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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
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: | 4 |
| Deposited By: | Menzies Institute for Medical Research |
| Deposited On: | 2013-06-20 |
| Last Modified: | 2017-11-06 |
| Downloads: | 0 |
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