Miki Suzuki

Transient Increase in Zn2+ in Hippocampal CA1 Pyramidal Neurons Causes Reversible Memory Deficit

The translocation of synaptic Zn2+ to the cytosolic compartment has been studied to understand Zn2+ neurotoxicity in neurological diseases. However, it is unknown whether the moderate increase in Zn2+ in the cytosolic compartment affects memory processing in the hippocampus. In the present study, the moderate increase in cytosolic Zn2+ in the hippocampus was induced with clioquinol (CQ), a zinc ionophore. Zn2+ delivery by Zn-CQ transiently attenuated CA1 long-term potentiation (LTP) in hippocampal slices prepared 2 h after i.p. injection of Zn-CQ into rats, when intracellular Zn2+ levels was transiently increased in the CA1 pyramidal cell layer, followed by object recognition memory deficit. Object recognition memory was transiently impaired 30 min after injection of ZnCl2 into the CA1, but not after injection into the dentate gyrus that did not significantly increase intracellular Zn2+ in the granule cell layer of the dentate gyrus. Object recognition memory deficit may be linked to the preferential increase in Zn2+ and/or the preferential vulnerability to Zn2+ in CA1 pyramidal neurons. In the case of the cytosolic increase in endogenous Zn2+ in the CA1 induced by 100 mM KCl, furthermore, object recognition memory was also transiently impaired, while ameliorated by co-injection of CaEDTA to block the increase in cytosolic Zn2+. The present study indicates that the transient increase in cytosolic Zn2+ in CA1 pyramidal neurons reversibly impairs object recognition memory.

Impairment of recognition memory and hippocampal long-term potentiation after acute exposure to clioquinol

Clioquinol (CQ) was associated with cases of transient global amnesia and with the neurodegenerative syndrome subacute myelo-optico-neuropathy (SMON) in humans. However, CQ forms lipophilic chelates with cations and has the potential as a scientific and clinical tool used for selective modulation of histochemically reactive zinc pools. The relationship among transient lack of synaptic zinc release, hippocampal long-term potentiation (LTP) induction and cognitive memory is poorly understood. To evaluate the role of synaptic zinc release, in the present study, hippocampal LTP induction and cognitive behavior were examined in young rats after i.p. injection of CQ (30 mg/kg). Intracellular zinc detected by Timms stain and extracellular (synaptic cleft) zinc detected by ZnAF-2 were significantly decreased in the hippocampus 6 h after CQ injection. The molecular layer of the dentate gyrus, in which perforant path-granule cell synapses exist, was most responsive to CQ injection. Dentate gyrus LTP was induced similarly to the control 2 h after CQ injection, while significantly attenuated 6-24 h after CQ injection. In the training trial of the object recognition memory 2 h after CQ injection, there was no significant difference in learning behavior between the control and CQ-treated rats. In the test trial, CQ-treated rats showed normal recognition memory 1 h after the training, whereas recognition memory deficit 24 h after the training unlike the control rats. These results indicate that acute exposure to CQ impairs long-term (24 h) memory in the hippocampus of young rats. The CQ-mediated attenuation of dentate gyrus LTP, which may be associated with the transient lack of zinc release from zincergic neurons, seems to be involved in the impairment of the long-term memory.

Excess influx of Zn(2+) into dentate granule cells affects object recognition memory via attenuated LTP.

