| Ion channels are a series of hydrophilic proteins. They locate at cell membrane and organelle, and are the basis of cardiac electrical activity. Multiple studies have demonstrated that the distribution of many ion channels is heterogeneous, and therefore generate a transmural gradient between epicardial myocytes (EPI) and endocardial myocytes (ENDO). The transmural gradients of different ion channels play an important role in maintaining cardiac electrical stability. Under the pathological state, the transmural gradients of ion channels are vanished, and as a consequence, people will suffer a sudden death caused by malignant arrhythmias. So the study about how the transmural gradients of ion channels are generated, modulated and changed under pathological state has important clinical values.Calcium (Ca2+) is the most important ion in cardiac activity. Besides its key role in excitation-contraction coupling (ECC), Ca+participates in the transcriptional regulation for gene expression as a second messenger. Ca2+-dependent transcriptional regulation is named as excitation-transcription coupling (ETC). It indicates the progress that Ca2+regulates gene expression through phosphatase or kinase. Ca2+/calmodulin-dependent protein kinase II (CaMKII) plays a critical role in ETC. Previous studies showed that CaMKII and its downstream signaling pathways take part in the modulation of ion channel expression, but it is still unclear whether and how these Ca2+-dependent transcriptional regulation act on cardiac transmural ion flow.In present study, we combine molecular biology and cell electrophysiology to illuminate the role and underlying mechanisms of CaMKII-induced ETC on the formation or regulation of L-type Ca2+current (ICa, L) and Na+/Ca2+exchanger current (INCX) transmural grandients. This study will discuss the relationship between transmembrane ion flow and arrhythmia, and provide a new theoretical basis to support that CaMKII can be a good target in reversing electrical remodeling under pathological state.Purpose:1. To explore the effects of CaMKII activity on the formation of ICa, Ltransmural grandient between EPI and ENDO;2. To explore the effects of CaMKII activity on the remodeling of INCX transmural grandient between EPI and ENDO in failing heart.Methods:Cardiac EPI and ENDO cells were isolated from male mice hearts (C57BL/6) by enzyme digestion. Voltage-clamp technique was used to record ICa, L, the steady-state activation and inactivation of ICa, L and the recovery from inactivation. Mice heart failure model was induced by arota binding. INCX was recorded in normal heart and failing heart. In addition, western blot was used to detecte the protein expression of Cav1.2(encoding a subunit of L-type Ca2+channel), NCX1(encoding a subunit of Na+/Ca2+exchanger), total CaMKII, phosphorylated-CaMKII, p65(a subtype of NFκB) and et al, between EPI and ENDO.Results:1. ICa, L density is obviously higher in EPI than that in ENDO. Except that the fast and slow components of ICa, L decay are different, no difference is detected in the channel kinetic properties of ICa, L between EPI and ENDO.2. CaMKII activity is higher in ENDO that in EPI, and therefore caused more nuclear translocation of p65, which induces a less Cav1.2expression in ENDO than that in EPI.3. INCX density is obviously higher in EPI than that in ENDO, and NCX1expression shows a transmural grandient.4. In failing heart, INCX transmural grandient is vanished. The NCX1expression is up-regulated in both EPI and ENDO, especially in ENDO. This is consistent with the change of CaMKII activity in EPI and ENDO.Conclusions:1. The formation of ICa, L transmural grandient is related to a higher activity of CaMKII/p65signaling in ENDO.2. The formation of INCX transmural grandient has no relationship to CaMKII activity, but the remodeling of INCX in failing heart is caused by the increased CaMKII activity in EPI and ENDO. |
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