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Studies on the Expression of Amino Acid Transporters in the Brain of Pentylenetetrazole Kindled Epileptic Rats and Its Correlation with BDNF

Author: LiangYi
Tutor: LiHongLi
School: Third Military Medical University
Course: Human Anatomy and Embryology
Keywords: Amino acid transporter Brain-derived neurotrophic factor GABA transporter -1 Glutamate transporter -1 Tyrosine kinase receptor B Pentylenetetrazol Astrocytes GAT-1 GLT-1 p-Trk BDNF
CLC: R742.1
Type: Master's thesis
Year: 2007
Downloads: 108
Quote: 0
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Abstract

Objective 1 observation pentylenetetrazol (PTZ) ignite chronic epileptic rat brain amino acid transporter γ-aminobutyric acid transporter -1 (GAT-1), glutamate transporter -1 (GLT-1) expression and brain derived neurotrophic factor (BDNF) signaling pathway Trk protein phosphorylation changed. Two observation BDNF on cultured astrocytes GLT-1 expression and function, combined with the detection of phosphorylated Trk protein, amino acid transporter investigate regulation of BDNF on its role in the process of the significance of chronic epilepsy. Methods and Results 1 to establish a stable model of PTZ kindled rats with chronic epilepsy. SD rats who had been made of grade Ⅲ, is considered fully lit, through improved experimental conditions, 90% success kindled rats. 2 using immunohistochemical techniques, observation of the normal and PTZ ignite chronic epileptic seizures in rats at different time points (7, 14 days) hippocampus and temporal cortex area GAT-1 and GFAP expression changed. Using image analysis software positive reaction in each group the mean optical density value and calculate GAT-1 and glial fibrillary acidic protein (GFAP) in the epilepsy group and the normal group ratio between. The results showed that PTZ ignite chronic epileptic rats GFAP, GAT-1 expression in the normal control group, there were significant differences in the hippocampus and temporal cortex region showed a significant increase, lit 7 days group increased the most significant; which GAT-1 in the epilepsy group and the normal group were significantly higher than the ratio of GFAP. 3 Western blotting was used to detect normal and attack seven days hippocampus and temporal cortex area GAT-1, GFAP, GLT-1 and phospho - tyrosine kinase receptor (p-Trk) expression. The results showed that ignited the hippocampus and temporal cortex area GAT-1, GFAP, GLT-1, p-Trk compared with normal controls were significantly higher existence. Calculating the hippocampus and temporal cortex area of ??the molecules in the epilepsy group and the normal control group absorbance ratio, GAT-1 ratio was significantly greater than the other three molecules. Tip seizures variety of amino acid transporter expression after that there are obvious changes, GAT-1 is increased in glial cells may exist outside source. 4 Immunofluorescence double staining was observed in normal and epileptic seizures in rat brain after 7 days p-Trk and GFAP expression and positional relationship. It was found that kindling group GFAP and p-Trk double positive cells was significantly higher than the normal control group. Tip seizures expressing p-Trk after astrocytes increased the proportion. 5 isolated and cultured and purified astrocytes, identified by GFAP immunostaining of astrocytes ratio greater than 95%. 6 Western blot experiments, measured at different concentrations of BDNF (50,100,200 ng / l) for different time (1,3,7,14 days) were cultured astrocytes GLT-1 and p-Trk expression. BDNF action group astrocytes GLT-1, p-Trk expression compared with the control group, there were significant differences, in which the concentration of BDNF (100ng / l) most significant difference, GLT-1 to 3 days BDNF action most obvious difference between groups , p-Trk BDNF effect seven days to the most obvious differences between groups. Tip BDNF can promote cultured astrocytes glutamate transporter GLT-1 expression increased and may BDNF / p-Trk pathway. 7 Application isotope incorporation experiments, astrocytes in culture systems (BDNF action group and the control group) were added DL-[2,3 - ^{3 H] glutamate, incubate 20 minutes After the determination of astrocytes DL-[2,3 - ^{3 H] glutamate. The results showed that the role of BDNF astrocytes DL-[2,3 - ^{3 H] glutamate was significantly higher than the normal control group, in which BDNF concentration (100ng / l) increased the most obvious. Tip BDNF enhances glutamate uptake of astrocytes. 8 Application Ca ^{2 fluorescence imaging techniques, confocal laser scanning microscopy to detect the role of BDNF cultured astrocytes Ca ^{2 real-time dynamic changes. The results showed that BDNF stimulates astrocyte Ca ^{2 concentrations were significantly increased, and showed cell heterogeneity. Tip BDNF astrocytes can be widely involved in functional activities. Conclusion The results of this study showed that rats kindled brain, especially the hippocampus and temporal cortex neurons exist amino acid transporter GAT-1 and GLT-1 upregulation and BDNF signal pathway enhanced Trk protein phosphorylation, while BDNF can significantly enhance cultured astrocytes GLT-1 expression and function, and possibly via Ca ^{2 TrkB signaling pathways involved. Tip BDNF astrocytes can regulate the transport and uptake of amino acids and other sectors involved in neural plasticity processes; PENTYLENETETRAZOL lit in the course of chronic epilepsy upregulation of BDNF may antagonize excitatory neurotransmitter and restore the role of change .}}}}}}}

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