The antinociceptive and analgesic effects of muscarinic receptor ligands in human and nonhuman species have been evident for more than half a century. In this review, we describe the current understanding of the roles of different muscarinic subtypes in pain modulation and their mechanism of action along the pain signaling pathway, including peripheral nociception, spinal cord pain processing, and supraspinal analgesia. Extensive preclinical and clinical validation of these mechanisms points to the development of selective muscarinic agonists as one of the most exciting and promising avenues toward novel pain medications.