Ritchie J. Feuers

Effect of chronic caloric restriction on physiological variables related to energy metabolism in the male Fischer 344 rat

In the present study, a number of physiological and behavioral measures that are related to metabolism were continuously monitored in 19-month-old male Fischer 344 rats that were fed ad libitum or fed a caloric restricted diet. Caloric restricted rats ate fewer meals but consumed more food during each meal and spent more time eating per meal than did rats fed ad libitum. Therefore, the timing and duration of meals as well as the total number of calories consumed may be associated with life extension. Average body temperature per day was significantly lower in restricted rats but body temperature range per day and motor activity were higher in restricted rats. Dramatic changes in respiratory quotient, indicating rapid changes in metabolic pathway and lower temperature, occurred in caloric restricted rats when carbohydrate reserves were depleted. Lower body temperature and metabolism during this time interval may result in less DNA damage, thereby increasing the survival potential of restricted rats. Nighttime feeding was found to synchronize physiological performance between ad libitum and caloric restricted rats better than daytime feeding, thereby allowing investigators to distinguish the effects of caloric restriction from those related solely to the time-of-day of feeding.

Effect of chronic caloric restriction on hepatic enzymes of intermediary metabolism in the male Fischer 344 rat

It is well established that caloric restriction extends life span and significantly retards the rate of occurrence of most age-associated degenerative disease processes. A paucity of data exists relative to the mechanisms by which caloric restriction accomplishes these events. We have examined the effect of caloric restriction in rats on several hepatic enzymes of intermediary metabolism. The activities of glycolytic and supporting enzymes including lactate dehydrogenase, pyruvate kinase, sorbitol dehydrogenase, and alcohol dehydrogenase were all decreased in response to caloric restriction. Fructose 1-phosphate aldolase and creatine phosphokinase were not altered. Likewise, enzymes associated with lipid metabolism (malic enzyme and glycerokinase) were reduced (fatty acid synthetase was reduced, but not to a statistically significant degree). Activities of enzymes supporting gluconeogenesis (glutamate oxaloacetate transaminase, tyrosine aminotransferase, glutamate pyruvate transaminase, glutamate dehydrogenase, amino acid oxidase, malate dehydrogenase, and glucose 6-phosphatase) were either unchanged or increased significantly by caloric restriction. Glucagon levels were decreased. Comparisons between young ad libitum fed and older calorically restricted rats revealed similar but not identical metabolic activity. These results suggest that caloric restriction produces an effect on intermediary metabolism, favoring the role of glucagon and glucose synthesis; but limiting the role of insulin and glucose catabolism in the liver. The former observation provides for the efficient support of peripheral tissues and the latter a level of energy production necessary only for self maintenance. Limited lipid metabolism suggests decreased potential for fatty acid epoxide formation and free radical damage to cellular macromolecules. Additionally, caloric restriction may delay the progressive age associated changes in the activities of some of the enzymes investigated.

MPP+-induced neurotoxicity in mouse is age-dependent: evidenced by the selective inhibition of complexes of electron transport

1-Methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP), has been demonstrated to cause selective neurotoxicity by inhibiting complex I in mitochondria, through its toxic metabolite 1-methyl-4-phenylpyridine (MPP+) which is formed during the bioactivation of MPTP by monoamine oxidase B. In this report, we have evaluated the effect of MPP+ on the 4 mitochondrial respiratory chain complexes by incubating brain mitochondria of mice at 3 different age groups with MPP+ (200 microM) and monitoring enzyme activities of complexes I, II, III, and IV at 5, 10, 15, 30, 60, and 120 min. Complexes I, III, and IV showed significant inhibition within 15 min in all the age groups studied, followed by some recovery in enzyme activities upon further incubation for complexes I and IV. However, complex II was not affected by MPP+ at any age. Our data suggest that inhibition of complexes I, III, and IV by MPP+ efficiently restrict the transport of electrons down the respiratory chain which ultimately leads to decreased ATP production. This could further aggravate oxidative stress as ATP is required for the synthesis of glutathione (GSH), one of the important scavengers of free radicals. In this study, inhibition was more severe in mitochondrial preparations from older rather than younger mice. Additionally, young animals showed faster recovery following inhibition than old animals for complex I. Impaired respiratory chain function in older animals compared to younger ones supports the hypothesis of accumulation of age-related mitochondrial DNA mutations which partly encode for subunits of complexes I, III, and IV. From this study, it seems that inhibition of complexes I, III, and IV may be the underlying cause of neurotoxicity due to MPP+ which could be intensified by age-associated dysfunction of electron transport.

