Alterations in neurobiological stress systems such as the autonomic nervous system (ANS) and hypothalamic-pituitary-adrenal (HPA) axis contribute to the development and maintenance of psychological and behavioral problems after traumatic experiences. Investigating neurobiological parameters and how these relate to each other may provide insight into the complex mechanisms at play. Whereas the preponderance of studies focuses on either the ANS or the HPA axis separately, the current study is the first to evaluate relations between posttraumatic stress and both basal activity during rest and stress reactivity of the ANS as well as the HPA axis in a sample of traumatized adolescents and healthy controls. The traumatized sample (n = 77), based on clinical levels of posttraumatic stress, was a convenience sample that was recruited within residential institutions, was compared to a healthy control sample (n = 48) recruited within the general community. For the ANS, we expected increased SNS and decreased PNS activity during rest and increased SNS and decreased PNS reactivity to social stress among traumatized adolescents compared to healthy controls. Regarding the HPA axis, we expected increased basal cortisol levels and decreased cortisol reactivity to stress in the traumatized sample. Compared to healthy controls, traumatized adolescents exhibited significantly higher sympathetic and lower parasympathetic activation during rest and increased sympathetic reactivity to acute stress (ANS parameters). Outcomes on the HPA axis (i.e. cortisol) indicated that traumatized adolescents showed increased cortisol levels during rest and blunted cortisol reactivity to acute stress. Implications for clinical relevance and trauma-focused treatment purposes are discussed.