'Pressure-flow'-triggered intracellular Ca2+ transients in rat cardiac myocytes: possible mechanisms and role of mitochondria.

Cardiac myocytes, in the intact heart, are exposed to shear/fluid forces during each cardiac cycle. Here we describe a novel Ca(2+) signalling pathway, generated by 'pressurized flows' (PFs) of solutions, resulting in the activation of slowly developing ( approximately 300 ms) Ca(2+) transients lasting approximately 1700 ms at room temperature. Though subsequent PFs (applied some 10-30 s later) produced much smaller or undetectable responses, such transients could be reactivated following caffeine- or KCl-induced Ca(2+) releases, suggesting that a small, but replenishable, Ca(2+) pool serves as the source for their activation. PF-triggered Ca(2+) transients could be activated in Ca(2+)-free solutions or in solutions that block voltage-gated Ca(2+) channels, stretch-activated channels (SACs), or the Na(+)-Ca(2+) exchanger (NCX), using Cd(2+), Gd(3+), or Ni(2+), respectively. PF-triggered Ca(2+) transients were significantly smaller in quiescent than in electrically paced myocytes. Paced Ca(2+) transients activated at the peak of PF-triggered Ca(2+) transients were not significantly smaller than those produced normally, suggesting functionally separate Ca(2+) pools for paced and PF-triggered transients. Suppression of nitric oxide (NO) or IP(3) signalling pathways did not alter the PF-triggered Ca(2+) transients. On the other hand, mitochondrial metabolic uncoupler FCCP, in the presence of oligomycin (to prevent ATP depletion), reversibly suppressed PF-triggered Ca(2+) transients, as did the mitochondrial Ca(2+) uniporter (mCU) blocker, Ru360. Reducing agent DTT and reactive oxygen species (ROS) scavenger tempol, as well as mitochondrial NCX (mNCX) blocker CGP-37157, inhibited PF-triggered Ca(2+) transients. In rhod-2 AM-loaded and permeabilized cells, confocal imaging of mitochondrial Ca(2+) showed a transient increase in Ca(2+) on caffeine exposure and a decrease in mitochondrial Ca(2+) on application of PF pulses of solution. These signals were strongly suppressed by either Na(+)-free or CGP-37157-containing solutions, implicating mNCX in mediating the Ca(2+) release process. We conclude that subjecting rat cardiac myocytes to pressurized flow pulses of solutions triggers the release of Ca(2+) from a store that appears to access mitochondrial Ca(2+).