Digital imaging of free calcium changes and of spatial gradients in growing processes in single, mammalian central nervous system cells.

Abstract

Intracellular free calcium levels have been measured in cultured central nervous system (CNS) cells by using the fluorescent indicator fura-2 and digital imaging techniques. Cells were plated from rat embryo diencephalon (embryonic day 17 or 18), with nearly all of the cells surviving dissociation having undergone final mitosis within the previous 24 hr. The initially spherical cells were observed within the first 24 hr in culture when they were extending processes but had not established a network of fibers that would prevent the identification of the origin of a given fiber. Cells that were rapidly extending showed high Ca2+ levels in the regions of growth. Where processes had just emerged from the soma or where growth was proceeding from more than one pole, Ca2+ levels were uniform and estimated levels of 500 nM were commonly seen. In active growth cones distant from the soma, Ca2+ levels exceeded 200 nM, whereas the soma levels were in the 60-80 nM range. Nonextended and extended cells that had stalled had uniform Ca2+ levels in the range of 30-70 nM. The results show that high Ca2+ levels are at least a correlate of extension in CNS cells and that under some conditions the region of high calcium can be localized to a small part of the cell.