Frédéric Hervant

Metabolic rate and oxidative stress in insects exposed to low temperature thermal fluctuations

Fluctuating temperatures are a predominant feature of the natural environment but their effects on ectotherm physiology are not well-understood. The warm periods of fluctuating thermal regimes (FTRs) provide opportunities for repair leading to increased survival, but there are also indications of negative effects of warm exposure. In this study, we examined respiration and oxidative stress in adult Alphitobius diaperinus exposed to FTRs and to constant low temperatures. We hypothesized that cold exposure will cause oxidative stress and that FTRs would reduce the amount of chill injuries, via activation of the antioxidant system. We measured V˙CO2, activities of super oxide dismutase (SOD), amounts of total (GSHt) and oxidized glutathione (GSSG) during cold and warm periods of FTRs. Increased severity of cold exposure caused a decrease in the glutathione pool. SOD levels increased during the recovery period in the more severe FTR. The antioxidant response was sufficient to counter the reactive oxygen species production, as the GSH:GSSG ratio increased. We conclude that cold stress causes oxidative damage in these beetles, and that a warm recovery period activates the antioxidant system allowing repair of cold-induced damage, leading to the increased survival previously noted in beetles exposed to fluctuating versus constant temperatures.

COMPARATIVE STUDY ON THE BEHAVIORAL, VENTILATORY, AND RESPIRATORY RESPONSES OF HYPOGEAN AND EPIGEAN CRUSTACEANS TO LONG-TERM STARVATION AND SUBSEQUENT FEEDING

Abstract Survival, oxygen consumption, locomotory activity and ventilatory activity were recorded during a 180-day starvation period and a subsequent 15-day feeding phase in 3 hypogean crustaceans, Niphargus rhenorhodanensis, Niphargus virei, and Stenasellus virei. For comparison, these parameters were also recorded during a 28-day starvation period and a subsequent 7-day feeding phase in two morphologically close epigean crustaceans, Gammarus fossarum and Asellus aquaticus. Hypogean crustaceans were better adapted to lack of food than epigean ones and all crustaceans previously studied, with survival times largely longer than 200 days. During long-term starvation, the locomotory, ventilatory, and respiratory rates were drastically lowered in subterranean species, whereas surface species showed lower decreases in these rates and responded by a marked and transitory hyperactivity. The higher reduction in metabolic rate by hypogean species would ensure their survival during prolonged periods of food deprivation. We propose an energy strategy for food-limited hypogean crustaceans involving the ability 1) to withstand long-term starvation, and 2) to use the consumed food very efficiently. Resistance to starvation would probably involve a state of temporary torpor during which the subterranean crustaceans subsist on a high energy reserve, such as lipid stores.

The importance of fluctuating thermal regimes for repairing chill injuries in the tropical beetle Alphitobius diaperinus (Coleoptera: Tenebrionidae) during exposure to low temperature

Abstract.  In this study, the impact of acclimation (1 month at 15 °C vs. breeding at 30 °C) and fluctuating thermal regimes (daily transfers from low temperatures to various higher temperatures for 2 h) on the cold tolerance of the tropical beetle, Alphitobius diaperinus Panzer (Coleoptera: Tenebrionidae) was examined. Acclimation increased significantly the duration of survival (Lt50) at a constant 5 °C (7.7 ± 0.3 days to 9.7 ± 0.5 days). Survival of acclimated and nonacclimated beetles increased slightly at alternating temperatures of 5 °C/10 °C or 5 °C/15 °C. When daily transfer to 20 °C was applied, survival (Lt50) was improved markedly (nonacclimated: 15.5 ± 0.7 days, acclimated: 19.6 ± 0.6 days). The higher temperatures may allow progressive repair of injuries, and the effects of chilling may be repaired completely at 25 and 30 °C, a phenomenon recorded here for the first time. It is estimated that the theoretical upper threshold of chill injury (Th) of nonacclimated beetles is 15.1 °C whereas it is shifted down to 11.2 °C in acclimated beetles, which might enable this temperature to allow effective repair of injury.