The influx of extracellular Zn(2+) into dentate granule cells is nonessential for dentate gyrus long-term potentiation (LTP) and the physiological significance of extracellular Zn(2+) dynamics is unknown in the dentate gyrus. Excess increase in extracellular Zn(2+) in the hippocampal CA1, which is induced with excitation of zincergic neurons, induces memory deficit via excess influx of Zn(2+) into CA1 pyramidal cells. In the present study, it was examined whether extracellular Zn(2+) induces object recognition memory deficit via excess influx of Zn(2+) into dentate granule cells. KCl (100 mM, 2 µl) was locally injected into the dentate gyrus. The increase in intracellular Zn(2+) in dentate granule cells induced with high K(+) was blocked by co-injection of CaEDTA and CNQX, an extracellular Zn(2+) chelator and an AMPA receptor antagonist, respectively, suggesting that high K(+) increases the influx of Zn(2+) into dentate granule cells via AMPA receptor activation. Dentate gyrus LTP induction was attenuated 1 h after KCl injection into the dentate gyrus and also attenuated when KCl was injected 5 min after the induction. Memory deficit was induced when training of object recognition test was performed 1 h after KCl injection into the dentate gyrus and also induced when KCl was injected 5 min after the training. High K(+)-induced impairments of LTP and memory were rescued by co-injection of CaEDTA. These results indicate that excess influx of Zn(2+) into dentate granule cells via AMPA receptor activation affects object recognition memory via attenuated LTP induction. Even in the dentate gyrus where is scarcely innervated by zincergic neurons, it is likely that extracellular Zn(2+) homeostasis is strictly regulated for cognition.

Zinc differentially acts on components of long-term potentiation at hippocampal CA1 synapses

Long-term potentiation (LTP) at hippocampal CA1 synapses consists of N-methyl-d-aspartate (NMDA) receptor-dependent and NMDA receptor-independent forms. The action of divalent heavy metals, which are NMDA receptor antagonists, was examined focusing on the evidence that CA1 LTP induced by a 100-Hz tetanus for 1s is abolished in the presence of 2-amino-5-phosphonovalerate (APV), a NMDA receptor antagonist. Only ZnCl2 (5microM) of heavy metals tested potentiated CA1 LTP. CA1 LTP induced by repeated 100-Hz tetanus (1s, 6 times, 10min interval), which reached a plateau in magnitude, was abolished in the presence of 50microM APV. In this case, CA1 LTP after the first tetanus was potentiated in the presence of 5microM ZnCl2, whereas CA1 LTP after the last tetanus was not potentiated. These results indicate that the magnitude of NMDA receptor-dependent CA1 LTP can be positively shifted with 5microM ZnCl2 in the range of the maximum magnitude. CA1 LTP induced by a 200-Hz tetanus for 1s was not potentiated in the presence of 5microM ZnCl2 and was partially inhibited in the presence of APV. Furthermore, CA1 LTP induced by a 200-Hz tetanus for 1s in the presence of APV was not potentiated in the presence of 5microM ZnCl2, indicating that NMDA receptor-independent CA1 LTP is not potentiated with 5microM ZnCl2. The present study suggests that zinc differentially acts on CA1 LTP components.

Influx of extracellular Zn2+ into the hippocampal CA1 neurons is required for cognitive performance via long-term potentiation

Physiological significance of synaptic Zn(2+) signaling was examined in the CA1 of young rats. In vivo CA1 long-term potentiation (LTP) was induced using a recording electrode attached to a microdialysis probe and the recording region was locally perfused with artificial cerebrospinal fluid (ACSF) via the microdialysis probe. In vivo CA1 LTP was inhibited under perfusion with CaEDTA and ZnAF-2DA, extracellular and intracellular Zn(2+) chelators, respectively, suggesting that the influx of extracellular Zn(2+) is required for in vivo CA1 LTP induction. The increase in intracellular Zn(2+) was chelated with intracellular ZnAF-2 in the CA1 1h after local injection of ZnAF-2DA into the CA1, suggesting that intracellular Zn(2+) signaling induced during learning is blocked with intracellular ZnAF-2 when the learning was performed 1h after ZnAF-2DA injection. Object recognition was affected when training of object recognition test was performed 1h after ZnAF-2DA injection. These data suggest that intracellular Zn(2+) signaling in the CA1 is required for object recognition memory via LTP. Surprisingly, in vivo CA1 LTP was affected under perfusion with 0.1-1μM ZnCl2, unlike the previous data that in vitro CA1 LTP was enhanced in the presence of 1-5μM ZnCl2. The influx of extracellular Zn(2+) into CA1 pyramidal cells has bidirectional action in CA1 LTP. The present study indicates that the degree of extracellular Zn(2+) influx into CA1 neurons is critical for LTP and cognitive performance.

