The Safety Limits Of An Extended Fast: Lessons from a Non-Model Organism

Abstract

While safety of fasting therapy is debated in humans, extended fasting occurs routinely and safely in wild animals. To do so, food deprived animals like breeding penguins anticipate the critical limit of fasting by resuming feeding. To date, however, no molecular indices of the physiological state that links spontaneous refeeding behaviour with fasting limits had been identified. Blood proteomics and physiological data reveal here that fasting-induced body protein depletion is not unsafe "per se". Indeed, incubating penguins only abandon their chick/egg to refeed when this state is associated with metabolic defects in glucose homeostasis/fatty acid utilization, insulin production and action, and possible renal dysfunctions. Our data illustrate how the field investigation of "exotic" models can be a unique source of information, with possible biomedical interest.

Figure 1. Uncoupling between the metabolic and behavioural shifts in fasting Adélie penguins.

The transition from phase 2 to phase 3 of fasting is characterised by a metabolic shift reflected in changes in body mass and plasma metabolic profiles, including uric acid levels that reflect protein breakdown (a). Penguin body masses (variations not shown for better viewing) ranged from 4.9 ± 0.28 kg (beginning of phase 2) to 3.35 ± 0.11 kg (late phase 3). For captive animals, measurements were achieved from 6 birds monitored from phase 2 to the late phase 3 of fasting. For breeding wild birds (i.e. incubating males), % of departing birds were calculated over four austral summers (b), means ± SEM; n = 92–105/year). Departure to sea was either consecutive to relief by females or to nest abandonment. These data show that the spontaneous behavioural shift toward nest abandonment, which likely corresponds to the safety limit of fasting, occurs during phase 3 of fasting and is therefore not coupled with the metabolic shift occurring at the breakpoint from phase 2 to phase 3. Penguin drawings were made by Fabrice Bertile.

Figure 2. Metabolic profiles of male Adélie penguins in the fed and fasted states.

The metabolic shift from phase 2 to phase 3 is characterized by plasma metabolite and hormone profiles, which drive changes in the respective contribution of fuels to energy needs. Results are means ± SEM (n = 6 per group, except for wild birds in phase 3 departing after relief where n = 3). To minimize disturbance of free-ranging birds and for relevance of the experimental protocol, only captive animals were equipped with a pedometer and weighed daily. Hence, Student’s t-test was used to compare the rate of daily body mass loss (dm/m.dt) and locomotor activity levels in captive birds (P < 0.05) and ANOVA followed by Tukey post-hoc tests were used for all other parameters in all birds (P < 0.05). Values that do not share the same letter are significantly different (P < 0.05). au: arbitrary unit. NEFA: non-esterified fatty acids; βOHB: β-hydroxybutyrate.

Figure 3. Differential plasma proteomics in wild animals characterizes the safety limit of an extended fast.

A total of 36 unique avian proteins from 30 excised “individual or grouped (i.e. from isoforms; boxed areas) gel spots” were identified (a; see also Tables S1 and S2). The intensity of 10 protein spots (a, blue numbers), containing 11 distinct proteins, differed significantly between groups (b). The relative plasma abundance of these 10 proteins (c) is illustrated for phase 3 penguins with reference to corresponding phase 2 birds (green dashed line; the variation on reference levels in phase 2 penguins is given as green error bars). Data are means ± SEM (n = 6 per group, except for wild birds in phase 3 departing after relief where n = 3). Comparisons between captive 3 birds were performed using Student’s t-tests, while comparisons between breeding birds were performed using ANOVA followed by Tukey post-hoc tests. Significance (*) between groups was set at P < 0.05.

Figure 4. Protein wasting is not the only determinant of the safety limit of fasting.

The low levels of adiponectin, complement factor D, and vitamin D binding protein only found in the plasma of phase 3 animals after nest abandonment show that fasting limits are achieved only when the rise in protein breakdown is associated with functional impairments and metabolic alterations (see text for details). The reasons for which only some penguins in phase 3 of fasting exhibit dysfunctions that, in addition to protein wasting, lead to nest abandonment remain to be determined.