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Ordered Assembly of the Adhesive and Electrochemical Connections within Newly Formed Intercalated Disks in Primary Cultures of Adult Rat Cardiomyocytes

Division of Pediatric Cardiology, Department of Pediatrics and Communicable Diseases, University of Michigan, 1150 West Medical Center Dr., Ann Arbor, MI 48109, USA.
BioMed Research International (Impact Factor: 2.71). 05/2010; 2010:624719. DOI: 10.1155/2010/624719
Source: PubMed

ABSTRACT The intercalated disk (ID) is a complex structure that electromechanically couples adjoining cardiac myocytes into a functional syncitium. The integrity of the disk is essential for normal cardiac function, but how the diverse elements are assembled into a fully integrated structure is not well understood. In this study, we examined the assembly of new IDs in primary cultures of adult rat cardiac myocytes. From 2 to 5 days after dissociation, the cells flatten and spread, establishing new cell-cell contacts in a manner that recapitulates the in vivo processes that occur during heart development and myocardial remodeling. As cells make contact with their neighbors, transmembrane adhesion proteins localize along the line of apposition, concentrating at the sites of membrane attachment of the terminal sarcomeres. Cx43 gap junctions and ankyrin-G, an essential cytoskeletal component of voltage gated sodium channel complexes, were secondarily recruited to membrane domains involved in cell-cell contacts. The consistent order of the assembly process suggests that there are specific scaffolding requirements for integration of the mechanical and electrochemical elements of the disk. Defining the relationships that are the foundation of disk assembly has important implications for understanding the mechanical dysfunction and cardiac arrhythmias that accompany alterations of ID architecture.

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    • "Important work by Delmar and colleagues in recent years has shed light on the degree to which Na v 1.5 channels are integrated into the macromolecular complex at the ID. They have demonstrated that localization of mechanical junction proteins, mainly PKP2 (a desmosomal protein), precedes recruitment of Cx43 gap junctions and AnkG (a sub-membrane adapter protein involved in localizing Na v 1.5 in the membrane) to sites of cell-to-cell contact in neonatal rat ventricular myocytes (Geisler et al., 2010). Functionally, they have demonstrated that both regulation of the sodium current PKP2 and AnkG and linked mutations in both proteins to Brugada syndrome, an inherited arrhythmia syndrome resulting from loss of sodium current density (Cerrone et al., 2013; Hashemi et al., 2009). "
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    ABSTRACT: Cardiac conduction is the propagation of electrical excitation through the heart and is responsible for triggering individual myocytes to contract in synchrony. Canonically, this process has been thought to occur electrotonically, by means of direct flow of ions from cell to cell. The intercalated disk (ID), the site of contact between adjacent myocytes, has been viewed as a structure composed of mechanical junctions that stabilize the apposition of cell membranes and gap junctions which constitute low resistance pathways between cells. However, emerging evidence suggests a more active role for structures within the ID in mediating intercellular electrical communication by means of non-canonical ephaptic mechanisms. This review will discuss the role of the ID in the context of the canonical, electrotonic view of conduction and highlight new, emerging possibilities of its playing a more active role in ephaptic coupling between cardiac myocytes.
    Cell Communication & Adhesion 04/2014; DOI:10.3109/15419061.2014.905932 · 1.52 Impact Factor
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    • "Dr. Mario Delmar and colleagues have provided evidence suggesting that Cx43 and Na v 1.5 participate in a macromolecular complex at the intercalated disk which includes the desmosomal protein plakophilin-2 (PKP2) as well as ankyrin-G, a submembrane adapter protein involved in localizing cardiac sodium channels (Na v 1.5) in the membrane. In primary cultures of neonatal rat ventricular myocytes, they found that Cx43 gap junctions and ankyrin-G (AnkG) are recruited to sites of cell-to-cell contact following the localization of mechanical adhesion proteins [22]. Subsequently, they demonstrated regula-tion of the sodium current by both PKP2 and AnkG [10] [53] [54] and mutations in both proteins have been associated with Brugada syndrome, an inherited arrhythmia syndrome characterized by decreased sodium current density [9] [25]. "
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    ABSTRACT: Cardiac conduction is the process by which electrical excitation is communicated from cell to cell within the heart, triggering synchronous contraction of the myocardium. The role of conduction defects in precipitating life-threatening arrhythmias in various disease states has spurred scientific interest in the phenomenon. While the understanding of conduction has evolved greatly over the last century, the process has largely been thought to occur via movement of charge between cells via gap junctions. However, it has long been hypothesized that electrical coupling between cardiac myocytes could also occur ephaptically, without direct transfer of ions between cells. This review will focus on recent insights into cardiac myocyte intercalated disk ultrastructure and their implications for conduction research, particularly the ephaptic coupling hypothesis.
    FEBS letters 01/2014; 588(8). DOI:10.1016/j.febslet.2014.01.026 · 3.34 Impact Factor
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    • "What remains is the challenge to integrate the implicated cytoskeletal proteins in an overall model to address how Cx43 gets its ''postal address'', or where in the subcellular trafficking pathway from Golgi exit to channel delivery does Cx43 become destined to arrive at the GJ plaque. Studies of membrane Cx43 localization typically utilize cultured cardiomyocytes that can undergo internal rearrangement once in vitro [31] [32] [33], or cultured non-cardiac cells with various geometries and different numbers of cellular contacts. This variability in the shape and contacts of cultured cells limits the isolation of a particular behavior or event that may be important to Cx43 transport from the Golgi apparatus to a point of cell–cell contact. "
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    ABSTRACT: The precise expression and timely delivery of connexin 43 (Cx43) proteins to form gap junctions are essential for electrical coupling of cardiomyocytes. Growing evidence supports a cytoskeletal-based trafficking paradigm for Cx43 delivery directly to adherens junctions at the intercalated disc. A limitation of Cx43 localization assays in cultured cells, in which cell-cell contacts are essential, is the inability to control for cell geometry or reproducibly generate contact points. Here we present a micropatterned cell pairing system well suited for live microscopy to examine how the microtubule and actin cytoskeleton confer specificity to Cx43 trafficking to precisely defined cell-cell junctions. This system can also be adapted for other cell types and used to study dynamic intracellular movements of other proteins important for cell-cell communication.
    FEBS letters 01/2014; 588(8). DOI:10.1016/j.febslet.2014.01.002 · 3.34 Impact Factor
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