OBJECTIVE Fibromyalgia (FM) is a syndrome associated with widespread pain and various other signs and symptoms. Several of these multisystem features could be explained on the basis of autonomic nervous system (ANS) dysfunction. METHODS The aim of the present study was to evaluate ANS dysfunction in FM based on time-domain heart rate variability (HRV) analysis and serum neuropeptide Y (NPY) levels in 51 patients with FM, 25 patients with systemic sclerosis (SSc), and 15 healthy controls (NHS). RESULTS Compared with the SSc and NHS groups, the FM group had significantly higher NPY levels, and in the FM subgroup subjected to HRV analysis (25/51 patients, 49%), certain HRV indices were significantly reduced. In this subgroup, NPY was significantly correlated with the SDANN index and the NN50, but neither NPY or HRV parameters showed any significant correlation with clinical aspects of the FM. CONCLUSIONS These findings suggest that autonomic dysfunction and NPY are crucial elements in the pathophysiology of FM. Additional studies are necessary to define the complex roles played by NPY and ANS in modulating pain and immunological functions of different diseases.