Original article Dyssynchronous (non-uniform) Ca 2+ release in myocytes from streptozotocin-induced diabetic rats Chun-Hong Shao a , George J. Rozanski b , Kaushik P. Patel b , Keshore R. Bidasee a, ⁎ a Department of Pharmacology and Experimental Neuroscience, University of Nebraska Medical Center, Omaha, NE 68198-5800, USA b Department of Cellular and Integrative Physiology, University of Nebraska Medical Center, Omaha, NE 68198-5850, USA Received 14 March 2006; received in revised form 3 August 2006; accepted 28 August 2006 Available online 6 October 2006 Abstract Using biochemical/pharmacological approaches, we previously showed that type 2 ryanodine receptors (RyR2) become dysfunctional in hearts of streptozotocin-induced type 1 diabetic rats. However, the functional consequence of this observation remains incompletely understood. Here we use laser confocal microscopy to investigate whether RyR2 dysfunction during diabetes alters evoked and spontaneous Ca 2+ release from the sarcoplasmic reticulum (SR). After 7–8 weeks of diabetes, steady-state levels of RyR2 remain unchanged in hearts of male Sprague-Dawley rats, but the number of functional receptors decreased by >37%. Interestingly, residual functional RyR2 from diabetic rat hearts exhibited increased sensitivity to Ca 2+ activation (EC 50activation decreased from 80 μM to 40 μM, peak Ca 2+ activation decreased from 425 μM to 160 μM). When field stimulated, intracellular Ca 2+ release in diabetic ventricular myocytes was dyssynchronous (non-uniform) and this was independent of L-type Ca 2+ currents. Time to peak Ca 2+ increased 3.7-fold. Diabetic myocytes also exhibited diastolic Ca 2+ release and 2-fold higher frequency of spontaneous Ca 2+ sparks, albeit at a lower amplitude. The amplitude of caffeine-releasable Ca 2+ was also lower in diabetic myocytes. RyR2 from diabetic rat hearts exhibited increased phosphorylation at Ser2809 and contained reduced levels of FKBP12.6 (calstablin2). Collectively, these data suggest that RyR2 becomes leaky during diabetes and this defect may be responsible to the reduced SR Ca 2+ load. Diastolic Ca 2+ release could also serve as a substrate for delayed after-depolarizations, contributing to the increased incidence of arrhythmias and sudden cardiac death in type 1 diabetes. © 2006 Elsevier Inc. All rights reserved. Keywords: Diabetes; Ryanodine receptors; Dysregulation; Ca 2+ sparks; Ca 2+ transients; Rat 1. Introduction Type 1 diabetes (T1D) is the most common metabolic syndrome in children under the age of 18 years [1]. It is a polygenic, multifactorial autoimmune disease in which T lymphocytes infiltrate into the pancreas and selectively destroy the insulin-producing β-cells in the islet of Langerhans. Polymorphic regions of HLA class II alleles account for 40% of genetic susceptibility to T1D [2]. Environmental factors including enteroviruses, cow's milk proteins, wheat gliadin, N-nitroso compounds and vitamin D deficiency serve as triggers for the destruction of the β-cells [3]. Although T1D accounts for about 10% of the total diabetic population, its impact is especially devastating since a high rate of complication can occur at a relatively young age. One of these complications is heart failure. Individuals with T1D have heart failure at rates three to five times higher than that of the general population and this can occur independent of arteriosclerosis and/or hypertension [4]. This diabetic cardiomyopathy (DC) or “diabetic heart muscle disease” starts as an asymptomatic slowing in relaxation kinetics (diastolic dysfunction) [5]. As the syndrome progresses, systolic function (ejection fraction, fractional shortening and stroke volume) also becomes compromised, increasing morbidity and mortality [6]. To date, the etiology underlying myocardial contractility loss during T1D remains incompletely understood. However, studies suggest that this defect stems in part from perturbation of sarcoplasmic reticular Ca 2+ cycling [7–10]. Cardiac muscle contraction depends critically on the timely and coordinated release of Ca 2+ from the internal SR. Following Journal of Molecular and Cellular Cardiology 42 (2007) 234 – 246 www.elsevier.com/locate/yjmcc ⁎ Corresponding author. 985800 Nebraska Medical Center, Durham Research Center, DRC 3047, Omaha, NE 68198-5800, USA. Tel.: +1 402 559 9018; fax: +1 402 559 7495. E-mail address: kbidasee@unmc.edu (K.R. Bidasee). 0022-2828/$ - see front matter © 2006 Elsevier Inc. All rights reserved. doi:10.1016/j.yjmcc.2006.08.018