NIH 그랜트 AG027297, 켄터키 척수 및 머리 부상 연구 신뢰에서 수상하고, 클레 버그 재단의 선물에 의해 지원 작동합니다.

<ol>
	<li>Norris, C. M., Halpain, S., Foster, T. C. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Alterations+in+the+balance+of+protein+kinase/phosphatase+activities+parallel+reduced+synaptic+strength+during+aging.">Alterations in the balance of protein kinase/phosphatase activities parallel reduced synaptic strength during aging.</a> J Neurophysiol. 80, 1567-1570 (1998).</li><li>Norris, C. M., Korol, D. L., Foster, T. C. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Increased+susceptibility+to+induction+of+long-term+depression+and+long-term+potentiation+reversal+during+aging.">Increased susceptibility to induction of long-term depression and long-term potentiation reversal during aging.</a> J Neurosci. 16, 5382-5392 (1996).</li><li>Norris, C. M. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Hippocampal+'zipper'+slice+studies+reveal+a+necessary+role+for+calcineurin+in+the+increased+activity+of+L-type+Ca(2+)+channels+with+aging.">Hippocampal 'zipper' slice studies reveal a necessary role for calcineurin in the increased activity of L-type Ca(2+) channels with aging.</a> Neurobiol Aging. 31, 328-338 (2010).</li><li>Burke, S. N., Barnes, C. A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Senescent+synapses+and+hippocampal+circuit+dynamics.">Senescent synapses and hippocampal circuit dynamics.</a> Trends Neurosci. 33, 153-161 (2010).</li><li>Foster, T. C. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Calcium+homeostasis+and+modulation+of+synaptic+plasticity+in+the+aged+brain.">Calcium homeostasis and modulation of synaptic plasticity in the aged brain.</a> Aging Cell. 6, 319-325 (2007).</li><li>Foster, T. C., Norris, C. M. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Age-associated+changes+in+Ca(2+)-dependent+processes:+relation+to+hippocampal+synaptic+plasticity.">Age-associated changes in Ca(2+)-dependent processes: relation to hippocampal synaptic plasticity.</a> Hippocampus. 7, 602-612 (1997).</li><li>Thibault, O., Gant, J. C., Landfield, P. W. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Expansion+of+the+calcium+hypothesis+of+brain+aging+and+Alzheimer's+disease:+minding+the+store.">Expansion of the calcium hypothesis of brain aging and Alzheimer's disease: minding the store.</a> Aging Cell. 6, 307-317 (2007).</li><li>Demuro, A., Parker, I., Stutzmann, G. E. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Calcium+signaling+and+amyloid+toxicity+in+Alzheimer+disease.">Calcium signaling and amyloid toxicity in Alzheimer disease.</a> J Biol Chem. 285, 12463-12468 (2010).</li><li>Green, K. N., LaFerla, F. M. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Linking+calcium+to+Abeta+and+Alzheimer's+disease.">Linking calcium to Abeta and Alzheimer's disease.</a> Neuron. 59, 190-194 (2008).</li><li>Kumar, A., Thinschmidt, J. S., Foster, T. C., King, M. A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Aging+effects+on+the+limits+and+stability+of+long-term+synaptic+potentiation+and+depression+in+rat+hippocampal+area+CA1.">Aging effects on the limits and stability of long-term synaptic potentiation and depression in rat hippocampal area CA1.</a> J Neurophysiol. 98, 594-601 (2007).</li><li>Norris, C. M., Halpain, S., Foster, T. C. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Reversal+of+age-related+alterations+in+synaptic+plasticity+by+blockade+of+L-type+Ca2++channels.">Reversal of age-related alterations in synaptic plasticity by blockade of L-type Ca2+ channels.</a> J Neurosci. 18, 3171-3179 (1998).</li><li>Norris, C. M., Scheff, S. W. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Recovery+of+afferent+function+and+synaptic+strength+in+hippocampal+CA1+following+traumatic+brain+injury.">Recovery of afferent function and synaptic strength in hippocampal CA1 following traumatic brain injury.</a> J Neurotrauma. 26, 2269-2278 (2009).</li><li>Sama, M. A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Interleukin-1beta-dependent+signaling+between+astrocytes+and+neurons+depends+critically+on+astrocytic+calcineurin/NFAT+activity.">Interleukin-1beta-dependent signaling between astrocytes and neurons depends critically on astrocytic calcineurin/NFAT activity.</a> J Biol Chem. 283, 21953-21964 (2008).</li><li>Ye, H., Jalini, S., Mylvaganam, S., Carlen, P. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Activation+of+large-conductance+Ca(2+)-activated+K(+)+channels+depresses+basal+synaptic+transmission+in+the+hippocampal+CA1+area+in+APP+(swe/ind)+TgCRND8+mice.">Activation of large-conductance Ca(2+)-activated K(+) channels depresses basal synaptic transmission in the hippocampal CA1 area in APP (swe/ind) TgCRND8 mice.</a> Neurobiol Aging. 31, 591-604 (2010).</li><li>Bear, M. F., Malenka, R. C. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Synaptic+plasticity:+LTP+and+LTD.">Synaptic plasticity: LTP and LTD.</a> Curr Opin Neurobiol. 4, 389-399 (1994).</li><li>Foster, T. C. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Involvement+of+hippocampal+synaptic+plasticity+in+age-related+memory+decline.">Involvement of hippocampal synaptic plasticity in age-related memory decline.</a> Brain Res Brain Res Rev. 30, 236-249 (1999).</li><li>Foster, T. C., Kumar, A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Susceptibility+to+induction+of+long-term+depression+is+associated+with+impaired+memory+in+aged+Fischer+344+rats.">Susceptibility to induction of long-term depression is associated with impaired memory in aged Fischer 344 rats.</a> Neurobiol Learn Mem. 87, 522-535 (2007).</li><li>Hsu, K. S. