The hypothalamus is a limbic structure involved in the emergence and persistence of major depressive disorder symptoms. Previous studies have indicated that major depressive disorder patients exhibited dysregulation between the hypothalamus and cerebral regions. However, it is still unclear about the exact hypothalamic functional connectivity patterns with other brain regions based on resting-state functional MRI in major depressive disorder. Here, we investigated the whole-brain voxel-based hypothalamic resting-state functional connectivity in 55 patients with major depressive disorder and 40 age sex-matched healthy controls. The results showed that major depressive disorder patients had a significant decrease in resting-state functional connectivity of the bilateral hypothalamus with the right insula, superior temporal gyrus, inferior frontal gyrus, and Rolandic operculum compared with healthy controls. This study suggests that the pathophysiology of major depressive disorder might be associated with the abnormal hypothalamic resting-state functional connectivity.