Involvement of glucocorticoid-mediated Zn2+ signaling in attenuation of hippocampal CA1 LTP by acute stress.

Glucocorticoid-glutamatergic interactions have been proposed as a potential model to explain stress-mediated impairment of cognition. However, it is unknown whether glucocorticoid-zincergic interactions are involved in this impairment. Histochemically reactive zinc (Zn(2+)) is co-released with glutamate from zincergic neurons. In the present study, involvement of synaptic Zn(2+) in stress-induced attenuation of CA1 LTP was examined in hippocampal slices from young rats after exposure to tail suspension stress for 30s, which significantly increased serum corticosterone. Stress-induced attenuation of CA1 LTP was ameliorated by administration of clioquinol, a membrane permeable zinc chelator, to rats prior to exposure to stress, implying that the reduction of synaptic Zn(2+) by clioquinol participates in this amelioration. To pursue the involvement of corticosterone-mediated Zn(2+) signal in the attenuated CA1 LTP by stress, dynamics of synaptic Zn(2+) was checked in hippocampal slices exposed to corticosterone. Corticosterone increased extracellular Zn(2+) levels measured with ZnAF-2 dose-dependently, as well as the intracellular Ca(2+) levels measured with calcium orange AM, suggesting that corticosterone excites zincergic neurons in the hippocampus and increases Zn(2+) release from the neuron terminals. Intracellular Zn(2+) levels measured with ZnAF-2DA were also increased dose-dependently, but not in the coexistence of CaEDTA, a membrane-impermeable zinc chelator, suggesting that intracellular Zn(2+) levels is increased by the influx of extracellular Zn(2+). Furthermore, corticosterone-induced attenuation of CA1 LTP was abolished in the coexistence of CaEDTA. The present study suggests that corticosterone-mediated increase in postsynaptic Zn(2+) signal in the cytosolic compartment is involved in the attenuation of CA1 LTP after exposure to acute stress.

Preventive effect of theanine intake on stress-induced impairments of hippocamapal long-term potentiation and recognition memory.

Theanine, γ-glutamylethylamide, is one of the major amino acid components in green tea. On the basis of the preventive effect of theanine intake after birth on mild stress-induced attenuation of hippocamapal CA1 long-term potentiation (LTP), the present study evaluated the effect of theanine intake after weaning on stress-induced impairments of LTP and recognition memory. Young rats were fed water containing 0.3% theanine for 3 weeks after weaning and subjected to water immersion stress for 30min, which was more severe than tail suspension stress for 30s used previously. Serum corticosterone levels were lower in theanine-administered rats than in the control rats even after exposure to stress. CA1 LTP induced by a 100-Hz tetanus for 1s was inhibited in the presence of 2-amino-5-phosphonovalerate (APV), an N-methyl-d-aspartate (NMDA) receptor antagonist, in hippocampal slices from the control rats and was attenuated by water immersion stress. In contrast, CA1 LTP was not significantly inhibited in the presence of APV in hippocampal slices from theanine-administered rats and was not attenuated by the stress. Furthermore, object recognition memory was impaired in the control rats, but not in theanine-administered rats. The present study indicates the preventive effect of theanine intake after weaning on stress-induced impairments of hippocampal LTP and recognition memory. It is likely that the modification of corticosterone secretion after theanine intake is involved in the preventive effect.

Advantageous effect of theanine intake on cognition

Abstract Theanine, γ-glutamylethylamide, is one of the major amino acid components in green tea. On the basis of the preventive effect of theanine intake after weaning on stress-induced impairment of recognition memory, the advantageous effect of theanine intake on recognition memory was examined in young rats, which were fed water containing 0.3% theanine for 3 weeks after weaning. The rats were subjected to object recognition test. Object recognition memory was maintained in theanine-administered rats 48 hours after the training, but not in the control rats. When in vivo dentate gyrus long-term potentiation (LTP) was induced, it was more greatly induced in theanine-administered rats than in the control rats. The levels of brain-derived neurotropic factor and nerve growth factor in the hippocampus were significantly higher in theanine-administered rats than in the control rats. The present study indicates the advantageous effect of theanine intake after weaning on recognition memory. It is likely that theanine intake is of advantage to the development of hippocampal function after weaning.