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Alterations+in+the+balance+of+protein+kinase+and+phosphatase+activities+and+age-related+impairments+of+synaptic+transmission+and+long-term+potentiation.">Alterations in the balance of protein kinase and phosphatase activities and age-related impairments of synaptic transmission and long-term potentiation.</a> Hippocampus. 12, 787-802 (2002).</li><li>Vouimba, R. M., Foy, M. R., Foy, J. G., Thompson, R. F. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=17beta-estradiol+suppresses+expression+of+long-term+depression+in+aged+rats.">17beta-estradiol suppresses expression of long-term depression in aged rats.</a> Brain Res Bull. 53, 783-787 (2000).</li><li>Abdul, H. M., Furman, J. L., Sama, M. A., Mathis, D. M., Norris, C. M. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=NFATs+and+Alzheimer's+Disease.">NFATs and Alzheimer's Disease.</a> Mol Cell Pharmacol. 2, 7-14 (2010).</li><li>Abdul, H. M. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Cognitive+decline+in+Alzheimer's+disease+is+associated+with+selective+changes+in+calcineurin/NFAT+signaling.">Cognitive decline in Alzheimer's disease is associated with selective changes in calcineurin/NFAT signaling.</a> J Neurosci. 29, 12957-12969 (2009).</li><li>Anantharaman, M. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Beta-amyloid+mediated+nitration+of+manganese+superoxide+dismutase:+implication+for+oxidative+stress+in+a+APPNLH/NLH+X+PS-1P264L/P264L+double+knock-in+mouse+model+of+Alzheimer's+disease.">Beta-amyloid mediated nitration of manganese superoxide dismutase: implication for oxidative stress in a APPNLH/NLH X PS-1P264L/P264L double knock-in mouse model of Alzheimer's disease.</a> Am J Pathol. 168, 1608-1618 (2006).</li><li>Gengler, S., Hamilton, A., Holscher, C. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Synaptic+plasticity+in+the+hippocampus+of+a+APP/PS1+mouse+model+of+Alzheimer's+disease+is+impaired+in+old+but+not+young+mice.">Synaptic plasticity in the hippocampus of a APP/PS1 mouse model of Alzheimer's disease is impaired in old but not young mice.</a> PLoS One. 5, e9764-e9764 (2010).</li><li>Stringer, J. L., Lothman, E. W. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=In+vitro+effects+of+extracellular+calcium+concentrations+on+hippocampal+pyramidal+cell+responses.">In vitro effects of extracellular calcium concentrations on hippocampal pyramidal cell responses.</a> Exp Neurol. 101, 132-146 (1988).</li><li>Landfield, P. W., Pitler, T. A., Applegate, M. D. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=The+effects+of+high+Mg2+-to-Ca2++ratios+on+frequency+potentiation+in+hippocampal+slices+of+young+and+aged+rats.">The effects of high Mg2+-to-Ca2+ ratios on frequency potentiation in hippocampal slices of young and aged rats.</a> J Neurophysiol. 56, 797-811 (1986).</li><li>Ames, A., Sakanoue, M., Endo, S. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=NA,+K,+CA,+MG,+AND+C1+CONCENTRATIONS+IN+CHOROID+PLEXUS+FLUID+AND+CISTERNAL+FLUID+COMPARED+WITH+PLASMA+ULTRAFILTRATE.">NA, K, CA, MG, AND C1 CONCENTRATIONS IN CHOROID PLEXUS FLUID AND CISTERNAL FLUID COMPARED WITH PLASMA ULTRAFILTRATE.</a> J Neurophysiol. 27, 672-681 (1964).</li><li>Gant, J. C., Sama, M. M., Landfield, P. W., Thibault, O. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Early+and+simultaneous+emergence+of+multiple+hippocampal+biomarkers+of+aging+is+mediated+by+Ca2+-induced+Ca2++release.">Early and simultaneous emergence of multiple hippocampal biomarkers of aging is mediated by Ca2+-induced Ca2+ release.</a> J Neurosci. 26, 3482-3490 (2006).</li><li>Thibault, O., Hadley, R., Landfield, P. W. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Elevated+postsynaptic+[Ca2+]i+and+L-type+calcium+channel+activity+in+aged+hippocampal+neurons:+relationship+to+impaired+synaptic+plasticity.">Elevated postsynaptic [Ca2+]i and L-type calcium channel activity in aged hippocampal neurons: relationship to impaired synaptic plasticity.</a> J Neurosci. 21, 9744-9756 (2001).</li><li>Thibault, O., Landfield, P. W. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Increase+in+single+L-type+calcium+channels+in+hippocampal+neurons+during+aging.">Increase in single L-type calcium channels in hippocampal neurons during aging.</a> Science. 272, 1017-1020 (1996).</li><li>Stutzmann, G. E., Caccamo, A., LaFerla, F. M., Parker, I. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Dysregulated+IP3+signaling+in+cortical+neurons+of+knock-in+mice+expressing+an+Alzheimer's-linked+mutation+in+presenilin1+results+in+exaggerated+Ca2++signals+and+altered+membrane+excitability.">Dysregulated IP3 signaling in cortical neurons of knock-in mice expressing an Alzheimer's-linked mutation in presenilin1 results in exaggerated Ca2+ signals and altered membrane excitability.</a> J Neurosci. 24, 508-513 (2004).</li><li>Stutzmann, G. E. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Enhanced+ryanodine+receptor+recruitment+contributes+to+Ca2++disruptions+in+young,+adult,+and+aged+Alzheimer's+disease+mice.">Enhanced ryanodine receptor recruitment contributes to Ca2+ disruptions in young, adult, and aged Alzheimer's disease mice.</a> J Neurosci. 26, 5180-5189 (2006).</li><li>Lee, H. K., Min, S. S., Gallagher, M., Kirkwood, A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=NMDA+receptor-independent+long-term+depression+correlates+with+successful+aging+in+rats.">NMDA receptor-independent long-term depression correlates with successful aging in rats.</a> Nat Neurosci. 8, 1657-1659 (2005).</li><li>McNaughton, B. L., Douglas, R. M., Goddard, G. V. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Synaptic+enhancement+in+fascia+dentata:+cooperativity+among+coactive+afferents.">Synaptic enhancement in fascia dentata: cooperativity among coactive afferents.</a> Brain Res. 157, 277-293 (1978).</li></ol>

