DMA对正常大鼠离体心脏和乳头肌的正性变力作用及其机制

目的研究二甲基氨氯吡咪(dimethylamiloride,DMA)对正常大鼠离体心脏和乳头肌的正性变力作用,并探讨其机制。方法采用Langendorff心脏灌流和离体乳头肌灌流方法观察DMA的变力性作用,并应用L-型钙通道阻滞药和Na+/Ca2+交换体(sodium calcium exchanger,NCX)抑制剂探讨其作用机制。结果①1~20μmol.L-1DMA对正常大鼠离体心脏产生正性变力作用,即增强左室主动收缩压(LVSP-LVDP)、左室压最大上升速率(+dp/dtmax)、左室压最大下降速率(-dp/dtmax);增强乳头肌收缩功能,即升高发展张力(TC),缩短收缩至最大速率的时间(T-dp/dtmax)。与用药前相比,差异有显著性(P<0.05或P<0.01)。②DMA的正性变力作用不能被L-型钙通道阻滞药尼卡地平(nicardipine)阻断。③NCX抑制剂氯化镍(NiCl2)可阻断DMA的正性变力作用。结论①(1~20)μmol.L-1DMA对正常大鼠离体心脏和乳头肌产生正性变力作用。②DMA正性变力作用与L-型钙通道无关。③DMA通过激动NCX产生正性变力作用。     

Phorbol ester‐evoked Ca2+signaling in human platelets is via autocrine activation of P2X1 receptors,not a novel non‐capacitative Ca2+entry

Summary. Background: Platelets are reported to possess a protein kinase C (PKC)‐dependent non‐capacitative Ca²⁺entry (NCCE) pathway. The phorbol ester, phorbol, 12‐myristate, 13‐acetate (PMA) has been suggested to stimulate platelet NCCE. Recently we demonstrated important roles in store‐operated Ca²⁺entry in human platelets for Na⁺/Ca²⁺ exchangers (NCXs) and autocrine signaling between platelets after dense granule secretion. As PMA evokes dense granule secretion, we have investigated the role of NCXs and autocrine signaling in PMA‐evoked Ca²⁺entry. Objectives: To investigate the roles of NCXs and dense granule secretion in PMA‐evoked Ca²⁺signaling in human platelets. Methods: Fura‐2‐ or sodium‐binding benzofuran isophthalate (SBFI)‐loaded platelets were used to monitor cytosolic Ca²⁺or Na⁺ concentrations. Dense granule secretion was monitored as ATP release using luciferin–luciferase. Results: The NCX inhibitors KB‐R7943 or SN‐6, and removal of extracellular Na⁺, significantly reduced PMA‐evoked Ca²⁺entry. PMA‐evoked dense granule secretion was almost abolished by pretreatment with the PKC inhibitor Ro‐31‐8220 and significantly slowed by KB‐R7943. The P_2X1 antagonists Ro‐0437626 or MRS‐2159, or desensitization of P_2X1 receptors by prior treatment with α,β‐Methylene‐ATP or omitting apyrase from the medium, reduced PMA‐evoked Ca²⁺entry. Ro‐0437626 or chelation of extracellular Ca²⁺ slowed but did not abolish PMA‐evoked ATP release, indicating that PMA‐evoked dense granule secretion does not require P_2X1 receptor activation but is accelerated by P_2X1‐mediated Ca²⁺entry. The presence of NCX3 in human platelets was demonstrated by Western blotting. Conclusion: PMA‐evoked Ca²⁺entry results from an NCX3‐dependent dense granule secretion and subsequent P_2X1 receptor activation by secreted ATP, rather than activation of a novel NCCE pathway.     

