Analytical note
Multi-elemental analysis of brain tissue from healthy Wistar rats using sector field inductively coupled plasma mass spectrometry

https://doi.org/10.1016/j.sab.2004.06.008Get rights and content

Abstract

The normal distribution of a range of elements in the brain tissue of healthy Wistar rats was established using sector field inductively coupled plasma mass spectrometry. A protocol was developed to determine concentrations of Ag, Cd, Hg, Pb, Bi, U, V, Cr, Mn, Fe, Co, Ni, Cu, Zn, As and Se in specific brain regions. The concentrations of these elements were determined in the range of 2±1 (e.g. Cr in diencephalon) to 7558±450 ng ml−1 (e.g. Fe in olfactory bulb). The detection limits of the sixteen elements ranged between 5 and 300 pg ml−1, with U yielding the lowest and Fe the highest value. The validity of the protocol was assessed by the analysis of SRM 1577B Bovine Liver and brain tissue spike recoveries. A principal component analysis was used to reveal elemental patterns of the brain regions.

Introduction

The mammalian brain is extremely complex and is a highly specialized organ of the body. Specific regions of the brain have been shown to have unequal distribution of metabolic enzymes [1], neurotransmitter substances [2] and trace elements [3], [4], [5], [6]. In the case of the latter, there is a need to focus on the quantification of essential and toxic elements in the central nervous system under nutritional, biomedical and environmental controlled conditions in order to elucidate the effects of these elements on the mammalian brain. Earlier studies have shown that many trace elements are delicately balanced and play a pivotal metabolic role [7], [8], [9], [10], [11], [12]. If these balances are actively disturbed, such as from the ingestion or inhalation of high dosages over long periods of time, selected trace elements may become toxic. Due to the increasing heavy metal pollution experienced by our modern societies, neurological studies of trace elements is of paramount importance, since it will not only shed knowledge on the appropriateness of the environment toward human and animals habitation, but also on possible causes for neurological disorders [13].

There have been only two previous investigations of tissue from specific brain regions of rats using inductively coupled plasma mass spectrometry and both have centered on using low-resolution instruments. Fujimura et al. [14] measured Al, Sc, V, Cr, Mn, Cu, Rb, Sr and Mo in cerebellum, brain stem regions, hypothalamus, mid-brain combined with thalamus, striatum, hippocampus and cerebral cortex, using male Wistar rats exposed to one to three periods of restraint conditions, namely 15, 30 and 60 min stress periods. Moreover, Saito et al. [15] measured Cu, Mg, Mo, Rb and Zn in the same brain regions using Long-Evans Cinnamon and Long-Evans agouti rats.

Sector field inductively coupled plasma mass spectrometry (SF-ICP-MS) is a high-resolution technique, which can resolve spectral-based interferences [16]. It is capable of performing rapid and simultaneous measurements on a wide range of elements. Additionally, this technique is highly sensitive and so requires only small amounts of sample (e.g. typically low mg level for biopsies [17]). It is therefore ideally suited for determining most trace metal elements in biological matrices.

As an extension of our earlier work in developing protocols for SF-ICP-MS in order to quantify trace elements in biological matrices [18], we wish to detail a general protocol for the simultaneous SF-ICP-MS determination of some essential and toxic trace elements in specific brain regions of five healthy male Wistar rats. The brain regions have been specifically chosen (e.g. olfactory bulb, telencephalon, pons and medulla, superior/inferior colliculus, diencephalon, hippocampus and cerebellum), since they can be readily identified and enable unambiguous extraction of tissue. This is in contrast to the two previous low-resolution ICP-MS studies [14], [15], which have extracted tissue from combined regions of the brain (e.g. brain stem regions and mid-brain combined with thalamus). There has been a high-resolution investigation of human brain tissue [19]. Moreover, more pertinent to this study there have been no investigations of specific brain regions of Wistar rats (which may be employed as a benchmark for future studies, since their genetic diversity is eliminated via in-breeding). To further highlight possible synergies across brain regions, a characteristic elemental pattern of each tissue in this study has also been revealed by employing a principal component analysis (PCA).

Section snippets

Laboratory ware, reference material, reagents and standards

All solutions were prepared with polypropylene laboratory ware [pipette tips (Rainin edp3™ electronic pipettes), volumetric flasks (Kartell) and sample tubes (Cetac)]. Standard solutions were prepared from Perkin Elmer 10 μg ml−1 multi-element ICP-MS standards and 1000 μg ml−1 single-element ICP-MS standards (Rh, Te and Ge). High purity HNO3 (65% v/v, Suprapur®, Merck) and H2O2 (30% v/v, Suprapur®, Merck) were used as received. Reagent grades of mannitol (>98%, Sigma), Na2HPO4 and NaH2PO4 (98%,

Results and discussion

The trace elements that were determined can be divided into two basic categories, namely: essential (i.e. those elements that have established physiological roles, namely: V, Cr, Mn, Fe, Co, Ni, Cu, Zn, Se) and non-essential (i.e. those elements that are toxic or have unclear physiological roles, namely: Ag, Cd, Hg, Pb, Bi, U, As) [7], [8], [9], [10], [11], [12]. Moreover, the essential trace elements listed are also those that will be monitored in terms of experience-dependent plasticity

Conclusions

The investigation of the brain tissue of healthy Wistar rats has now been established using SF-ICP-MS. Tissue concentrations of As, Ag, Bi, Cd, Co, Cr, Cu, Fe, Hg, Mn, Ni, Pb, Se, U, V and Zn in the olfactory bulb, telencephalon, pons and medulla, superior/inferior colliculus, diencephalon, hippocampus and cerebellum brain regions have been investigated and elemental patterns have been revealed using PCA. The proposed analytical method and analysis provides an effective probe in measuring the

Acknowledgments

We would like to acknowledge the support of the Advanced Mass Spectrometry Unit for the generous allocations of time on the SF-ICP-MS. We wish to acknowledge support from the National Health and Medical Research Council, the Hunter Medical Research Institute, the University of Newcastle Central Animal Facility, the University of Newcastle Research Program and Traveling Grants schemes. MCP wishes to acknowledge support from a University of Newcastle Research scholarship.

References (25)

  • S. Leong et al.

    Energy-metabolizing enzymes in brain regions of adult and aging rats

    J. Neurochem.

    (1981)
  • J. Lai et al.

    Brain regional distribution of glutamic acid decarboxylase, chlorine acetyltransferase, and acetylcholinesterase in the rat: effect of chronic manganese chloride administration after two years

    J. Neurochem.

    (1981)
  • Cited by (0)

    View full text