이 비디오 문서의 생산은 Leica Microsystems에 의해 후원되었다.

이 프로토콜에 제시된 단계는 뇌의 해부가 젊은 성인 쥐처럼, ​​노인 최소한으로 신속하게 효율적으로 수행하는 보험에 가입하는 데 도움이됩니다. 우리는 또한 LTP와 LTD에 자신의 슬라이스 연구를 설정하는 초보자를위한 충분한 세부 사항을 제공합니다. 노화와 신경 기능과 소성의 광고 변경 추가 탐사가 귀하의 목표 중 하나가있다면, 더 배려를받을 자격이 상기에 언급한 건 요즘 적어도 두 개의 다른 방법 론적 문제가있다. 첫째, 여러 실험실 보여주 그 칼슘 2 + : Mg2 ACSF가 hippocampal 조각 2,10,24,25의 유도 시냅스 소성에 표시된 영향을 미칠 수있는 기록에 + 비율. 포유류의 CSF에서 CA 2 + : Mg2 + 비율이 약 (예 : 26를 참조)입니다. 그러나, ACSF 칼슘 2 + : Mg2 + 가까이 2 비율이 일반적으로 시냅스 기능과 소성의 슬라이스 연구에 사용됩니다. 초기 연구에서는 이러한 행위는 아마 LTP의 유도를 최적화하기 위해 적응되었다, 그때 이후 모든 소성 연구를위한 루틴이되었습니다. 그러나, 이러한 행위 때문에 칼슘의 연결 2 + 규제에 잘 특징 차이 고령화 및 광고 연구에 문제가있을 수 있습니다. 특히, CA 2 + 유입 및 / 또는 칼슘 2 + 유도된 칼슘 2 + - 릴리스의 연결을 활성화 3,27-31 동안 세 쥐 및 / 또는 광고 모델 마우스에서 고가입니다. LTD의 유도는 ACSF 칼슘 2 + 수준의 미묘한 변화에 특히 민감합니다. Mg2 가까이 두 + 비율이 성인에 강력한 회사를 감찰하러 온 적도에서 : 2mM 칼슘 2 + 2 MM Mg2 +, 노인,하지만이 성인 동물 젊은 없습니다 LTD에서 일반적으로 결과 중에 칼슘 2를 사용하는 연구 +를 사용하여 우리의 프로토콜, 32 세 쥐에서 감소 회사와 함께 나이 차이 2,10 또는 부재. 이러한 관찰은 노인과 젊은 성인 동물 + 종속 소성 칼슘 2 비교하면 조심스럽게 ACSF 칼슘 2 + 및 Mg2 + 수준을 고려하는 필요성을 강조 표시합니다. 두 번째 방법 론적 문제는 postsynaptic 탈분극 33 시냅스 강도 가능 노화 / 유전자형의 차이에 LTP의 강한 의존성에 관한. 전형적인 LTP 실험, 기본 및 LTP의 자극 강도는 일반적으로 반 최대한의 (또는 3 분기 최대한) EPSP 진폭을 생산하도록 조정됩니다. 잠재적인 문제는 세 쥐와 APP/PS1 쥐가 일반적으로베이스 라인 EPSP 가치도 세 쥐와 APP/PS1 생쥐에 작은 것입니다 즉, 자신의 젊은 및 / 또는 야생 타입 대응에 상대적으로 시냅스 강도를 감소 보여주는 것입니다. 작은 EPSPs는 LTP 33 유도를위한 확률 감소의 결과로, LTP의 자극 동안 적은 탈분극로 번역 수도 있습니다. 때문에 잠재적인 혼란, 그것이 동물들이 처리량 적자, 소성 적자 또는 둘 모두를 전시 여부를 확인하기가 어렵습니다. 즉, 세 및 / 또는 APP/PS1 생쥐에서 LTP 유도 메커니즘 (NO 소성 적자) 기능 그대로 수 있지만 충분 이러한 조건 하에서 (처리량 적자) 자극. 처리량 및 소성에 대한 메커니즘에 대한 메커니즘은 특정 pharmacologic 치료에 매우 다르게 반응 수도 있으므로이 구분은 중요합니다. 우리는 이전 LTP의 자극에 대한 모든 조각에 걸쳐 동일한 수준 (예 : 1 MV)로 EPSP 진폭을 정규화하여 LTP 유도에 감소 처리량의 영향을 최소화하려고합니다. 다른 전략뿐만 아니라 효과 (LTP의 자극 동안 그룹 전체의 막 잠재력을 맞춰야 전압 또는 전류 클램프의 예 사용), 이러한 동물 모델에서 LTP를 조사 때 고려해야 수 있습니다.