Blockade of intracellular Zn2+ signaling in the basolateral amygdala affects object recognition memory via attenuation of dentate gyrus LTP

&NA; Hippocampus‐dependent memory is modulated by the amygdala. However, it is unknown whether intracellular Zn2+ signaling in the amygdala is involved in hippocampus‐dependent memory. On the basis of the evidence that intracellular Zn2+ signaling in dentate granule cells (DGC) is necessary for object recognition memory via LTP at medial perforant pathway (PP)‐DGC synapses, the present study examined whether intracellular Zn2+ signaling in the amygdala influences object recognition memory via modulation of LTP at medial PP‐DGC synapses. When ZnAF‐2DA (100 &mgr;M, 2 &mgr;l) was injected into the basolateral amygdala (BLA), intracellular ZnAF‐2 locally chelated intracellular Zn2+ in the amygdala. Recognition memory was affected when training of object recognition test was performed 20 min after ZnAF‐2DA injection into the BLA. Twenty minutes after injection of ZnAF‐2DA into the BLA, LTP induction at medial PP‐DGC synapses was attenuated, while LTP induction at PP‐BLA synapses was potentiated and LTP induction at BLA‐DGC synapses was attenuated. These results suggest that intracellular Zn2+ signaling in the BLA is involved in BLA‐associated LTP and modulates LTP at medial PP‐DGC synapses, followed by modulation of object recognition memory.

Adrenergic β receptor activation in the basolateral amygdala, which is intracellular Zn2+-dependent, rescues amyloid β1-42-induced attenuation of dentate gyrus LTP

&NA; On the basis of the evidence that the basolateral amygdala (BLA) modulates hippocampal memory processes via synaptic plasticity, here we report that adrenergic &bgr; receptor activation in the BLA rescues amyloid &bgr;1‐42 (A&bgr;1‐42)‐induced attenuation of long‐term potentiation (LTP) at perforant pathway‐dentate granule cell (DGC) synapses. When 500 &mgr;M isoproterenol (2 &mgr;l), an adrenergic &bgr; receptor agonist, was injected into the BLA 20 min before LTP induction, LTP was enhanced. Isoproterenol‐mediated enhancement of LTP was blocked by co‐injection with 100 &mgr;M ZnAF‐2DA, an intracellular Zn2+ chelator, suggesting that intracellular Zn2+ is required for the intracellular signaling cascade after adrenergic &bgr; receptor activation in the BLA. A&bgr;1‐42‐induced attenuation of LTP, which was induced by A&bgr;1‐42 injection into the dentate gyrus 60 min before LTP induction, was rescued by isoproterenol injection into the BLA 20 min before LTP induction, but not by 500 &mgr;M phenylephrine (2 &mgr;l), an adrenergic &agr;1 receptor agonist, injection into the BLA, which did not enhance LTP unlike the case of isoproterenol injection. Interestingly, A&bgr;1‐42‐induced attenuation of LTP was also rescued by 100 &mgr;M isoproterenol injection into the BLA 20 min before LTP induction, which did not enhance LTP. The present study demonstrates that adrenergic &bgr; receptor activation in the BLA, which is linked with intracellular Zn2+ signaling, rescues A&bgr;1‐42‐induced attenuation of dentate gyrus LTP. It is likely that adrenergic &bgr; receptor activation in the BLA is a strategy for rescuing A&bgr;1‐42‐induced cognitive decline that is associated with hippocampal synaptic plasticity. HighlightsAdrenergic &bgr; receptor activation requires intracellular Zn2+ in the basolateral amygdala (BLA).Adrenergic &bgr; receptor activation in the BLA rescues amyloid &bgr;1‐42‐induced attenuation of dentate gyrus LTP.Adrenergic &bgr; receptor activation in the BLA may be a strategy for rescuing A&bgr;1‐42‐induced cognitive decline.