Behavioral, Ventilatory, and Metabolic Responses to Severe Hypoxia and Subsequent Recovery of the Hypogean Niphargus rhenorhodanensis and the Epigean Gammarus fossarum (Crustacea: Amphipoda)

Two aquatic amphipod crustaceans were investigated: Niphargus rhenorhodanensis (a hypogean species, which has to cope with severe hypoxic conditions during about 6 mo per year during the hydrological cycle) and Gammarus fossarum (an epigean species, which lives continuously in well-oxygenated water). The lethal times for 50% of the population (LT50) for these closely) related species were 46. 7 and 6.3 h, respectively, in very severe hypoxia (<0.2 Torr) at 11°C. Therefore, the aim of this study was to examine some possible reasons why the subterranean Niphargus survived severe hypoxia longer than Gammarus and numerous other epigean crustaceans. During severe hypoxia Niphargus reduced its energetic expenditures compared with Gammarus. This reduction was associated with changes in locomotion and ventilation. In both species, anaerobic metabolism was fueled in severe hypoxia by the breakdown of glycogen and arginine phosphate. In Niphargus, however glutamate was also utilized. This resulted in the production of l-lactate as the major end product in both species. Alanine was found to be a minor end product in Gammarus, while alanine and succinate were found to be the minor end products in Niphargus. Compared to Gammarus, and to most other epigean crustaceans, Niphargus showed high amounts of stored glycogen and arginine phosphate. During severe hypoxia, both organisms excreted lactate into the medium, which is unusual for crustaceans. The differences in the stores of phosphagen and glycogen, and in behavioral and ventilatory responses, are probably the main reasons for their different resistance to severe hypoxia. During recovery, both species displayed a characteristic hyperventilation and their metabolism was predominantly aerobic. Gammarus excreted a great part of the lactate accumulated during severe hypoxia, whereas Niphargus remetabolized it, the latter possessing a higher glyconeogenesis capacity from lactate. After 24 h recovery, energy charge in both amphiods was not completely restored, and only Niphargus showed a total removal of the accumulated end products. Niphargus showed a faster replenishment of ATP concentration.

Behavioral, Ventilatory, and Metabolic Responses of the Hypogean Amphipod Niphargus virei and the Epigean Isopod Asellus aquaticus to Severe Hypoxia and Subsequent Recovery

The locomotory and ventilatory activities and the intermediary and energy metabolism modifications of the hypogean amphipod Niphargus virei and the epigean isopod Asellus aquaticus were compared during severe hypoxia (<0.03 kPa) and subsequent recovery. The aims of this study were (1) to determine why the subterranean species displayed a greater tolerance of hypoxia than A. aquaticus and numerous other epigean crustaceans, (2) to confirm previous results obtained with the hypogean amphipod Niphargus rhenorhodanensis and the epigean amphipod Gammarus fossarum, (3) to provide an interspecific comparison of epigean species in order to see if responses showed by epigean amphipods during hypoxia and recovery can be extended to epigean isopods, and (4) to better understand the ecological problems of the hypogean organisms survival and perennation in subterranean habitats. Both organisms responded to long-term experimental severe hypoxia with classical anaerobic metabolism, characterized by a decrease in ATP and phosphagen, the use of glycogen and glutamate, and the accumulation of lactate (with some alanine). In addition, some accumulation of succinate was found in N. virei. Lactate (and succinate for N. virei) was also largely excreted by both amphipods, which is unusual for the crustacea in general Compared with A. aquaticus and most other epigean crustaceans, N. virei showed large amounts of stored glycogen and arginine phosphate. These differences in glycogen and phosphagen stores, and the ability to reduce glycolytic flux and energetic expenditures linked to locomotion and ventilation, extended the survival of hypogean crustaceans under experimental anaerobiosis (LT50 was 52.1 h for N. virei and 19. 7 h for A. aquaticus during severe hypoxia at 11° C). During recovery, both species displayed characteristic hyperventilation, slow locomotory activity, and predominantly aerobic metabolism. Asellus aquaticus excreted a large part of the lactate accumulated during severe hypoxia, whereas N. virei remetabolized it, as it had a higher lactate-derived glyconeogenesis capacity. The disposal of endproducts and replenishing of glycogen, ATP, and phosphagen required more than 24 h for both organisms. Niphargus virei showed a faster and more complete replenishment of ATP and arginine phosphate levels than A. aquaticus. Data concerning locomotory, ventilatory, and metabolic responses to hypoxia and subsequent recovery in N. virei and A. aquaticus are similar to those obtained with N. rhenorhodanensis and G. fossarum.

