Abstract: Norepinephrine (NE)-stimulated 3H-inositol phosphate (3H-InsP) formation in primary glial cell cultures is thought to be due to α1-adrenergic receptor activation. Surprisingly, the α1-selective agonists phenylephrine and methoxamine showed only 12–21% of the intrinsic activity of NE in activating this response. Although the α2-selective agonist UK 14,304 was itself inactive, inclusion of UK 14,304 increased the response to the α1-selective agonists by about threefold. This increase was concentration-dependent and occurred at all time points examined. 6-Fluoro-NE and α-methyl-NE mimicked the effect of NE in glial cultures, although with lower potencies. However, several partial agonists were ineffective in activating this response, in both the presence and absence of UK 14,304. Synergistic interactions were not observed for α1-mediated responses in slices of rat cerebral cortex, either for formation of 3H-InsPs or potentiation of isoproterenol- or adenosine-stimulated cyclic AMP accumulation. Both UK 14,304 and phenylephrine inhibited NE-stimulated 3H-InsP formation in concentrations similar to those necessary to activate this response directly. These results suggest that NE activates 3H-InsP formation in primary glial cultures by synergistic actions on both α1- and α2-adrenergic receptors. The agonists UK 14,304 and phenylephrine also can act to inhibit the response to NE competitively.