Topographical Analysis of Reactive Zinc in the Central Nervous System of Adult Zebrafish (Danio Rerio)

Abstract

Reactive zinc (Zn) is crucial for neuronal signaling and is largely distributed within presynaptic vesicles of some axon terminals of distinct vertebrates. However, the distribution of reactive Zn throughout the central nervous system (CNS) is not fully explored. We performed a topographical study of CNS structures containing reactive Zn in the adult zebrafish (Danio rerio). Slices of CNS from zebrafish were stained by Neo-Timm and/or cresyl violet. The Zn specificity of Neo-Timm was evaluated with Zn chelants, N,N,N¢,N¢-Tetrakis(2-pyridylmethyl)ethylenediamine (TPEN), sodium diethyldithiocarbamate (DEDTC), Zn sulfide washing solution, and hydrochloric acid (HCl). Unfixed slices were also immersed in the fluorescent Zn probe (zinpyr-1). Yellowto-brown-to-black granules were revealed by Neo-Timm in the zebrafish CNS. Telencephalon exhibited slightly stained regions, while rhombencephalic structures showed high levels of staining. Although stained granules were found on the cell bodies, rhombencephalic structures showed a neuropil staining profile. The TPEN produced a mild reduction in Neo-Timm staining, while HCl and mainly DEDTC abolished the staining, indicating a large Zn content. This result was also confirmed by the application of a Zn probe. The present topographical study revealed reactive Zn throughout the CNS in adult zebrafish that should be considered in future investigation of Zn in the brain on a larger scale.

Brain zinc chelation by diethyldithiocarbamate increased the behavioral and mitochondrial damages in zebrafish subjected to hypoxia

“…Specifically, the PGz is a cerebral area of optic tectum with abundant and well-located content of chelatable Zn 7 , being related to visual functions of zebrafish. In fact, chelatable Zn has been identified as important neuromodulator in visual pathway of this species 18 .…”

“…The chelatable Zn content from PGz was histochemically stained by Neo-Timm method, as performed in zebrafish brain 7 . The PGz was evaluated due to its abundant content of chelatable Zn and its intrinsic relation with optic tectum, which plays a role in zebrafish visual processing.…”

“…In particular, the diethyldithiocarbamate (DEDTC) is a chelator highly selective for Zn 7 able to cross the blood-brain barrier 8 . Previous studies showed that DEDTC is beneficial against several disorders 9 10 .…”

Modulation of the chelatable Zn pool in the brain by diethyldithiocarbamate is associated with behavioral impairment in adult zebrafish

“…Histochemical staining of reactive Zn was performed using the Neo-Timm method previously described for zebrafish brains. 26 After the exposure period, brains were excised and immersed in 3% glutaraldehyde in 0.1 M phosphate buffer ( pH 7.4) for 24 h, and then transferred to a sodium sulfide solution (1% Na 2 S) in 0.12 M Millonig's buffer for 24 h. Coronal (30 μmthick) slices were cut in a vibroslicer (VTS-1000; Leica), and mounted on slides. For histochemical staining of reactive Zn, the slices were incubated in a solution containing silver nitrate.…”

“…24 Particularly, the cellular Zn transporters of zebrafish embryos have been recently characterized, showing a high degree of similarity to their mammalian counterparts. 25 Moreover, a reactive pool of Zn is present throughout the CNS of zebrafish, 26 suggesting that Zn could play a role in brain homeostasis in this species. These characteristics make the zebrafish an appropriate model to investigate the behavioral changes caused by DEDTC via modulation of the Zn pool in the CNS.…”