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Keywords:

  • aging;
  • behavior;
  • dietary restriction;
  • Drosophila;
  • functional senescence;
  • lifespan;
  • negative geotaxis;
  • olfactory

Summary

  1. Top of page
  2. Summary
  3. Introduction
  4. Results
  5. Discussion
  6. Experimental procedures
  7. Acknowledgments
  8. References

Dietary restriction extends lifespan substantially in numerous species including Drosophila. However, it is unclear whether dietary restriction in flies impacts age-related functional declines in conjunction with its effects on lifespan. Here, we address this issue by assessing the effect of dietary restriction on lifespan and behavioral senescence in two wild-type strains, in our standard white laboratory stock, and in short-lived flies with reduced expression of superoxide dismutase 2. As expected, dietary restriction extended lifespan in all of these strains. The effect of dietary restriction on lifespan varied with genetic background, ranging from 40 to 90% extension of median lifespan in the seven strains tested. Interestingly, despite its robust positive effects on lifespan, dietary restriction had no substantive effects on senescence of behavior in any of the strains in our studies. Our results suggest that dietary restriction does not have a global impact on aging in Drosophila and support the hypothesis that lifespan and behavioral senescence are not driven by identical mechanisms.


Introduction

  1. Top of page
  2. Summary
  3. Introduction
  4. Results
  5. Discussion
  6. Experimental procedures
  7. Acknowledgments
  8. References

Aging is a complex, multifaceted process that manifests as a progressive reduction in functional capacity and an increased probability of dying. Interventions that ameliorate the negative consequences of aging would be extremely valuable. Arguably the most widely accepted intervention that extends lifespan is dietary restriction, a manipulation that reduces total nutrient intake without leading to a malnourished state (Partridge et al., 2005b).

Dietary restriction extends lifespan in numerous species (Merry, 2005; Partridge et al., 2005b; Piper et al., 2005a; Pletcher et al., 2005). Dietary restriction also mitigates many age-related functional declines and delays the onset or lessens the severity of various pathological states in rodents. For example, rats maintained on a restricted diet exhibit attenuated age-related learning and motor impairments (Ingram et al., 1987; Stewart et al., 1989) and reduced incidence of cancer (Mattson, 2005). Dietary restriction, however, does not positively affect senescence of all functions in rodents (Campbell & Gaddy, 1987; Bond et al., 1989) and can actually accelerate age-related declines in oxidative stress and starvation stress in Drosophila (Burger et al., 2007). These studies raise the possibility that dietary restriction might not positively impact all aspects of aging.

Here, we report studies that address whether dietary restriction positively impacts behavioral senescence in conjunction with its effects on lifespan. We find that while dietary restriction significantly extends lifespan, it does not substantially influence age-related declines in locomotor or olfactory behavior. Our studies suggest that dietary restriction does not alter all aspects of aging equally and support the hypothesis that distinct mechanisms influence different facets of aging in flies.

Results

  1. Top of page
  2. Summary
  3. Introduction
  4. Results
  5. Discussion
  6. Experimental procedures
  7. Acknowledgments
  8. References

Dietary restriction in Drosophila is typically achieved by reducing the concentration of sucrose and yeast in the food medium (Mair et al., 2005; Partridge et al., 2005a; Piper et al., 2005a; Min & Tatar, 2006). We used this approach for restricting the diet of flies and focused on females in our studies because the effect of dietary restriction on lifespan appears to be larger in this sex (Magwere et al., 2004). To determine whether genetic background influences the effect of dietary restriction on lifespan, we assessed survival of our standard laboratory stock (w[cs]) and two wild-type strains (Oregon-R and Canton-S) when maintained on food medium with a range of sugar and yeast (SY) concentrations. Survival of females from all three strains was significantly enhanced at low SY concentrations as compared to flies on high SY food (Fig. 1). Dietary restriction extended mean and median lifespan by 38–40% in w[cs] and Oregon-R, while it increased mean and median lifespan by 80–90% in Canton-S. Genetic background therefore significantly impacts the magnitude of the lifespan extension seen in Drosophila.

image

Figure 1. Dietary restriction extends lifespan in standard laboratory strains. Survival of w[cs] (A), Oregon-R (B) and Canton-S (C) adult females housed on different sugar-yeast (SY) media. Concentrations of sugar and yeast are shown as percentages. In all cases, flies maintained on 5% SY medium were longer lived than flies on higher percentage media (P < 0.0001, log-rank test). Data are representative of two or more independent experiments with 150–300 adults per group.

