||Effects of exercise training on the learning and memory behavior and the expression of IAP in the mouse hippocampus
||Department of Physiology
passive avoidance learning task
許多流行病學的研究指出，規律的運動可以改善認知功能。而動物實驗的研究中也指出運動可以增加許多不同種類的神經滋養因子，以及增進學習記憶行為的表現。而在大鼠腦中的Integrin-associated protein (IAP) mRNA 的表現已被發現與記憶的形成相關。Nuclear respiratory factor 1 (NRF-1) 是IAP的轉錄因子，目前已被證實至少能藉由促進調節下游IAP基因進而增加神經分支的長度。但是IAP以及其上游轉錄因子NRF-1是否涉及運動增進學習與記憶的能力仍然未知。因此，我們使用雄性十二週齡的BALB/c 小鼠來探討這個問題。小鼠被隨機分到運動組以及對照組。運動組的動物接受兩週或四週的長期跑步機運動訓練，並且在最後一次運動訓練後不同的時間點犧牲。IAP 的蛋白質表現和在腦區中分布情形將分別利用西方點漬法(Western blotting)以及免疫組織螢光染色法來驗證。運動是否能促進NRF-1調控 IAP 基因表現的活性則是利用電泳遷移率實驗(electrophoresis mobility shift assay : EMSA)來證實。實驗結果顯示經過長期運動訓練後，1) 單次抑制躲避學習測試(one-trial inhibitory avoidance learning task)的行為表現獲得改善；2) 在最後一次運動後海馬迴中 IAP 的蛋白質表現上升；3) NRF-1 與 IAP 啟動子的結合活性在四週運動後零小時與一小時和對照組相比沒有差異。因此我們推論長期運動能改善恐懼學習記憶能力，同時能刺激 IAP 的表現上升，但不是透過 NRF-1的調控。
Results from epidemiological studies suggest that regular exercise improves cognitive function. Animal studies have shown that exercise increases levels of various neurotrophic factors and enhances learning and memory behavioral performance. Integrin-associated protein (IAP) expression is associated with memory formation in rats. Nuclear respiratory factor 1 (NRF-1), a transcription factor for IAP, can increase neurite outgrowth, which is mediated, at least in part, by upregulating its downstream IAP gene. Whether NRF-1 and IAP are involved in exercise-facilitated learning and memory capacity is unclear. Therefore, we used male BALB/c (3-month-old) mice as an animal model to answer this question. Mice were randomly divided into exercise and control groups. Animals in the exercise group received 4 weeks of treadmill exercise training and sacrificed at different time course after the last run. IAP protein expression and distribution were determined by Western blotting and immunohistochemistry, respectively. Whether exercise activates NRF-1 to upregulate IAP gene expression was clarified by electrophoresis mobility shift assay (EMSA). Our results demonstrated that after chronic exercise training, 1) one-trial inhibitory avoidance learning performance was improved; 2) the hippocampal IAP protein level was increased after last run of chronic exercise; and 3) no significant difference in NRF-1 binding activity was found after last run 0h and 1h of 4-wk exercise training. These results suggest that chronic exercise training could improve learning and memory in PA and enhance IAP expression in hippocampus, but not through activate NRF-1.
第一章 緒論 4
第二章 實驗材料與方法 10
第三章 實驗結果 25
第四章 討論 31
第五章 結論 43
Arami S, Jucker M, Schachner M, and Welzl H. (1996) The effect of continuous intraventricular infusion of L1 and NCAM antibodies on spatial learning in rats. Behav Brain Res 81: 81-87.
Arber S and Caroni P. (1995) Thrombospondin-4, an extracellular matrix protein expressed in the developing and adult nervous system promotes neurite outgrowth. J Cell Biol 131: 1083-1094.
Brown E, Hopper L, Ho T, and Gresham H. (1990) Integrin-associated protein: a 50-kD plasma membrane antigen physically and functionally associated with integrins. J Cell Biol 111: 2785-2794.
Brown EJ and Frazier WA. (2001) Integrin-associated protein (CD47) and its ligands. Trends Cell Biol 11: 130-135.
Chang HP, Lindberg FP, Wang HL, Huang AM, and Lee EHY. (1999) Impaired memory retention and decreased long-term potentiation in integrin-associated protein-deficient mice. Learn Mem 6: 448-457.
Chang HP, Ma YL, Wan FJ, Tsai LY, Lindberg FP, and Lee EHY. (2001) Functional blocking of integrin-associated protein impairs memory retention and decreases glutamate release from the hippocampus. Neuroscience 102: 289-296.
Chang WT, Chen HI, Chiou RJ, Chen CY, and Huang AM. (2005) A novel function of transcription factor α-Pal/NRF-1:Increasing neurite outgrowth. Biochem Biophys Res Commun 334: 199-206.
Chang WT and Huang AM. (2004) α-Pal/NRF-1 regulates the promoter of the human integrin-associated protein/CD47 gene. J Biol Chem 279: 14542-14550.
Clark EA and Brugge JS. (1995) Integrins and signal transduction pathways: the road taken. Science 268: 233-239.
