The large intestine plays a major role in the regulation of whole-body fluid and salt homeostasis. Every day 1-1.5 l of fluid enters the colon with only 0.1-0.2 l being lost in the faeces under normal conditions. The balance between Na+ absorption and Cl- secretion underpin intestinal fluid movement1. Oestrogen, (17β-estradiol, E2), produces a female sex-specific, anti-secretory effect in rat and human distal colon2. This response involves the inhibition of KCNQ1:KCNE3 potassium channel activity via PKCδ signalling3. The aim of this work was to determine the molecular mechanisms underpinning E2 modulation of colonic electrolyte homeostasis using genetically modified mouse models. Experiments were carried out on wild-type (WT) and genetically modified IK-1-KO and PKCδ dominant-negative mice (DN-PKCδ) bred on a C57Bl/6J background aged 2-4 months. All experimental procedures were approved by the CECs Institutional Animal Care and Use Committee (IACUC). In electrophysiological experiments on distal colonic epithelium in WT mice, we demonstrated an antisecretory effect of E2 (100nM) which was female sex-specific and estrus-cycle dependent (n=6). This effect was selective for Cl- secretion sensitive to cAMP. E2 inhibited KCNQ1:KCNE3 ionic currents, without effecting IK-1 potassium channel (n=4) or apical CFTR chloride channel currents (n=5). The E2 antisecretory effect was strongly inhibited in DN-PKCδ female mice (n=6), demonstrating the key role of PKCδ activity in determining sexual dimorphism of the E2 anti-secretory response. Oestrogen also exerted a pro-absorptive effect on Na+ absorption via activation of the ENaC sodium channel. E2 injection (1µg/kg BW s.c 24h) in ovariectomised female WT mice enhanced the ENaC current, revealing a pro-absortive effect which was suppressed using the oestrogen receptor antagonist ICI 182,780 (5mg/kg BW s.c during 3 days) (n=3). The stimulatory effect of E2 on ENaC currents was suppressed in the DN-PKCδ female mice. Here, we demonstrate, for the first time, a combined anti-secretory and pro-absorptive response to oestradiol in colonic epithelium. We propose that the combined anti-secretory and pro-absorptive responses to E2 are the molecular basis for fluid retention in females under high oestrogen states.