Effects Of Long-term Caloric Restriction Research Articles

Effect of long-term caloric restriction on telomere length in healthy adults: CALERIE™ 2 trial analysis.

Caloric restriction (CR) modifies lifespan and aging biology in animal models. The Comprehensive Assessment of Long-Term Effects of Reducing Intake of Energy (CALERIE™) 2 trial tested translation of these findings to humans. CALERIE™ randomized healthy, nonobese men and premenopausal women (age 21-50y; BMI 22.0-27.9 kg/m2), to 25% CR or ad-libitum (AL) control (2:1) for 2 years. Prior analyses of CALERIE™ participants' blood chemistries, immunology, and epigenetic data suggest the 2-year CR intervention slowed biological aging. Here, we extend these analyses to test effects of CR on telomere length (TL) attrition. TL was quantified in blood samples collected at baseline, 12-, and 24-months by quantitative PCR (absolute TL; aTL) and a published DNA-methylation algorithm (DNAmTL). Intent-to-treat analysis found no significant differences in TL attrition across the first year, although there were trends toward increased attrition in the CR group for both aTL and DNAmTL measurements. When accounting for adherence heterogeneity with an Effect-of-Treatment-on-the-Treated analysis, greater CR dose was associated with increased DNAmTL attrition during the baseline to 12-month weight-loss period. By contrast, both CR group status and increased CR were associated with reduced aTL attrition over the month 12 to month 24 weight maintenance period. No differences were observed when considering TL change across the study duration from baseline to 24-months, leaving it unclear whether CR-related effects reflect long-term detriments to telomere fidelity, a hormesis-like adaptation to decreased energy availability, or measurement error and insufficient statistical power. Unraveling these trends will be a focus of future CALERIE™ analyses and trials.

Effect of long-term caloric restriction on DNA methylation measures of biological aging in healthy adults from the CALERIE trial

The geroscience hypothesis proposes that therapy to slow or reverse molecular changes that occur with aging can delay or prevent multiple chronic diseases and extend healthy lifespan1–3. Caloric restriction (CR), defined as lessening caloric intake without depriving essential nutrients4, results in changes in molecular processes that have been associated with aging, including DNA methylation (DNAm)5–7, and is established to increase healthy lifespan in multiple species8,9. Here we report the results of a post hoc analysis of the influence of CR on DNAm measures of aging in blood samples from the Comprehensive Assessment of Long-term Effects of Reducing Intake of Energy (CALERIE) trial, a randomized controlled trial in which n = 220 adults without obesity were randomized to 25% CR or ad libitum control diet for 2 yr (ref. 10). We found that CALERIE intervention slowed the pace of aging, as measured by the DunedinPACE DNAm algorithm, but did not lead to significant changes in biological age estimates measured by various DNAm clocks including PhenoAge and GrimAge. Treatment effect sizes were small. Nevertheless, modest slowing of the pace of aging can have profound effects on population health11–13. The finding that CR modified DunedinPACE in a randomized controlled trial supports the geroscience hypothesis, building on evidence from small and uncontrolled studies14–16 and contrasting with reports that biological aging may not be modifiable17. Ultimately, a conclusive test of the geroscience hypothesis will require trials with long-term follow-up to establish effects of intervention on primary healthy-aging endpoints, including incidence of chronic disease and mortality18–20.

Differential Effects of Long-Term Caloric Restriction and Dietary Protein Source on Bone and Marrow Fat of the Aging Rat.

Long-term caloric restriction (CR) has been shown to be beneficial to various tissues and organs. In contrast, CR exerts differential effects on bone, which could be due in part to the nature of the protein regime utilized. Male Sprague Dawley rats (8-month-old) were subjected for 12 months to 40% CR in macronutrients and compared with rats fed ad libitum for the same period. Casein- and soy-fed groups were compared. There was a significant decrease in bone quality in both CR groups, which was independent of the source of protein in the diet. In contrast, the group fed soy protein ad libitum showed better bone quality and higher levels of bone formation compared with casein-fed animals. Notably, bone marrow adipocytes were not mobilized upon CR as demonstrated by an absence of change in adipocyte number and tissue expression of leptin. This study demonstrates that the negative effect of CR on bone quality could not be prevented by the most common protein regimes.