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Dietary restriction extends lifespan in numerous species and also forestalls the onset of at least some age-related pathophysiological states in rodents (Mattson, 2005). The effect of dietary restriction on behavioral senescence in Drosophila, however, has not been vigorously investigated. To determine whether dietary restriction positively impacts age-related behavioral declines in flies, we assessed negative geotaxis (a locomotor behavior) and odor avoidance (an olfactory behavior) across age in w[cs], Oregon-R and Canton-S flies maintained on food with a range of SY concentrations. Consistent with previous reports (Cook-Wiens & Grotewiel, 2002; Goddeeris et al., 2003; Gargano et al., 2005), negative geotaxis (Fig. 2A–C) and odor avoidance (Fig. 2D) declined with age. Negative geotaxis performance across age in w[cs] and Oregon-R (Fig. 2A,B) was lowest in the 25% SY flies, whereas diet had no measurable effect on this behavior in Canton-S flies (Fig. 2C). Diet also had no demonstrable effect on odor avoidance across age in w[cs] flies (Fig. 2D). In contrast to its robust enhancement of survival (Fig. 1), dietary restriction had modest effects on negative geotaxis and odor avoidance measured across age.

image

Figure 2. Diet does not substantially alter behavioral senescence in standard laboratory flies. Senescence of negative geotaxis (A–C) and odor avoidance (D) in flies maintained on different sugar-yeast (SY) media. Concentrations of sugar and yeast are indicated as percentages. Overall, there was a significant effect of age on negative geotaxis [two-way analysis of variance (anova), P < 0.0001, n = 10] in w[cs] (A), Oregon-R (B) and Canton-S flies (C). Diet significantly affected negative geotaxis measured across age (P ≤ 0.002) in w[cs] and Oregon-R, but not Canton-S (ns, non-significant). Tukey's honestly significant difference (HSD) tests indicated that negative geotaxis in w[cs] and Oregon-R flies was reduced on 25% relative to 5% SY medium (P < 0.05). (D) Senescence of olfaction [measured as avoidance of methylcyclohexanol (MCH)] in w[cs] flies declined significantly with age (two-way anova, P ≤ 0.0001, n = 20), but was not affected by SY media (ns). Data (mean ± SEM) are compiled from two to three independent experiments.

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Although the effect of dietary restriction on negative geotaxis and odor avoidance across age appeared to be minimal, we postulated that a more detailed analysis of the data might reveal a consistent effect of dietary restriction on specific parameters related to behavioral senescence. We addressed this issue by determining total behavior across age (measured as the area under the curve) and decline times (DT, the time required for the behaviors to decline to prescribed levels) (Martin et al., 2005; Martin & Grotewiel, 2006) in flies subjected to dietary restriction. For negative geotaxis, flies maintained on 5% SY medium tended to have enhanced total behavior and extended DTs (Supplementary Fig. S1A–F). These effects, however, were small and inconsistently observed. Dietary restriction also tended to increase total odor avoidance, but again this effect was not statistically significant (Supplementary Fig. S1G). These analyses indicate that although there is a trend for dietary restriction to increase total behavior across age and extend DTs for behavior in standard laboratory flies, these effects are small and inconsistently observed.

