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Intracellular dialysis disrupts Zn2+ dynamics and enables selective detection of Zn2+ influx in brain slice preparations

Citation

Aiba, I and West, AK and Sheline, CT and Shuttleworth, CW, Intracellular dialysis disrupts Zn2+ dynamics and enables selective detection of Zn2+ influx in brain slice preparations, Journal of Neurochemistry, 125, (6) pp. 822-831. ISSN 0022-3042 (2013) [Refereed Article]

Copyright Statement

2013 International Society for Neurochemistry

DOI: doi:10.1111/jnc.12246

Abstract

We examined the impact of intracellular dialysis on fluorescence detection of neuronal intracellular Zn2+ accumulation. Comparison between two dialysis conditions (standard; 20 min, brief; 2 min) by standard whole-cell clamp revealed a high vulnerability of intracellular Zn2+ buffers to intracellular dialysis. Thus, low concentrations of zinc-pyrithione generated robust responses in neurons with standard dialysis, but signals were smaller in neurons with short dialysis. Release from oxidation-sensitive Zn2+ pools was reduced by standard dialysis, when compared with responses in neurons with brief dialysis. The dialysis effects were partly reversed by inclusion of recombinant metallothionein-3 in the dialysis solution. These findings suggested that extensive dialysis could be exploited for selective detection of transmembrane Zn2+ influx. Different dialysis conditions were then used to probe responses to synaptic stimulation. Under standard dialysis conditions, synaptic stimuli generated significant FluoZin-3 signals in wild-type (WT) preparations, but responses were almost absent in preparations lacking vesicular Zn2+ (ZnT3-KO). In contrast, under brief dialysis conditions, intracellular Zn2+ transients were very similar in WT and ZnT3-KO preparations. This suggests that both intracellular release and transmembrane flux can contribute to intracellular Zn2+ accumulation after synaptic stimulation. These results demonstrate significant confounds and potential use of intracellular dialysis to investigate intracellular Zn2+ accumulation mechanisms.

Item Details

Item Type:Refereed Article
Keywords:hippocampal slice, metallothionein, pyrithione, whole-cell, Zinc, ZnT3
Research Division:Medical and Health Sciences
Research Group:Neurosciences
Research Field:Central Nervous System
Objective Division:Health
Objective Group:Clinical Health (Organs, Diseases and Abnormal Conditions)
Objective Field:Nervous System and Disorders
Author:West, AK (Professor Adrian West)
ID Code:85242
Year Published:2013
Web of Science® Times Cited:2
Deposited By:Menzies Institute for Medical Research
Deposited On:2013-06-20
Last Modified:2017-11-06
Downloads:0

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