TY - JOUR

T1 - RXP-E: a connexin43-binding peptide that prevents action potential propagation block

AU - Lewandowski, Rebecca

AU - Procida, Kristina

AU - Vaidyanathan, Ravi

AU - Coombs, Wanda

AU - Jalife, José

AU - Nielsen, Morten S

AU - Taffet, Steven M

AU - Delmar, Mario

N1 - Keywords: Acids; Action Potentials; Animals; Biological Transport; Carrier Proteins; Cells, Cultured; Connexin 43; Drug Design; Gap Junctions; Heptanol; Hydrogen-Ion Concentration; Myocytes, Cardiac; Patch-Clamp Techniques; Potassium Channels, Inwardly Rectifying; Protein Binding; Rats; Sodium Channels

PY - 2008

Y1 - 2008

N2 - Gap junctions provide a low-resistance pathway for cardiac electric propagation. The role of GJ regulation in arrhythmia is unclear, partly because of limited availability of pharmacological tools. Recently, we showed that a peptide called "RXP-E" binds to the carboxyl terminal of connexin43 and prevents chemically induced uncoupling in connexin43-expressing N2a cells. Here, pull-down experiments show RXP-E binding to adult cardiac connexin43. Patch-clamp studies revealed that RXP-E prevented heptanol-induced and acidification-induced uncoupling in pairs of neonatal rat ventricular myocytes. Separately, RXP-E was concatenated to a cytoplasmic transduction peptide (CTP) for cytoplasmic translocation (CTP-RXP-E). The effect of RXP-E on action potential propagation was assessed by high-resolution optical mapping in monolayers of neonatal rat ventricular myocytes, containing approximately 20% of randomly distributed myofibroblasts. In contrast to control experiments, when heptanol (2 mmol/L) was added to the superfusate of monolayers loaded with CTP-RXP-E, action potential propagation was maintained, albeit at a slower velocity. Similarly, intracellular acidification (pH(i) 6.2) caused a loss of action potential propagation in control monolayers; however, propagation was maintained in CTP-RXP-E-treated cells, although at a slower rate. Patch-clamp experiments revealed that RXP-E did not prevent heptanol-induced block of sodium currents, nor did it alter voltage dependence or amplitude of Kir2.1/Kir2.3 currents. RXP-E is the first synthetic molecule known to: (1) bind cardiac connexin43; (2) prevent heptanol and acidification-induced uncoupling of cardiac gap junctions; and (3) preserve action potential propagation among cardiac myocytes. RXP-E can be used to characterize the role of gap junctions in the function of multicellular systems, including the heart.

AB - Gap junctions provide a low-resistance pathway for cardiac electric propagation. The role of GJ regulation in arrhythmia is unclear, partly because of limited availability of pharmacological tools. Recently, we showed that a peptide called "RXP-E" binds to the carboxyl terminal of connexin43 and prevents chemically induced uncoupling in connexin43-expressing N2a cells. Here, pull-down experiments show RXP-E binding to adult cardiac connexin43. Patch-clamp studies revealed that RXP-E prevented heptanol-induced and acidification-induced uncoupling in pairs of neonatal rat ventricular myocytes. Separately, RXP-E was concatenated to a cytoplasmic transduction peptide (CTP) for cytoplasmic translocation (CTP-RXP-E). The effect of RXP-E on action potential propagation was assessed by high-resolution optical mapping in monolayers of neonatal rat ventricular myocytes, containing approximately 20% of randomly distributed myofibroblasts. In contrast to control experiments, when heptanol (2 mmol/L) was added to the superfusate of monolayers loaded with CTP-RXP-E, action potential propagation was maintained, albeit at a slower velocity. Similarly, intracellular acidification (pH(i) 6.2) caused a loss of action potential propagation in control monolayers; however, propagation was maintained in CTP-RXP-E-treated cells, although at a slower rate. Patch-clamp experiments revealed that RXP-E did not prevent heptanol-induced block of sodium currents, nor did it alter voltage dependence or amplitude of Kir2.1/Kir2.3 currents. RXP-E is the first synthetic molecule known to: (1) bind cardiac connexin43; (2) prevent heptanol and acidification-induced uncoupling of cardiac gap junctions; and (3) preserve action potential propagation among cardiac myocytes. RXP-E can be used to characterize the role of gap junctions in the function of multicellular systems, including the heart.

U2 - 10.1161/CIRCRESAHA.108.179069

DO - 10.1161/CIRCRESAHA.108.179069

M3 - Journal article

C2 - 18669919

VL - 103

SP - 519

EP - 526

JO - Circulation Research

JF - Circulation Research

SN - 0009-7330

IS - 5

ER -