Our results indicated that long term treatment of manual acupuncture, 2 Hz and 100 Hz at GV20 and GV16 can reduce mossy fiber sprouting and also can reduce astrogliosis 6 weeks after KA administration in SD rats, suggesting manual acupuncture, 2 Hz and 100 Hz may prevent the development of epileptic focus formation, i.e. anti-epileptogenesis. In addition, 2 Hz EA may reduce neuron loss of hippocampus, whether or not 2 Hz may produce a greater effect in protection of neuron, further study is needed.
SD Rat 250-350 g
The epileptic seizures were induced by intra-peritoneal administration of kainic acid in the Spragrue-Dawley Rats. The epileptic seizure rats were divided randomly into control group (Control) without acupuncture treatment, MA group (MA) with manual acupuncture treatment, EA2 group (EA2) with 2 Hz EA treatment, EA100 group (EA100) with 100 Hz EA treatment, whereas normal group (Normal) with phosphate buffer saline (PBS) treatment only (without kainic acid treatment). Acupuncture or electroacupuncture treatment (three times per week) begins one week after KA administration, and the rats were sacrificed at 6 weeks after kainic acid administration.
Figure 2. The effect of acupuncture on mossy fiber sprouting.
The optical density difference of Timm’s staining in hippocampus increased 6 weeks after kainic acid administration (Control) in the Spragrue-Dawley rats, these increase was reduced by manual acupuncture (MA), 2 Hz
Normal
Control
MA
EA2
EA100
IML
GL
Hilus
H
CA3c
Figure 3. The effect of acupuncture on the mossy fiber sprouting in the kainic acid treated rats.
The optic density difference increased 6 weeks after kainic acid administration. These increases were reduced by manual cupuncture, 2 Hz electroacupuncture and 100 Hz electroacupuncture treatments. Normal:
normal group; Control: control group; MA: manual acupuncture group; EA2:
2 Hz electroacupuncture group; EA100: 100 Hz electroacupuncture group.
***p<0.001 compared with Normal; ###p<0.001 compared with Control.
Figure 4. The effect of acupuncture on the NeuN immunoreactive cells in
the kainic acid treated rats.The NeuN immunoreactive cells reduced in the CA3c region of hippocampus 6 weeks after kainic acid administration. Manual cupuncture, 2
CA3c
CA3c
Normal
Control
MA
EA2
EA100
40X 100X 400X
Figure 5. The effect of acupuncture on the NeuN immunoreactive cells in
the kainic acid treated rats.The NeuN immunoreactive cells reduced in the CA3c region of hippocampus 6 weeks after kainic acid administration. Manual cupuncture, 2 Hz electroacupuncture and 100 Hz electroacupuncture treatment could not increased NeuN immunoreactive cells. Normal: normal group; Control:
control group; MA: manual acupuncture group; EA2: 2 Hz electroacupuncture group; EA100: 100 Hz electroacupuncture group. Arrow:
NeuN immunoreactive cells. ***p<0.001 compared with Normal.
Figure 6. The effect of acupuncture on the NeuN immunoreactive cells in
the kainic acid treated rats.The NeuN immunoreactive cells reduced in the CA1 region of hippocampus 6 weeks after kainic acid administration. 2 Hz electroacupuncture and 100
CA1
CA3
CA2 CA1
Normal
Control
MA
EA2
EA100
40X 100X 400X
Figure 7. The effect of acupuncture on the NeuN immunoreactive cells in
the kainic acid treated rats.The NeuN immunoreactive cells reduced in the CA1 region of hippocampus 6 weeks after kainic acid administration. 2 Hz electroacupuncture and 100 Hz electroacupuncture treatment increased NeuN immunoreactive cells.
Normal: normal group; Control: control group; MA: manual acupuncture group; EA2: 2 Hz electroacupuncture group; EA100: 100 Hz electroacupuncture group. ***p < 0.001 compared with Normal; ###p <
0.001 compared with Control.
J J
＊
＊ ＊
SO SR
Normal
Control
MA
EA2
EA100
40X 100X 400X
acupuncture group; EA2: 2 Hz electroacupuncture group; EA100: 100 Hz electroacupuncture group. Arrow: GFAP immunoreactive cells; CA1 region of hippocampus; SO: striatum oriens region of hippocampus; SR:stratum radiate region of hippocampus; arrow: GFAP immunoreactive cells, less reactive; Arrow head: GFAP immunoreactive, reactive astrocytes; Asterisk:
area with abundant GFAP immunoreactive cells. 40X; microscopic field 40 amplifiers;100X:microscopic field 100 amplifiers; 400X microscopic field 400 amplifiers.
