The Drosophila insulin receptor independently modulates lifespan and locomotor senescence

The Insulin/IGF-like signalling (IIS) pathway plays an evolutionarily conserved role in ageing. In model organisms reduced IIS extends lifespan and ameliorates some forms of functional senescence. However, little is known about IIS in nervous system ageing and behavioural senescence. To investigate this role in Drosophila melanogaster, we measured the effect of reduced IIS on senescence of two locomotor behaviours, negative geotaxis and exploratory walking. Two long-lived fly models with systemic IIS reductions (daGAL4/UAS-InRDN (ubiquitous expression of a dominant negative insulin receptor) and d2GAL/UAS-rpr (ablation of insulin-like peptide producing cells)) showed an amelioration of negative geotaxis senescence similar to that previously reported for the long-lived IIS mutant chico. In contrast, exploratory walking in daGAL4/UAS-InRDN and d2GAL/UAS-rpr flies declined with age similarly to controls. To determine the contribution of IIS in the nervous system to these altered senescence patterns and lifespan, the InRDN was targeted to neurons (elavGAL4/UAS-InRDN), which resulted in extension of lifespan in females, normal negative geotaxis senescence in males and females, and detrimental effects on age-specific exploratory walking behaviour in males and females. These data indicate that the Drosophila insulin receptor independently modulates lifespan and age-specific function of different types of locomotor behaviour. The data suggest that ameliorated negative geotaxis senescence of long-lived flies with systemic IIS reductions is due to ageing related effects of reduced IIS outside the nervous system. The lifespan extension and coincident detrimental or neutral effects on locomotor function with a neuron specific reduction (elavGAL4/UAS-InRDN) indicates that reduced IIS is not beneficial to the neural circuitry underlying the behaviours despite increasing lifespan

Exploratory walking senescence in daGAL4/UAS-InR^DN male and female flies.

<p>(<b>A-B</b>) Representative images of the exploratory walking track of an individual w^<b>Dah</b> control female fly at 1 week old (<b>A</b>) and 7 weeks old (<b>B</b>) during a 15 minute observation period. (<b>C</b>) Survival of daGAL4/UAS-InR^<b>DN</b> once mated female flies compared to daGAL4/+ and UAS-InR^<b>DN</b>/+ controls. Median lifespans and sample sizes were: daGAL4/UAS-InR^<b>DN</b> = 56 days, N = 69; daGAL4/+ = 48 days, N = 99; and UAS-InR^<b>DN</b>/+ = 44 days, N = 60. Survival curves were compared using nonparametric log rank tests and p values calculated. daGAL4/UAS-InR^<b>DN</b> showed an increased survival compared to both controls (P<0.0001). (<b>D-M</b>) Exploratory walking senescence for a cohort of female flies of the indicated genotypes run in parallel with the survival experiment shown in (C). Data are shown as mean value for each parameter ±SEM, N = 12 for the indicated genotype. (<b>D</b>) Female mean distance walked (mm) vs age. (<b>I</b>) Female Total Function of mean distance walked (mm). (<b>E</b>) Female Mean velocity (mm/sec) vs age. (<b>J</b>) Female Total Function of mean velocity (mm/sec). (<b>F</b>) Female Mean walking duration (secs) vs age. (<b>K</b>) Female Total Function of mean walking duration (secs). (<b>G</b>) Female Mean frequency of rotations (change in walking direction) vs age. (<b>L</b>) Female Total Function of mean Rotation Frequency. (<b>H</b>) Female Mean Duration in Central Zone (secs) vs age. (<b>M</b>) Female Total Function of mean duration in central Zone (secs). (<b>N-W</b>) Exploratory walking senescence for a cohort of male flies of the indicated genotypes run in parallel with the survival experiment shown in <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0125312#pone.0125312.g001" target="_blank">Fig 1G</a>. Data are shown as mean value for each parameter ±SEM, N = 12 for the indicated genotype. (<b>N</b>) Male mean distance walked (mm) vs age. (<b>T</b>) Male Total Function of mean distance walked (mm). (<b>O</b>) Male Mean velocity (mm/sec) vs age. (<b>T</b>) Male Total Function of mean velocity (mm/sec). (<b>P</b>) Mean walking duration (secs) vs age. (<b>U</b>) Male Total Function of mean walking duration (secs). (<b>Q</b>) Mean frequency of rotations (change in walking direction) vs age. (<b>V</b>) Male Total Function of mean Rotation Frequency. (<b>R</b>) Male Mean Duration in Central Zone (secs) vs age. (<b>W</b>) Total Function of mean duration in central Zone (secs). Data were analysed by two way ANOVA and genotype and age found to be the main effects (p<0.05). Data at individual time points or total function data were analysed by one way ANOVA followed by post hoc means comparisons using Tukey HSD, and * indicates significant differences (p<0.05) between daGAL4/UAS-InR^<b>DN</b> flies and both controls.</p

