We hypothesized that in preeclampsia (PE) contribution of endothelium-derived hyperpolarizing factor (EDHF) and mechanisms of its action differ from that in normal pregnancy (NP). We aimed to assess endothelial function and morphology in arteries from NP and PE with particular focus on EDHF. Arteries (200µm) were dissected from subcutaneous fat biopsies obtained from women undergoing cesarean section. Using wire-myography, responses to the endothelium-dependent agonist bradykinin (BK) were determined before and after inhibition of pathways relevant to EDHF activity. The overall responses to BK in arteries from PE (n=13) and NP (n=17) were similar. However, in PE EDHF-mediated relaxation was reduced (P<0.05). All women within PE group were divided into two subgroups: with more (1) or less (2) than 50% reduction of EDHF-typed responses after 18--glycyrrhetinic acid (an inhibitor of myoendothelial gap junctions, MEGJs). The division showed that: 1) MEGJs are principally involved when the EDHF contribution is reduced; 2) when EDHF contribution is similar to that in women without PE, the H₂O₂ and/or CYP450 epoxygenase product of arachidonic acid (AA), along with MEGJs, confer EDHF-mediated relaxation. In contrast, MEGJs were the main pathway for EDHF in NP. The abundant presence of MEGJs in arteries from NP but deficiency of them in PE was observed using transmission electron microscopy. We conclude that PE is associated with heterogeneous contribution of EDHF, and the mechanism behind EDHF-typed responses is mediated either by MEGJs alone or in combination with H₂O₂ or CYP450 epoxygenase metabolites of AA.