低钾加重大鼠心肌细胞再灌注损伤的机制

目的:观察低钾灌流液对大鼠心肌细胞再灌注时钙震荡的影响并探讨其作用机制. 方法:Ca2 荧光指示剂Fura-2标记心肌细胞,在代谢抑制并低氧25 min后,改用低钾台氏液([K ]=3.0 mmol/L)灌流,记录细胞钙震荡的变化. 以(再灌注末期细胞长度-缺血末期细胞长度)/(缺血前细胞长度-缺血末期细胞长度)×100%反映再灌注后细胞长度的恢复状况. 结果:与含正常钾浓度的灌流液对照组([K ]=5.4 mmol/L)相比,在再灌注10 min内,低钾灌流液组出现钙震荡的次数显著增加(P＜0.01,低K 组:138.80±9.54 vs对照组:82.30±8.16,n=6),心肌细胞长度的恢复显著被抑制[P＜0.01,低K 组:(24.30±6.01)% vs对照组:(54.50±6.56)%,n=6];再灌注期间给予钠-钙交换体反向交换模式抑制剂KB-R7943,可显著抑制低钾溶液对钙震荡和细胞长度的影响[P＜0.01 vs低K 组,n=6; 钙震荡:27.40±6.76和细胞长度恢复:(58.90±7.30)%]. 结论:低钾再灌注液通过增强反向钠-钙交换体活性加重钙震荡,进而引发心肌损伤.     

ORM-10103: a significant advance in sodium-calcium exchanger pharmacology?

The sodium-calcium exchanger (NCX) is an electrogenic transporter that is widely expressed in different tissues. In the heart, the NCX plays important roles in calcium ion homeostasis, excitation-contraction coupling and the electrophysiological properties of cardiac myocytes. Precise determination of the roles of the NCX has somewhat been hampered by a lack of selective small molecule inhibitors. In this issue of the BJP, Jost and colleagues present data on a new NCX inhibitor, ORM-10103, which has submicromolar EC₅₀ values against cardiac forward and reverse exchange activity. The compound exhibits improved selectivity over existing small molecule NCX inhibitors and, in particular, appears to be without effect on L-type calcium channels at high concentrations. ORM-10103 could therefore have significant value for studies of the (patho)physiological roles of the NCX in the heart. Further pharmacological studies are required to investigate the actions of ORM-10103 on cardiac cells and tissues and to determine its effects on non-cardiac NCX isoforms.

Linked Article

This article is a commentary on Jost et al., pp. 768–778 of this issue. To view this paper visit http://dx.doi.org/10.1111/bph.12228     

ORM-10103, a novel specific inhibitor of the Na+/Ca2+ exchanger,decreases early and delayed afterdepolarizations in the canine heart

BACKGROUND AND PURPOSE

At present there are no small molecule inhibitors that show strong selectivity for the Na⁺/Ca²⁺ exchanger (NCX). Hence, we studied the electrophysiological effects of acute administration of ORM-10103, a new NCX inhibitor, on the NCX and L-type Ca²⁺ currents and on the formation of early and delayed afterdepolarizations.

EXPERIMENTAL APPROACH

Ion currents were recorded by using a voltage clamp technique in canine single ventricular cells, and action potentials were obtained from canine and guinea pig ventricular preparations with the use of microelectrodes.

KEY RESULTS

ORM-10103 significantly reduced both the inward and outward NCX currents. Even at a high concentration (10 μM), ORM-10103 did not significantly change the L-type Ca²⁺ current or the maximum rate of depolarization (dV/dt_max), indicative of the fast inward Na⁺ current. At 10 μM ORM-10103 did not affect the amplitude or the dV/dt_max of the slow response action potentials recorded from guinea pig papillary muscles, which suggests it had no effect on the L-type Ca²⁺ current. ORM-10103 did not influence the Na⁺/K⁺ pump or the main K⁺ currents of canine ventricular myocytes, except the rapid delayed rectifier K⁺ current, which was slightly diminished by the drug at 3 μM. The amplitudes of pharmacologically- induced early and delayed afterdepolarizations were significantly decreased by ORM-10103 (3 and 10 μM) in a concentration-dependent manner.

CONCLUSIONS AND IMPLICATIONS

ORM-10103 is a selective inhibitor of the NCX current and can abolish triggered arrhythmias. Hence, it has the potential to be used to prevent arrhythmogenic events.

LINKED ARTICLE

This article is commented on by Terracciano and Hancox, pp. 765–767 of this issue. To view this commentary visit http://dx.doi.org/10.1111/bph.12299     

Activation of cardiac M3 muscarinic acetylcholine receptors has cardioprotective effects against ischaemia-induced arrhythmias

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Impaired SERCA function contributes to cardiomyocyte dysfunction in insulin resistant rats

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