Oxygen Consumption and Ventilation in Declining Oxygen Tension and Posthypoxic Recovery in Epigean and Hypogean Crustaceans

ABSTRACT Respiratory and ventilatory responses to declining O2 tension and posthypoxic recovery were investigated in 3 hypogean and in 2 epigean aquatic crustaceans. The aims of this study were to determine how these species reacted to declining O2 tension, to investigate their changes in ventilation and metabolic rate during progressive hypoxia, and to extend our knowledge on the fate of anaerobic end products during subsequent recovery. Hypogean organisms had normoxic O2 consumption rates 1.7―3.5 times lower than the epigean species. All 5 crustaceans were able to maintain their O2 consumption rates (1) at relatively constant levels and (2) independent of Po2 between normoxia and the critical Po2. Hypogean species also possessed lower critical Po2 than epigean ones, which may indicate that these organisms are better adapted to low O2 content and are better equipped to stay aerobic under hypoxia. For all species, posthypoxic recovery resulted in a high O2 debt. The payment of this debt was smaller in hypogean than in epigean species. The main explanations of the lower 0, debt shown by hypogean organisms are the lower energetic expenditures noticed during hypoxia, partly due to a decrease in locomotory and ventilatory activities.

Metabolic responses to cold in subterranean crustaceans

SUMMARY Changes in polyol, sugar and free amino acid (FAA) body contents were investigated in the aquatic, subterranean (i.e. hypogean) crustaceans Niphargus rhenorhodanensis and Niphargus virei and in a morphologically close aquatic, surface-dwelling (i.e. epigean) crustacean Gammarus fossarum acclimated to 12°C, 3°C and –2°C. With decreasing temperature, G. fossarum significantly increased its alanine and glutamine levels, while trehalose body content was found to increase above control levels only at –2°C. N. virei showed moderate increases of alanine and glycine, and no change in trehalose level was observed in this species. By contrast, N. rhenorhodanensis was the only species showing a significant rise in its total FAA pool, mainly explained by alanine, glycine, arginine and glutamine accumulations. This species also gradually increased its trehalose body content with decreasing temperature. Several cold-hardy ectotherms show metabolic responses to cold that are identical to those observed in N. rhenorhodanensis. A previous comparative study showed that the hypogean N. rhenorhodanensis exhibited a survival time (Lt50) at– 2°C that was 26.3 times and 2.6 times higher than the hypogean N. virei and the epigean G. fossarum, respectively. Thus, crustacean levels of FAA and trehalose were correlated with their respective cold tolerances. Such differences in metabolic responses to cold in both hypogean organisms were unexpected since they both live in thermally buffered biotopes. Considering the current distribution areas of the two subterranean crustaceans studied, we assume that the cold hardiness found in the hypogean N. rhenorhodanensis could be correlated with its biogeography history during the quaternary glaciations.