Figure 9. The effect of acupuncture on the glial fibrillary acid protein
(GFAP) immunoreactive cells in the SO and SR in the kainic acid treated rats.The GFAP immunoreactive cells increased hippocampus 6 weeks after
CA1
SO
SR
＊＊＊
Normal
Control
MA
EA2
EA100
40X 100X 400X
radiate region of hippocampus; arrow: GFAP immunoreactive cells, less reactive; Arrow head: GFAP immunoreactive, reactive astrocytes; Asterisk:
area with abundant GFAP immunoreactive cells. 40X; microscopic field 40 amplifiers;100X:microscopic field 100 amplifiers; 400X microscopic field 400 amplifiers.
Figure 10. The effect of acupuncture on the glial fibrillary acid protein
(GFAP) immunoreactive cells in the kainic acid treated rats.The GFAP immunoreactive cells increased hippocampus 6 weeks after kainic acid administration. Manual acupuncture; 2 Hz electroacupuncture and 100 Hz electroacupuncture treatment reduced GFAP immunoreactive cells. Normal: normal group; Control: control group; MA: manual acupuncture group; EA2: 2 Hz electroacupuncture group; EA100: 100 Hz electroacupuncture group. Hilus: hilus region of hippocampus; SO: striatum oriens region of hippocampus; SR:stratum radiate region of hippocampus.
***p < 0.001 compared with Normal; #p < 0.05, ##p < 0.01, ###p < 0.001 compared with Control
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Abstract
Acupuncture has been used for epilepsy treatment for a long period of time in China. However, to our knowledge, the efficacy of acupuncture for preventing the development of epileptic focus formation is not yet elucidated until now. Therefore, the purpose of the present study was to investigate the effect of acupuncture on the development of epileptic focus formation. We establish a chronic epileptic model by using intraperitoneal administration of kainic acid (12 mg/kg, KA) in Spragrue-Dawley (SD) rats for 6 weeks. The rats were randomly divided into groups in which they receive the manual acupuncture (MA), 2 Hz electroacupuncture (EA), 100 Hz EA on Baihui acupoint (GV20) and Fengfu acupoint (GV16), respectively, one week after KA administration. The rats were sacrificed 6 weeks after KA administration and the brain tissue was removed for section. The Timm’s staining was done for mossy fiber sprouting, NeuN staining for neuron, and glial fibrillary acidic protein (GFAP) staining for astrogliosis. The results indicated that MA, 2 Hz EA and 100 Hz EA reduced optical density difference of Timm’s
hippocamous, and 2 Hz EA and 100 Hz EA reduced GFAP immunoreactive cells in the striatum oriens and stratum radiate regions of hippocampus.
In conclusion, our results indicated that MA, 2 Hz EA and 100 Hz EA at GV20 and GV16 for 5 weeks can reduce mossy fiber sprouting and also can reduce astrogliosis 6 weeks after KA administration in SD rats,
suggesting that MA, 2 Hz EA and 100 Hz EA may prevent the development of epileptic focus formation, i.e. anti-epileptogenesis. In addition, 2 Hz EA may reduce neuron loss of hippocampus, whether or not 2 Hz may produce a greater effect in protection of neuron, further study is needed.
Key words: Manual acupuncture; Electroacupunctur; Kainic acid;
Mossy fiber sprouting; Astrogliosis
謝辭
兩年的實驗室生涯，總覺得有學不完的東西，總是在重重失敗當中，尋 求那美麗的成功的瞬間。這麼多的風風雨雨，讓兩年的時間，有著厚重 的質感。很幸運的，實驗室的前輩們，從不猶豫在我最需要的時候拉我 一把。剛進實驗室時一無所知，蒲曉韻學姊及方姿琪學姊一步一步的帶 著我練習、操作，並且在實驗失敗的時候，比我還擔憂的急著尋找原因，
我可以說這篇論文的一半以上，有著他們的身影。博士班的程錦宜及蘇 珊玉醫師，給了我很多實驗設計及論文撰寫的指導，並且在假日週末颱 風天時，毫不意外的出現在實驗室一起奮鬥。杜寧漪學姊，李怡欣同學 把我當作自己的夥伴，不吝惜提供儀器，熱心的幫助我解決許多的疑難 雜症。英文超強的潘玉苹同學，在農曆年放大假的時候，默默的帶著我 的論文回家修改，提供我許多寶貴的建議，並且幫我紀錄了許多口試委 員的意見。說到口試，就要感謝帶給實驗室超多活力的蔡青芳醫師，幫
我打點大大小小的雜事，真的讓我完全無後顧之憂的準備。精通Word
各種功能的古欣平小姐，在最後上傳論文的關頭，讓我的困難豁然而 解。當然，更要感謝謝慶良老師及林景彬老師的諄諄教誨及鼓勵，讓我