<table border="1"><tbody><tr><td> 시약의 이름 </td><td> 회사 </td><td> 카탈로그 번호 </td></tr><tr><td> NaCl </td><td> 어부 </td><td> BP358 - 1 </td></tr><tr><td> KCl </td><td> 어부 </td><td> BP366 - 500 </td></tr><tr><td> KH 2 PO 4 (일염기의) </td><td> 시그마 </td><td> P5379 - 100G </td></tr><tr><td> MgSO 4 </td><td> 시그마 </td><td> M2643 - 500G </td></tr><tr><td> CaCl 2 (이수화물) </td><td> 시그마 </td><td> C3306 - 250G </td></tr><tr><td> NaHCO 3 </td><td> 어부 </td><td> S233 - 500 </td></tr><tr><td> C 6 H 12 O 6 (덱스 트로 오스) </td><td> 어부 </td><td> BP350 - 1 </td></tr></tbody></table> 표 1. 시약이 필요 <table border="1"><tbody><tr><td> 장비의 명칭 </td><td> 회사 </td><td> 카탈로그 번호 </td><td> 댓글 </td></tr><tr><td> Erlenmeyer Flasks </td><td> 어부 </td><td> FB - 500-2000 FB - 500-1000 </td><td></td></tr><tr><td> 수족관 버블러 </td><td></td><td></td><td> oxygenating 미디어에 사용됩니다. 대부분의 애완 동물 상점에서 사용 가능 </td></tr><tr><td> 50 ML 유리 비이커 </td><td> 어부 </td><td> 02 - 540G </td><td> ACSF의 뇌 storarge에 대한 </td></tr><tr><td> Parafilm </td><td> 어부 </td><td> 13-374-10 </td><td></td></tr><tr><td> 스몰 애니멀 기요틴 </td><td> 세계 정밀 계측기 (WPI) </td><td> DCAP - M </td><td></td></tr><tr><td> 플랫 종이 타월 </td><td></td><td></td><td></td></tr><tr><td> # 11 피더서 외과 블레이드 </td><td> 어부 </td><td> 08-916 - 5B </td><td></td></tr><tr><td> 비비 뼈 가위 </td><td> 파인 과학 도구 (FST) </td><td> 16044-10 </td><td></td></tr><tr><td> Lempert Rongeurs </td><td> Roboz </td><td> RS - 8321 </td><td> 쥐에 사용 </td></tr><tr><td> 프리드먼 - 피어슨 Rongeurs </td><td> FST </td><td> 16020-14 </td><td> 마우스 또는 쥐에 사용 </td></tr><tr><td> 해마 도구 </td><td> FST </td><td> 10099-15 </td><td></td></tr><tr><td> 숟가락 </td><td></td><td></td><td> 플라스틱 찻숟가락 할 것입니다 </td></tr><tr><td> 주걱 </td><td> 어부 </td><td> 21-401 - 25A </td><td> 주걱 </td></tr><tr><td> 외과 홍채 가위 </td><td> FST </td><td> 14058-09 </td><td></td></tr><tr><td> 플라스틱 전송 pipets </td><td> 어부 </td><td> 13-711-43 </td><td></td></tr><tr><td> 110mm 왓먼 필터 용지 </td><td> 어부 </td><td> 09 - 805E </td><td> 왓먼 고양이. 1001-110 </td></tr><tr><td> 유리 페트리 접시 </td><td> 어부 </td><td></td><td></td></tr><tr><td> Leica VT1000P 매뉴얼 마이크로톰 진동 </td><td> Vibratome </td><td></td><td></td></tr><tr><td> 0.1mm FA ​​- 10 깃털 S 블레이드 </td><td> 테드 펠라 </td><td> 121-9 </td><td> 0.1mm FA ​​- 10 깃털 S 블레이드 </td></tr><tr><td> Borosilicate 유리 파스퇴르 Pipet (고무 전구 포함) </td><td> 어부 </td><td> 13-678 - 20A </td><td> 조각 전송 : 도움말은 큰 여는 부러 및 열 광택입니다 </td></tr><tr><td> 35mm Polysterine 문화 요리 </td><td> 코닝 </td><td> 430588 </td><td> 절개 후 슬라이스를 수집하기 위해 사용 </td></tr></tbody></table> 표 2. 