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Abstract Mindfulness has been incorporated into several treatment approaches for psychopathology. Despite the popularity of this approach, relatively few empirical investigations have examined the relationship between mindfulness and autonomic indicators of flexible emotion regulation, such as heart rate variability (HRV). Generalized anxiety disorder (GAD) has been associated with both low levels of mindfulness and HRV. In this investigation, we examined the relationship between HRV and mindfulness in the context of elevated generalized anxiety (GA) symptoms—an analog for GAD—by examining whether GA level moderated this relationship. HRV was collected while participants completed self-report measures of GA and trait mindfulness. GA level interacted with mindfulness in the prediction of HRV; in the high GA, but not low GA group, mindfulness was positively associated with HRV. This suggests that for individuals with high GA, mindfulness may enhance parasympathetic influences on the heart rate. We address the limitations of the current investigation and suggest avenues for future research on mindfulness-related changes in tonic and phasic HRV over time.