Lead uptake in brain capillary endothelial cells: activation by calcium store depletion.
The mechanism by which lead crosses the blood-brain barrier is not known. Brain capillary endothelial cells, which form tight junctions with each other, are an important component of the blood-brain barrier. Lead must traverse these cells to reach the brain. In the present study, uptake of lead was followed in primary cultures of bovine brain capillary endothelial cells. Lead uptake into cells was measured by monitoring the fluorescence of cells loaded with indo-1 at a wavelength where indo-1 fluorescence is independent of calcium but quenched by binding of lead. Lead uptake was visualized with digital images recorded with a fluorescence microscope. Lead added to the extracellular medium caused fluorescence quench over time which was reversed upon addition of a membrane permeant heavy metal chelator. Lead uptake by cells in suspension, measured by fluorescence spectroscopy, exhibited time and concentration dependence. Lead uptake was enhanced following depletion of intracellular Ca2+ stores by the addition of thapsigargin, cyclopiazonic acid, or tert-butylhydroquinone, inhibitors of the sarco/endoplasmic reticulum calcium ATPase. SK&F 96365, which blocks store-operated calcium channels, inhibited the stimulation of lead uptake by thapsigargin. These results indicate that indo-1 fluorescence quench is a useful method for investigation of lead uptake in brain capillary endothelial cells. Furthermore, entry of lead into these cells is activated by the depletion of intracellular Ca2+ stores and may occur via store-operated cation channels.
Kerper
L E
LE
Department of Pharmacology and Physiology and the Cancer Center, University of Rochester School of Medicine and Dentistry, Rochester, New York 14642, USA.
Hinkle
P M
PM
D000818:Animals; D001812:Blood-Brain Barrier; D001921:Brain / Q000098:blood supply* / Q000378:metabolism; D002118:Calcium / Q000378:metabolism; D015220:Calcium Channels / Q000502:physiology; D002196:Capillaries / Q000378:metabolism; D002417:Cattle; D002478:Cells, Cultured; D004730:Endothelium, Vascular / Q000378:metabolism*; D007854:Lead / Q000493:pharmacokinetics*
D015220:Calcium Channels; D007854:Lead; D002118:Calcium