Fura‐2 calcium transients in frog skeletal muscle fibres.
Fura‐2 calcium transients in frog skeletal muscle fibres.
复制标题
青蛙骨骼肌纤维中的 Fura-2 钙瞬变。
DOI：
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发表时间：
1988
期刊：
影响因子：
--
通讯作者：
S. Hollingworth
中科院分区：
文献类型：
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作者：
S. Baylor;S. Hollingworth
1.将完整的蛙肌肌纤维以长肌节间距(3.5-4.2微米)固定在光学台上，用于测量肌浆内注射钙离子指示剂Fura-2和安替比拉偶氮III后的吸光度和荧光信号。在16-17℃的静止和电刺激下测量纤维中与染料有关的信号，以产生单个动作电位或简短的动作电位序列。2.Fura-2在肌浆中的表观扩散常数DAPP是由Fura-2荧光测量的时间和距离染料注入部位的函数估算的。平均而言(N=7)，DAPP为0.36×10~(-6)cm~2·S~(-1)，如果所有的Fura-2都自由溶解在肌浆液中，这个值比预期的要小近3倍。如果大约60%-65%的Fura-2分子结合到肌浆中相对静止的位置，就可以解释DAPP的小价值。3.在静息纤维中，Fura-2以钙结合形式存在的比例很小，平均(N=11)0.06。然而，由于Fura-2的很大一部分不能自由溶解在肌浆中，并且采用间接方法估计钙结合染料，因此从钙结合染料组分中校准肌浆游离钙([Ca~(2+)])是不可靠的。4.在单个动作电位的作用下，Fura-2的荧光强度(F)和吸光度(A)都发生了很大的变化，这两种变化具有相同的时程。正如预期的那样，这些瞬变的方向与钙离子-染料复合体的增加相对应。在波长380到460 nm之间，峰增量A(Lambda)与体外校准中测定的Fura-2的钙-染料差光谱非常相似。用比耳定律来校准在活动过程中形成的钙-染料络合物的浓度(增量[CaFura-2])，根据增量A(Lambda)信号。峰值增量[CaFura-2]在0.01到0.4 mm之间变化，这取决于注射的Fura-2([Fura-2T])的总浓度，最高可达0.9 mm。5.在峰值[CaFura-2]小于0.06 mm的纤维中，[CaFura-2]的极限最小半宽为50-60ms。然而，随着峰值[CaFura-2]的增加(高达0.3-0.4 mm)，[CaFura-2]的半宽明显延长(高达150-200ms)，这表明大浓度的Fura-2对潜在的[Ca+]瞬变([CaFura])有很强的缓冲作用。
1. Intact single twitch fibres from frog muscle were mounted at long sarcomere spacing (3.5‐4.2 microns) on an optical bench apparatus for the measurement of absorbance and fluorescence signals following the myoplasmic injection of either or both of the Ca2+ indicator dyes Fura‐2 and Antipyrylazo III. Dye‐related signals were measured at 16‐17 degrees C in fibres at rest and stimulated electrically to give a single action potential or brief train of action potentials. 2. The apparent diffusion constant of Fura‐2 in myoplasm, Dapp, was estimated from Fura‐2 fluorescence measured as a function of time and distance from the site of dye injection. On average (N = 7), Dapp was 0.36 x 10(‐6) cm2 s‐1, a value nearly 3‐fold smaller than expected if all the Fura‐2 was freely dissolved in the myoplasmic solution. The small value of Dapp is explained if approximately 60‐65% of the Fura‐2 molecules were bound to relatively immobile sites in myoplasm. 3. In resting fibres the fraction of Fura‐2 in the Ca2+‐bound form was estimated to be small, on average (N = 11) 0.06 of total dye. However, because of the large fraction of Fura‐2 not freely dissolved in myoplasm, and the indirect method employed for estimating Ca2+‐bound dye, calibration of the resting level of myoplasmic free Ca2+ ([Ca2+]) from the fraction of Ca2+‐bound dye was not considered reliable. 4. In response to a single action potential, large changes in Fura‐2 fluorescence (delta F) and absorbance (delta A) were detected, which had identical time courses. As expected, the directions of these transients corresponded to an increase in Ca2+‐dye complex. For wavelengths, lambda, between 380 and 460 nm, peak delta A(lambda) was closely similar to the Ca2+‐dye difference spectrum for Fura‐2 determined in in vitro calibrations. Beer's law was used to calibrate the concentration of Ca2+‐dye complex formed during activity (delta[CaFura‐2]) from the delta A(lambda) signal. Peak delta[CaFura‐2] was found to vary between 0.01 and 0.4 mM, depending on the total concentration of injected Fura‐2 ([Fura‐2T]), which ranged as high as 0.9 mM. 5. In fibres in which peak delta[CaFura‐2] was less than 0.06 mM, delta[CaFura‐2] had a limiting minimal half‐width of 50‐60 ms. However, as peak delta[CaFura‐2] increased (up to 0.3‐0.4 mM), delta[CaFura‐2] half‐width became markedly prolonged (up to 150‐200 ms), indicative of a strong buffering action of large concentrations of Fura‐2 on the underlying [Ca2+] transient (delta[Ca2+]).(ABSTRACT TRUNCATED AT 400 WORDS)
影响因子：
3.4
作者：
CANNELL, MB;ALLEN, DG
通讯作者：
ALLEN, DG
影响因子：
5.5
作者：
BAYLOR, SM;CHANDLER, WK;MARSHALL, MW
通讯作者：
MARSHALL, MW
注射 Azo1（一种四羧酸 Ca2 指示剂）的青蛙骨骼肌纤维中的钙瞬变。
DOI：
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发表时间：
1986
期刊：
Society of General Physiologists series
影响因子：
--
作者：
Hollingworth,S;Baylor,SM
通讯作者：
Baylor,SM