Studies in adults have demonstrated a relationship between lowered heart rate variability (HRV) and poor health. However, less is known about the role of autonomic arousal in children’s well-being. The aim of the current study was to examine resting HRV in children with chronic pain compared to healthy control children and, further, to examine children’s HRV following a series of acute experimental pain tasks in both groups. Participants included 104 healthy control children and 48 children with chronic pain aged 8–17 years. The laboratory session involved a 5-minute baseline electrocardiogram followed by four pain induction tasks: evoked pressure, cold pressor, focal pressure, and a conditioned pain modulation task. After the tasks were complete, a 5-minute post-task electrocardiogram recording was taken. Spectral analysis was used to capture high-frequency normalized power and the ratio of low-to-high frequency band power, signifying cardiac vagal tone and sympathetic balance, respectively. Results revealed that children with chronic pain had significantly lower resting HRV (signified by low high-frequency normalized power and high ratio of low-to-high frequency band power) compared to healthy children; moreover, a significant interaction between groups and time revealed that children with chronic pain displayed a static HRV response to the pain session compared to healthy children, whose HRV was reduced concomitant with the pain session. These findings suggest that children with chronic pain may have a sustained stress response with minimal variability in response to new acute pain stressors.