Astrocytes are characterized by extensive gap junctional intercellular communication (GJIC) mediated by channels composed primarily of connexin43. To examine some of the functions of this intercellular communication in glial cells, we have used three approaches. The first involves transfection of glioma cells, which are deficient in connexin expression and gap junctional communication, with connexin cDNAs to examine changes in cellular phenotype following increased gap junctional communication. Using differential display, we have identified several genes which appear to be regulated by GJIC. The second is to study astrocytes cultured from embryonic mice with a null mutation in the connexin43 gene. These homozygous null astrocytes are devoid of connexin43 and also deficient in intercellular dye transfer. Markers of glial differentiation appear similar in all genotypes. Measurement of intercellular calcium concentration following mechanical stimulation of confluent astrocytes revealed that the number of cells affected by a rise in intracellular calcium was reduced in homozygous cultures compared to wild type. The growth rate of astrocytes lacking connexin43 was reduced compared to wild-type astrocytes. The third approach employs the use of gap junction blockers in a model of neuronal and glial differentiation, namely P19 mouse embryonal carcinoma cells treated with retinoic acid. In this case, blocking GJIC during the differentiation protocol prevents the appearance of neuronal and astrocytic phenotypes. Taken together, these data suggest an important role for GJIC in glial function and differentiation.