Locomotory, ventilatory and metabolic responses of the subterranean Stenasellus virei (Crustacea, Isopoda) to severe hypoxia and subsequent recovery

The locomotory and ventilatory activities and the intermediary and energy metabolism modifications of the hypogean aquatic isopod crustacean Stenasellus virei were investigated in severe hypoxia (PO2 < 0.03 kPa) and subsequent recovery. The aims of this study were i) to determine why the subterranean species displayed a greater tolerance of hypoxia than numerous other epigean crustaceans, ii) to confirm previous results obtained with four hypogean and epigean crustaceans, iii) to compare the responses to severe hypoxia in hypogean amphipods and isopods, and iv) to better understand the ecological problems of the hypogean organisms survival in subterranean habitats. S. virei responded to experimental long-term, severe hypoxia with classical anaerobic metabolism mainly characterized by a decrease in adenosine triphosphate (ATP) and phosphagen, utilization of glycogen and glutamate, and accumulation of lactate and alanine. Lactate was also largely excreted by this organism, which is unusual for crustaceans in general. Compared to most other epigean crustaceans, the isopod S. virei showed high amounts of stored glycogen and arginine phosphate. These differences in glycogen and phosphagen stores, and the ability to reduce energetic expenditures linked to locomotion and ventilation, extended the survival of S. virei under experimental anaerobiosis. During recovery, the isopod S. virei showed a higher capacity for glyconeogenesis from lactate and a faster and total replenishment of ATP and arginine phosphate levels than epigean crustaceans. Data concerning responses to hypoxia and subsequent recovery in S. virei are similar to those previously obtained with two other hypogean amphipods, except that this isopod did not synthesize succinate in anaerobiosis.

Ecology of the Hybridogenetic Rana esculenta Complex: Differential Oxygen Requirements of Tadpoles

Because of intrinsic demographic load induced by hybridogenesis (infertility of homotypic hybrid matings), the maintenance of hybrid lineages supposes that they present better performances (heterosis) than their host species which allows them to coexist on a long-term basis. However, this necessity of high fitness can be relaxed if a relative niche partitioning occurs between the taxa, each of them differing in their ecological optima. In the waterfrog hybridogenetic complex (Rana esculenta complex), recent studies have revealed that hybrids show intermediate distribution between parental species across a gradient of river influence (that is related to a gradient of oxygen levels), and intermediate performances of their tadpoles with regard to oxygen availability (hypoxia). In investigating oxygen consumption rates, survival time in anoxia, and metabolite contents in the three forms of the complex, the present study confirms intermediate characteristics of hybrid tadpoles (R. esculenta) when compared to both parental lineages (R. lessonae and R. ridibunda). Whereas R. ridibunda tadpoles were the most sensitive to anoxia, R. lessonae tadpoles were the most tolerant. Because oxygen requirements of the hybrid proved to be intermediate, no heterosis was detected. These results confirm the hypothesis of the intermediate niche hypothesis to explain the coexistence of R. lessonae and R. esculenta and the success of the hybridogens.

Comparative study of the metabolic responses during food shortage and subsequent recovery at different temperatures in the adult lesser mealworm, Alphitobius diaperinus (Coleoptera: Tenebrionidae)

Metabolic responses to prolonged food shortage (35 days) and subsequent re‐feeding (14 days) were investigated in adults of an introduced beetle, Alphitobius diaperinus Panzer, as a function of temperature (12, 16, 20 and 24 °C). Various qualitative and quantitative changes that greatly vary according to the temperature experienced occurred in metabolite levels during prolonged starvation. Whereas levels of protein and ATP did not change significantly, triglycerides decreased markedly and glycogen changed little. Metabolite levels were differently affected by temperature, with triglycerides being less rapidly degraded at 20 than at 24 °C and almost completely depleted at 12 and 16 °C; in contrast to higher temperatures, glycerol is accumulated at 12 °C. Physiological adaptation to starvation and low temperatures are highly linked and energy allocation for starvation vs. temperature acclimation must be strictly regulated, both being essential for insect survival. Re‐synthesis rates during recovery are probably highly temperature‐dependent for all metabolites. The proteins retained during starvation and the preferential degradation of lipids allowed a rapid recovery. Above 16 °C, adult A. diaperinus regained locomotory activity rapidly and the triglyceride, glycerol and glycogen reserves were restored. This tropical species may be able to colonize other environments such as natural and/or artificial biotopes where conditions are close to those of its natural habitat.