Original article Transgenic overexpression of SUR1 in the heart suppresses sarcolemmal K ATP Thomas P. Flagg a, *, Maria Sara Remedi a , Ricard Masia a , Jefferson Gomes a , Meredith McLerie b , Anatoli N. Lopatin b , Colin G. Nichols a a Department of Cell Biology and Physiology, Washington University School of Medicine, 660 S. EuclidAvenue, Box 8228, St. Louis, MO 63110, USA b Department of Physiology, University of Michigan Medical School, Ann Arbor, MI 48109, USA Received 28 January 2005; received in revised form 29 April 2005; accepted 6 June 2005 Available online 15 August 2005 Abstract The lack of pathological consequences of cardiac ATP-sensitive potassium channel (K ATP ) channel gene manipulation is in stark contrast to the effect of similar perturbations in the pancreatic b-cell. Because the pancreatic and cardiac channel share the same pore-forming subunit (Kir6.2), the different effects of genetic manipulation likely reflect, at least in part, the tissue-specific expression of the regulatory subunit (SUR1 in pancreas vs. SUR2A in heart) of the bipartite channel complex. To examine this, we have generated transgenic (TG) mice that overexpress epitope-tagged SUR1 or SUR2A under the transcriptional control of the a-myosin heavy chain promoter. Western blot and real time RT-PCR analysis confirm transgene expression in the heart, and variable levels of SUR1 RNA and protein, in 16 viable founder lines. Surprisingly, activation of channels by either pharmacological agents (diazoxide and pinacidil) or metabolic inhibitors (oligomycin and 2-deoxyglucose) reveals a suppression of total K ATP conductance in high expressing TG mice. Moreover, K ATP channel activity was signifi- cantly reduced in excised cardiac patches from TG myocytes that overexpress either SUR1 or SUR2A. Using a recombinant cell system, we show that overexpression of either SUR1 or Kir6.2 suppresses the functional expression of K ATP from optimized dimeric SUR1-Kir6.2. Thus, the graded effect of SUR1 expression in the intact heart appears to demonstrate an in vivo requirement for 1:1 expression ratio of Kir6.2 and SURx. © 2005 Elsevier Ltd. All rights reserved. Keywords: K ATP ; SUR1; Cardiac; Stoichiometry; Transgenic 1. Introduction ATP-sensitive potassium channels (K ATP ) channels are expressed in a diverse set of tissues, including the heart [1] and the pancreas [2]. By responding to physiological changes in the ratio of [ADP] to [ATP], b-cell K ATP channels control the membrane potential, entry of Ca 2+ through voltage- dependent channels, and hence insulin secretion. Consistent with this fundamental role in metabolism-secretion cou- pling, activating mutations in the genes encoding the K ATP channel cause permanent neonatal diabetes in both mice [3] and humans [4]. In cardiac myocytes, however, the physi- ological role of K ATP is less established. Under normal meta- bolic conditions, K ATP channels are not significantly open and thus do not contribute to action potential repolarization and excitation–contraction coupling [5,6]. When myocytes are exposed to a severe metabolic stress (anoxia, hypoxia, meta- bolic inhibition (MI), ischemia, etc.) K ATP channels open, causing action potential shortening and contractile failure [7–9]. However, the precise trigger for K ATP activation, the timing of channel opening and physiological consequences in vivo remain unclear. Action potential duration and myo- cardial contractility in mice that lack K ATP (Kir6.2–/–) are similar to the APD and contractility in wild type mice [8]. The principal phenotypes of the Kir6.2–/– mice are loss of action potential shortening in response to MI, ischemia and stress [9,10], and abolition of ischemic preconditioning [9]. It is well established that K ATP channels are multiprotein complexes that consist of at least two principal subunits [11]. A member of the inward rectifying K + (Kir) channel protein family forms the potassium selective pore. A member of the * Corresponding author. Tel.: +1 314 362 6629; fax: +1 314 362 7463. E-mail address: tflagg@cellbiology.wustl.edu (T.P. Flagg). Journal of Molecular and Cellular Cardiology 39 (2005) 647–656 www.elsevier.com/locate/yjmcc 0022-2828/$ - see front matter © 2005 Elsevier Ltd. All rights reserved. doi:10.1016/j.yjmcc.2005.06.003