The role of protein kinase C (PKC) in contraction of the human myometrium induced by endothelin-1 (ET-1) was investigated at the end of pregnancy. The expression and subcellular distribution of PKC isoforms were examined by Western blot analysis using isoform-specific antibodies. At least three conventional PKC isoforms (cPKC; α, β1, and β2), two novel PKC isoforms (ε and δ), and an atypical PKC isoform (ζ) were detected in pregnant myometrium. Quantitative immunoblotting revealed that all these isoforms were mainly distributed in the particulate fraction. The lack of a calcium chelator to modify the particulate sequestration of cPKC suggests an interaction with an anchoring protein such as receptor-activated C kinase-1, which is evidenced in the particulate fraction of the pregnant myometrium. Of the six isoforms, only PKCβ1, PKCβ2, PKCδ, and PKCζ were translocated to the particulate fraction, and PKCε to the cytoskeletal fraction, after stimulation with ET-1. Involvement of PKC in the ET-1-induced contractile response is supported by the inhibition caused by the PKC inhibitor calphostin C. However, we demonstrated that the selective cPKC isoform inhibitor, Gö 6976, as well as the substantial depletion of PKCβ1 and PKCε and the partial depletion of PKCα and PKCδ by a long-term treatment with phorbol 12,13-dibutyrate did not prevent ET-1-induced contraction. Accordingly, our results suggest that PKCδ and PKCζ activation mediated ET-1-induced contraction, whereas cPKC isoforms were not implicated in the human pregnant myometrium.