Expression of NF-κB and downstream antioxidant genes in skeletal muscle of hibernating ground squirrels, Spermophilus tridecemlineatus.

Many small mammals survive the winter by hibernating, entering long periods of cold torpor that are interspersed with brief periods of arousal back to euthermia. This cycling is accompanied by wide changes in oxygen consumption, perfusion of tissues and ATP turnover, and the arousal period in particular is challenging because of oxidative stress associated with the huge increase in oxygen consumption needed to support thermogenesis by brown adipose tissue and skeletal muscle. Well-developed antioxidant defences are needed. The present study analyses responses of the redox-sensitive transcription factor, NF-κB, in skeletal muscle over six points on the torpor-arousal cycle to gain insight into its regulation and role during hibernation. Immunoblotting was used to analyse NF-κB p50 and p65 subunit levels, nuclear versus cytoplasmic localization and DNA-binding activity as well as levels and phosphorylation state of the IκBα inhibitor and the kinase IKK that phosphorylates IκBα to trigger its dissociation from NF-κB. The data were generally consistent with an activation of NF-κB during the entrance into torpor with responses including an auto-up-regulation of p50 subunits seen during early torpor and elevated IκBα protein during arousal. Protein levels of two downstream antioxidant targets showed differential regulation, Mn-superoxide dismutase (MnSOD) rising during early torpor versus heme oxygenase 1 (HO-1) increasing during early arousal. The mRNA transcript levels of p50, p65, HO-1 and MnSOD also showed differential expression over the torpor-arousal cycle. The results suggest that antioxidant defences are up-regulated at specific phases of the torpor-arousal cycle and that NF-κB mediates such protective responses.