Exploratory walking senescence in elavGAL4/UAS-InR^DN male flies.

<p>(A) Survival of elavGAL4/UAS-InR^<b>DN</b> male flies compared to elavGAL4/+ and UAS-InR^<b>DN</b>/+ controls. Survival curves were compared using nonparametric log rank tests and p values calculated. Median lifespans and sample sizes were: elavGAL4/UAS-InR^<b>DN</b> = 56 days, N = 105; elavGAL4/+ = 53 days, N = 89; and UAS-InR^<b>DN</b>/+ = 56 days, N = 95. elavGAL4/UAS-InR^<b>DN</b> showed no difference in survival compared to both controls (Log Rank test, p>0.05). (<b>B-K</b>) Exploratory walking senescence for a cohort of male flies of the indicated genotypes run in parallel with the survival experiment shown in (A). Data are shown as mean value for each parameter ±SEM, and N = 30 for the indicated genotype. (<b>B</b>) Mean distance walked (mm) vs age. (<b>C</b>) Total Function of mean distance walked (mm). (<b>D</b>) Mean velocity (mm/sec) vs age. (<b>E</b>) Total Function of mean velocity (mm/sec). (<b>F</b>) Mean walking duration (secs) vs age. (<b>G</b>) Total Function of mean walking duration (secs). (<b>H</b>) Mean frequency of rotations (change in walking direction) vs age. (<b>I</b>) Total Function of mean Rotation Frequency. (<b>J</b>) Mean Duration in Central Zone (secs) vs age. (<b>K</b>) Total Function of mean duration in central Zone (secs Walking data were analysed by two way ANOVA and genotype and age found to be the main effects (p<0.05). Data at individual time points or total function data were analysed by one way ANOVA followed by post hoc means comparisons using Tukey HSD, and * indicates significant differences (p<0.05) between elavGAL4/UAS-InR^<b>DN</b> flies and both controls.</p

The Effect of InR^DN Expression on <i>dilp</i> Expression.

<p><b>(A-D)</b> The effect of UAS-InR^<b>DN</b> expression in CNS neurons on <i>dilp</i> expression in 10 day old adult male and female heads and bodies was measured by quantitative RT-PCR, and N = 3 (groups of 20 heads or bodies) for each genotype. (<b>A</b>) <i>dilp 2</i>, <i>3</i>, <i>5</i> and <i>6</i> relative transcript levels in elavGAL/UAS-InR^<b>DN</b> and control female heads. (<b>B</b>) <i>dilp 4</i>, <i>5</i>, <i>6</i> and <i>7</i> relative transcript levels in elavGAL/UAS-InR^<b>DN</b> and control female bodies. (<b>C</b>) <i>dilp 2</i>, <i>3</i>, <i>5</i> and <i>6</i> relative transcript levels in elavGAL/UAS-InR^<b>DN</b> and control male heads. (<b>D</b>) <i>dilp 4</i>, <i>5</i>, <i>6</i> and <i>7</i> relative transcript levels in elavGAL/UAS-InR^<b>DN</b> and control male bodies. Data are shown as mean relative expression level ±SEM (N = 3).</p