Stimulation of histamine H₁ receptors produced a marked activation of inositol phospholipid hydrolysis, intracellular calcium mobilization, and stimulation of the c-fos promoter in CHO-H1 cells expressing the H₁ receptor at a level of 3 pmol/mg protein. The latter response was determined using a luciferase-based reporter gene (pGL3). This response to histamine was not sensitive to inhibition by pertussis toxin but could be completely attenuated by the protein kinase C (PKC) inhibitor Ro-31-8220, or by 24-h pretreatment with the phorbol esters phorbol 12,13-dibutyrate or phorbol-12-myristate-13-acetate. Several isoforms of PKC can be detected in CHO-H1 cells (α, δ, ε, μ, ι, ζ) but only PKCα and PKCδ were down-regulated by prolonged treatment with phorbol esters. Of the two isoforms that were down-regulated, only protein kinase Cα was translocated to CHO-H1 cell membranes after stimulation with either histamine or phorbol esters. The PKC inhibitor Gö6976, which inhibits PKCα but not PKCδ, was also able to significantly attenuate the c-fos-luciferase response to histamine. The mitogen-activated protein kinase kinase inhibitor PD 98059 markedly inhibited the response to histamine, suggesting that the likely major target for PKCα was the mitogen-activated protein kinase pathway. These data suggest that the histamine H₁ receptor can signal to the nucleus via PKCα after activation of phospholipase Cβ.