Alterations of both arterial and ion channel function have been reported, not only in human and animal models of diabetes, but also in subjects with insulin resistance. However, few such studies have been carried out in the Zucker Diabetic Fatty (ZDF) rat, a model of spontaneous non-insulin dependent diabetes. We have therefore investigated the role of Ca²⁺ -activated K⁺ channels (BK_Ca) in small mesenteric arteries of these animals. Arterial tension changes were examined in humanely killed 17-20 week old male ZDF (fa/fa) animals with approximately 7-10 weeks of established diabetes (glucosuria), together with age-matched lean (fa/+) control animals. Vessel tension was measured in small (<300 μm) mesenteric arteries in the presence of indomethacin (10 μM) and L-nitroarginine (100 μM). Possible changes in BK_Ca activity were evaluated firstly, using cumulative phenylephrine concentration-response curves, in the presence and absence of 100nM iberiotoxin (IbTX) and secondly, using sharp-microelectrode recordings of myocyte membrane potential in the absence and presence of the BK_Ca opener, NS1619. Non-linear regression curve-fit (GraphPad Prism 4) comparison of phenylephrine concentration-response curves from vessels of ZDF and lean animals in the absence of IbTX showed no differences. However, in vessels from control animals IbTX produced a small but significant leftward shift (lean log EC₅₀ -6.38; lean+IBTX log EC₅₀ -6.68, p<0.003, ANOVA, n = 11 and 8, respectively) in the phenylephrine concentration-response curve. In contrast no differences in log EC₅₀ values were seen in arteries from ZDF animals in the absence or presence of IbTX (ZDF v ZDF+IbTX). Sharp-microelectrode experiments indicated that resting membrane potentials of lean and ZDF arteries were similar (lean 54.2± 0.5mV v ZDF -54.7 ± 0.5mV, mean ± SEM). Application of NS1619 (33 μM) induced a significantly larger hyperpolarisation in lean (Δ-17.2 ± 0.5mV, n = 8) compared with ZDF arteries (Δ-9.8 ± 1.1mV, n = 6). These results demonstrate that BK_Ca activity in ZDF animals is reduced compared to lean controls. The impairment of BK_Ca seen upon addition of NS1619 suggests that diabetes affects channel function directly rather than the coupling between phenylephrine stimulation and activation of BK_Ca.