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Trauma is associated with increased risk for anxiety disorders such as posttraumatic stress disorder (PTSD). To further understand biologic mechanisms of PTSD, we examined the dark-enhanced startle response, a psychophysiological correlate of anxiety, and heart rate variability (HRV) in traumatized individuals with and without PTSD. The associations of these measures with PTSD may be sex-specific because of their associations with the bed nucleus of the stria terminalis, a sexually dimorphic brain structure in the limbic system that is approximately 2.5 times larger in men than in women.The study sample (N = 141) was recruited from a highly traumatized civilian population seeking treatment at Grady Memorial Hospital in Atlanta, Georgia. Psychophysiological responses during a dark-enhanced startle paradigm task included startle magnitude, assessed by eyeblink reflex, and measures of high-frequency HRV, during light and dark phases of the startle session.The startle magnitude was higher during the dark phase than the light phase (mean ± standard error = 98.61 ± 10.68 versus 73.93 ± 8.21 μV, p < .001). PTSD was associated with a greater degree of dark-enhanced startle in women (p = .03) but not in men (p = .38, p interaction = .48). Although HRV measures did not differ between phases, high-frequency HRV was greater in men with PTSD compared with men without PTSD (p = .02).This study demonstrates that the dark-enhanced paradigm provides novel insights into the psychophysiological responses associated with PTSD in traumatized civilian sample. Sex differences in altered parasympathetic and sympathetic function during anxiety regulation tasks may provide further insight into the neurobiological mechanisms of PTSD.