도구 및 해부를위한 자료 <table border="1"><tbody><tr><td> 장비 / 재료 </td><td> 회사 </td><td> 댓글 (옵션) </td></tr><tr><td> 홀딩 챔버 </td><td> 사용자가 내장 </td><td></td></tr><tr><td> P - 97 수평 피펫 풀러 </td><td> 서터 악기 (주) </td><td></td></tr><tr><td> 진동 절연 테이블 </td><td> 기술 제조 주식 회사 (TMC) </td><td></td></tr><tr><td> 패러데이 케이지 </td><td> 사용자가 만든 </td><td></td></tr><tr><td> 히프 권총과 Pyrex 흡인기 바틀 (제품 # 1220 - 1L) </td><td> 코닝 </td><td></td></tr><tr><td> 레귤레이터와 중력 - contolled IV 세트 (제품 # 2C8891) </td><td> 백스터 </td><td></td></tr><tr><td> 중앙 진공 라인 </td><td></td><td> 가장 현대적인 실험실에서 사용 가능 </td></tr><tr><td> 95% O 5분의 2 %의 CO 2 기체 혼합물 </td><td> 스코틀랜드 사람T - 그로스 주식 회사 </td><td></td></tr><tr><td> TygonTM 랩 튜브 O 2 / CO 2 전달 </td><td> 피셔 사이 언티픽 </td><td> 무독성, 비 산화, 다양한 크기의 제공됩니다. </td></tr><tr><td> 이클립스 E600FN 현미경 </td><td> 니콘 </td><td> infared 필터 및 GFP, DS - Red2 필터 근처 10X와 40x 목표와 </td></tr><tr><td> 쿨 스냅 ES 디지털 카메라 </td><td> Photometrics </td><td> 쿨 스냅 ES 디지털 카메라 </td></tr><tr><td> 형광 조명기을 X - 인용 </td><td> EXFO </td><td> 형광 조명기을 X - 인용 </td></tr><tr><td> 현미경 플랫폼 </td><td> 시스키유 </td><td> 사용자가 조립 </td></tr><tr><td> RC - 22 잠수함 기록 챔버 (제품 # 64-0228) </td><td> 워너 인 스트 루먼트 (WI) </td><td> P - 1 플랫폼 무대 어댑터 (워너에서 제품 # 64-0277)이 필요합니다 </td></tr><tr><td> TC2BIP 2/3Ch 온도 컨트롤러 </td><td> 셀 Microcontrols </td><td> TC2BIP 2/3Ch 온도 컨트롤러 </td></tr><tr><td> 4 축 매뉴얼 미니의 속이는 사람 </td><td> 시스키유 </td><td></td></tr><tr><td> 플래티넘 이리듐 와이어 (0.002)을 (항목 # PTT0203) </td><td> WPI </td><td></td></tr><tr><td> A365 자극 아이 솔 레이터 </td><td> WPI </td><td> A365 자극 아이 솔 레이터 </td></tr><tr><td> 700b 앰프를 Multiclamp </td><td> 액슨 인 스트 루먼트 </td><td></td></tr><tr><td> Digidata 1322A A / D 변환기 </td><td> 액슨 인 스트 루먼트 </td><td></td></tr><tr><td> PClamp 소프트웨어 </td><td> 액슨 인 스트 루먼트 </td><td></td></tr><tr><td> 개인용 컴퓨터 (펜티엄 4) </td><td> 작은 골짜기 </td><td></td></tr></tbody></table> 표 3. 전기 생리학 장비 및 재료

