Major depressive disorder is associated with lowered mood, anxiety, anhedonia, sleep problems, and cognitive impairments. Many of these functions are regulated by μ-opioid receptor (MOR) system. Preclinical, in vivo, and post-mortem studies have however yielded inconclusive results regarding the role of the MOR in depression and anxiety. Moreover, it is not known whether alterations in MOR are already present in subclinical depression and anxiety. In a large-scale retrospective cross-sectional study we pooled data from 135 (113 males and 22 females) healthy subjects whose brain’s MOR availability was measured with positron emission tomography (PET) using an agonist radioligand [C]carfentanil that has high affinity for MORs. Depressive and anxious symptomology was addressed with BDI-II and STAI-X questionnaires, respectively. Both anxiety and depression scores in the subclinical range were negatively associated with MOR availability in cortical and subcortical areas, notably in amygdala, hippocampus, ventral striatum, and orbitofrontal and cingulate cortices. We conclude that dysregulated MOR availability is involved in altered mood and pathophysiology of depression and anxiety disorders.