TSA Increases The Excitatory Synaptic Transmission And Its Mechanism

In biology, the term Epigenetics refers to changes in gene expression that are stable between cell divisions, and sometimes between generations, but do not involve changes in the underlying DNA sequence of the organism. Acetylation is one of the most important kinds of Epigenetics. A large number of research about histone acetylation modification shows that the modification involved in the irreversibility of gene transcription and regulation of the chromosome activity,which were regarded as the important mechanism of many diseases, t such as many tumors, chronic obstructive pulmonary diseases and inflammatory autoimmune diseases.Protease enzyme histone acetylase and the histone deacetylase, the mutual antagonisms, respectively play the role of histone acetylation and deacetylation and commonly control the function of histone acetylation . The histone with positive charge alkaline amino acids, posseses high affinity with DNA. Therefore, the DNA and the histone closely bind together, thus impeding the basic units of protein transcription complexes to enter the promoter binding site, leading to the inhibition of transcriptional function. N-terminal of histone protein outstretches from the nuclear core body, its specific location lysine can be acetylated and neutralizes its positive charge, which weaken electrostatic attraction between alkaline amino acids of the nucleosome and DNA, reduce the aggregation of adjacent nuclear bodys, increase access of transcription factors to DNA, thereby promoting gene transcription. In normal cells, the enzyme histone acetylation and deacetylation enzymes are in dynamic equilibrium, and precisely control chromatin structure and gene transcription. But not all HDACs only role in the histone protein , HDACs also role some non-histone substrates. A series of transcription factors, such as p53, GATA-1 and GATA-2, GATA-3, EKLF, UBF (HMG box architectural actor), AML1 and sex hormone nuclear hormone receptors, can be regulated by acetylation. In addition, recent studies also found that acetylation affected physiological functions, such as cell movement, protein transportation and apoptosis , by regulated plasma protein ,such as tubulin, the wrong folding protein and K70.As early as the 1980s, people began to explore the role of acetyl modification on the ion channels.P protein ( a cation selective channel protein in the Pseudomonas aeruginosa outer membrane), PhoE channel in the outer membrane of Escherichia coli Bacillus can be acetylated,resulting the change of their ion permeability characteristics. At present, there are a lot of literature showing that acetylation can affect learning and memory by regulation of gene transcription. And hippocampal synaptic transmission and plasticity is recognized as the learning and memory mechanisms. But so far we don’t know whether acetylation can affect neuronal synaptic transmission through regulation of the ion channel function. This study used whole-cell patch-clamp technique to record PSC, EPSC and mEPSC in primary cultured rat hippocampal neurons to study the impact of acetylation on synaptic transmission.The cells we chose in the experiment were primarily cultured for 8-10 days, and werehealthy typical hippocampal pyramidal neurons. Accordance to the amplitude,the PSC could be divided into two categories for analysis: one category is less than 100 pA (PSC_S), the other is greater than 100 pA (PSC_L). The experimental results showed that, without TSA in external solution, the frequency of PSC_S (n=10) has the significant run-down effect after 10min. It was 2.953±0.809Hz at 0min, and dropped to 1.936±0.556Hz at 10min.However, the frequency of the PSC increased significantly after adding TSA.The frequency of PSC_S was 0.437±0.128Hz (n = 15) before adding TSA. 10-30 minutes after adding TSA,the frequency increased significantly. it was 0.791±0.212Hz after 10 minutes, rose to 1.274±0.334Hz after 20 minutes, and become 2.412±0.957 Hz at 30 minutes. Compared to that before adding TSA, the frequencie of PSC_L did not increase significantly 10-20 minutes after the drug, but after 30 minutes it increased significantly (n = 14).The amplitude of both PSC_S (n= 14) and PSC_L(n= 5)didn’t change significantly. Thus, TSA significantly increased the frequency of the PSC_S and PSC_L, but had no effect on their amplitude. Therefore TSA can increase synaptic transmission of cultured hippocampal pyramidal neurons.EPSC was divided into two categories, EPSC_S and EPSC_L Adding GABA_A receptor antagonist bicuculline to the external solution to record excitatory ion channel-mediated currents. Compared to control recorded before TSA, the frequency of EPSC_S significantly increased to 404% 20 minutes after TSA (n=13). At the same time, the frequency of EPSC_L significantly increased 3.4-fold and 7-fold after 10 minutes sand 20 minutes(n=15). The amplitude of EPSC_S(n=13) and EPSC_L(n=9) didn’t change.This suggests that acetylation can enhance the frequency of EPSC, and then leaded to the increase of synaptic transmission.Adding GABAA receptor antagonist bicuculline and excitatory ion channel blockers TTX to the external solution to record mEPSC. Experimental results showed that the frequency(n=6) and amplitude(n=7) of mEPSC had no significant difference between before and after adding TSA. The enhancement of synaptic transmission was almost completely blocked, which proved that the enhancement of excitatory synaptic transmission caused by TSA is not due to the change of synapse of its own, such as increasing of presynaptic neurotransmitter release and postsynaptic glutamate receptors , but because of the increase of excitability.In summary, our results show that acetylation increased the excitatory synaptic transmission through the enhancement of excitability in the hippocampal pyramidal neurons.