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Silencing of Cav1.2 gene in neonatal cardiomyocytes by lentiviral delivered shRNA.

Abstract：

:Cav1.2 (alpha1C) and Cav1.3 (alpha1D) L-type Ca channels are co-expressed in the heart. To date, there are no pharmacological or biophysical tools to separate alpha1D from alpha1C Ca currents (I(Ca-L)) in cardiomyocytes. Here, we established a physiological model to study alpha1D I(Ca-L) in native myocytes using RNA interference. Transfection of rat neonatal cardiomyocytes (RNC) with alpha1C specific siRNA resulted in low silencing efficiency (50-60%) at the mRNA and protein levels. The use of lentivirus shRNA resulted in 100% transfection efficiency and 92% silencing of the alpha1C gene by real-time PCR and Western blot. Electrophysiological experiments showed that the total I(Ca-L) was similarly reduced by 80% in lentivirus transfected cells. Both biochemical and functional data demonstrated high transfection and silencing efficiency in the cardiomyocytes using lentiviral shRNA. This novel approach allows for the assessments of the roles of alpha1C and alpha1D Ca channels in native myocytes and could be used to examine their roles in physiological and pathological settings.

journal_name

Biochem Biophys Res Commun

journal_title

Biochemical and biophysical research communications

authors

Karnabi E,Qu Y,Mancarella S,Yue Y,Wadgaonkar R,Boutjdir M

doi

10.1016/j.bbrc.2009.04.150

subject

Has Abstract

pub_date

2009-07-10 00:00:00

pages

409-14

issue

4

eissn

0006-291X

issn

1090-2104

pii

S0006-291X(09)00746-3

journal_volume

384

pub_type

杂志文章

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