A new method to detect rapid oxygen changes around cells: How quickly do calcium channels sense oxygen in cardiomyocytes?
A new method to detect rapid oxygen changes around cells: How quickly do calcium channels sense oxygen in cardiomyocytes?
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DOI：
10.1152/japplphysiol.00770.2013
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发表时间：
2013-12-01
影响因子：
3.3
通讯作者：
Cleemann, Lars
中科院分区：
文献类型：
--
作者：
Scaringi, John A.;Rosa, Angelo Oscar;Cleemann, Lars
急性缺氧被认为会触发保护性反应，在心脏和颈动脉体等组织中，包括Ca2+和K+通道的快速（5-10秒）抑制。为了深入了解心脏l型Ca2+通道抑制的机制，我们测量了电压钳住的心脏细胞附近的O-2依赖性荧光，这些细胞受到不同O-2张力溶液的快速交换。这是通过一个带有玻璃底的实验室来完成的，该实验室用作光导，用于激发薄的钌基o -2敏感ORMOSIL涂层。荧光成像显示，在电磁控制的溶液“泵”中，稳定状态的PO2在整个流中得到了很好的控制，但是在流的外围（tau（1/2）类似于500 ms）的变化比立即在电压钳住的肌细胞（tau（1/2）类似于225 ms）周围的变化要慢，而在那里，牢固附着的细胞又产生了额外的50-100 ms的局部延迟。同时进行电压钳和O-2测量，我们发现急性缺氧逐渐和可逆地抑制Ca2+通道（Ca-V 1.2）。以Ba2+为载流子，在1.5 s后抑制显著，2.5 s后抑制幅度接近10%，5 s后几乎完全可逆。所描述的荧光测量提供了检查和微调溶液泵的手段，并表明PO2的变化可以在类似的200 ms内完成。缺氧条件下钡电流的快速可逆抑制与心脏Ca2+通道直接由O-2调节的观点是一致的。
Acute hypoxia is thought to trigger protective responses that, in tissues like heart and carotid body, include rapid (5-10 s) suppression of Ca2+ and K+ channels. To gain insight into the mechanism for the suppression of the cardiac L-type Ca2+ channel, we measured O-2-dependent fluorescence in the immediate vicinity of voltage-clamped cardiac cells subjected to rapid exchange of solutions with different O-2 tensions. This was accomplished with an experimental chamber with a glass bottom that was used as a light guide for excitation of a thin ruthenium-based O-2-sensitive ORMOSIL coating. Fluorescence imaging showed that steady-state PO2 was well controlled within the entire stream from an electromagnetically controlled solution "puffer" but that changes were slower at the periphery of the stream (tau(1/2) similar to 500 ms) than immediately around the voltage-clamped myocyte (tau(1/2) similar to 225 ms) where, in turn, firmly attached cells produced an additional local delay of 50-100 ms. Performing simultaneous voltage clamp and O-2 measurements, we found that acute hypoxia gradually and reversibly suppressed the Ca2+ channel (Ca-V 1.2). Using Ba2+ as charge carrier, the suppression was significant after 1.5 s, reached similar to 10% after 2.5 s, and was nearly completely reversible in 5 s. The described fluorescence measurements provide the means to check and fine tune solution puffers and suggest that changes in PO2 can be accomplished within similar to 200 ms. The rapid and reversible suppression of barium current under hypoxia is consistent with the notion that the cardiac Ca2+ channel is directly modulated by O-2.