Connexin-43 in relation to gap junctions between astrocytes and between other cell types in rat spinal cord was investigated immunohistochemically. In gray matter, connexin-43 was distributed thoughout all laminae, but was more concentrated in the substantia gelatinosa and around the central canal. Ultrastructurally, immunostaining was present in the cytoplasm of, and at gap junctions between, fine astrocytic processes, most of which ensheathed neuronal elements. In white matter, connexin-43 was localized to somata of fibrous astrocytes, their glial fibrillary acidic protein-positive processes running parallel to myelinated axons, and at gap junctions between these processes. Labelling was also evident in thick radially-directed astrocytic processes displaying pockets of staining near immunopositive gap junctions. Near the cord surface, staining was present in cell bodies of subpial astrocytes and at gap junctions between their tangential processes which formed most of the glia limitans. Radially-directed processes of subpial astrocytes formed symmetrically- and asymmetrically-labelled gap junctions with each other and extended fine branches into surrounding white matter where they made contact and often formed gap junctions with oligodendrocytic processes at the outer surface of myelinated fibres. Immunopositive astrocyte processes also made heterologous gap junctions with unstained oligodendrocyte cell bodies. Ependymal cells lining the central canal exhibited apical cytoplasmic labelling, as well as symmetrically-labelled gap junctions at their apices. Ependymal cells also formed asymmetrically-labelled gap junctions at which the junctional membranes of unlabelled cells, presumed to be tanycytes, were unstained. The results indicate the expression of connexins in addition to connexin-43 at asymmetrically-labelled gap junctions between some astrocytic processes, between astrocytes and oligodendrocytes and between some ependymal cells. The presence of gap junctions between astrocyte and oligodendrocyte processes at the outer surface of myelin suggests incorporation of the latter into the extensive gap junctionally-coupled astrocytic syncytium.