ATP-sensitive inward-rectifying K+ (KATP) channels play a vital role in coupling cell metabolic activity to the electrical activity of the plasma membrane (Ashcroft & Rorsman 1990). In cells expressing these channels, which include pancreatic beta cells and neurons, increases in ATP levels reduce the KATP activity triggering membrane depolarization and a range of cellular responses, including release of insulin from beta cells. Conversely, reduced cellular metabolism opens KATP channels, leading to a hyperpolarization of the membrane potential and suppression of electrical activity. One class of drug, the sulfonylureas, which are commonly used as anti-diabetic drugs, akin to ATP close the channel. However, chronic sulfonylurea treatment has been shown to cause progressive long-term insulin secretory failure (Matthews et al. 1998; Remedi & Nichols 2008) . Interestingly, patients with mutations that render KATP channels overactive display a higher frequency of secondary treatment failure (Sesti et al. 2006). These studies indicate that activity of KATP channels may play a role in the homeostatic regulation of intrinsic cellular excitability. In this study we have investigated the effect of chronic sulfonylurea treatment on cellular electrophysiological properties in two different KATP expressing cell-lines. Cultures of the rat insulinoma cell line (INS-1) or the mouse hypothalamic cell line (GT1-7) were pre-incubated with either; vehicle (Cntl) or Glibenclamide (Glib,100 nM) for ~48 hrs. Whole-cell patch-clamp recordings were then made at 33 ± 1 °C in the absence of any drug. The pipette solution lacked ATP. To characterize both leak and voltage-gated current components voltage steps of 100 ms duration were applied from a holding potential of -70 mV to between -80 and +60 mV in 10 mV increments. In INS-1 cells, pre-treatment with glibenclamide significantly depolarised the resting membrane potential (VREST) (mV: -73.3±5.2 (Cntl, n=17) vs. -58.2±5.1 (Glib, n=18), p = 0.046) and decreased the normalised conductance of the background leak current (nS/pF: 0.8±0.18 (Cntl, n=17) vs. 0.3±0.07 (Glib, n=18), p = 0.02). Conversely, in GT1-7 cells a significant depolarisation in VREST was not observed after chronic administration of glibenclamide (mV: -91.6±1.29 (Cntl, n=13) vs. -87±2.04 (Glib, n=16), p = 0.08). However, similar to INS-1 cells, there was a reduction in the capacitance-normalized conductance of the leak current (nS/pF: 1.02±0.19 (Cntl, n=13) vs. 0.52±0.07 (Glib, n=16), p = 0.026). Depolarization-activated outward current components that activate positive from ~-40 mV were not changed by glibenclamide pretreatment. These data suggest that a period of chronic block of KATP channels produces lasting changes to intrinsic membrane properties that could affect both cellular physiology and drug efficacy.