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Since the laser has been invented it has been highly instrumental in ablating different parts of the cell to test their functionality. Through induction of damage in a defined sub-micron region in the cell nucleus, laser microirradiation technique is now established as a powerful real-time and high-resolution methodology to investigate mechanisms of DNA damage response and repair, the fundamental cellular processes for the maintenance of genomic integrity, in mammalian cells. However, irradiation conditions dictate the amounts, types and complexity of DNA damage, leading to different damage signaling responses. Thus, in order to properly interpret the results, it is important to understand the features of laser-induced DNA damage. In this review, we describe different types of DNA damage induced by the use of different laser systems and parameters, and discuss the mechanisms of DNA damage induction. We further summarize recent advances in the application of laser microirradiation to study spatiotemporal dynamics of cellular responses to DNA damage, including factor recruitment, chromatin modulation at damage sites as well as more global damage signaling. Finally, possible future application of laser microirradiation to gain further understanding of DNA damage response will be discussed. 

Astrocytes are known to respond to various perturbations with oscillations of calcium, including to cellular injury. Less is known about astrocytes’ ability to detect DNA/nuclear damage. This study looks at changes in calcium signaling in response to laser-induced nuclear damage using a NIR Ti:Sapphire laser. Primary astrocytes derived from genetically engineered mice expressing G6Campf genetically encoded calcium indicator were imaged in response to laser induced injury. Combining laser nanosurgery with calcium imaging of primary astrocytes allow for spatial and temporal observation of the astrocyte network in response to nuclear damage. Nuclear damage resulted in a significant increase in calcium peak frequency, in nuclear damaged cells and astrocytes directly attached to it. The increase in calcium event frequency observed in response to damage and the transfer to neighboring cells was not observed in cytoplasm damaged cells. Targeted astrocytes and attached neighboring cells treated with Poly (ADP-ribose) polymerase inhibitor have a significantly lower peak frequency following laser damage to the nucleus. These results indicate the increase in calcium peak frequency following nuclear damage is poly (ADP-ribose) polymerase dependent.