Slow Ca(2+) wave stimulation using low repetition rate femtosecond pulsed irradiation

We demonstrated stimulation of Ca(2+) in living cells by near-infrared laser pulses operated at sub-MHz repetition rates. HeLa cells were exposed to focused 780 nm femtosecond pulses, generated by a titanium-sapphire laser and adjusted by an electro-optical modulator. We found that the laser-induced Ca(2+) waves could be generated over three orders of magnitude in repetition rates, with required laser pulse energy varying by less than one order of magnitude. Ca(2+) wave speed and gradients were reduced with repetition rate, which allows the technique to be used to modulate the strength and speed of laser-induced effects. By lowering the repetition rate, we found that the laser-induced Ca(2+) release is partially mediated by reactive oxygen species (ROS). Inhibition of ROS was successful only at low repetition rates, with the implication that ROS scavengers may in general be depleted in experiments using high repetition rate laser irradiation.