Nuclear Ca²⁺/calmodulin translocation activated by μ-opioid (OP₃) receptor

Previous evidence has suggested a role for calmodulin (CAM) in opioid receptor signaling. We demonstrate here that morphine stimulation of the μ- opioid (OP₃) receptor causes rapid CaM translocation to the nucleus in OP₃- transfected human embryonic kidney (HEK)-293 cells and in SH-SY5Y human neuroblastoma cells. Ca²⁺ influx into the cells resulting from OP₃ receptor activation was required for nuclear CaM translocation. Moreover, in HEK-OP₃ and SH-SY5Y cells, increased nuclear CaM content was associated with enhanced phosphorylation of the nuclear transcription factor cyclic AMP- responsive element-binding protein. This appeared to be mediated by Ca²⁺/CaM kinases and also by a pathway involving protein kinase C. CaM was previously shown to bind directly to the OP₃ receptor and to be released from the plasma membrane on agonist stimulation. To test whether OP₃- mediated CaM release contributes to nuclear CaM signaling, we used a mutant OP₃ receptor (K273A) with reduced affinity for CaM that fails to release CaM from the plasma membrane. K273A-OP₃ activated Ca²⁺ influx to a similar extent as wild-type OP₃; however, CaM translocation to the nucleus was attenuated. These results indicate that OP₃-stimulated Ca²⁺ influx results in nuclear CaM translocation, which appears to be enhanced by simultaneous CaM release by OP₃ wild-type receptor from plasma membranes. These results suggest a novel Ca²⁺/CaM signaling pathway of opioid receptors in the regulation of transcriptional activity.