preparation of acute hippocampal slices from rats and transgenic mice for the study of synaptic alterations during aging and amyloid pathology

쥐 hippocampal 슬라이스 준비 아마 포유류의 시냅스 기능과 소성 조사를위한 가장 광범위하게 사용되는 도구입니다. 해마는 쥐와 생쥐에서 빠르고 쉽게 추출하여 슬라이스는 산소 인공 뇌척수의 시간 동안 가능한 유지하실 수 있습니다. 또한, 기본 electrophysisologic 기술은 쉽게 hippocampal 조각의 시냅스 기능의 조사에 적용하고 인지적 손상에 가장 적합한 생체의 일부를 제공하고 있습니다. hippocampal 슬라이스는 학습과 기억에 관련된 신경 소성 메카니즘 연구 특히 인기가있다. hippocampal 조각 (또는 부족 그것의)에서 시냅스 효능의 장기 potentiation과 우울증 (LTP와 LTD)의 유도에 변경이 자주 cognitively 장애인 동물 및 / neurologic 표현형을 설명하기 위해 또는 행동의 메커니즘을 평가하는 데 사용됩니다 nootropic 화합물. 이 문서는 우리가 두뇌 노화와 알츠하이머 질환 (AD) 1-3과 관련된 시냅스 변경의 연구를위한 유전자 변형 쥐 및 생쥐에서 hippocampal 조각 준비 사용하는 절차를 설명합니다. 세 쥐 및 광고 모델 마우스의 사용은 그들의 연구에 젊은 쥐 및 / 또는 마우스를 사용하여 익숙한 연구자 도전의 고유한 집합을 표시할 수 있습니다. 세 쥐들이 두뇌 추출 및 / 또는 절개를 지연하고 따라서 시냅스 기능과 소성의 실제 나이 차이를 부정하거나 과장 수있는 젊은 쥐 및 생쥐보다 두꺼운 두개골과 강하다는 결합 조직을 보유하고 있습니다. 노화와 아밀로이드 병리도 다시 physiologic 평가에서 그려진 모든 inferences을 복잡 절개 절차 동안 지속 hippocampal 손상을 더욱 심하게하다 수 있습니다. 여기, 우리는 이러한 문제를 최소화하기 위해 절개 절차 동안 촬영한 단계에 대해 설명합니다. &quot;건강&quot;과 쥐 및 생쥐에서 &quot;건강에 해로운 &#39;조각 인수 시냅스 응답의 예는 물론 대표적인 시냅스 소성 실험으로 제공됩니다. 이러한 동물 모델 (예 : 레코딩 솔루션 구성 요소, 자극 매개 변수)에 시냅스 기능에 다른 방법 론적 요소의 가능한 영향도 설명합니다. 이 문서의 초점은 생리를 슬라이스에 세 쥐와 유전자 변형 마우스, 초보자의 사용에있는 동안 쥐 모델의 다양한 사용하여 자신의 연구를 시작하려면 여기를 충분한 세부 사항을 찾을 수 있어야합니다.

Watch this Scientific Journal Video about Preparation of Acute Hippocampal Slices from Rats and Transgenic Mice for the Study of Synaptic Alterations during Aging and Amyloid Pathology at JoVE.com

Preparation of Acute Hippocampal Slices from Rats and Transgenic Mice for the Study of Synaptic Alterations during Aging and Amyloid Pathology

이 문서는 뇌의 노화 및 알츠하이머 질환 같은 나이와 관련된 neurodegenerative 질병와 관련된 시냅스 변경의 연구를위한 유전자 변형 쥐 및 생쥐에서 hippocampal 조각 준비 절차를 설명합니다.

노화와 아밀로이드 병리 동안 시냅틱 얼터 레이션의 연구에 대한 고양이와 트렌스 제닉 마우스의 급성 Hippocampal 조각의 준비

The rodent hippocampal slice preparation is perhaps the most broadly used tool for investigating mammalian synaptic function and plasticity. The ...

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Neuroscience

93.7K 조회수.  University of Kentucky College of Public Health. 이 문서는 뇌의 노화 및 알츠하이머 질환 같은 나이와 관련된 neurodegenerative 질병와 관련된 시냅스 변경의 연구를위한 유전자 변형 쥐 및 생쥐에서 hippocampal 조각 준비 절차를 설명합니다.

