Effects And Mechanisms Of Aquaporin-4and Asymmetric Dimethylarginine On Synaptic Plasticity And Learning Memory In Limbic System

Part I Effects and mechanisms of aquaporin-4knockout on amygdala long-term potentiation and fear memoryAims:Astrocytes have been generally believed to have a mainly supportive function for neurons. However, increasing evidence suggests that astrocytes also play important roles in information processing, signal transmission and regulation of neural and synaptic plasticity. Aquaporin-4(AQP4) is the major water channel in adult brain and is primarily expressed in astrocytes. A growing body of evidence indicates that AQP4is a potential molecular target for the regulation of astrocytic function. However, little is known about the role of AQP4in synaptic plasticity in the amygdala. Therefore, we evaluated long-term potentiation (LTP) in the lateral amygdala (LA) and associative fear memory of AQP4knock-out (KO) and wild-type (WT) mice.Methods:RT-PCR and Western blotting were used to detect the expression of AQP4mRNA, AQP4protein and glutamate transporter-1(GLT-1) protein in lateral amygdala (LA) region of WT and KO mice, respectively. Field potentials recording and whole-cell recording methods were used to record the field excitatory postsynaptic potentials (fEPSPs) and Excitatory postsynaptic currents (EPSCs) in the thalamo-LA pathway, respectively. Cued fear conditioning test was used to evaluate amygdala-dependent fear memory in WT and KO mice. Open field and elevated plus maze test were employed to detect locomotive activity and anxiety-related behavior of mice, respectively. Two-way repeated-measure (RM)-ANOVA followed by Student’s t test and Student’s t test and/or ANOVA followed by Newman-Keuls post hoc test were used to statistically evaluate the data. Results:AQP4mRNA and AQP4protein were readily detected in LA of WT mice. AQP4deficiency impaired LTP without alter basal synaptic trastission in the thalamo-LA pathway. AQP4KO mice exhibited an impairement of associative fear memory and showed normal locomotive activity and anxiety-related behavior. Furthermore, AQP4deficiency significantly down-regulated GLT-1expression and selectively increased N-methyl-D-aspartate receptors (NMDARs)-mediated EPSCs in the LA. However, low concentration of NMDAR antagonist D-APV reversed the impairment of LTP in KO mice. Up-regulating GLT-1expression by chronic treatment with ceftriaxone (Cef) also reversed the impairment of LTP and fear memory in AQP4KO mice.Conclusion:These findings imply a role for AQP4in synaptic plasticity and associative fear memory in the amygadala by regulating GLT-1expression. Part Ⅱ Effects and mechanisms of asymmetric dimethylarginine on hippocampal long-term potentiation and learning memoryAims:Nitric oxide (NO), a gaseous signal molecule, is synthesized from L-arginine by nitric oxide synthases (NOS). NO plays important roles in the regulation of vascular function, synaptical plasticity and memory. NOS has three isforms:neuronal NOS (nNOS), inducible NOS (iNOS) and endothelial NOS (eNOS). Endogenous NOS inhibitors play an important role in the regulation of NO synthesis. Asymmetric dimethylarginine (ADMA), a endogenous NOS inhibitor, is synthesized in the process of degrading methylated proteins with arginine residues. Humans generate approximately300μM (～60mg) ADMA per day. Large amounts of ADM A are found in rat neurons, suggesting its importance in modulating neuronal activity. ADMA competitively inhibits the three isoforms of NOS, to reduce the NO synthesis. Elevated ADMA levels and decreased cognitive function were found in cerebral ischemia, Alzheimer’s disease (AD), aging, diabetes and other diseases. Howerver, Whether there is a correlation between ADMA and cognitive function is still unclear. This study investigated the effects of ADMA and NO donors on hippocampal long term potentiation (LTP) and learning memory in rats, to provide new ideas for improvement of cognitive founction in patients with cerebrovascular disease and neurodegenerative diseases.Methods:Field excitatory postsynaptic potentials (fEPSPs) were recorded using electrophysiological technique to the evaluate the effects of ADMA, S-nitroso-N-acetylpenicillamine (SNAP) and sodium nitroprusside (SNP) on long-term potentiation (LTP) in rats hippocampal CA1region. Contextual fear conditioning and inhibitory avoidance tasks were performed to evaluate the effects of ADMA and SNAP on hippocampus-dependent memory in rats. Two-way repeated-measure (RM)-ANOVA followed by Student’s t test and Student’s t test and/or ANOVA followed by Newman-Keuls posthoc test were used to statistically evaluate the data.Results:Pretreatment with ADMA50or100μM30min before high-frequency stimulation (HFS) significantly impaired the LTP in rats hippocampal CA1region, while ADMA10μM had no effect on the LTP. Posttreatment with ADMA100μM after HFS also had no effect on the LTP. Interestingly, pretreatment with SNAP100μM or SNP300u.M10min before HFS significantly increased the LTP in normal slices, and reversed the ADMA-induced impairment of LTP. In addition, intrahippocampal injection ADMA5μg and10μg30min before training significantly impaired the acquisition and consolidation of contextual fear memory, ADMA10μg also impaired short-term and long-term memory in inhibitory avoidance tasks, while ADMA1μg had no effect on fear memory. Intrahippocampal injection ADMA10μg after training also had no effect on fear memory and inhibitory avoidance memory. Intrahippocampal injection SNAP10μg10min before training significantly revesed ADMA-induced impairment of memory in contextual fear conditioning and inhibitory avoidance tasks.Conclusion:ADMA impairs the hippocampal long-term potentiation and hippocampus-dependent memory in rats by inhibiting NO synthesis.