Is rapid cold-hardening an aerobic process? Characterization of changes in metabolic activity during its induction and effects of anoxia in flesh fly.

Rapid cold-hardening (RCH) is a type of phenotypic plasticity that promotes a swift improvement of cold tolerance in insects. A brief exposure to mild cold dramatically increases insect survival to a subsequent cold exposure that would be lethal otherwise. In adult male flesh fly, Sarcophaga bullata, as little as 15 min at 5 °C significantly improved organismal survival at -7°C from 0 to 66.7 ± 11.1%. In this study, we investigated whether this RCH response is an aerobic process in S. bullata by characterizing changes in metabolic activity during its induction. At the level of whole organism, CO2 production continued at a level above our detection limit, and a relatively greater rate was observed during the early phase before it stabilized after ~1 h of the RCH induction. Similarly, in isolated flight muscle tissues, those maintained at 5 °C for 10 min exhibited significantly greater rates of oxygen consumption, compared to those maintained at 5 °C for 1 h (2.82 ± 0.29 vs. 1.36 ± 0.22 µl O2 mg-1 DM h-1). When these tissues were exposed to LaCl3, a treatment that should inhibit RCH ex vivo, oxygen consumption rates of the muscles were reduced significantly to a level similar to those that had been maintained at 5 °C for 1 h. Interestingly, however, the RCH response was still evident among individuals exposed to chilling under anoxia. Compared to those exposed to anoxia for 30 min only at 25 °C, flies exposed to 5 °C for 2 h under anoxia following the initial exposure exhibited a significantly greater level of cold tolerance at -7.5 °C (41.7 ± 7.1 vs. 91.8 ± 3.9%). Our results suggest that while relatively greater rates of metabolic activity are associated with the early phase of the RCH induction, it can proceed under the anoxic condition, thereby suggesting its independence to aerobic respiration.