Preventive effect of Ninjin-yoei-to, a Kampo medicine, on amyloid ß1-42-induced neurodegeneration via intracellular Zn2+ toxicity in the dentate gyrus.

Ninjin-yoei-to (NYT), a Kampo medicine, has ameliorative effects on cognitive dysfunction via enhancing cholinergic neuron activity. To explore an efficacy of NYT administration for prevention and cure of Alzheimer's disease, here we examined the effect of NYT on amyloid ß1-42 (Aß1-42)-induced neurodegeneration in the dentate gyrus. A diet containing 3% NYT was administered to mice for 2 weeks and human Aß1-42 was intracerebroventricularly injected. Neurodegeneration in the dentate granule cell layer of the hippocampus, which was determined 2 weeks after the injection, was rescued by administration of the diet for 4 weeks. Aß staining (uptake) was not modified in the dentate granule cell layer by pre-administration of the diet for 2 weeks, while Aß1-42-induced increase in intracellular Zn2+ was reduced, suggesting that pre-administration of NYT prior to Aß injection is effective for reducing Aß1-42-induced Zn2+ toxicity in the dentate gyrus. As a matter of fact, Aß1-42-induced neurodegeneration in the dentate gyrus was rescued by pre-administration of NYT. Interestingly, the level of metallothioneins, intracellular Zn2+-binding proteins, which can capture Zn2+ from Zn-Aß1-42 complexes, was elevated in the dentate granule cell layer by pre-administration of NYT. The present study suggests that pre-administration of NYT prevents Aß1-42-mediated neurodegeneration in the dentate gyurs by induced synthesis of metallothioneins, which reduces intracellular Zn2+ toxicity induced by Aß1-42.

Dehydroeffusol Pprevents Amyloid ß1-42-mediated Hippocampal Neurodegeneration via Reducing Intracellular Zn2+ Toxicity.

Dehydroeffusol, a phenanthrene isolated from Juncus effusus, is a Chinese medicine. To explore an efficacy of dehydroeffusol administration for prevention and cure of Alzheimer's disease, here we examined the effect of dehydroeffusol on amyloid ß1-42 (Aß1-42)-mediated hippocampal neurodegeneration. Dehydroeffusol (15 mg/kg body weight) was orally administered to mice once a day for 6 days and then human Aß1-42 was injected intracerebroventricularly followed by oral administration for 12 days. Neurodegeneration in the dentate granule cell layer, which was determined 2 weeks after Aß1-42 injection, was rescued by dehydroeffusol administration. Aß staining (uptake) was not reduced in the dentate granule cell layer by pre-administration of dehydroeffusol for 6 days, while increase in intracellular Zn2+ induced with Aß1-42 was reduced, suggesting that pre-administration of dehydroeffusol prior to Aß1-42 injection is effective for Aß1-42-mediated neurodegeneration that was linked with intracellular Zn2+ toxicity. As a matter of fact, pre-administration of dehydroeffusol rescued Aß1-42-mediated neurodegeneration. Interestingly, pre-administration of dehydroeffusol increased synthesis of metallothioneins, intracellular Zn2+-binding proteins, in the dentate granule cell layer, which can capture Zn2+ from Zn-Aß1-42 complexes. The present study indicates that pre-administration of dehydroeffusol protects Aß1-42-mediated neurodegeneration in the hippocampus by reducing intracellular Zn2+ toxicity, which is linked with induced synthesis of metallothioneins. Dehydroeffusol, a novel inducer of metallothioneins, may protect Aß1-42-induced pathogenesis in Alzheimer's disease.

Retention Period of Amyloid ß1-42 in the Brain Extracellular Fluid as the Toxicological Determinant in Freely Moving Rats.