Initial age-associated metabolic alterations in white and brown fat depots: effect of long-term caloric restriction.

Initial age-associated metabolic alterations in white and brown fat depots: effect of long-term caloric restriction.

Brain region-specific effects of long-term caloric restriction on redox balance of the aging rat

Caloric restriction (CR) is the most effective intervention to improve health span and extend lifespan in preclinical models. This anti-aging effect of CR is related to attenuation of oxidative damage in various tissues, with divergent results in the brain. We addressed how brain oxidoreductive balance would be modulated in male Sprague-Dawley (SD) rats submitted to a 40% CR from 8 to 19 months of age, by reference to ad libitum-fed (AL) rats at 2 and 19 months of age. Four brain structures were compared: hippocampus, striatum, parietal cortex, cerebellum. Our CR diet elicits significant prevention of oxidative damages with the upregulation of antioxidant defenses (levels of glutathione [GSH], mRNAs of clusterin and of three key antioxidant enzymes) as compared to age-matched AL controls, in a strikingly region-specific pattern. CR also prevented a drastic rise of the glial fibrillary acidic protein in the hippocampus of old AL rats. Besides, the CR effects at age 19 months mainly consist in improving endogenous defenses before the onset of age-related redox alterations. These effects are more prominent in the hippocampus.

Caloric restriction promotes functional changes involving short-chain fatty acid biosynthesis in the rat gut microbiota

Caloric restriction (CR) is known to promote health and longevity, likely via modification of the gut microbiota (GM). However, functional and metabolic changes induced in the GM during CR are still unidentified. Here, we investigated the short- and long-term effects of CR on the rat GM using a metaproteogenomic approach. We show that a switch from ad libitum (AL) low fat diet to CR in young rats is able to induce rapid and deep changes in their GM metaproteomic profile, related to a reduction of the Firmicutes/Bacteroidetes ratio and an expansion of lactobacilli. Specifically, we observed a significant change in the expression of the microbial enzymes responsible for short-chain fatty acid biosynthesis, with CR boosting propionogenesis and limiting butyrogenesis and acetogenesis. Furthermore, these CR-induced effects were maintained up to adulthood and started to be reversed after a short-term diet change. We also found that CR alters the abundance of an array of host proteins released in stool, mainly related to epithelial barrier integrity and inflammation. Hence, our results provide thorough information about CR-induced modifications to GM and host functional activity, and might constitute the basis for novel GM-based approaches aimed at monitoring the effectiveness of dietary interventions.

Abstract 270: Long term Caloric Restriction and Myocardial Aging in Non Human Primates

Caloric restriction (CR) has been shown to decrease aging and increase the lifespan of several model organisms. However, in two on-going independent, long-term CR studies in non-human primates (NHP) at the National Institute of Aging (NIA) and University of Wisconsin, Madison (UW) the outcome has been more complex. While CR monkeys in the UW study have experienced a significant increase in lifespan relative to their study controls, this was not the case for the NIA study. There, CR did not increase lifespan compared to the already long-lived controls. Yet, in both studies CR was associated with a decrease in the occurrence of age-related diseases. Left ventricle (LV) samples were provided from the rhesus monkeys from both studies to investigate the effect of long-term CR on myocardial pathology and aging in these two cohorts of non-human primates (NHPs). Although CR did not reduce cardiac fibrosis or hypertrophy in either study, there was a 5.9-fold increase in fibrosis (p<0.0001) in LV tissue from the UW study compared to NIA, irrespective of CR treatment. Myocardial mTOR levels were reduced with CR in the UW study, only. These findings indicate that the severity of cardiac fibrosis may not be susceptible to the mechanisms by which CR is affecting disease progression, and myocardial mTOR is apparently differently affected dependent on the study paradigm. Interestingly, the diet administered by the UW study contained a much higher sucrose content (28.5% sucrose by weight), compared to that of the NIA diet (3.9% sucrose by weight). This raises the intriguing possibility that myocardial fibrosis in NHPs may be driven by a high sucrose diet. Abbreviations: CR: Caloric Restriction NHP: Non-human primate mTOR: mechanistic Target of Rapamycin NIA: National Institute on Aging UW: University of Wisconsin, Madison