The w[cs], Canton-S and Oregon-R strains have maximal lifespans of ~60–100 days (Fig. 1) and exhibit negative geotaxis behavior for the first several weeks of adulthood (Fig. 2). Because we observed no substantive effect of diet on behavioral senescence in these control strains, we thought it possible that dietary restriction might have more pronounced effects on behavioral senescence in shorter-lived flies with accelerated age-related decline in negative geotaxis. We explored this possibility by assessing senescence of negative geotaxis in flies with Gal4-driven expression of an inverted repeat (IR) transgene against superoxide dismutase 2 (SOD2), a major mitochondrial antioxidant enzyme (Landis & Tower, 2005). Ubiquitous expression of Sod2-IR transgenes induces RNA interference (RNAi) for this locus, and thereby severely curtails SOD2 expression, lifespan and resistance to oxidative stress (Kirby et al., 2002). We confirmed that lifespan was dramatically shortened by ubiquitous expression of Sod2-IR driven by da-Gal4 (Fig. 3A) and found that these flies also had greatly accelerated age-related decline of negative geotaxis (Fig. 3B).

image

Figure 3. Ubiquitous expression of Sod2-IR shortens lifespan and accelerates age-related decline in negative geotaxis. (A) Mean and median lifespan in flies expressing Sod2-IR via da-Gal4 (da-Gal4; Sod2-IR) are reduced by 75 and 85%, respectively. (B) Age-related decline in negative geotaxis is accelerated in flies expressing Sod2-IR via da-Gal4 [analysis of variance (anova), P < 0.0001, n = 5 vials of 25 adult females per genotype].

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Lifespan was sensitive to SY concentration in short-lived flies ubiquitously expressing Sod2-IR transgenes driven by da-Gal4 (Fig. 4A–B) or Actin5c-Gal4 (Fig. 4C–D). Mean lifespan was maximized at 15% SY when Sod2-IR was driven by da-Gal4 (Fig. 4E) and at 5% SY when Sod2-IR was driven by Actin5c-Gal4 (Fig. 4F). Although we do not currently understand why lifespan was maximized at different food concentrations in flies expressing Sod2-IR via da-Gal4 and Actin-Gal4, it is possible that this is due to differences in the magnitude of Gal4 expression levels or differences in the temporal or spatial patterns of Gal4 expression. Importantly, however, these results are consistent with the expected effects of dietary restriction on lifespan.

image

Figure 4. Dietary restriction enhances survival of short-lived flies expressing Sod2-RNAi. (A–D) Survival of flies expressing Sod2-inverted repeat (IR) transgenes on a range of sugar-yeast (SY) media. For clarity, survival data beyond 20 days are not shown. SY medium had a significant effect on survival (log-rank test, P ≤ 0.0001) when either of two Sod-IR transgenes (IR-15, A and C; IR-24, B and D) were driven by da-Gal4 (A and B) or Actin5c-Gal4 (C and D). Data are representative of two independent experiments with each Gal4-Sod2-IR combination using 100–200 flies in each group. Mean lifespan varied with SY medium in flies expressing Sod2-IR transgenes via da-Gal4 (E) or Actin5c-Gal4 (F).

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Senescence of negative geotaxis in the short-lived Sod2-IR flies was also somewhat sensitive to SY concentration. Interestingly, however, negative geotaxis across age was unexpectedly lower in flies maintained on the lowest percentage SY medium (i.e. the longer-lived flies) in three out of the four strains tested (Fig. 5). More detailed analyses revealed that only da-Gal4::Sod2-IR24 (Fig. 5B) and Actin5c-Gal4::Sod2-IR15 (Fig. 5C) flies had significantly reduced total negative geotaxis on the lower percentage SY medium (Supplementary Fig. S2A,C,E,G). Additionally, only the da-Gal4::Sod2-IR24 (Fig. 5B) and Actin5c-Gal4::Sod2-IR24 (Fig. 5D) flies had reduced decline times on the lower percentage SY diet (Supplementary Fig. S2B,D,F,H). In contrast to its effects on lifespan, therefore, dietary restriction appears to have subtle, negative consequences for locomotor senescence in short-lived Sod2-IR flies.

