Objective The aim of the present study was to investigate the cardiovascular autonomic control in clinically definite multiple sclerosis (MS) patients with a standardised battery of cardiovascular tests and to correlate these findings with the brain magnetic resonance imaging (MRI) lesion load. Methods Fifty-one patients with MS and 50 healthy controls were studied. Brain MRI was performed in all patients showing typical MS lesions. The cardiovascular tests were carried out using a standardised battery. Results Heart rate (HR) responses to deep breathing (P<0.05) and tilt table testing (P<0.001) were significantly decreased in MS patients when compared to those of the controls. Blood pressure (BP) responses in the tilt table test were also impaired in MS patients (diastolic P<0.001, systolic P<0.05). Of the different brain areas investigated the total volume of the midbrain MRI lesions (P<0.05) was the one most clearly associated with the impaired BP responses. Conclusions MS results in both reduced HR variation and decreased BP reactions indicating disturbed cardiovascular regulation. In particular, the midbrain lesions found in MS are associated with cardiovascular dysfunction.