Phosphoinositide-3-kinase/akt - dependent signaling is required for maintenance of [Ca2+]i,ICa, and Ca2+ transients in HL-1 cardiomyocytes

Graves, Simerly Thomas, Li Chuanfu, Williams, Wondergem, Wondergem Robert

Description

The phosphoinositide 3-kinases (PI3K/Akt) dependent signaling pathway plays an important role in cardiac function, specifically cardiac contractility. We have reported that sepsis decreases myocardial Akt activation, which correlates with cardiac dysfunction in sepsis. We also reported that preventing sepsis induced changes in myocardial Akt activation ameliorates cardiovascular dysfunction. In this study we investigated the role of PI3K/Akt on cardiomyocyte function by examining the role of PI3K/Akt-dependent signaling on [Ca 2+ ] i , Ca 2+ transients and membrane Ca 2+ current, I Ca , in cultured murine HL-1 cardiomyocytes. LY294002 (1–20 μM), a specific PI3K inhibitor, dramatically decreased HL-1 [Ca 2+ ] i , Ca 2+ transients and I Ca . We also examined the effect of PI3K isoform specific inhibitors, i.e. α (PI3-kinase α inhibitor 2; 2–8 nM); β (TGX-221; 100 nM) and γ (AS-252424; 100 nM), to determine the contribution of specific isoforms to HL-1 [Ca 2+ ] i regulation. Pharmacologic inhibition of each of the individual PI3K isoforms significantly decreased [Ca 2+ ] i , and inhibited Ca 2+ transients. Triciribine (1–20 μM), which inhibits AKT downstream of the PI3K pathway, also inhibited [Ca 2+ ] i , and Ca 2+ transients and I Ca . We conclude that the PI3K/Akt pathway is required for normal maintenance of [Ca 2+ ] i in HL-1 cardiomyocytes. Thus, myocardial PI3K/Akt-PKB signaling sustains [Ca 2+ ] i required for excitation-contraction coupling in cardiomyoctyes.

Sujets

• Calcium
• Fura-2
• Electrophysiology

Informations

• Publié par: biomed
• Publié le: 01 janvier 2012
• Nombre de lectures: 8
• Langue: English
• Poids de l'ouvrage: 1 Mo

Extrait

Graveset al. Journal of Biomedical Science2012,19:59 http://www.jbiomedsci.com/content/19/1/59

R E S E A R C HOpen Access Phosphoinositide3kinase/akt  dependent 2+ signaling is required for maintenance of [Ca] i, 2+ I, and Catransients in HL1 cardiomyocytes Ca 1 21 12,3* Bridget M Graves , Thomas Simerly , Chuanfu Li , David L Williamsand Robert Wondergem

Abstract The phosphoinositide 3kinases (PI3K/Akt) dependent signaling pathway plays an important role in cardiac function, specifically cardiac contractility. We have reported that sepsis decreases myocardial Akt activation, which correlates with cardiac dysfunction in sepsis. We also reported that preventing sepsis induced changes in myocardial Akt activation ameliorates cardiovascular dysfunction. In this study we investigated the role of PI3K/Akt on 2+ 2+ cardiomyocyte function by examining the role of PI3K/Aktdependent signaling on [Ca] i transients and, Ca 2+ membrane Cacurrent,I Ca , in cultured murine HL1 cardiomyocytes. LY294002 (1–20μM), a specific PI3K inhibitor, 2+ 2+ dramatically decreased HL1 [Ca] i , Catransients andI Ca . We also examined the effect of PI3K isoform specific inhibitors,i.e.α(PI3kinaseαinhibitor 2; 2–8 nM);β(TGX221; 100 nM) andγ(AS252424; 100 nM), to determine the 2+ contribution of specific isoforms to HL1 [Ca] i regulation. Pharmacologic inhibition of each of the individual PI3K 2+ 2+ isoforms significantly decreased [Ca] i transients. Triciribine (1, and inhibited Ca–20μM), which inhibits AKT 2+ 2+ downstream of the PI3K pathway, also inhibited [Ca] i , and Catransients andI Ca . We conclude that the PI3K/Akt 2+ pathway is required for normal maintenance of [Ca] i in HL1 cardiomyocytes. Thus, myocardial PI3K/AktPKB 2+ signaling sustains [Ca] i required for excitationcontraction coupling in cardiomyoctyes. Keywords:Calcium, Fura2, Phosphoinositide3kinase/Akt, HL1 cardiomyocytes, Wholecell voltage clamp, Electrophysiology

Background The phosphoinositide 3kinases (PI3K) are a conserved family of signal transduction enzymes that are involved in regulating cellular proliferation and survival [1,2]. The PI3Ks and the downstream serine/threonine kinase Akt (also known as protein kinase B; PKB) regulate cellular activation, inflammatory responses, chemotaxis and apoptosis [1]. We [3] and others [4] have demonstrated that activation of PI3K/Akt dependent signaling attenu ates the proinflammatory phenotype and increases sur vival outcome in sepsis. We have also reported that sepsis decreases myocardial Akt activation [5], which correlates with cardiac dysfunction in sepsis. In the same

* Correspondence: [e-mail] 2 Departments of Biomedical Science, James H. Quillen College of Medicine, East Tennessee State University, Johnson City, TN 37614, USA 3 Department of Physiology/Biomedical Science, James H. Quillen College of Medicine, East Tennessee State University, P.O. Box 70,576, Johnson City, TN 376141708, USA Full list of author information is available at the end of the article

report, we demonstrated that preventing sepsisinduced changes in myocardial Akt activation correlates with prevention of cardiac dysfunction [5]. PI3K/Akt/PKB may play a role in cardiomyocyte cal cium regulation; however, the precise mechanisms by which this occurs have not been fully elucidated. Yano and colleagues employed a transgenic mouse model overexpressing PI3K p110αin the heart [6], which resulted in increased left ventricular pressure, increased 2+ levels of Ltype Cachannels, ryanodine receptors and 2+ sarcoplasmic reticulum CaATPase 2a [6]. In a subse quent report, Lu et al. demonstrated that genetic abla tion of PI3K p110αresulted in reduced numbers of 2+ voltagedependent Ltype Cachannels in isolated car 2+ diomyocytes, reduced inward Cacurrent and a defect in contractile function [7]. Taken together the results above indicate that PI3k/Akt signaling plays a critical role in normal cardiac function and in maintaining

© 2012 Graves et al.; licensee BioMed Central Ltd. This is an Open Access article distributed under the terms of the Creative Commons Attribution License (http://creativecommons.org/licenses/by/2.0), which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited.