In many species, territorial behavior is limited to the breeding season and is tightly coupled to circulating gonadal steroid levels. In contrast, both male and female red squirrels (Tamiasciurus hudsonicus) are highly aggressive in both the breeding and non-breeding seasons in defense of food stores on their individual territories throughout the boreal and northern forests of North America. Dehydroepiandrosterone (DHEA), an androgen precursor, is secreted from the adrenal cortex in some mammals, and DHEA has been linked to aggression in non-breeding songbirds. Here, we examined plasma DHEA levels in a natural population of red squirrels in the Yukon, Canada. Plasma DHEA levels in both males and females reached high concentrations (up to 16.952 ng/ml in males and 14.602 ng/ml in females), markedly exceeding plasma DHEA concentrations in laboratory rats and mice and similar to plasma DHEA concentrations in some primates. Circulating DHEA levels showed both seasonal and yearly variation. Seasonal variation in male plasma DHEA levels was negatively correlated with testes mass. Yearly variation in male DHEA levels was positively correlated with population density. In both males and females, circulating DHEA rapidly increased after ACTH treatment, implying an adrenal origin. This is the first examination of plasma DHEA concentrations in a wild rodent and the first field experiment on the regulation of plasma DHEA in any wild mammal. These data lay the foundation for future studies on the role of DHEA in non-breeding territoriality in this species and other mammals.