May 2008
Volume 49, Issue 13
ARVO Annual Meeting Abstract  |   May 2008
Internalization of L-Type Ca Channels Protects Cells Against Glutamate-Induced Excitotoxicity in Salamander Retina
Author Affiliations & Notes
  • A. Akopian
    Ophthalmology, New York Univ Sch of Medicine, New York, New York
  • F. Mizuno
    Ophthalmology, New York Univ Sch of Medicine, New York, New York
  • Footnotes
    Commercial Relationships  A. Akopian, None; F. Mizuno, None.
  • Footnotes
    Support  NIH Grant EY12497
Investigative Ophthalmology & Visual Science May 2008, Vol.49, 5800. doi:
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      A. Akopian, F. Mizuno; Internalization of L-Type Ca Channels Protects Cells Against Glutamate-Induced Excitotoxicity in Salamander Retina. Invest. Ophthalmol. Vis. Sci. 2008;49(13):5800.

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      © ARVO (1962-2015); The Authors (2016-present)

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Purpose: : Glutamate-induced excitotoxicity has been implicated in retinal diseases that lead to retinal ischemia, common cause of visual impairment and blindness. The intracellular Ca2+ overload resulting from increased Ca2+ influx through both receptor-operated and voltage-activated L-type Ca channels is believed to be the key trigger for a loss of retinal amacrine and ganglion cells. In the present study we investigated whether of glutamate-induced L-type channel internalization as a protective mechanism against excitotoxicity.

Methods: : Immunocytochemistry, live/dead viability assay and whole-cell patch clamp technique were applied to study the effect of L-type Ca channel internalization on glutamate or kainate-induced excitotoxicity in dissociated cells and slice preparations.

Results: : Exposure of dissociated cells for 10 min to 100 µM glutamate resulted internalization of Cav1.3 L-type channels. This process was attenuated when cells were pretreated with BAPTA-AM, indicating that effect of glutamate was mediated by elevation of intracellular [Ca2+]. To study the role of Ca channel internalization in glutamate-induced excitotoxicity, we used retinal slices, incubating them for various times in 100-200 µM kainate. Quantification with live/dead viability assay showed that the number of dead cells in the GCL of slices incubated in Ringer solution and those exposed to kainate were 1.4±0.4 cells/slice and 5.3±0.9 cells/slice, respectively. Dynamin-inhibitory peptide (DIP), suppressed glutamate-induced internalization of Cav1.3 channels and significantly increased the number of dead cells to 13.8±1.2 cells/slice. We reported earlier that F-actin disrupter cytochalasin D (cytD) promotes internalization of Cav1.3 L-type channels (Cristofanilli et al 2007). To probe a link between F-actin dynamics and excitotoxicity, we pretreated retinal slices with either jasplakinolide or cytD, prior to exposing them to glutamate or kainate. Jasplakinolide increased, whereas cytD reduced, the number of dead ganglion cells induced by kainate application. Jasplakinolide also attenuated the glutamate-induced internalization of Cav1.3 channels. In patch clamp experiments, treatment of retinal slices with DIP or jasplakinolide suppressed the inhibitory effect of glutamate on L-type Ca current in ganglion cells.

Conclusions: : These data suggest that glutamate-induced Ca-mediated internalization of L-type channels may serve as a negative feedback mechanism to protect cells against excitotoxicity.

Keywords: calcium • cell survival • cytoskeleton 

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