The use of opioids, which achieve therapeutic analgesia through activation of μ-opioid receptors, are limited in the management of chronic pain by adverse effects including tolerance and addiction. Optogenetics is an emerging approach of designing molecular targets that can produce cell-specific receptor-mediated analgesia with minimal side effects. Here we report the design and functional characterization of a chimeric μ-opioid receptor that could be photoactivated to trigger intracellular signaling. A prototype optoactive μ-opioid receptor (optoMOR) was designed by replacing the intracellular domains from rhodopsin with those of the native μ-opioid receptor and was transiently expressed in human embryonic kidney (HEK293) cells. Expression and distribution of the protein were confirmed by immunocytochemistry. The signal-transduction mechanisms induced by photoactivation of the optoMOR were evaluated and compared with the native μ-opioid receptor stimulation by an agonist, D-Ala2, N-MePhe4, Gly-ol-enkephalin (DAMGO). Cells were depolarized by extracellular potassium and the depolarization-induced calcium (Ca2+) influx was quantified by using Fura-2 imaging. The forskolin-stimulated adenylate cyclase/cAMP cascade was evaluated by ELISA or western blotting of brain-derived neurotrophic factor (BDNF) and the phosphorylation of cAMP response element binding protein (CREB). The optoMOR protein distribution was observed intracellularly and on the plasma membrane similar to the native μ-opioid receptor in HEK293 cells. Photoactivation of optoMOR decreased the Ca2+ influx and inhibited the forskolin-induced cAMP generation, activation of CREB, and BDNF levels in optoMOR-expressing cells similar to the activation of native μ-opioid receptor by DAMGO. Thus the current study has accomplished the design of a prototype optoMOR and characterized the cellular signaling mechanisms activated by light stimulation of this receptor.