Sarcoplasmic reticulum Ca2+ release channel ryanodine receptor (RyR2) plays a crucial role in aconitine-induced arrhythmias.
The present study established a model of RyR(2) knockdown cardiomyocytes and elucidated the role of RyR(2) in aconitine-induced arrhythmia. Cardiomyocytes were obtained from hearts of neonatal Sprague-Dawley rats. siRNAs were used to down-regulate RyR(2) expression. Reduction of RyR(2) expression was documented by RT-PCR, western blot, and immunofluorescence. Ca(2+) signals were investigated by measuring the relative intracellular Ca(2+) concentration, spontaneous Ca(2+) oscillations, caffeine-induced Ca(2+) release, and L-type Ca(2+) currents. In normal cardiomyocytes, steady and periodic spontaneous Ca(2+) oscillations were observed, and the baseline [Ca(2+)](i) remained at the low level. Exposure to 3 microM aconitine increased the frequency and decreased the amplitude of Ca(2+) oscillations; the baseline [Ca(2+)](i) and the level of caffeine-induced Ca(2+) release were increased but the L-type Ca(2+) currents were inhibited after application of 3 microM aconitine for 5 min. In RyR(2) knockdown cardiomyocytes, the steady and periodic spontaneous Ca(2+) oscillations almost disappeared, but were re-induced by aconitine without affecting the baseline [Ca(2+)](i) level; the level of caffeine-induced Ca(2+) release was increased but L-type Ca(2+) currents were inhibited. Alterations of RyR(2) are important consequences of aconitine-stimulation and activation of RyR(2) appear to have a direct relationship with aconitine-induced arrhythmias. The present study demonstrates a potential method for preventing aconitine-induced arrhythmias by inhibiting Ca(2+) leakage through the sarcoplasmic reticulum RyR(2) channel.
Fu
Min
M
School of Medicine and Department of Biological Sciences and Biotechnology, Tsinghua University, Beijing 100084, People's Republic of China.
Li
Ru-Xin
RX
Fan
Li
L
He
Guo-Wei
GW
Thornburg
Kent L
KL
Wang
Zhao
Z
D000157:Aconitine / Q000633:toxicity*; D000818:Animals; D001145:Arrhythmias, Cardiac / Q000139:chemically induced* / Q000378:metabolism*; D002110:Caffeine / Q000494:pharmacology; D020746:Calcium Channels, L-Type / Q000378:metabolism; D020013:Calcium Signaling / Q000187:drug effects / Q000502:physiology; D002478:Cells, Cultured; D015870:Gene Expression / Q000187:drug effects; D020868:Gene Silencing; D008564:Membrane Potentials / Q000502:physiology; D032383:Myocytes, Cardiac / Q000187:drug effects* / Q000378:metabolism; D012333:RNA, Messenger / Q000187:drug effects / Q000378:metabolism; D034741:RNA, Small Interfering / Q000378:metabolism; D051381:Rats; D017207:Rats, Sprague-Dawley; D019837:Ryanodine Receptor Calcium Release Channel / Q000235:genetics / Q000378:metabolism*; D012519:Sarcoplasmic Reticulum / Q000187:drug effects / Q000378:metabolism*; D014162:Transfection
D020746:Calcium Channels, L-Type; D012333:RNA, Messenger; D034741:RNA, Small Interfering; D019837:Ryanodine Receptor Calcium Release Channel; D002110:Caffeine; D000157:Aconitine