Caloric restriction, aging, and antioxidant enzymes

The basic mechanisms of aging and its retardation by caloric restriction (CR) remain unclear. One suggested means by which CR could retard aging is based on production of mitochondrial free radicals, and efficiency of their subsequent metabolism. Currently, there is little information concerning the influences of age and CR on the rates of in vivo mitochondrial free radical production. However, evidence for CR-induced modulation of free radical detoxification capacities is mounting. The direction of the influence of CR on free radical detoxification is tissue-specific. These effects are broad and appear to provide positive advantage.

The Effects of Dietary Restriction on Mitochondrial Dysfunction in Aginga

ABSTRACT: Age‐associated alterations in the mitochondrial electron transport system (ETS) may lead to free radical generation and contribute to aging. The complexes of the ETS were screened spectrophotometrically in gastrocnemius of young (10 month) as well as older (20 and 26 month) B6C3F1 female mice fed an ad libitum (AL) diet or a restricted (DR) in total calories diet (40% less food than AL mice). The activities of complexes I, III, and IV decreased significantly by 62%, 54%, and 74%, respectively, in old AL mice (AL20) compared to young AL mice (AL10). Complexes I, III, and IV from DR10 mice had activities that were significantly lower than those seen in AL10 mice (suggesting a lower total respiratory rate or improved efficiency). By contrast, complex II activity did not decrease with age (actually increased, but not significantly) in AL20 mice. Complex II was decreased across age in DR mice. Km for ubiquinol‐2 of complex III was significantly increased in AL10 animals (0.33 mM vs. 0.26 mM in DR10 mice) and was further increased with aging (0.44 mM in AL20 vs. 0.17 mM in DR20 mice). This suggests obstruction of binding, inhibition of electron flow in aging, which could yield premature product release as a free radical. Total complex IV by Vmax was highest in AL10 mice, but the proportion of complex as high‐affinity sites was lower (69%) than in either DR10 (80%) or DR20 (80%). The percentage of high‐affinity sites decreased to only 45% in AL20 mice, and Vmax was reduced by 75 percent. In AL26 mice high‐affinity sites decreased to 33 percent. At physiologic concentration of reduced cytochrome c, significant dysfunction of complex IV in AL20 or AL26 mice would be expected with obstruction of overall electron transport. The age‐associated loss of activity and function of complexes I, III, and IV may contribute to increased free radical production. Lack of sufficient DNA repair in mitochondria and juxtaposition to the ETS adds to susceptibility and accumulation of mtDNA and other mitochondrial macromolecular damage. DR seems to retard this deterioration of mitochondrial respiratory function by preserving enzymatic activities and function.

Chronic caloric restriction induces stress proteins in the hypothalamus of rats

The induction of stress proteins (sps) in the hypothalamus of female Fischer 344 rats in response to caloric restriction (CR) and to heat stress was investigated. Caloric restriction was found to elicit sps 27, 34, 70, and 90 in the hypothalamus of both young and old rats while none was found in the hypothalamus of ad libitum (AL) fed controls. Heat stress initiated heat shock proteins (hsps/sps) 27, 70, and 90 in the hypothalamus of the young (AL) fed animals, the same proteins evoked by feeding stress. The same sps were induced in the old (AL) rats although the expression showed substantial decline with age. This reduction was less marked, however, with the old CR rats. Stress protein 34, an infrequently reported protein, was related to feeding and was not induced by heat shock. Recent reports point to the important role sps play in the cellular reaction to stress, as well as their involvement in the higher functions. The findings reported here suggest that sps are involved in the regulatory mechanisms allowing CR animals to tolerate stress related to metabolic substrate deprivation.

The effects of different levels of dietary restriction on aging and survival in the Sprague-Dawley rat: Implications for chronic studies

A study was undertaken to determine the effects of incremental levels of dietary restriction (DR) in rats. Survival, growth, reproductive, and dietary intake (DI) variables were monitored in a chronic study in which male Sprague Dawley (SD) rats (NCTR colony) were fed their ration ad libitum (AL), or DR. The main objectives were to determine if low levels of DR could be used to increase the survival rate of SD rats in the chronic bioassay, and to identify the survival characteristics of a long-lived SD rat strain (NCTR colony). The average life span of AL rats was 115 months. At 104 weeks on study (110 weeks of age), the survival rate for the AL and 10%, 25%, and 40% DR groups was 63.4, 87.5, 87.5, and 97.5%, respectively. The largest increase in survival (24.1%) occurred between AL and 10% DR, indicating that very low levels of DR have a significant effect on survival. Whole-body, liver, prostate, and epididymis weights and body length were decreased by DR, whereas brain weight, testicular weight, and skull length were not altered by DR. Rats from the NCTR colony were found to be ideal for chronic studies because they are much longer-lived than other SD stocks. Although the 104-week survival rate for these SD, non-obese AL rats exceeds the FDA’s “Redbook” survival guideline (> 50%) for chronic bioassays, the use of DR is advocated because it reduces individual variability in body weight.

