Fs-laser-induced Ca2+ concentration change during membrane perforation for cell transfection

Area of Science:

• Cell biology
• Biophysics
• Laser-based cell manipulation

Background:

• Fs-laser opto-perforation offers gentle gene transfer into sensitive cells.
• Existing methods face challenges including cell stress and unclear mechanisms.
• Understanding cellular responses to laser-induced membrane perforation is critical.

Purpose of the Study:

• Investigate calcium (Ca2+) homeostasis during laser-induced cell membrane perforation.
• Elucidate the role of extracellular calcium in cellular responses to opto-perforation.
• Determine the impact of laser parameters on calcium dynamics.

Main Methods:

• Utilized fs-laser opto-perforation for cell membrane manipulation.
• Monitored cytosolic Ca2+ concentration changes in response to laser exposure.
• Varied extracellular calcium concentrations and laser parameters (repetition rate, irradiation time).

Main Results:

• Opto-perforation induced an increase in cytosolic Ca2+ concentration.
• This increase was dependent on extracellular calcium availability.
• Extracellular Ca2+ uptake varied with laser repetition rate and irradiation time, potentially triggering Ca2+-induced Ca2+ release.
• Localized perforations could induce instantaneous intracellular Ca2+ release.

Conclusions:

• Extracellular calcium influx is a key factor in cellular responses to opto-perforation.
• Laser parameters significantly influence calcium dynamics during the procedure.
• Performing opto-perforation in calcium-free solutions may mitigate negative side effects.