Calcium Signaling and Contractility in Cardiac Myocyte of Wolframin Deficient Rats.

Michal Cagalinec,Ludovit Paulis,Matej Hot’Ka,Ivan Zahradník,Jana Pavelková,Simona Kureková,Mario Plaas,Marta Novotová,Alexandra Zahradníková,Dominika Kováčová

doi:10.3389/fphys.2019.00172

Abstract

Wolframin (Wfs1) is a membrane protein of the sarco/endoplasmic reticulum. Wfs1 mutations are responsible for the Wolfram syndrome, characterized by diabetic and neurological symptoms. Although Wfs1 is expressed in cardiac muscle, its role in this tissue is not clear. We have characterized the effect of invalidation of Wfs1 on calcium signaling-related processes in isolated ventricular myocytes of exon5-Wfs1 deficient rats (Wfs1-e5/-e5) before the onset of overt disease. Calcium transients and contraction were measured in field-stimulated isolated myocytes using confocal microscopy with calcium indicator fluo-3 AM and sarcomere length detection. Calcium currents and their calcium release-dependent inactivation were characterized in whole-cell patch-clamp experiments. At 4 months, Wfs1-e5/-e5 animals were euglycemic, and echocardiographic examination revealed fully compensated cardiac function. In field-stimulated isolated ventricular myocytes, both the amplitude and the duration of contraction of Wfs1-e5/-e5 animals were elevated relative to control Wfs1+/+ littermates. Increased contractility of myocytes resulted largely from prolonged cytosolic calcium transients. Neither the amplitude of calcium currents nor their voltage dependence of activation differed between the two groups. Calcium currents in Wfs1-e5/-e5 myocytes showed a larger extent of inactivation by short voltage prepulses applied to selectively induce calcium release-dependent inactivation of calcium current. Neither the calcium content of the sarcoplasmic reticulum, measured by application of 20 mmol/l caffeine, nor the expression of SERCA2, determined from Western blots, differed significantly in myocytes of Wfs1-e5/-e5 animals compared to control ones. These experiments point to increased duration of calcium release in ventricular myocytes of Wfs1-e5/-e5 animals. We speculate that the lack of functional wolframin might cause changes leading to upregulation of RyR2 channels resulting in prolongation of channel openings and/or a delay in termination of calcium release.

Highlights

Wolfram syndrome (WS; OMIM 222300) is a rare hereditary disorder, first identified as a clinical entity separate from the juvenile type of diabetes mellitus by Wolfram and Wagener (1938)
Since calcium ions are the sole trigger for myocyte contraction and Wfs1 is strongly involved in calcium metabolism, in this work we evaluated calcium transients and contractility of left ventricular myocytes freshly isolated from the exon5-Wfs1 deficient (Wfs1−e5/−e5) rats
Tests for postprandial blood glucose showed slightly but significantly higher glycemia in Wfs1−e5/−e5 (7.79 ± 0.47 mmol/l, n = 8) than in Wfs1+/+ animals (6.10 ± 0.16, n = 6, P = 0.009) but no hyperglycemia in either group, in agreement with the original study for this age group (Plaas et al, 2017). This confirms that the cohort of experimental animals used in this study was not biased by specific or unspecific side effects related to breeding

Summary

Introduction

Wolfram syndrome (WS; OMIM 222300) is a rare hereditary disorder, first identified as a clinical entity separate from the juvenile type of diabetes mellitus by Wolfram and Wagener (1938). WS is characterized by symptoms including diabetes insipidus (DI), diabetes mellitus (DM), optical atrophy (OA), and deafness (D), and it is termed DIDMOAD (Barrett et al, 1995). The Wfs protein is highly expressed in the brain, heart and pancreatic β-cells (Inoue et al, 1998; Yamada et al, 2006); pancreas and brain represent the crucial organs responsible for most of clinical symptoms in WS. Cardiac symptoms of WS were not originally recognized; emerging clinical findings include heart malformations as well as sinus tachycardia, atrial or ventricular arrhythmias (Medlej et al, 2004; Fabbri et al, 2005; Ganie et al, 2011; Korkmaz et al, 2016). The high expression of Wfs in the heart tissue, and the cardiac symptoms identified until now suggest functional importance of Wfs in the heart

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