Effects of caloric restriction on rodent drug and carcinogen metabolizing enzymes: implications for mutagenesis and cancer

Caloric restriction in rodents results in increased longevity and a decreased rate of spontaneous and chemically induced neoplasia. The low rates of spontaneous neoplasia and other pathologies have made calorically restricted rodents attractive for use in chronic bioassays. However, caloric restriction also alters hepatic drug metabolizing enzyme (DME) expression and so may also alter the biotransformation rates of test chemicals. These alterations in DME expression may be divided into two types: (1) those that are the direct result of caloric restriction itself and are detectable from shortly after the restriction is initiated; (2) those which are the result of pathological conditions that are delayed by caloric restriction. These latter alterations do not usually become apparent until late in the life of the organism. In rats, the largest direct effect of caloric restriction on liver DMEs is an apparent de-differentiation of sex-specific enzyme expression. This includes a 40-70% decrease in cytochrome P450 2C11 (CYP2C11) expression in males and a 20-30% reduction of corticosterone sulfotransferase activity in females. Changes in DME activities that occur late in life in calorically restricted rats include a stimulation of CYP2E1-dependent 4-nitrophenol hydroxylase activity and a delay in the disappearance of male-specific enzyme activities in senescent males. It is probable that altered DME expression is associated with altered metabolic activation of chemical carcinogens. For example the relative expression of hepatic CYP2C11 in ad libitum-fed or calorically restricted rats of different ages is closely correlated with the amount of genetic damage in 2-acetylaminofluorene- or aflatoxin B1-pretreated hepatocytes isolated from rats of the same age and caloric intake. This suggests that altered hepatic drug and carcinogen metabolism in calorically restricted rats can influence the carcinogenicity of test chemicals.

Chronic Caloric Restriction in Old Female Mice: Changes in the Circadian Rhythms of Physiological and Behavioral Variables

One of the most frequently reported observations in the gerontological literature is the significant increase in life span (Sacher 1977; McCay et al. 1935) and the prevention or delayed onset of various types of chronic diseases (Walford et al. 1974; Maeda et al. 1985) associated with chronic caloric restriction (CR). CR has also been shown to be effective in inhibiting various types of spontaneous tumors (Sarkar et al. 1985) and chemically induced lesions (Ruggeri et al. 1987; Kritchevsky et al. 1984) in rats and mice. However, little is known about the primary mechanisms by which CR interacts with environmental factors to alter the timing of the biological clock. One hypothesis that may account for the slowing down of age-related physiological and disease processes is that the synchronization of organisms to their surroundings is controlled or modified by qualitative or quantitative changes in nutritional parameters and that changes in 24-h (circadian) rhythms that are a direct result of CR may ultimately alter primary mechanisms of aging. Several studies in which total calories were restricted to 76% of ad libitum (AL) by limiting access to food to a few hours daily (single meal feeding) (Nelson et al. 1975; Philippens et al. 1977) or when intake was decreased by reducing the total weight of the food pellets fed to the caloric restricted group to 60% of the AL level (Duffy et al. 1989a; McCarter et al. 1985) have reported significant alterations in the circadian rhythms of various physiological and biochemical parameters. However, in other studies, no significant difference in longevity was found in CR mice fed a single meal during the early light phase, a single meal during the dark phase, or fed multiple meals (six times) during the dark phase (Nelson and Halberg 1986a). While CR in itself was found to prolong life, the added imposition of frequent photoperiod schedule shifting had no statistically significant effect on mean survival time, regardless of whether the meal schedule reinforced or opposed shifts in the photoperiod (Nelson and Halberg 1986b). This may mean that varying the dietary regimen may have no effect on longevity if the total amount of food consumed is maintained at a constant restricted calorie level.

Influence of body fat distribution on oxygen uptake and pulmonary performance in morbidly obese females during exercise

Objective: The aim of this study was to determine the effects of fat distribution on aerobic and ventilatory response to exercise testing in morbidly obese (MO) females.