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Part Number Referenced: |
24-Well Plates |
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| TR |
TITLE |
AUTHORS |
KEYWORDS |
MATERIALS & METHODS
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MICROSCOPIC
TECHNIQUES |
SPECIES |
MORPHOLOGY |
CELL
LINE |
| 625 |
Preferential
Zn21 influx through Ca21-permeable AMPAykainate
channels triggers prolonged mitochondrial superoxide production |
Stefano
L. Sensi, Hong Z. Yin, Sean G. Carriedo, Shyam S. Rao, John H. Weiss |
N-methyl-D-asparte,
kainate, hydroethidine oxidation, reactive oxygen
species (ROS), mitochondrial electron transport blocker rotenone |
Briefly, dissociated mixed neocortical
cell suspensions were prepared...on previously established astrocytic monolayers in either 24-well
plates or glass-bottomed dishes (Plastek Cultureware, Ashland,MA).
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epifluorescence
microscopy, inverted microscopy |
mouse |
neuronal |
mixed
neocortical
cell suspensions |
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Synaptically released Zn21 can enter and cause injury
to postsynaptic neurons. Microfluorimetric studies using the Zn21-sensitive probe, Newport green,
examined levels of [Zn21]i attained in cultured cortical neurons on exposure to N-methyl-D-asparte,
kainate, or high K1 (to activate voltage-sensitive Ca21 channels) in the presence of 300 mM
Zn21. Indicating particularly high permeability through Ca21-permeable a-amino3-hydroxy-5-methyl-4-
isoxazolepropionic-acidykainate (Ca-AyK) channels, micromolar [Zn21]i rises were observed only
after kainate exposures and only in neurons expressing these channels [Ca-Ay K(1) neurons].
Further studies using the oxidation-sensitive dye, hydroethidine, revealed Zn21-dependent reactive
oxygen species (ROS) generation that paralleled the [Zn21]i rises, with rapid oxidation observed
only in the case of Zn21 entry through Ca-AyK channels. Indicating a mitochondrial source of
this ROS generation, hydroethidine oxidation was inhibited by the mitochondrial electron transport
blocker, rotenone. Additional evidence for a direct interaction between Zn21 and mitochondria
was provided by the observation that the Zn21 entry through Ca-AyK channels triggered rapid
mitochondrial depolarization, as assessed by using the potentialsensitive dye tetramethylrhodamine
ethylester. Whereas Ca21 influx through Ca-AyK channels also triggers ROS production, the [Zn21]i
rises and subsequent ROS production are of more prolonged duration |
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