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Intrasarcomere [Ca²⁺] gradients and their spatio-temporal relation to Ca²⁺ sparks in rat cardiomyocytes

Hikaru Tanaka, Toshiyuki Sekine, Toru Kawanishi *, Ryu Nakamura ¹ and Koki Shigenobu

1. Line-scan analyses of spontaneous Ca²⁺ sparks, non-propagating local rises in Ca²⁺ concentration, and the early phase of Ca²⁺ transients in cardiomyocytes were performed with a rapid-scanning laser confocal microscope (Nikon RCM8000) and fluo-3.

2. On electrical stimulation, points at which rise in Ca²⁺ began earliest were observed at regular spacings of 1·82 ± 0·26 µm (mean ± s.d.) along the longitudinal axis of the cell. The points were heavily stained with di-2-ANEPEQ, which stains the T-tubules, indicating that they were at the Z-line.

3. The points where spontaneous Ca²⁺ sparks originated coincided with the points which showed faster Ca²⁺ elevation, i.e. the Z-line.

4. In some cases where a Ca²⁺ spark had occurred within about 30 ms before the evoked Ca²⁺ transient, fast elevation of Ca²⁺ was not observed at the corresponding Z-line, indicating the presence of a refractory period in Ca²⁺ release from the SR.

5. The present results provide visual evidence for Ca²⁺ release from the junctional sarcoplasmic reticulum in cardiomyocytes. The presence of a refractory period in Ca²⁺ release after Ca²⁺ sparks provided new evidence that the normal Ca²⁺ transient may be the summation of Ca²⁺ sparks.

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