Staining of c-fos-like-immunoreactivity (CFIR) in neurones was used to study neuronal activation within subdivisions of periaqueductal gray (PAG), and in locus coeruleus and ventral tegmental area during opiate withdrawal in awake and anaesthetised, morphine-dependent rats. The number of CFIR containing neurones was significantly increased during naloxone-precipitated withdrawal in lateral and ventrolateral, particularly the caudal ventrolateral PAG. No changes were observed in dorsal-intermediate or dorsal-caudal PAG. In awake rats, a similar but more generalised increase in CFIR was observed in PAG following naloxone-precipitated withdrawal. Increases in ventral tegmental area and locus coeruleus during naloxone-precipitated withdrawal under anaesthesia varied greatly between animals. Induction of c-fos in lateral and ventrolateral PAG during withdrawal is consistent with known functions of these regions, involving the integration of autonomic and somatic components of defensive and escape behaviours which are characteristic signs of opiate withdrawal.