The role of metallothionein (MT) in the brain in heavy metal detoxification is relatively unexplored. Brain copper (Cu) elevation although unusual in chronic Cu poisoning in sheep, has been shown to occur after treatment with the chelating agent ammonium tetrathiomolybdate (TTM). The aim of this study was to investigate the expression of MT in TTM Cu-enhanced sheep brains, with immunohistochemical techniques. Brains from TTM-treated, Cu-poisoned sheep were examined for MT immunolabelling with a mouse monoclonal antibody (E9) for MT1 and MT2, and compared with brains from untreated sheep. Brain samples were analysed separately for Cu and zinc (Zn) by atomic absorption spectrophotometry. Markedly increased MT immunoreactivity was found in astrocytes of the cerebellum, thalamus/hypothalamus, cerebrum and medulla oblongata of the high-Cu brains, corresponding to the regional Cu elevations. MT immunolabelling was also found in the pia mater, choroid plexus and ependymal cells. Neurons were rarely labelled. MT induction within astrocytes and at the blood-brain barrier suggests that these are sites of stabilization and possibly transport for Cu and supports the hypothesis that the astrocyte compartment modulates metal homeostasis, conferring protection on vulnerable neurons and effecting damage limitation.