image

Figure 5. Effect of diet on senescence of negative geotaxis in short-lived flies expressing Sod2-RNAi. Senescence of negative geotaxis in flies expressing Sod2-IR15 (A and C) or Sod2-IR24 (B and D) driven by da-Gal4 (A and B) or Actin5c-Gal4 (C and D) maintained on sugar-yeast (SY) media. Negative geotaxis declined with age in all groups [two-way analysis of variance (anova), P ≤ 0.0001, n = 5] and SY medium had a significant effect on negative geotaxis measured across age (P ≤ 0.004) in all panels except A (da-Gal4; Sod-IR15). Tukey's honestly significant difference (HSD) tests indicated that negative geotaxis in flies housed on the 25% SY medium was greater overall than in flies fed the 15% (B and D) or 5% (C and D) SY media (P < 0.05). Data are mean ± SEM and are representative of two independent experiments with each group.

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Discussion

  1. Top of page
  2. Summary
  3. Introduction
  4. Results
  5. Discussion
  6. Experimental procedures
  7. Acknowledgments
  8. References

Dietary restriction extends lifespan in a number of species (Merry, 2005; Partridge et al., 2005b; Piper et al., 2005a; Pletcher et al., 2005). Whether dietary restriction impacts all aspects of aging, however, is much less clear. Here, we report the results of experiments that explored whether dietary restriction alters senescence of negative geotaxis (a locomotor behavior) and odor avoidance (an olfactory behavior). Our results indicate that dietary restriction, despite robustly extending lifespan in several independent fly strains, has minimal effects on senescence of behavior in Drosophila.

Dietary restriction extended mean lifespan in our standard laboratory and two wild-type strains. Low concentration SY food medium extended lifespan most (~80–90%) in Canton-S, while it extended lifespan ~40% in two other strains, Oregon-R and w[cs]. Dietary restriction also extended the lifespan of short-lived flies that express Sod2-IR ubiquitously. These results confirm that dietary restriction had the expected effect on lifespan in all of our studies. Furthermore, these results reinforce the need to carefully control for genetic background in studies on dietary restriction, both within a single experiment and between experiments that are interpreted collectively.

Although dietary restriction extended lifespan in standard laboratory strains and in short-lived flies ubiquitously expressing Sod2-IR, diet had minimal effects on senescence of negative geotaxis (a locomotor behavior) and odor avoidance (an olfactory behavior). Interestingly, standard laboratory flies on dietary restriction had subtle improvements in behavioral function across age, while flies with reduced Sod2 expression on dietary restriction had subtle impairments in behavioral senescence. Given that the effects of dietary restriction on behavioral senescence were (i) subtle at best, (ii) somewhat inconsistent between similar strains, and (iii) had opposing effects in standard laboratory strains and short-lived flies with reduced Sod2 expression, dietary restriction appears to have essentially no measurable effect on age-related behavioral declines in flies. It seems possible, however, that discernable effects of dietary restriction on Drosophila behavioral senescence might be revealed in studies that investigate other dietary regimens, other strains, or even other behaviors. Nevertheless, the simplest interpretation of our studies with seven different strains is that dietary restriction does not have a substantive impact on behavioral senescence in flies. This interpretation is consistent with the results of a recent study, which found that dietary restriction led to age-dependent reductions in stress resistance in flies (Burger et al., 2007). Taken together, our data and those of Burger and colleagues (2007) demonstrate that the effects of dietary restriction on age-related traits in Drosophila are quite complex. A low concentration diet can have positive effects (lifespan extension), negative consequences (stress resistance), or essentially no effect (behavioral senescence) in flies.

