Prolonged opioid treatment leads to a comprehensive cellular adaptation mediated by opioid receptors, a basis to understand the development of opioid tolerance and dependence. However, the molecular mechanisms underlying opioid-induced cellular adaptation remain obscure. Recent advances in opioid receptor trafficking and signaling in cells have extensively increased our insight into the network of intracellular signal integration. This review focuses on those important intracellular biochemical processes that play critical roles in the development of opioid tolerance and dependence after opioid receptor activation, and tries to explain what happens after opioid receptor activation, and how the cellular adaptation develops from cell membrane to nucleus. Decades of research have delineated a network on opioid receptor trafficking and signaling, but the challenge remains to explain opioid tolerance and dependence from a single cellular signal network.