Modulation of focal and global Ca2+ release in calsequestrin-overexpressing mouse cardiomyocytes

ABSTRACT

• 1. 

  Focal and global Ca²⁺ releases were monitored in voltage-clamped control and hypertrophied calsequestrin (CSQ)-overexpressing mouse cardiomyocytes, dialysed with fluo-3, using rapid (120-240 frames s⁻¹) two-dimensional confocal imaging.
• 2. 

  Spontaneous focal Ca²⁺ releases (Ca²⁺ sparks) were absent or significantly reduced in frequency in hypertrophied myocytes of CSQ-overexpressing mice compared to their age-matched controls. Sporadic Ca²⁺ sparks seen in CSQ-overexpressing myocytes had intensities and durations similar to those of controls although quantitative analysis showed a trend towards more diffuse focal releases.
• 3. 

  Activation of Ca²⁺ current (I_Ca) failed to produce the typical sarcomeric Ca²⁺ striping pattern consistently seen in control myocytes. Instead, focal Ca²⁺ releases appeared as a disorganized patchwork of diffuse or 'woolly' fluorescence signals, resulting in slowly developing and reduced global Ca²⁺ transients.
• 4. 

  Although the density of I_Ca in CSQ-overexpressing myocytes was only slightly smaller than that of controls, the inactivation kinetics of the current were greatly reduced, consistent with the much smaller rate of rise of cytosolic Ca²⁺.
• 5. 

  Enhancement of I_Ca by elevation of [Ca²⁺]_o from 2 to 10 mM or addition of 3 μM isoproterenol (isoprenaline) failed to normalize the frequency of spontaneous Ca²⁺ sparks at rest or the pattern and the magnitude of I_Ca-gated Ca²⁺ transients. Isoproterenol was somewhat more effective than elevation of [Ca²⁺]_o.
• 6. 

  In sharp contrast, low (0·5 mM) caffeine concentrations that produced no measurable effects on I_Ca or Ca²⁺ transients in control myocytes, re-established spontaneous focal Ca²⁺ releases in CSQ-overexpressing cells, triggered large I_Ca-gated cellular Ca²⁺ transients, and strongly enhanced the kinetics of inactivation of I_Ca.
• 7. 

  Our data suggest that impaired Ca²⁺ signalling in CSQ-overexpressing myocytes results from reduced co-ordination and decreased frequency of Ca²⁺ sparks. The impaired Ca²⁺ signalling could not be restored by procedures that increased I_Ca, but was mostly restored in the presence of caffeine, which may alter the Ca²⁺ sensitivity of the ryanodine receptor.