Our results raise several intriguing possibilities. First, our data suggest that dietary restriction does not have a global impact on aging in Drosophila. Although dietary restriction appears to positively influence several age-related functional declines in rodents (Idrobo et al., 1987; Ingram et al., 1987; Means et al., 1993; Mattson et al., 2001; Mattson, 2005; Mattson & Magnus, 2006), the positive effects of dietary restriction do not apply to all functional declines in these animals (Campbell & Gaddy, 1987; Bond et al., 1989) and dietary restriction in flies can lead to age-dependent reductions in stress resistance in Drosophila (Burger et al., 2007). Thus, it will be important to further delineate the heterogeneous effects of diet on lifespan and various aspects of functional senescence in flies and other species. Second, our data suggest that the mechanisms influencing lifespan and behavioral senescence in flies are at least somewhat distinct as suggested by behavioral aging studies in long-lived methuselah mutants (Cook-Wiens & Grotewiel, 2002). The dissociation between mechanisms that influence lifespan and functional aging is particularly striking in the present studies because dietary restriction extended lifespan up to 90% with no substantive positive effect on behavioral senescence. Importantly, however, a number of interventions that extend lifespan also delay senescence of certain behaviors in flies. Flies aged at lower temperatures have longer lifespans (Helfand & Rogina, 2000) and slower locomotor aging (Grotewiel et al., 2005). Mutations in myospheroid ameliorate senescence of negative geotaxis and extend mean lifespan (Goddeeris et al., 2003), and long-lived chico mutants (Clancy et al., 2001; Tu et al., 2002) have delayed senescence of negative geotaxis (Gargano et al., 2005; Martin & Grotewiel, 2006). Overexpression of peptide methionine sulfoxide reductase A (MSRA) also extends lifespan and delays behavioral senescence (Ruan et al., 2002). Together, these studies suggest that there are several common and some distinct mechanisms that influence lifespan and functional senescence in flies. Third, our data showing no substantial effect of diet on age-related behavioral declines, in conjunction with data demonstrating increased egg-laying on high concentration SY food medium (Piper et al., 2005b), strongly suggest that high concentration food is not toxic to flies. Therefore, the lifespan extending effects of food dilution are likely to be related to dietary restriction in the low concentration medium as opposed to toxic effects of high SY food medium.

Experimental procedures

  1. Top of page
  2. Summary
  3. Introduction
  4. Results
  5. Discussion
  6. Experimental procedures
  7. Acknowledgments
  8. References

Fly husbandry

Flies were reared to adulthood at 25 °C and 55% relative humidity under a 12-h light–dark cycle on a sugar : yeast : cornmeal : agar medium (10% : 2% : 3.3%: 1% w/v) supplemented with 0.2% Tegosept (Sigma Chemical Co., St. Louis, MO, USA) and active yeast. For all analyses, 1- to 3-day-old mated females were isolated with brief carbon dioxide (CO2) anesthesia and placed in groups of 25 into fresh food vials containing SY medium composed of 1.5% agar and 5–25% w/v sucrose (Sigma Chemical Co.) and yeast (Fisher Scientific, Hampton, NH, USA) throughout the duration of the experiment. SY food was autoclaved in 500 mL batches in glass beakers, cooled to 50 °C in a water bath, distributed in 10 mL aliquots into vials, plugged with cotton and kept at 4 °C until used. Canton-S flies were provided by Ron Davis (Baylor College of Medicine, Houston, TX, USA). Oregon-R and Actin5c-Gal4 were obtained from the Drosophila Stock Center (Bloomington, IN, USA). Our standard laboratory stock, w[cs] (Cook-Wiens & Grotewiel, 2002; Goddeeris et al., 2003; Gargano et al., 2005), harbors the w1118 allele backcrossed to Canton-S. The da-Gal4 and UAS-Sod2-R lines (Kirby et al., 2002) were generously supplied by John Phillips (University of Guelph, Guelph, ON, Canada).

Survival analyses

Cohorts of 150–250 adult females (housed at 25 females per vial) on SY food were aged at 25 °C and 55% relative humidity. Dead flies were tallied and surviving flies were transferred to fresh SY vials twice weekly. Survival data were analyzed with JMP 5.01 software (SAS Institute, Cary, NC, USA).