Preventive Effects of Long-Term Caloric Restriction on Aging Related In Vivo Bladder Dysfunction and Molecular Biological Changes in the Bladder and Dorsal Root Ganglia in Rats

We evaluated aging related bladder dysfunctions and biological changes in the bladder and dorsal root ganglia in rats. We also investigated whether long-term caloric restriction may have preventive effects on these changes. Male Fischer 344 rats were divided into a young group (age 6 months) and an old group (age 25 to 28 months), each with free access to normal food, and an old group (age 25 to 28 months) with food restricted to 3 days per week. Conscious cystometry, cDNA microarray analysis, immunohistochemistry and oxidative stress measurements of the bladder and dorsal root ganglia were performed. The old group with free access to normal food showed higher threshold pressure, more nonvoiding contractions and lower bladder compliance than the young group with free access to food. Old rats with free access showed greater post-void residual volume and lower voiding efficiency than old rats with caloric restriction and young rats. In the old group with free access 83 genes in the bladder and 48 in the L6 dorsal root ganglia were up-regulated compared with old rats with caloric restriction and young rats. These genes were mostly related to immune and inflammatory responses. Immunohistochemistry showed stronger expression of the immune response protease Gzm (granzyme) B and the collagenase Mmp13 (matrix metalloproteinase-13) in the bladder of old rats with free access vs old rats with caloric restriction and young rats. The level of malondialdehyde, an oxidative stress marker, was higher in the bladder of old rats with free access than in young rats but there was no difference between old rats with caloric restriction and young rats with free access to food. In rats aging leads to storage and voiding dysfunctions associated with immune and inflammatory related responses in the bladder and dorsal root ganglia, and with increased oxidative stress in the bladder. Caloric restriction reduced these aging related changes.

Caloric restriction may reverse age‐related autonomic decline in humans

Caloric restriction (CR) retards aging in laboratory rodents. No information is available on the effects of long-term CR on physiologic markers of aging and longevity in humans. Heart rate variability (HRV) is a marker for cardiac autonomic functioning. The progressive decline in HRV with aging and the association of higher HRV with better health outcomes are well established. Heart rate variability assessment is a reliable tool by which the effects of CR on autonomic function can be assessed. Time- and frequency-domain analyses compared 24-h HRV in 22 CR individuals aged 35-82 years and 20 age-matched controls eating Western diets (WD). The CR group was significantly leaner than the WD group. Heart rate was significantly lower, and virtually, all HRV values were significantly higher in the CR group than in the WD group (P < 0.002). Heart rate variability in the CR individuals was comparable with published norms for healthy individuals 20 years younger. In addition, when differences in heart rate (HR) and HRV between CR and WD were compared with previously published changes in HRV induced in healthy adults given atenolol, percent differences in each measure were generally similar in direction and magnitude and suggested declines in sympathetic and increases in parasympathetic modulation of HR and increased circadian variability associated with CR. These findings provide evidence that CR has direct systemic effects that counter the expected age-associated changes in autonomic function so that HRV indexes in CR individuals are similar to those of individuals 20 years younger eating WDs.

