Effects Of Aβ(10-21) On Potassium Currents Of Hippocampal Pyramidal Neurons & The Neuroprotective Function Of Lithium Chloride

Alzheimer's disease (AD), also called senile dementia, is one of themost difficult miscellaneous diseases in progressive old-age society. Due toinsufficient effective therapy and precautionary measures, AD has becomeone of the four killers (cardio-cerebral vascular disease, malignant tumor,apoplexia, AD) that kill human beings at present. Therefore, many developedcountries regard AD as one of the strategy problems which threaten publichealth and retard the development of economy and society, and bring it intostate plan to conquer this disease. Now it is stepping into old-age society inour country. The proportion of the elderly people is continually increasing.People suffering from senile dementia become more and more. This presentsan enormous burden on the families of those affected, as well as society ingeneral. So it is urgent to prevent and cure AD. This is not only a medicalproblem but a social problem which related to the development of the countryand the progress of the society. Thus, we must accelerate the research ofpathogenesis of AD and improve the level of treating and preventing AD. Onthe one hand, it can better the life of the elderly people, improve the healthlevel of the whole nation and prolong human being's average life span; on theother hand, it will have an important influence on conquering the mechanismof study and memory, and then discover the mystery of the brain. Therefore,this subject has potential economic and social benefits in addition to theacademic value.Chapterâ… of the dissertation is the summary and tentative plan forresearch. It introduced the recent development of AD research, mainly fromthe pathogenesis, diagnosis and other aspects. The emphasis is to summarizeand analyze the three hypotheses of Aβtheory. Accordingly, it is proposedthat the basis of the topic, way & what to research, and the associatedmethod.Chapterâ…¡introduced the function, sorts, properties of ion channels ofhippocampal neurons and record key points of patch-clamp. It mainly introduced the isolation technique of outward potassium current,current-voltage character curve, the formation of action potential andactivation, inactivation and reactivation of potassium channel. We choose theoptimum parameters for the experiments from the above. This part is thetheory basis and experiment condition of chapterâ…£andâ…¤.The topic of Chapterâ…¢is spectrum. In this chapter, it has beenresearched that the configuration & aggregation effects of two metal ions(Zn²⁺ and Cu²⁺) on Aβ(10-21) by adopting UV, fluorescence, CD and othertechniques. The study shows that the prepared Aβ(10-21) is unaggregated inthe absence of metal ions. And fluorescence spectrum shows that the additionof Zn²⁺ can make Aβ(10-21) aggregate rapidly in a concentration-dependentfashion. Resonant scattering spectrum also shows the aggregation formed.Circular dichroism clearly indicated the addition of Zn²⁺ can lead to distinctchange of secondary configuration of Aβ(10-21), and makeα-helix changefrom 83.03% to 43.10%;β-sheet and irregular curl, the percentage changefrom 16.97% to 56.9%. Similar to Zn²⁺, the addition of Cu²⁺ also make Aβ(10-21) aggregate from unaggregated state. Meanwhile, it can lead to thechange of secondary configuration of Aβ(10-21), and the percentage ofα-helix obviously reduced;β-sheet and irregular curl, the percentageobviously increased. The difference from Zn²⁺ is that, although resonantscattering spectrum shows the addition of Cu²⁺ also lead to the aggregation ofAβ(10-21), the buffer solution has more effect on the above change. As tohow does Zn²⁺/Cu²⁺ lead to the aggregation of Aβ(10-21), the exactmechanism shall be confirmed by further experiments.The topic of Chapterâ…£is electrophysiology. This part mainlyresearched the effects of Aβ(10-21) in two forms (unaggregated andaggregated) on rat hippocampal CAl pyramidal neurons outward potassiumcurrents by using the whole-cell patch-clamp technique to expound thenosogenesis of Aβfrom channel level. The study shows that unaggregatedAβ(10-21) selectively blocked the fast-activating & fast-inactivating outwardpotassium current (I_A) and made the