Behavioral assays

For all behavioral tests, adult female flies (25 flies per vial) were housed at 25 °C and 55% relative humidity and transferred to fresh SY food vials twice weekly. Negative geotaxis was assessed in rapid iterative negative geotaxis (RING) assays as described by Gargano et al. (2005). Age-matched cohorts of adult flies were transferred to negative geotaxis tubes, the tubes were placed in the RING apparatus, negative geotaxis was elicited by a sharp rapping of the apparatus and behavioral performance was captured 4 s later via digital photography. Data from digital images were extracted using Scion Image (Frederick, MD, USA). Odor avoidance was determined in T-mazes using standard procedures (Stoltzfus et al., 2003; Bhandari et al., 2006). Age-matched groups of flies were placed in a T-maze and allowed to choose between an arm containing fresh air and an opposing arm containing 4-methylcyclohexanol (MCH), an aversive odorant. Metrics or descriptors for negative geotaxis and odor avoidance data were obtained as described by Martin et al. (2005) and Martin and Grotewiel (2006). Analysis of variances and post hoc tests were performed with JMP 5.01 or Prism 4.03 (GraphPad Software, San Diego, CA, USA).

Acknowledgments

  1. Top of page
  2. Summary
  3. Introduction
  4. Results
  5. Discussion
  6. Experimental procedures
  7. Acknowledgments
  8. References

The authors thank Ron Davis and John Phillips for contributing fly stocks; Scott Pletcher for guidance with the food media preparation; and Devin Rhodenizer and Pretal Patel for expert technical assistance. This work was supported in part by grants to Michael S. Grotewiel from the National Institute on Aging and the National Institute of Mental Health.