Sexual dimorphism in cardiac triacylglyceride dynamics in mice on long term caloric restriction

Human studies indicate augmented myocardial lipid metabolism in females, and that sex and obesity interact to predict myocardial fatty acid oxidation and storage. Altered lipid dynamics precede cardiomyopathies, and many studies now address high fat diets. Conversely, caloric restriction (CR), is the most studied model for longevity and stress resistance, including protection against myocardial ischemia. However, no information exists on the effects of long-term caloric restriction (CR) on triacylglyceride (TAG) content and dynamics in the heart. This study explored the effects of CR, sex and age on TAG dynamics in mouse hearts. Male and female SVJ129 mice were fed either normal (ND) or CR diet for 3 or 10months. In 5-month-old mice, CR similarly decreased cardiac TAG in males (ND: 25.5±4.5nmol/mg protein; CR: 12.6±2.7, P<0.05) and females (ND: 30.1±4.4; CR: 13.7±1.2) (no significant differences in TAG content were seen between sexes). CR reduced the contribution of exogenous palmitate to oxidative metabolism in males and females, by 15% and 11% respectively, versus ND, without affecting cardiac workload. CR also induced a larger reduction in TAG turnover in male (68%) than female hearts (38%). Interestingly, in 5month old male mice, CR reproduced the lower TAG turnover rates of middle-aged males (ND 13-month-old male=423±76nmol/mgprotein/min). Thus, long term CR reduces TAG pool dynamics. Despite reduced content, hearts of female mice subjected to CR retained a more dynamic TAG pool than males, while males respond with greater metabolic remodeling of cardiac lipid dynamics.

Adipokine Profile and Insulin Sensitivity in Formerly Obese Women Subjected to Bariatric Surgery or Diet-Induced Long-term Caloric Restriction

To better understand the contribution of the fat mass to the effects of long-term caloric restriction in humans, we compared adipokine profile and insulin sensitivity in long-term calorically restricted formerly obese women (CRW) subjected to different interventions, bariatric surgery, or reducing diet, with age- and BMI-matched obese (OW) and normal-weight women (NW) eating ad libitum. Our key findings are that despite a considerably stronger weight loss induced by bariatric surgery, both long-term caloric restriction interventions improved insulin sensitivity to the same degree and led to significantly lower retinol-binding protein-4 and interleukin-6 serum levels than in OW, suggesting that lowering of these two adipokines contributes to the improved insulin sensitivity. Moreover, serum leptin was considerably lower in CRW than in OW as well as in NW, suggesting that CRW develop hypoleptinemia.

Caloric Restriction and Aging: Studies in Mice and Monkeys

It is widely accepted that caloric restriction (CR) without malnutrition delays the onset of aging and extends lifespan in diverse animal models including yeast, worms, flies, and laboratory rodents. The mechanism underlying this phenomenon is still unknown. We have hypothesized that a reprogramming of energy metabolism is a key event in the mechanism of CR (Anderson and Weindruch 2007). Data will be presented from studies of mice on CR, the results of which lend support to this hypothesis. Effects of long-term CR (but not short-term CR) on gene expression in white adipose tissue (WAT) are overt. In mice and monkeys, a chronic 30% reduction in energy intake yields a decrease in adiposity of approximately 70%. In mouse epididymal WAT, long-term CR causes overt shifts in the gene expression profile: CR increases the expression of genes involved in energy metabolism (Higami et al. 2004), and it down-regulates the expression of more than 50 pro-inflammatory genes (Higami et al. 2006). Whether aging retardation occurs in primates on CR is unknown. We have been investigating this issue in rhesus monkeys subjected to CR since 1989 and will discuss the current status of this project. A new finding from this project is that CR reduces the rate of age-associated muscle loss (sarcopenia) in monkeys (Colman et al. 2008).

