metabolic response during exercise and results in a reduced endurance performance. 4. Both iron deficiency and chronic muscle use represent contrasting experimental models for the study of mitochondrial function and biogenesis.

Mitochondrial biogenesis during pressure overload induced cardiac hypertrophy in adult rats.

Existing literature provides an equivocal picture of the behavior of mitochondrial synthesis during the time course of cardiac hypertrophy. Therefore, we examined the effect of cardiac hypertrophy on mitochondrial cytochrome c oxidase (CYTOX) activity, the content of CYTOX subunit VIc mRNA, and the expression of molecular chaperones. Adult male Sprague-Dawley rats were subjected to either abdominal aortic constriction to induce pressure overload (PO) or a sham operation (SH). Animals were studied 2, 4, 7, 14, 21, or 28 days after surgery. Aortic constriction resulted in a significant evaluation in arterial pressure by 4 days after surgery. Significant (p < 0.05) hypertrophy was attained by 4 days and was stabilized at 37% between 7 and 28 days. CYTOX activity (U/g) did not differ significantly between PO and SH animals at either early (< 7 days) or later time points, indicating that mitochondrial content increased in proportion to adaptive cellular hypertrophic growth. The concentration of the molecular chaperones HSP60 and GRP75 involved in mitochondrial protein import did not change with PO treatment. The levels of mRNAs encoding both CYTOX subunit VIc and HSP60 remained constant, in proportion to cardiac growth. This suggests that the accelerated synthesis of CYTOX and HSP60 during cardiac hypertrophy is regulated transcriptionally. The data help to resolve the controversy in the literature regarding mitochondrial biogenesis during moderate, stable cardiac hypertrophy, and they indirectly indicate that proportional mitochondrial synthesis relative to cellular hypertrophy is regulated at the transcriptional level.

Stability of high-energy substrates in fast- and slow-twitch muscle: comparison of enzymatic assay of biopsy with in vivo 31P nuclear magnetic resonance spectroscopy.

The purpose of this study was to investigate the stability of ATP and PCr levels in stored muscle samples and extracts. ATP and PCr levels were measured by fluorimetric analysis in freeze-clamped biopsies of soleus, extensor digitorum longus, and gastrocnemius muscles of the rat after storage in a freezer at -70 degrees C as (i) intact wet muscle, (ii) freeze-dried muscle, and (iii) an extract of freeze-dried muscle. Assays were performed within 24 h of taking the biopsy and after variable periods of storage from 1 to 4 weeks. The data for the gastrocnemius muscles were compared with those obtained, in the same rat, by in vivo 31P NMR spectroscopy. In the biopsies, the ATP levels were stable irrespective of the duration or method of storage. The PCr levels fell by 13-16% compared with the values obtained from the assay done within 24 h of taking the biopsy, irrespective of the method of storage, but could be corrected in the freeze-dried stored muscle by expressing the data in relation to the total creatine levels. The fluorimetrically measured PCr, in whole muscle extracts of the gastrocnemius, assayed within 24 h, were comparable to those obtained from 31P NMR spectroscopy. We concluded that PCr levels in muscle are not stable during storage at -70 degrees C and should be assayed within 24 h of taking a muscle biopsy to ensure that the values are the same as those obtained by 31P NMR.

Mitochondrial biogenesis in striated muscle.

Mitochondrial biogenesis (synthesis) has been observed to occur in skeletal muscle in response to chronic use. It also occurs in cardiac muscle during growth and hypertrophy, and it may be impaired during the aging process. This review summarizes the literature on the processes of mitochondrial biogenesis at the biochemical and molecular levels, with particular reference to striated muscles. Mitochondrial biogenesis involves the expression of nuclear and mitochondrial genes and the coordination of these two genomes, the synthesis of proteins and phospholipids and their import into the organelle, and the incorporation of these lipids and proteins into their appropriate locations within the matrix, inner or outer membranes. The emphasis is on the regulation of these events, with information derived in part from other cellular systems. Although descriptions of mitochondrial content changes in heart and skeletal muscle during altered physiological states are plentiful, much work is needed at the molecular level to investigate the regulatory processes involved. A knowledge of biochemical and molecular biology techniques is essential for continued progress in the field. This is a promising area, and potential new avenues for future research are suggested.