The chronic administration of morphine and related opioid drugs results in tolerance and dependence which limits the clinical utility of these agents. Neuronal plasticity is probably responsible in large part for tolerance and dependence. Ca²⁺/calmodulin-dependent protein kinase II (CaMKII) plays a crucial role in the neuroplastic events underlying memory formation and other phenomena. However, the role of this kinase in morphine tolerance remains unclear. To clarify this issue we explored mRNA and protein expression of CaMKIIα in spinal cord tissue from control and morphine treated mice using real-time polymerase chain reaction, Western blot analysis and confocal microscopy. Our chronic exposure paradigm involved the subcutaneous implantation of morphine pellets for 6 days prior to tissue analysis. The results indicate that the levels of CaMKIIα mRNA and protein were robustly increased in spinal cord tissue from morphine-treated mice. Confocal microscopy demonstrated that the increase in CaMKIIα expression was primarily localized to superficial laminae of the dorsal horn. In addition, the abundance of phosphorylated CaMKIIα was increased in spinal cord tissue from morphine-treated mice. We conclude that enhanced CaMKIIα expression and activity in spinal cord tissue may contribute to the development of morphine tolerance in mice. The involvement of this enzyme in opioid tolerance suggests other parallels may exist between the neuroplastic events related to memory formation and those related to opioid tolerance or pain.