The pathological significance of amyloid-ß1-42 (Aß1-42) dynamics is poorly understood in the brain extracellular compartment. Here we test which of the concentration or the retention is critical for Aß1-42 toxicity after injection of equal dose into dentate granule cell layer of freely moving rats. The toxicity of Aß1-42 (25 µM) was compared between injections at the rate of 0.25 µL/min for 4 min (fast injection) and 0.025 µL/min for 40 min (slow injection). Dentate gyrus long-term potentiation (LTP) was affected 1 and 2 h after the fast injection, but not 4 h. In contrast, LTP was affected even 72 h after the slow injection. Aß1-42 staining 5 min after finish of the slow injection was more intense in the dentate granule cell layer than of the fast injection. The present study indicates that the retention of Aß1-42 in the extracellular fluid is correlated with neuronal Aß1-42 uptake and plays a key role in Aß1-42 neurotoxicity. In the extracellular fluid of the dentate gyrus, the retention period of Aß1-42 is much more critical for Aß1-42 toxicity than Aß1-42 concentration. It is likely that Aß1-42 toxicity is accelerated by the disturbance of Aß1-42 metabolism in the dentate gyrus.

Adrenergic ß receptor activation reduces amyloid ß1-42-mediated intracellular Zn2+ toxicity in dentate granule cells followed by rescuing impairment of dentate gyrus LTP.

Adrenergic ß receptor activation prevents human soluble amyloid ß (Aß)-induced impairment of long-term potentiation (LTP) in slices. On the basis of the evidence that human Aß1-42-induced impairment of LTP is due to Aß1-42-mediated Zn2+ toxicity, we postulated that adrenergic ß receptor activation reduces Aß1-42-mediated intracellular Zn2+ toxicity followed by rescuing Aß1-42 toxicity. To test the effect of adrenergic ß receptor activation, LTP was recorded at perforant pathway-dentate granule cell synapses of anesthetized rats 60 min after Aß1-42 injection into the dentate granule cell layer. Human Aß1-42-induced impairment of LTP was rescued by co-injection of isoproterenol, an adrenergic ß receptor agonist, but not by co-injection of phenylephrine, an adrenergic α1 receptor agonist. Isoproterenol did not reduce Aß1-42 uptake into dentate granule cells, but reduced increase in intracellular Zn2+ in dentate granule cells induced by Aß1-42. In contrast, phenylephrine did not reduce both Aß1-42 uptake and increase in intracellular Zn2+ by Aß1-42. In the case of human Aß1-40 and rat Aß1-42, which do not increase intracellular Zn2+, human Aß1-40- and rat Aß1-42-induced impairments of LTP were not rescued by co-injection of isoproterenol. The present study indicates that adrenergic ß receptor activation reduces Aß1-42-mediated increase in intracellular Zn2+ in dentate granule cells, resulting in rescuing Aß1-42-induced impairment of LTP. It is likely that noradrenergic neuron activation by stimulating the locus coeruleus is effective for rescuing Aß1-42-induced cognitive decline that is caused by intracellular Zn2+ dysregulation in the hippocampus.

In vivo synaptic activity-independent co-uptakes of amyloid ß1-42 and Zn2+ into dentate granule cells in the normal brain.

Neuronal amyloid ß1-42 (Aß1-42) accumulation is considered an upstream event in Alzheimer's disease pathogenesis. Here we report the mechanism on synaptic activity-independent Aß1-42 uptake in vivo. When Aß1-42 uptake was compared in hippocampal slices after incubating with Aß1-42, In vitro Aß1-42 uptake was preferentially high in the dentate granule cell layer in the hippocampus. Because the rapid uptake of Aß1-42 with extracellular Zn2+ is essential for Aß1-42-induced cognitive decline in vivo, the uptake mechanism was tested in dentate granule cells in association with synaptic activity. In vivo rapid uptake of Aß1-42 was not modified in the dentate granule cell layer after co-injection of Aß1-42 and tetrodotoxin, a Na+ channel blocker, into the dentate gyrus. Both the rapid uptake of Aß1-42 and Zn2+ into the dentate granule cell layer was not modified after co-injection of CNQX, an AMPA receptor antagonist, which blocks extracellular Zn2+ influx, Both the rapid uptake of Aß1-42 and Zn2+ into the dentate granule cell layer was not also modified after either co-injection of chlorpromazine or genistein, an endocytic repressor. The present study suggests that Aß1-42 and Zn2+ are synaptic activity-independently co-taken up into dentate granule cells in the normal brain and the co-uptake is preferential in dentate granule cells in the hippocampus. We propose a hypothesis that Zn-Aß1-42 oligomers formed in the extracellular compartment are directly incorporated into neuronal plasma membranes and form Zn2+-permeable ion channels.