비디오: Preparation of Acute Hippocampal Slices from Rats and Transgenic Mice for the Study of Synaptic Alterations during Aging and Amyloid Pathology

Preparation of Oligomeric &beta;-amyloid1-42 and Induction of Synaptic Plasticity Impairment on Hippocampal Slices

Impairment of synaptic connections is likely to underlie the subtle amnesic changes occurring at the early stages of Alzheimer s Disease (AD). &beta;-amyloid (A&beta;), a peptide produced in high amounts in AD, is known to reduce Long-Term Potentiation (LTP), a cellular correlate of learning and memory. Indeed, LTP impairment caused by A&beta; is a useful experimental paradigm for studying synaptic dysfunctions in AD models and for screening drugs capable of mitigating or reverting such synaptic impairments. Studies have shown that A&beta; produces the LTP disruption preferentially via its oligomeric form. Here we provide a detailed protocol for impairing LTP by perfusion of oligomerized synthetic A&beta;1-42 peptide onto acute hippocampal slices. In this video, we outline a step-by-step procedure for the preparation of oligomeric A&beta;1-42. Then, we follow an individual experiment in which LTP is reduced in hippocampal slices exposed to oligomerized A&beta;1-42 compared to slices in a control experiment where no A&beta;1-42 exposure had occurred.

Preparation of Oligomeric &amp;#946;-amyloid1-42 and Induction of Synaptic Plasticity Impairment on Hippocampal Slices

One feature of Alzheimer's Disease is the elevation of A&beta;1-42 peptide. Here we provide a protocol for preparing synthetic A&beta;1-42 oligomers, which impairs hippocampal Long-Term Potentiation, a cellular correlate of memory. This procedure is useful for investigating mechanisms of A&beta;-induced pathology and drug screening.

β - 아밀로이드 Oligomeric의 준비 1-42</subHippocampal 조각에 시냅틱 소성 장애의>와 유도

Impairment of synaptic connections is likely to underlie the subtle amnesic changes occurring at the early stages of Alzheimer s Disease (AD). ...

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Vibrodissociation of Neurons from Rodent Brain Slices to Study Synaptic Transmission and Image Presynaptic Terminals

Mechanical dissociation of neurons from the central nervous system has the advantage that presynaptic boutons remain attached to the isolated neuron of interest. This allows for examination of synaptic transmission under conditions where the extracellular and postsynaptic intracellular environments can be well controlled. A vibration-based technique without the use of proteases, known as vibrodissociation, is the most popular technique for mechanical isolation. A micropipette, with the tip fire-polished to the shape of a small ball, is placed into a brain slice made from a P1-P21 rodent. The micropipette is vibrated parallel to the slice surface and lowered through the slice thickness resulting in the liberation of isolated neurons. The isolated neurons are ready for study within a few minutes of vibrodissociation. This technique has advantages over the use of primary neuronal cultures, brain slices and enzymatically isolated neurons including: rapid production of viable, relatively mature neurons suitable for electrophysiological and imaging studies; superior control of the extracellular environment free from the influence of neighboring cells; suitability for well-controlled pharmacological experiments using rapid drug application and total cell superfusion; and improved space-clamp in whole-cell recordings relative to neurons in slice or cell culture preparations. This preparation can be used to examine synaptic physiology, pharmacology, modulation and plasticity. Real-time imaging of both pre- and postsynaptic elements in the living cells and boutons is also possible using vibrodissociated neurons. Characterization of the molecular constituents of pre- and postsynaptic elements can also be achieved with immunological and imaging-based approaches.

This report demonstrates a technique for mechanical isolation of individual viable neurons retaining attached presynaptic boutons. Vibrodissociated neurons have the advantages of rapid production, excellent pharmacological control and improved space-clamp without influence from neighboring cells. This method can be used for imaging of synaptic elements and patch-clamp recording.

쥐 뇌 조각에서 뉴런의 Vibrodissociation는 시냅틱 전송 및 이미지 Presynaptic 터미널 공부

Mechanical dissociation of neurons from the central nervous system has the advantage that presynaptic boutons remain attached to the isolated neuron of ...

Preparation of Acute Subventricular Zone Slices for Calcium Imaging

The subventricular zone (SVZ) is one of the two neurogenic zones in the postnatal brain. The SVZ contains densely packed cells, including neural progenitor cells with astrocytic features (called SVZ astrocytes), neuroblasts, and intermediate progenitor cells. Neuroblasts born in the SVZ tangentially migrate a great distance to the olfactory bulb, where they differentiate into interneurons. Intercellular signaling through adhesion molecules and diffusible signals play important roles in controlling neurogenesis. Many of these signals trigger intercellular calcium activity that transmits information inside and between cells. Calcium activity is thus reflective of the activity of extracellular signals and is an optimal way to understand functional intercellular signaling among SVZ cells.
Calcium activity has been studied in many other regions and cell types, including mature astrocytes and neurons. However, the traditional method to load cells with calcium indicator dye (i.e. bath loading) was not efficient at loading all SVZ cell types. Indeed, the cellular density in the SVZ precludes dye diffusion inside the tissue. In addition, preparing sagittal slices will better preserve the three-dimensional arrangement of SVZ cells, particularly the stream of neuroblast migration on the rostral-caudal axis.
Here, we describe methods to prepare sagittal sections containing the SVZ, the loading of SVZ cells with calcium indicator dye, and the acquisition of calcium activity with time-lapse movies. We used Fluo-4 AM dye for loading SVZ astrocytes using pressure application inside the tissue. Calcium activity was recorded using a scanning confocal microscope allowing a precise resolution for distinguishing individual cells. Our approach is applicable to other neurogenic zones including the adult hippocampal subgranular zone and embryonic neurogenic zones. In addition, other types of dyes can be applied using the described method.