Caloric restriction and brain function

In addition to extending lifespan, animal research shows that specific diets benefit brain functioning. Indeed, it has been proven that caloric restriction prevents age-related neuronal damage. What are those mechanisms involved in the effects of caloric restriction on brain functioning? Could caloric restriction be proposed in the future to prevent or treat neurodegenerative disorders such as Alzheimer's disease? Is there a future for caloric restriction interventions in adults? Hypotheses linking caloric restriction to cognitive capability include anti-inflammatory mechanisms, reduction of neural oxidative stress, promotion of synaptic plasticity, induction of various stress and neurotrophic/neuroprotective factors. Caloric restriction may also prevent beta-amyloid neuropathology in Alzheimer transgenic models. Finally, both exercise and caloric restriction enhance neurogenesis via different mechanisms suggesting that their combination may decrease the risk of neurodegenerative disease. It is now well established that caloric restriction could be used to promote successful brain aging. Data from randomized controlled trials in humans are limited. No positive effect on cognitive impairment was found probably due to methodological limitations. The long-term effects of caloric restriction in adults must be clarified before engaging in such preventive strategy. Additional animal studies must be conducted in the future to test the effects of 'multidomain' interventions (caloric restriction plus regular exercise) on age-related cognitive decline.

Long-Term Effects of Caloric Restriction or Exercise on DNA and RNA Oxidation Levels in White Blood Cells and Urine in Humans

Excessive adiposity is associated with increased oxidative stress and accelerated aging. Weight loss induced by negative energy balance reduces markers of oxidation in experimental animals and humans. The long-term effects of weight loss induced by calorie restriction or increased energy expenditure induced by exercise on measures of oxidative stress and damage have not been studied in humans. The objective of the present study was to compare the effects of 20% caloric restriction or 20% exercise alone over 1 year on oxidative damage to DNA and RNA, as assessed through white blood cell and urine analyses. Eighteen men and women aged 50 to 60 years with a body mass index (BMI) between 23.5 to 29.9 kg/m(2) were assigned to one of two conditions--20% CR (n = 9) or 20% EX (n = 9)--which was designed to produce an identical energy deficit through increased energy expenditure. Compared to baseline, both interventions significantly reduced oxidative damage to both DNA (48.5% and 49.6% reduction for the CR and EX groups, respectively) and RNA (35.7% and 52.1% reduction for the CR and EX groups, respectively) measured in white blood cells. However, urinary levels of DNA and RNA oxidation products did not differ from baseline values following either 12-month intervention program. Data from the present study provide evidence that negative energy balances induced through either CR or EX result in substantial and similar improvements in markers of DNA and RNA damage to white blood cells, potentially by reducing systemic oxidative stress.

Long–term effects of caloric restriction on total and resting energy expenditure in healthy adults

The effects of long‐term caloric restriction (CR) on resting metabolic rate (RMR) and total energy expenditure (TEE) in humans is uncertain. Objective To examine the effects of a 30% CR regimen on TEE and RMR. Methods One year randomized controlled trial of 30% CR in 29 healthy overweight adults (mean±SD, age 35±5y; BMI 27.5±1.5 kg/m2). All food was provided for 6 mo, and self‐administered for the next 6 mo. Outcomes included TEE, RMR, and body fat and fat free mass (FFM). Results Absolute values for RMR decreased during the intervention compared to baseline but measured RMR during CR was not significantly different from RMR predicted from body composition, and RMR changes over time assessed at 3, 6 and 12 mo were not statistically significant after accounting for decreases in body fat and FFM. TEE also decreased significantly with CR but at 12 mo there was a greater reduction in TEE than that anticipated due to the loss of FFM and fat mass (P=0.018). Conclusions This study found no evidence for long‐term metabolic adaptation to weight loss in assessments of RMR, but an adaptive reduction in TEE was observed, which may serve to protect the body against weight loss and contribute to weight regain.Support: NIH grants: NGA‐3U01‐AG20480&amp; #H150001; USDA No. 58‐1950‐4‐401.

