Flash Photolysis of Caged IP3 to Trigger Intercellular Ca2+ Waves

Abstract

Caged IP3 is an inactive form of the second messenger IP3, consisting of the biologically active molecule linked to a cage group through a photolabile bond. This bond is cleaved by exposure to brief “flashes” of ultraviolet (UV) light, thereby releasing the active IP3 molecule. The protection of caged IP3 against metabolic transformation in combination with a defined time point of fast photoliberation of IP3 provides an efficient way to temporally and spatially control the cytosolic release of IP3 and subsequent increase of cytoplasmic Ca2+. These properties make it an ideal method for kinetic studies and also a well-suited procedure to initiate intercellular Ca2+ waves from a point source of IP3. This protocol describes cell loading with membrane impermeable caged IP3 and the UV flash illumination procedure.