Confocal microscopy of intracellular calcium dynamics during fertilization.

During fertilization in all animals investigated, fully developed eggs or meiotically immature oocytes must generate a proper change in their intracellular free calcium concentrations ([Ca]i) to develop normally (31). The first images of a fertilization-induced calcium response were obtained in fish oocytes that had been microinjected with the calcium-sensitive photoprotein, aequorin (4). Since then, aequorin has been used to monitor calcium dynamics in several other species (13). However, with the introduction of confocal laser scanning microscopy (CLSM) and a wide collection of calciumbinding fluorescent probes, there has been a rapid growth in the number of investigations that use CLSM to track changes in [Ca]i during fertilization (24). This column specifically addresses changes in [Ca2+]i that occur within fertilized eggs imaged by commonly available confocal laser scanning microscopes (i.e., those employing single-photon excitations provided by a low-power laser). It serves as an adjunct to more broadly based reviews covering confocal microscopy and calcium dynamics in eukaryotic cells subjected to diverse stimuli (15,27). Several advantages and potential drawbacks to consider when conducting CLSM studies of fertilization-induced calcium dynamics are discussed. In addition, a few of the major findings of such investigations are described, and some possible future directions for CLSM analyses are explored.