Sub-acute restraint stress progressively increases oxidative/nitrosative stress and inflammatory markers while transiently upregulating antioxidant gene expression in the rat hippocampus

Abstract

We have previously demonstrated that acute stress decreases neuronal nitric oxide synthase (NOS) expression in the hippocampus despite increased concentrations of nitric oxide which may indicate feedback inhibition of neuronal NOS expression via inducible NOS-derived nitric oxide. Moreover, the hippocampus undergoes an initial oxidative/nitrosative insult that is rapidly followed by upregulation of protective antioxidants, including the zinc-binding metallothioneins, in order to counter this and restore redox balance following acute stress exposure. In the present study, we have utilized indicators of oxidative/nitrosative stress, members of the nuclear factor (erythroid-derived 2)-like 2 (Nrf2) pathway, antioxidant metallothioneins, and neuroinflammatory markers to observe the changes occurring in the hippocampus following short term repeated stress exposure. Male Wistar rats were subjected to control conditions or 6 h of restraint stress applied for 1, 2, or 3 days (n = 8 per group) after which the hippocampus was isolated for redox assays and relative gene expression. The hippocampus showed increased oxidative stress, transient dys-homeostasis of total zinc, and increased expression of the Nrf2 pathway members. Moreover, repeated stress increased nitrosative status, nitric oxide metabolites, and 3-nitrotyrosine, indicative of nitrosative stress in the hippocampus. However, levels of neuronal NOS decreased over all stress treatment groups, while increases were observed in inducible NOS and xanthine dehydrogenase. In addition to inducible NOS, mRNA expression of other inflammatory markers including interleukin-6 and interleukin-1β also increased even in the presence of increased anti-inflammatory glucocorticoids. Together, these results demonstrate that despite increases in antioxidant expression, sub-acute stress causes an inflammatory phenotype in the hippocampus by inducing oxidative/nitrosative stress, zinc dys-homeostasis, and the accumulation of nitrotyrosinated proteins which is likely driven by increased inducible NOS signaling.