Increase in hippocampal cell death after treatment with kainate in zinc deficiency

https://doi.org/10.1016/j.neuint.2005.07.009Get rights and content

Abstract

Susceptibility to kainate-induced seizures is enhanced by zinc deficiency. To evaluate kainate-induced excitotoxicity in zinc deficiency, the relationship between kainate-induced seizures and hippocampal cell death was examined in control and zinc-deficient mice. Mice were fed a control and zinc-deficient diet for 4 weeks, and then intraperitoneally injected with 12 mg/kg kainate every 60 min three times. The rate of dead mice to the total mice was higher in zinc-deficient group than in control group 3 days after the last injection of kainate. In the survivals, which exhibited tonic convulsions in both control and zinc-deficient groups, kainate-induced hippocampal cell death was also analyzed by cresyl violet staining. Neuronal loss was more observed in the CA1, CA2 and CA3 pyramidal cell layers of zinc-deficient group than those of the control group. TUNEL-positive cells were significantly more detected in the CA1 and CA3 pyramidal cell layers of zinc-deficient group. These results demonstrate that kainate-induced hippocampal cell death occurs more easily in zinc deficiency. Extracellular zinc concentration detected with ZnAF-2 was significantly decreased in the hippocampal CA3 of zinc-deficient mice, in agreement with the previous data measured by in vivo microdialsysis. Synaptically released zinc may be less involved in kainate-induced hippocampal cell death in zinc deficiency.

Introduction

Dietary zinc deficiency not only retards the growth of humans and animals, but also affects brain function (Golub et al., 1995, Penland, 2000, Sandstead et al., 2000, Takeda, 2000). Approximately, 50% of the world population does not get adequate zinc (Brown et al., 2001). Ten percent of the North American population consumes less than half the recommended daily allowance for zinc (King et al., 2000).

Zinc is a component of over 300 proteins, such as DNA-binding proteins with zinc fingers and is important for the function of many enzymes and other proteins, including some unique to the brain and important to neurotransmission (Prohaska, 1987, Ebadi, 1991, Vallee and Falchuk, 1993). Dietary zinc deficiency affects zinc homeostasis in the brain (Takeda, 2001); zinc concentration in the presynaptic vesicle and extracelullar zinc concentration are decreased in 8-week-old rats fed a zinc-deficient diet for 4 weeks (Takeda et al., 2003a), in which susceptibility to kainate-induced seizures is more enhanced than in the control rats (Takeda et al., 2003b). The enhanced susceptibility to epileptic seizures is also observed in zinc-deficient EL mice (Fukahori and Itoh, 1990) and kindled cats (Sterman et al., 1986). The in vivo microdialysis experiment demonstrated an enhanced release of glutamate associated with a decrease in GABA concentrations as a possible mechanism for the enhanced susceptibility to kainate-induced seizures in zinc deficiency (Takeda et al., 2003b).

To evaluate kainate-induced excitotoxicity in zinc deficiency, in the present study, the relationship between kainate-induced seizures and hippocampal cell death was examined in control and zinc-deficient mice, which showed the same seizure severity scores. The observation of hippocampal cell death is important to evidence the vulnerability to glutamate excitotoxicity in zinc deficiency. In the hippocampal CA3 of zinc-deficient mice, furthermore, extracellular zinc and intracellular calcium levels were checked using their indicators.

Section snippets

Diets and chemicals

Control (44 mg Zn/kg) and zinc-deficient (2.7 mg Zn/kg) diets were purchased from Oriental Yeast Co. Ltd. (Yokohama, Japan). ZnAF-2, a membrane-impermeable zinc indicator, was kindly supplied from Daiichi Pure Chemicals Co., Ltd. (Tokyo, Japan). Fura-2 AM, a membrane-permeable calcium indicator, was purchased from Molecular Probes, Inc. (Eugene, OR).

Experimental animals

Male ddY mice (both 3-week-old) were purchased from Japan SLC (Hamamatsu, Japan). They were housed under the standard laboratory conditions (23 ± 1 °C,

Results

When 4-week-old mice were fed a zinc-deficient diet for 4 weeks, zinc concentration in the liver and serum was significantly decreased, while zinc concentration in the whole brain was not decreased (Fig. 1). However, zinc concentration in the hippocampus was significantly decreased by the zinc deficiency. Kainate was injected into the control and zinc-deficient mice and seizure severity of each mouse was semi-quantified according the seizure score scale of Racine (1972) and Sperk et al. (1985).

Discussion

Zinc homeostasis in the brain is altered by dietary zinc deficiency (Takeda, 2001) and its alteration may be associated with the etiology and manifestation of epileptic seizures (Sterman et al., 1988). When susceptibility to kainate-induced seizures was examined in 8-week-old mice and rats fed the zinc-deficient diet for 4 weeks, kainate-induced seizures were enhanced by dietary zinc deficiency (Takeda et al., 2003b). Furthermore, the mechanism of the enhanced seizure susceptibility was studied

References (32)

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