The Ischemic Heart

Caloric restriction has emerged as an effective strategy for lengthening lifespan in a variety of species.1 In mammals, one mechanism for this phenomenon may be the prevention of detrimental age-related alterations in cellular function1 and presumably subsequent improvement in organ function. The effects of caloric restriction on the heart, at least in rats and mice, involve a number of changes in gene expression that are beneficial to the aged cardiomyocyte2 and/or protect the heart from ischemic injury.3 Although it is likely that all of the beneficial mediators of caloric restriction have not been identified, a number of proteins in the mammalian sirtuin family may play key roles in the regulation of health and longevity.4 In addition, recent evidence has suggested that alterations in whole-body energy metabolism contribute to the beneficial effects of caloric restriction.5 Indeed, caloric restriction in mammals leads to loss of adipose tissue and dramatically alters the action of this endocrine organ.6 As such, caloric restriction contributes to changes in adipose tissue–derived hormone (adipokine) secretion, which can govern whole-body metabolism.7 Furthermore, studies using isolated cardiac myocytes suggest that these adipokines may exert direct end-organ effects that are independent from alterations in whole-body metabolism.8–10 One adipokine that is significantly increased during caloric restriction is adiponectin.11 Previous work has shown that adiponectin exerts a host of protective effects on the cardiovascular system12 and as such may be an essential component mediating the effects of caloric restriction. Article p 2809 The focus on adiponectin in the cardiovascular system has been due largely to the fact that in humans, circulating adiponectin levels are negatively correlated with increased body mass index.13 Because increased body mass index is associated with a number of obesity-linked disorders, including cardiovascular disease, the reduction in serum …

Life-Long Caloric Restriction Reveals Biphasic and Dimorphic Effects on Bone Metabolism in Rodents

Caloric restriction (CR) extends the lifespan of various organisms and slows the onset of age-related disorders; however, little is known about the long-term effects of CR per se on bone. In the present study, we have examined the effects of life-long CR vs. ad libitum (AD) feeding, mainly on the trabecular bone of proximal tibiae in male C57BL/6 mice and F344 rats. Micro-computed tomography scanning of tibiae revealed that CR for 3-9 months caused a substantial decrease in three-dimensional bone volume with structural derangements. Bone histomorphometry revealed the reduced bone mass was due mainly to suppression of bone formation. In db/db mice with defective leptin receptor, CR was unable to decrease bone mass and suppress bone formation. The effect of CR on bone mass was inhibited by administration of a beta-adrenergic blocker, propranolol. Thus, CR may regulate bone formation through leptin signaling and elevated sympathetic nervous tone. Interestingly, the difference in bone volume between the CR and AD groups disappeared after 1 yr of age, and mice and rats on an additional extension of CR to natural death maintained higher bone mass than the AD groups, with reduced bone turnover, suggesting that CR slows skeletal aging by regulating the rate of bone turnover. This is the first report, to our knowledge, that has examined the effects of lifelong CR on bone metabolism and trabecular microstructure and documents its contrasting effects during maturation vs. the postmaturational, involutional period.

Effect of long-term caloric restriction on oxygen consumption and body temperature in two different strains of mice

The hypothesis, that a decrease in metabolic rate mediates the life span prolonging effect of caloric restriction (CR), was tested using two strains of mice, one of which, C57BL/6, exhibits life span extension as a result of CR, while the other, DBA/2, shows little or no effect. Comparisons of the rate of resting oxygen consumption and body temperature were made between the strains after they were fed ad libitum (AL) or maintained under 40% CR, from 4 to 16 months of age. Ad libitum-fed mice of the two strains weighed the same when young and consumed similar amounts of food throughout the experiment; however, the C57BL/6 mice weighed 25% more than DBA/2 mice at 15 months of age. The rate of oxygen consumption was normalized as per gram body weight, lean body mass or organ weight as well as per animal. The body temperature and the rate of oxygen consumption, expressed according to all of the four criteria, were decreased in the DBA/2 mice following CR. The C57BL/6 mice also showed a CR-related decrease in body temperature and in the rate of oxygen consumption per animal and when normalized according to lean body mass or organ weight. The results of this study indicate that CR indeed lowers the rate of metabolism; however, this effect by CR does not necessarily entail the prolongation of the life span of mice.

