Background and Purpose: The role of peripheral mu-opioid receptors (MOPs) in chronic pain conditions is not well understood. Here, we used a combination of mouse genetics, behavioral assays, and pharmacological interventions to investigate the contribution of MOPs on primary afferent neurons to nociceptive, inflammatory, and neuropathic pain as well as to opioid analgesia. Experimental Approach: We generated conditional knockout mice in which MOPs were selectively deleted in primary sensory neurons. Inflammatory and neuropathic pain states were induced in mutant and control wild-type mice and their behavioral responses to noxious stimuli compared. Gross motor function was also evaluated. Immunohistochemistry was used to assess MOP expression in the dorsal root ganglia, periaqueductal gray, and small intestine. Behavioral assays and whole-cell patch-clamp recordings were used to evaluate the inhibitory effects of mu-opioid agonists, DALDA and morphine, on pain behavior and neurophysiologic effects in DRG neurons, respectively. Key Results: Conditional MOP knockouts and control mice exhibited similar behavioral responses to acute nociceptive stimuli and developed similar inflammation-induced hypersensitivity. Nerve injury in animals lacking peripheral MOPs induced enhanced, bilateral mechanical allodynia. DALDA administered subcutaneously was unable to decrease the hypersensitivity induced by inflammation and nerve injury in MOP knockout animals, and morphine’s antinociceptive effects were significantly attenuated in the absence of peripheral MOPs. Conclusion and Implication: MOPs in primary sensory neurons contribute to the modulation of neuropathic pain behavior and opioid analgesia. Our observations highlight the clinical potential of peripherally acting opioid agonists in the management of chronic inflammatory and neuropathic pain.