Journal: Journal of Molecular and Cellular Cardiology
Article Title: Inducing Ito,f and phase 1 repolarization of the cardiac action potential with a Kv4.3/KChIP2.1 bicistronic transgene
Figure Lengend Snippet: Increasing I to,f density with bicistronic Kv4.3/KChIP2.1 expression affects cardiomyocyte AP waveform. Panel A shows APs recorded from 2 day cultured (untransduced black), and transduced myocytes with increasing I to,f current densities at a cycle length of 1 s. Introduction of I to,f (lower panel traces show exemplar records for pulses to −30, +10 and + 40 mV) resulted in a prominent AP phase 1 in all transduced myocytes. As I to,f density increased, there was a progressive change in AP morphology that caused the AP to resemble that recorded in other species. B. Increasing density of I to,f (at +40 mV) decreased APD20. APD90 (right panel) decreased approximately exponentially (solid line) with increasing I to,f except when I to,f was between ~12 and ~ 17 pA/pF (deviation highlighted as dashed line) where emergence of a marked ‘spike-and-dome’ morphology developed (as shown center right in A). All trend curves drawn by eye. C. Representative traces of I to,f recorded in untransduced (Control) iPSC-CMs and iPSC-CMs expressing bicistronic Kv4.3-mCherry/KChIP2.1 under a cTnT promoter. After a brief step to −40 mV to inactivate Na + channels, subsequent membrane depolarizations to −30 mV, −10 mV, +10 mV, +30 mV and + 50 mV were used to elicit I to in control and transduced iPSC-CMs. D. Three examplar AP waveforms of iPSC-CMs without (black traces) or with expressed I to,f currents (green lower traces). I to,f densities in the exemplar transduced iPSC-CMs were 5–10 pA/pF at +40 mV. APs were elicited at a frequency of 1 Hz. (For interpretation of the references to colour in this figure legend, the reader is referred to the web version of this article.)
Article Snippet: Samples were then labelled with primary antibodies including anti-Caveolin 3 (anti-Cav3; BD Bioscience, San Jose, USA; 1/500 dilution) anti-KChIP2 (Alomone, Israel; 1/500 dilution) overnight at 4 °C.
Techniques: Expressing, Cell Culture