Subtropical hibernation in juvenile tegu lizards (Salvator merianae): insights from intestine redox dynamics

Abstract

Juvenile tegu lizards (Salvator merianae) experience gradual and mild temperature changes from autumn to winter in their habitat. This tropical/subtropical reptile enter a state of dormancy, with an 80% reduction in metabolic rate, that remains almost constant during winter. The redox metabolism in non-mammalian vertebrates that hibernate under such distinguished conditions is poorly understood. We analyzed the redox metabolism in the intestine of juvenile tegus during different stages of their first annual cycle. The effect of food deprivation (in spring) was also studied to compare with fasting during hibernation. Both winter dormancy and food deprivation caused decreases in reduced glutathione levels and glutathione transferase activity. While glutathione peroxidase and glutathione transferase activities decreased during winter dormancy, as well as glutathione (GSH) levels, other antioxidant enzymes (catalase, superoxide dismutase and glutathione reductase) remained unchanged. Notably, levels of disulfide glutathione (GSSG) were 2.1-fold higher in late autumn, when animals were in the process of depressing metabolism towards hibernation. This increased "oxidative tonus" could be due to a disruption in NADPH-dependent antioxidant systems. In dormancy, GSSG and lipid hydroperoxides were diminished by 60-70%. The results suggest that the entrance into hibernation is the main challenge for the redox homeostasis in the intestine of juvenile tegus.

Figure 1

Antioxidant enzymes activities in the intestine of tegu lizards at different metabolic states and under food deprivation. (A) Glutathione transferase (GST), (B) total glutathione peroxidase (tGPX), (C) total superoxide dismutase tSOD (total superoxide dismutase) and (D) manganese superoxide dismutase (MnSOD). N = 7–8. Values are shown as mean ± s.e.m. ‘Spring activity’ and ‘Fed Activity’ data are from the same animal group. Different letters indicate significant differences between groups (P < 0.05). Asterisks indicate significant differences between fed and unfed active lizards (P < 0.05).

Figure 2

Redox status in the intestine of tegu lizards at different metabolic states and under food deprivation. Glutathione parameters: (A) Total glutathione equivalents (GSH-eq), (B) reduced glutathione (GSH), (C) disulfide glutathione (GSSG), and (D) GSSG/GSH-eq ratio (%GSSG). N = 6–8. Values are shown as mean ± s.e.m. ‘Spring activity’ and ‘Fed Activity’ data are from the same animal group. Different letters indicate significant differences between groups (P < 0.05). Asterisks indicate significant differences between fed and unfed active lizards (P < 0.05).

Figure 3

Oxidative stress markers in the intestine of tegu lizards under different metabolic states and food deprivation. (A) Carbonyl proteins (Carbonyl), (B) lipid hydroperoxides (LOOH), and (C) thiobarbituric acid reactive substances (TBARS). N = 5–8. Values are shown as mean ± s.e.m. ‘Spring activity’ and ‘Fed Activity’ data are from the same animal group. Different letters indicate significant differences between groups (P < 0.05).

Figure 4

Schematic model illustrating hypothetical changes of the ‘oxidative tonus’ in the small intestine of juvenile tegu lizards, associated with seasonal changes in metabolic rate during their first annual cycle. (A) Circles depict the decrease of resting metabolic rates during the autumn and winter in neonate tegus, and the reactivation during arousal in spring (data from de Souza et al., 2004). Diamonds depict the ‘oxidative tonus’ of the intestinal tissue in the corresponding stages of the annual cycle, according to the current results. The tegus hatched during the summer, and early autumn samples were not available for this study. Late in the autumn, the highest GSSG levels indicate a redox imbalance and a more oxidized environment in intestinal tissue. During winter dormancy the concentration of LOOH decreased, indicating that proportional adjustments in the processes that produce and consume energy promote a lower steady state concentration of ROS. The reactivation of metabolism is gradual during arousal from dormancy, restoring the pre-hibernation redox balance in the intestinal tissue. (B) Hypothetical mechanism to explain the increased GSSG levels observed during late autumn in the tegu intestine. NADPH should be generated in large amounts through the pentose phosphate pathway (PPP) in the intestinal mucosa of actively growing tegus (in summer and early autumn). However, in late autumn metabolic rates slow down and NADPH supply should – hypothetically – decrease in the tissue. This may impair the enzymatic management of H₂O₂ (mainly by GPX and peroxiredoxins), leading to redox imbalance. LOOH, lipid hydroperoxides; GPX, glutathione peroxidase; GR, glutathione reductase; GSH, reduced glutathione; GSSG, disulfide glutathione; PRX, peroxiredoxin; Trx_(ox), oxidized thioredoxin; Trx_(red), reduced thioredoxin; TrxR, thioredoxin reductase.