Cotman CW and Berchtold NC. (2002) Exercise: a behavioral intervention to enhance brain health and plasticity. Trends Neurosci 25: 295-301.
Davis HP and Squire LR. (1984) Protein synthesis and memory: a review. Psychol Bull 96: 518-559.
DeFreitas MF, Yoshida CK, Frezler WA, Mendrick DL, Kypta RM, and Reichardt LF. (1995) Identification of Integrin as a Neuronal Thrombospondin Receptor Mediating Neurite Outgrowth. Neuron 15: 333-343.
Doyle E, Nolan PM, Bell R, and Regan CM. (1992) Intraventricular infusions of anti-neural cell adhesion molecules in a discrete posttraining period impair consolidation of a passive avoidance response in the rat. J Neurochem 59: 1570-1573.
Eadie BD, Redila VA, and Christie BR. (2005) Voluntary exercise alters the cytoarchitecture of the adult dentate gyrus by increasing cellular proliferation, dendritic complexity, and spine density. J Comp Neurol 486: 39-47.
Evans MJ and Scarpulla RC. (1989) Interaction of nuclear factors with multiple sites in the somatic cytochrome c promoter. Characterization of upstream NRF-1, ATF, and intron Sp1 recognition sequences. J Biol Chem 264: 14361-14368.
Farmer J, Zhao X, van Praag H, Wodtke K, Gage FH, and Christie BR. (2004) Effects of voluntary exercise on synaptic plasticity and gene expression in the dentate gyrus of adult male sprague-dawley rats in vivo. Neuroscience 124: 71-79.
Fujimoto T-T, Katsutani S, Shimomura T, and Fujimura K. (2003) Thrombospondin-bound integrin-associated protein (CD47) physically and functionally modifies integrin αIIbβ3 by its extracellular domain. J Biol Chem 278: 26655-26665.
Gao AG, Lindberg FP, Dimitry JM, Brown EJ, and Frazier WA. (1996a) Thrombospondin modulates αVβ3 function through integrin-associated protein. J Cell Biol 135: 533-544.
Gao AG, Lindberg FP, Finn MB, Blystone SD, Brown EJ, and Frazier WA. (1996b) Integrin-associated protein is a receptor for the C-terminal domain of thrombospondin. J Biol Chem 271: 21-24.
Goelet P, Castellucci VF, Schacher S, and Kandel ER. (1986) The long and the short of long-term memory--a molecular framework. Nature 322: 419-422.
Gómez-Cuadrado A, Martín M, Noël M, and Ruiz-Carrillo A. (1995) Initiation binding repressor, a factor that binds to the transcription initiation site of the histone h5 gene, is a glycosylated member of a family of cell growth regulators. Mol Cell Biol 15: 6670-6685.
Gómez-Pinilla F, So V, and Kesslak JP. (2001) Spatial learning induces neurotrophin receptor and synapsin I in the hippocampus. Brain Res 904: 13-19.
Huang AM, Jen CJ, Chen HF, Yu L, Kuo YM, and Chen HI. (2006) Compulsive exercise acutely upregulates rat hippocampal brain-derived neurotrophic factor. J Neural Transm 113: 803-811.
Huang AM, Wang HL, Tang YP, and Lee EHY. (1998) Expression of integrin-associated protein gene associated with memory formation in rats. J Neurosci 18: 4305-4313.
Jacob WF, Silverman TA, Cohen RB, and Safer B. (1989) Identification and characterization of a novel transcription factor participating in the expression of eukaryotic initiation factor 2α. J Biol Chem 264: 20372-20384.
Jiang P, Lagenaur CF, and Narayanan V. (1999) Integrin-associated protein is a ligand for the P84 neural adhesion molecule. J Biol Chem 274: 559-562.
Jones LS. (1996) Integrins: possible functions in the adult CNS. Trends Neurosci 19: 68-72.
Juliano RL and Haskill S. (1993) Signal transduction from the extracellular matrix. J Cell Biol 120: 577-585.
Kelly DP and Scarpulla RC. (2004) Transcriptional regulatory circuits controlling mitochondrial biogenesis and function. Genes Dev 18: 357-368.
Lee EHY, Hsieh YP, Yang CL, Tsai KJ, and Liu CH. (2000) Induction of integrin-associated protein (IAP) mRNA expression during memory consolidation in rat hippocampus. Eur J Neurosci 12: 1105-1112.
Lindberg FP, Gresham HD, Schwarz E, and Brown EJ. (1993) Molecular cloning of integrin-associated protein: an immunoglobulin family member with multiple membrane-spanning domains implicated in αVβ3-dependent ligand binding. J Cell Biol 123: 485-496.
Mi ZP, Jiang P, Weng WL, Lindberg FP, Narayanan V, and Lagenaur CF. (2000) Expression of a Synapse-Associated Membrane Protein, P84/SHPS-1, and Its Ligand, IAP/CD47, in Mouse Retina. J Comp Neurol 416: 335-344
Miyashita M, Ohnishi H, Okazawa H, Tomonaga H, Hayashi A, Fujimoto T-T, Furuya N, and Matozaki T. (2004) Promotion of neurite and filopodium formation by CD47: roles of integrins, Rac, and Cdc42. Mol Biol Cell 15: 3950-3963.
