The Effects Of Soluble Amyloid β Protein (25-35) On MRNA Expression Of Delayed Rectifier Potassium Channel In Pyramidal Neurons At Rat Hippocampal CA3

Object:Alzheimer's disease(AD)is a neurodegenerative disorder influencing functions of memory and cognition,and AD is the most common reason of dementia.Some researches indicate that the main pathological characteristic of AD is increased amyloidβ-peptide(Aβ),and the deposition of Aβin the brain has clear neurotoxicity. There are other researches showing that soluble Aβalso has neurotoxicity. Hippocampus is a key structure of learning and memory,which is directly related to the acquisition and foundation of learning and memory.Hippocampal CA3 area correlates to learning memory of spatial discrimination closely.While researches on the molecule and physiology mechanism of learning and memory show that potassium channel,PKC,memory-relating protein Cp20 and regulation of intracellular Ca²⁺play an important role in the mechanism of learning and memory, especially that potassium channel plays a key role,and this suggests that potassium channel has significant effects on AD.Some studies indicate that soluble Aβ_25-35can inhibit I_k of the neurons of hippocampal CA3 area distinctly in newborn rats.The effect is time-dependent and voltage-dependent,but the specific mechanism remains unknown.We investigate the effect of soluble Aβ_25-35on the expression of mRNA of several K⁺ channel of hippocampal CA3 area of newborn rats,using neurons of hippocampal CA3 area and taking the methods of patch clamp and single cell RT-PCR and research the mechanism of I_k change in the point of molecular biology. Our research provided theoretical proof to the mechanism of Aβin the AD disease and the precaution and cure of AD.Methods:Combining the methods of enzymic digestion and mechanical separation,we separated rat hippocampal CA3 pyramidal neurons from the slice of the brain tissue. After that,we use the patch clamp technique to get a single cell,and use the RT-PCR technique to amplify potassium channels' mRNA.Then we can record the effects of soluble Aβ_25-35on the mRNA expression of delayed rectifier potassium channel subtypes Kv1.2,Kv1.5 and Kv2.1.Results:1.In the rat hippocampal CA3 pyramidal neurons,delayed rectifier potassium channel Kv1.2,Kv1.5 and Kv2.1 mRNA are expressed in different levels,and Kv2.1 has a relatively high level expression.2.Compared to the control group,the expression level of kv1.2 mRNA fell 15.9% (P＜0.01),24.0%(P＜0.01)and 35.8%(P＜0.01)after addition of 1.0,2.5 and 5.0μM soluble Aβ_25-35for 1 minute.3.Compared to the control group,the expression level of kv1.5 mRNA fell 17.8% (P＜0.01),23.2%(P＜0.01)and 36.1%(P＜0.01)after addition of 1.0,2.5 and 5.0μM soluble Aβ_25-35for 1 minute.4.Compared to the control group,the expression level of kv2.1 mRNA fell 17.2% (P＜0.01),23.7%(P＜0.01)and 36.9%(P＜0.01)after addition of 1.0,2.5 and 5.0μM soluble Aβ_25-35for 1 minute.5.Compared to the control group,the expression level of kv2.1 mRNA fell 38.7% (P＜0.01),43.5%(P＜0.01),53.3%(P＜0.01),65.6%(P＜0.01),73.7%(P＜0.01) after addition of 5.0μM soluble Aβ_25-35for 1,3,5,8 and 10 minutes.Conclusion:1.In the rat hippocampal CA3 pyramidal neurons,delayed rectifier potassium channel Kv1.2,Kv1.5 and Kv2.1 mRNA are expressed in different levels,and Kv2.1 has a relatively high level expression.2.The expression of Kv1.2,Kv1.5 and Kv2.1 mRNA were obviously inhibited by soluble Aβ_25-35and the inhibition effect was concentration- and time-depended.3.The number of potassium channel reduced because of the block effect of soluble Aβ_25-35on rat hippocampal CA3 pyramidal neurons.And this may be one of the mechanisms how soluble Aβ_25-35can inhibit electric current of potassium.