activation curve and inactivation curve shift to negative potentials and prolong I_A reactivation time constant.Compared to unaggregated Aβ(10-21), the blockage of Aβ(10-21)aggregated by Zn²⁺/Cu²⁺ on outward potassium current I_A obviouslypotentiated. And concentration inhibited rate reached (45.93±0.99)ï¼….Aggregated Aβ(10-21) also shift the activation curve and inactivation curveof I_A to negative potentials, but the shifted amplitude is more than that ofunaggregated AD (10-21). Also the amplitude of aggregated Aβ(10-21)prolonging I_A reactivation time constant is more than that of unaggregated Aβ(10-21). The above results show that Aβ(10-21) has aggregation-dependenton the inhibition of I_A. The aggregation of Aβ(10-21) enhances itsmodulatory effects on I_A in every respects. Potassium channels play crucialroles in regulating membrane potential and excitability. Blockage ofpotassium channels can cause cell depolarization, increase cell excitability,accelerate Ca²⁺ influx and disruption of intracellular ion homeostasis, thusbeginning a cascade of subsequent cellular responses that eventually result inneuronal dysfunction and death. This can be one of the molecularmechanisms of Aβneurotoxicity.Moreover, it has been researched that electrostatic interactions betweenAβ(10-21) and potassium channel. Through electrostatic interactions, Aβ(10-21) binds to open channels and increases their rate of inactivation orbinds to closed channels and retards their rate of reactivation, thus quickeninginactivation and slowing reactivation of potassium current, and decreasingcurrent intensity. This is the main reason of Aβ(10-21) inhibiting potassiumcurrent.Chapterâ…¤is to probe into lithium neuroprotective properties. Lithiumis the traditional anti-manic medicine. Recently it is found that lithium hasneuroprotective functions, especially in reducing the phosphorylation of Tauof AD patients, which implying lithium has potential value to cure AD. Inthis chapter, the effects of lithium chloride on hippocampal pyramidalneurons outward potassium currents were studied by using the whole-cellpatch-clamp technique, which researched the neuroprotective functions of lithium from another aspect. The findings shows that lithium chloride canselectively increased I_A and the half enhancement concentration (EC₅₀) oflithium chloride was (22.8±5.45) umol/L. Lithium chloride shifted theactivation curve and inactivation curve of I_A to negative potentials, mainlyinfluencing the activation kinetics of I_A, and shorten I_A reactivation timeconstant. The advance of activation and the acceleration of reactivationjointly lead to the increase of I_A current, which lead to membranehyperpolarization, canceling or counteracting the depolarization caused byAβ(10-21), thus reducing the damage of cell excitability. But, at present, it isnot clear that how much I_A increases can cancel the depolarization caused byAβ(10-21) because it is very complex to control the channel, which needsconsidering many factors.Chapterâ…¥is the summary and prospect, which summarized what hasbeen researched and then described what to be expected for further research.In summary, in this dissertation, it has been demonstrated that theaggregation & configuration effects of two metal ions (Zn²⁺ and Cu²⁺) on Aβ(10-21) by adopting spectrum technique; it has been researched the effects ofAβ(10-21) in two forms (unaggregated and aggregated) on hippocampalpyramidal neurons outward potassium currents three processes (activation,inactivation and reactivation) by using the whole-cell patch-clamp technique,and expound the molecular mechanism of Aβneurotoxicity from channellevel and the key role of metal ions. It is suggested that Aβ(10-21) can beused as model fragment for the research of Aβaggregation induced by metalions and interaction with cell. It builds a platform for mutual communicationand intersectant research among chemistry, biology and electrophysiology. Inaddition, the dissertation researched the neuroprotective functions of lithiumchloride, and studied and analyzed the potential mechanism lithium chlorideto cure AD in a new aspect. These results are significant for the furtherstudy of Aβnosogenesis and the development of new type target drugs forthe treatment of AD.