Murata T, Ohnishi H, Okazawa H, Murata Y, Kusakari S, Hayashi Y, Miyashita M, Itoh H, Oldenborg P-A, Furuya N, and Matozaki T. (2006) CD47 promotes neuronal development through Src- and FRG/Vav2-mediated activation of Rac and Cdc42. J Neurosci 26: 12397-12407.
Myers SJ, Peters J, Huang Y, Comer MB, Barthel F, and Dingledine R. (1998) Transcriptional regulation of the GluR2 gene: neural-specific expression, multiple promoters, and regulatory elements. J Neurosci 18: 6723-6739.
Neeper SA, Gdmez-Pinilla F, Choi J, and Cotman C. (1995) Exercise and brain neurotrophins. Nature 373: 109.
Neeper SA, Gómez-Pinilla F, Choi J, and Cotman CW. (1996) Physical activity increases mRNA for brain-derived neurotrophic factor and nerve growth factor in rat brain. Brain Research 726: 49-56.
Numakawa T, Ishimoto T, Suzuki S, Numakawa Y, Adachi N, Matsumoto T, Yokomaku D, Koshimizu H, Fujimori KE, Hashimoto R, Taguchi T, and Kunugi H. (2004) Neuronal Roles of the Integrin-associated Protein (IAP/ CD47) in Developing Cortical Neurons. J Biol Chem 279: 43245-43253.
Ohnishi H, Kaneko Y, Okazawa H, Miyashita M, Sato R, Hayashi A, Tada K, Nagata S, Takahashi M, and Matozaki T. (2005) Differential localization of Src homology 2 domain-containing protein tyrosine phosphatase substrate-1 and CD47 and its molecular mechanisms in cultured hippocampal neurons. J Neurosci 25: 2702-2711.
Pilegaard H, Saltin B, and Neufer PD. (2003) Exercise induces transient transcriptional activation of the PGC-1α gene in human skeletal muscle. J Physiol 546: 851-858
Reichardt LF and Tomaselli KJ. (1991) Extracellular matrix molecules and their receptors: functions in neural development. Annu Rev Neurosci 14: 531-570.
Samorajski T, Delaney C, Durham L, Ordy JM, Johnson JA, and Dunlap WP. (1985) Effect of exercise on longevity, body weight, locomotor performance, and passive-avoidance memory of C57BL/6J mice. Neurobiol Aging 6: 17-24.
Schwartz MA, Brown EJ, and Fazeli B. (1993) A 50-kDa integrin-associated protein is required for integrin-regulated calcium entry in endothelial cells. J Biol Chem 268: 19931-19934.
Silva AJ, Rosahl TW, Chapman PF, Marowitz Z, Friedman E, Frankland PW, Cestari V, Cioffi D, Sudhof TC, and Bourtchuladze R. (1996) Impaired learning in mice with abnormal short-lived plasticity. Curr Biol 6: 1509-1518.
Srere PA. (1969) Citrate synthase. Methods Enzymol 13:3-8
St-Pierre J, Drori S, Uldry M, Silvaggi JM, Rhee J, Jäger S, Handschin C, Zheng K, Lin J, Yang W, Simon DK, Bachoo R, and Spiegelman BM. (2006) Suppression of reactive oxygen species and neurodegeneration by the PGC-1 transcriptional coactivators. Cell 127: 397-408.
Stäubli U, Chun D, and Lynch G. (1998) Time-dependent reversal of long-term potentiation by an integrin antagonist. J Neurosci 18: 3460-3469.
Tamkuna JW, DeSimonea DW, Fondaa D, Patela RS, Buckb C, Horwitzc AF, and Hynesa RO. (1986) Structure of integrin, a glycoprotein involved in the transmembrane linkage between fibronectin and actin. Cell 46: 271-282.
van Praag H, Christie BR, Sejnowski TJ, and Gage FH. (1999) Running enhances neurogenesis, learning, and long-term potentiation in mice. Proc Natl Acad Sci U S A 96: 13427-13431.
Vaynman S, Ying Z, and Gómez-Pinilla F. (2004) Hippocampal BDNF mediates the efficacy of exercise on synaptic plasticity and cognition. Eur J Neurosci 20: 2480-2490.
Virbasius CA, Virbasius JV, and Scarpulla RC. (1993) NRF-1, an activator involved in nuclear-mitochondrial interactions, utilizes a new DNA-binding domain conserved in a family of developmental regulators. Genes Dev 7: 2431-2445.
Wright DC, Han D-H, Garcia-Roves PM, Geiger PC, Jones TE, and Holloszy JO. (2007) Exercise-induced mitochondrial biogenesis begins before the increase in muscle PGC-1α expression. J Biol Chem 282: 194-199.
Wu Z, Puigserver P, Andersson U, Zhang C, Adelmant G, Mootha V, Troy A, Cinti S, Lowell B, Scarpulla RC, and Spiegelman BM. (1999) Mechanisms controlling mitochondrial biogenesis and respiration through the thermogenic coactivator PGC-1. Cell 98: 115-124.