Measurement of intracellular free zinc concentrations accompanying zinc-induced neuronal death.
Toxic zinc influx may contribute to selective neuronal death after transient global ischemia. We previously used the high-affinity (K(D) = 27 nm) fluorescent dye mag-fura-5 to detect initial increases in neuronal intracellular free Zn(2+) ([Zn(2+)](i)) associated with brief Zn(2+) exposure. Here we used the specific low-affinity Zn(2+) indicator Newport Green (K(D) = 1 microm) to measure the peak levels of [Zn(2+)](i) attained during prolonged, toxic exposures to extracellular Zn(2+). Murine cortical cell cultures exposed for 5-10 min to 300 microm Zn(2+) in the presence of kainate or elevated extracellular K(+) developed widespread neuronal death over the next 24 hr. Such Zn(2+) exposure under depolarizing conditions was accompanied by a large increase in [Zn(2+)](i) reaching several hundred nanomolar, which gradually recovered over the next 20-40 min after termination of Zn(2+) exposure. Both the level of [Zn(2+)](i) elevation and the extent of subsequent neuronal death depended on the concentration of extracellular Zn(2+) between 30 microm and 1 mm. In contrast, exposure to 300 microm Zn(2+) in the presence of 300 microm NMDA resulted in little increase in [Zn(2+)](i) and little neuronal death, suggesting that NMDA receptor-gated channels are less important as a route of toxic Zn(2+) entry than voltage-gated calcium channels.
Canzoniero
L M
LM
Center for the Study of Nervous System Injury and Department of Neurology, Washington University School of Medicine, St. Louis, Missouri 63110, USA.
Turetsky
D M
DM
Choi
D W
DW
D000818:Animals; D016923:Cell Death; D002478:Cells, Cultured; D018690:Excitatory Amino Acid Agonists / Q000494:pharmacology; D005110:Extracellular Space / Q000378:metabolism; D005333:Fetus; D005456:Fluorescent Dyes; D007424:Intracellular Fluid / Q000378:metabolism; D007608:Kainic Acid / Q000494:pharmacology; D051379:Mice; D018613:Microscopy, Confocal; D016202:N-Methylaspartate / Q000494:pharmacology; D019579:Neocortex / Q000166:cytology / Q000378:metabolism; D009457:Neuroglia / Q000166:cytology / Q000378:metabolism; D009474:Neurons / Q000166:cytology / Q000187:drug effects / Q000378:metabolism*; D011188:Potassium / Q000494:pharmacology; D015032:Zinc / Q000378:metabolism* / Q000494:pharmacology
D018690:Excitatory Amino Acid Agonists; D005456:Fluorescent Dyes; D016202:N-Methylaspartate; D015032:Zinc; D011188:Potassium; D007608:Kainic Acid