Long-Term Caloric Restriction Ameliorates the Decline in Diastolic Function in Humans

We determined whether caloric restriction (CR) has cardiac-specific effects that attenuate the established aging-associated impairments in diastolic function (DF). Caloric restriction retards the aging process in small mammals; however, no information is available on the effects of long-term CR on human aging. In healthy individuals, Doppler echocardiography has established the pattern of aging-associated DF impairment, whereas little change is observed in systolic function (SF). Diastolic function was assessed in 25 subjects (age 53 +/- 12 years) practicing CR for 6.5 +/- 4.6 years and 25 age- and gender-matched control subjects consuming Western diets. Diastolic function was quantified by transmitral flow, Doppler tissue imaging, and model-based image processing (MBIP) of E waves. C-reactive protein (CRP), tumor necrosis factor-alpha (TNF-alpha), and transforming growth factor-beta1 (TGF-beta1) were also measured. No difference in SF was observed between groups; however, standard transmitral Doppler flow DF indexes of the CR group were similar to those of younger individuals, and MBIP-based, flow-derived DF indexes, reflecting chamber viscoelasticity and stiffness, were significantly lower than in control subjects. Blood pressure, serum CRP, TNF-alpha, and TGF-beta(1) levels were significantly lower in the CR group (102 +/- 10/61 +/- 7 mm Hg, 0.3 +/- 0.3 mg/l, 0.8 +/- 0.5 pg/ml, 29.4 +/- 6.9 ng/ml, respectively) compared with the Western diet group (131 +/- 11/83 +/- 6 mm Hg, 1.9 +/- 2.8 mg/l, 1.5 +/- 1.0 pg/ml, 35.4 +/- 7.1 ng/ml, respectively). Caloric restriction has cardiac-specific effects that ameliorate aging-associated changes in DF. These beneficial effects on cardiac function might be mediated by the effect of CR on blood pressure, systemic inflammation, and myocardial fibrosis.

Long‐term caloric restriction abrogates the age‐related decline in skeletal muscle aerobic function

The purpose of this study was to determine the effect of long-term caloric restriction (CR) on the age-associated decline of skeletal muscle aerobic function. Skeletal muscle maximal aerobic performance (VO2max) was assessed in ad libitum (AL) and CR rats aged 8-10 months and 35 months using a pump-perfused hindlimb model to match oxygen delivery to muscle mass between groups. Whereas there was a 46% decline in muscle mass-specific VO2max between 8-10 mo (524+/-13 micromol x min(-1) x 100 g(-1); mean+/- SE) and 35 mo (281+/-54 micromol x min(-1) x 100 g(-1)) in AL rats, not only did CR rats begin at the same point in 8-10 mo old rats (490+/-42 micromol x min(-1) x 100 g(-1)), we found no decline in 35 mo old CR animals (484+/-49 micromol x min(-1) x 100 g(-1)). Interestingly, although most markers of oxidative capacity began at a lower point in young adult CR animals, CR rats exhibited a higher in situ activity of complex IV at VO2max. This activity allows the young adult CR animals to exhibit normal aerobic capacity despite the lower oxidative enzyme activities. In stark contrast to the 19-41% decline in activities of citrate synthase, complexes I-III, and complex IV in homogenates prepared from the plantaris muscle and mixed region of gastrocnemius muscle with aging in AL rats, no age-related decline was found in CR animals. Thus, our results showed that CR preserves aerobic function in aged skeletal muscles by facilitating a higher in situ function of complex IV and by preventing the age-related decline in mitochondrial oxidative capacity.