References

  1. Top of page
  2. Summary
  3. Introduction
  4. Results
  5. Discussion
  6. Experimental procedures
  7. Acknowledgments
  8. References
  • Bhandari P, Gargano JW, Goddeeris MM, Grotewiel MS (2006) Behavioral responses to odorants in Drosophila require nervous system expression of the beta integrin gene myospheroid. Chem. Senses 31, 627639.
  • Bond NW, Everitt AV, Walton J (1989) Effects of dietary restriction on radial-arm maze performance and flavor memory in aged rats. Neurobiol. Aging 10, 2730.
  • Burger JM, Hwangbo DS, Corby-Harris V, Promislow DE (2007) The functional costs and benefits of dietary restriction in Drosophila. Aging Cell 6, 6371.
  • Campbell BA, Gaddy JR (1987) Rate of aging and dietary restriction: sensory and motor function in the Fischer 344 rat. J. Gerontol. 42, 154159.
  • Clancy DJ, Gems D, Harshman LG, Oldham S, Stocker H, Hafen E, Leevers SJ, Partridge L (2001) Extension of life-span by loss of CHICO, a Drosophila insulin receptor substrate protein. Science 292, 104106.
  • Cook-Wiens E, Grotewiel MS (2002) Dissociation between functional senescence and oxidative stress resistance in Drosophila. Exp. Gerontol. 37, 13471357.
  • Gargano JW, Martin I, Bhandari P, Grotewiel MS (2005) Rapid iterative negative geotaxis (RING): a new method for assessing age-related locomotor decline in Drosophila. Exp. Gerontol. 40, 386395.
  • Goddeeris MM, Cook-Wiens E, Horton WJ, Wolf H, Stoltzfus JR, Borrusch M, Grotewiel MS (2003) Delayed behavioural aging and altered mortality in Drosophilaβ integrin mutants. Aging Cell 2, 257264.
  • Grotewiel MS, Martin I, Bhandari P, Cook-Wiens E (2005) Functional senescence in Drosophila melanogaster. Ageing Res. Rev. 4, 372397.
  • Helfand SL, Rogina B (2000) Regulation of gene expression during aging. In The Molecular Genetics of Aging, vol. 29 (SHekimi ed.). Berlin Heidelberg: Springer-Verlag, pp. 6780.
  • Idrobo F, Nandy K, Mostofsky DI, Blatt L, Nandy L (1987) Dietary restriction: effects on radial maze learning and lipofuscin pigment deposition in the hippocampus and frontal cortex. Arch. Gerontol. Geriatr. 6, 355362.
  • Ingram DK, Weindruch R, Spangler EL, Freeman JR, Walford RL (1987) Dietary restriction benefits learning and motor performance of aged mice. J. Gerontol. 42, 7881.
  • Kirby K, Hu J, Hilliker AJ, Phillips JP (2002) RNA interference-mediated silencing of Sod2 in Drosophila leads to early adult-onset mortality and elevated endogenous oxidative stress. Proc. Natl Acad. Sci. USA 99, 1616216167.
  • Landis GN, Tower J (2005) Superoxide dismutase evolution and life span regulation. Mech. Ageing Dev. 126, 365379.
  • Magwere T, Chapman T, Partridge L (2004) Sex differences in the effect of dietary restriction on life span and mortality rates in female and male Drosophila melanogaster. J. Gerontol. A Biol. Sci. Med. Sci. 59, 39.
  • Mair W, Piper MD, Partridge L (2005) Calories do not explain extension of life span by dietary restriction in Drosophila. PLoS Biol. 3, e223.
  • Martin I, Gargano JW, Grotewiel MS (2005) A proposed set of descriptors for functional senescence data. Aging Cell 4, 161164.
  • Martin I, Grotewiel MS (2006) Distinct genetic influences on locomotor senescence in Drosophila revealed by a series of metrical analyses. Exp. Gerontol. 41, 877881.
  • Mattson MP (2005) Energy intake, meal frequency, and health: a neurobiological perspective. Annu. Rev. Nutr. 25, 237260.
  • Mattson MP, Duan W, Lee J, Guo Z (2001) Suppression of brain aging and neurodegenerative disorders by dietary restriction and environmental enrichment: molecular mechanisms. Mech. Ageing Dev. 122, 757778.
  • Mattson MP, Magnus T (2006) Ageing and neuronal vulnerability. Nat. Rev. Neurosci. 7, 278294.
  • Means LW, Higgins JL, Fernandez TJ (1993) Mid-life onset of dietary restriction extends life and prolongs cognitive functioning. Physiol. Behav. 54, 503508.
  • Merry BJ (2005) Dietary restriction in rodents – delayed or retarded ageing? Mech. Ageing Dev. 126, 951959.
  • Min KJ, Tatar M (2006) Drosophila diet restriction in practice: do flies consume fewer nutrients? Mech. Ageing Dev. 127, 9396.
  • Partridge L, Piper MD, Mair W (2005a) Dietary restriction in Drosophila. Mech. Ageing Dev. 126, 938950.
  • Partridge L, Pletcher SD, Mair W (2005b) Dietary restriction, mortality trajectories, risk and damage. Mech. Ageing Dev. 126, 3541.
  • Piper MD, Mair W, Partridge L (2005a) Counting the calories: the role of specific nutrients in extension of life span by food restriction. J. Gerontol. A Biol. Sci. Med. Sci. 60, 549555.
  • Piper MD, Skorupa D, Partridge L (2005b) Diet, metabolism and lifespan in Drosophila. Exp. Gerontol. 40, 857862.
  • Pletcher SD, Libert S, Skorupa D (2005) Flies and their golden apples: the effect of dietary restriction on Drosophila aging and age-dependent gene expression. Ageing Res. Rev. 4, 451480.
  • Ruan H, Tang XD, Chen ML, Joiner ML, Sun G, Brot N, Weissbach H, Heinemann SH, Iverson L, Wu CF, Hoshi T, Joiner MA (2002) High-quality life extension by the enzyme peptide methionine sulfoxide reductase. Proc. Natl Acad. Sci. USA 99, 27482753.
  • Stewart J, Mitchell J, Kalant N (1989) The effects of life-long food restriction on spatial memory in young and aged Fischer 344 rats measured in the eight-arm radial and the Morris water mazes. Neurobiol. Aging 10, 669675.
  • Stoltzfus JR, Horton WJ, Grotewiel MS (2003) Odor-guided behavior in Drosophila requires calreticulin. J. Comp. Physiol. A Neuroethol. Sens. Neural. Behav. Physiol. 189, 471483.
  • Tu MP, Epstein D, Tatar M (2002) The demography of slow aging in male and female Drosophila mutant for the insulin-receptor substrate homologue chico. Aging Cell 1, 7580.