A method to load subventricular zone (SVZ) cells with calcium indicator dyes for recording calcium activity is described. The postnatal SVZ contains tightly packed cells including neural progenitor cells and neuroblasts. Rather than using bath loading we injected the dye by pressure inside the tissue allowing better dye diffusion.

칼슘 이미징에 대한 급성 Subventricular 지대 슬라이스의 작성

The subventricular zone (SVZ) is one of the two neurogenic zones in the postnatal brain. The SVZ contains densely packed cells, including neural ...

Dual Electrophysiological Recordings of Synaptically-evoked Astroglial and Neuronal Responses in Acute Hippocampal Slices

Astrocytes form together with neurons tripartite synapses, where they integrate and modulate neuronal activity. Indeed, astrocytes sense neuronal inputs through activation of their ion channels and neurotransmitter receptors, and process information in part through activity-dependent release of gliotransmitters. Furthermore, astrocytes constitute the main uptake system for glutamate, contribute to potassium spatial buffering, as well as to GABA clearance. These cells therefore constantly monitor synaptic activity, and are thereby sensitive indicators for alterations in synaptically-released glutamate, GABA and extracellular potassium levels. Additionally, alterations in astroglial uptake activity or buffering capacity can have severe effects on neuronal functions, and might be overlooked when characterizing physiopathological situations or knockout mice. Dual recording of neuronal and astroglial activities is therefore an important method to study alterations in synaptic strength associated to concomitant changes in astroglial uptake and buffering capacities. Here we describe how to prepare hippocampal slices, how to identify stratum radiatum astrocytes, and how to record simultaneously neuronal and astroglial electrophysiological responses. Furthermore, we describe how to isolate pharmacologically the synaptically-evoked astroglial currents.

The preparation of acute brain slices from isolated hippocampi, as well as the simultaneous electrophysiological recordings of astrocytes and neurons in stratum radiatum during stimulation of schaffer collaterals is described. The pharmacological isolation of astroglial potassium and glutamate transporter currents is demonstrated.

급성 Hippocampal 슬라이스에 Synaptically-evoked Astroglial와 Neuronal 응답의 듀얼 Electrophysiological 레코딩

Astrocytes form together with neurons tripartite synapses, where they integrate and modulate neuronal activity. Indeed, astrocytes sense neuronal inputs ...

Stereotaxic Infusion of Oligomeric Amyloid-beta into the Mouse Hippocampus

Alzheimer&#8217;s disease is a neurodegenerative disease affecting the aging population. A key neuropathological feature of the disease is the over-production of amyloid-beta and the deposition of amyloid-beta plaques in brain regions of the afflicted individuals. Throughout the years scientists have generated numerous Alzheimer&#8217;s disease mouse models that attempt to replicate the amyloid-beta pathology. Unfortunately, the mouse models only selectively mimic the disease features. Neuronal death, a prominent effect in the brains of Alzheimer&#8217;s disease patients, is noticeably lacking in these mice. Hence, we and others have employed a method of directly infusing soluble oligomeric species of amyloid-beta - forms of amyloid-beta that have been proven to be most toxic to neurons - stereotaxically into the brain. In this report we utilize male C57BL/6J mice to document this surgical technique of increasing amyloid-beta levels in a select brain region. The infusion target is the dentate gyrus of the hippocampus because this brain structure, along with the basal forebrain that is connected by the cholinergic circuit, represents one of the areas of degeneration in the disease. The results of elevating amyloid-beta in the dentate gyrus via stereotaxic infusion reveal increases in neuron loss in the dentate gyrus within 1 week, while there is a concomitant increase in cell death and cholinergic neuron loss in the vertical limb of the diagonal band of Broca of the basal forebrain. These effects are observed up to 2 weeks. Our data suggests that the current amyloid-beta infusion model provides an alternative mouse model to address region specific neuron death in a short-term basis. The advantage of this model is that amyloid-beta can be elevated in a spatial and temporal manner.

Here, we present a protocol for direct stereotaxic brain infusion of amyloid-beta. This methodology provides an alternative in vivo mouse model to address the short-term effects of amyloid-beta on brain neurons.

마우스 해마에 올리고머 아밀로이드 - 베타의 정위 주입

Alzheimer&#8217;s disease is a neurodegenerative disease affecting the aging population. A key neuropathological feature of the disease is the ...

Investigation of Synaptic Tagging/Capture and Cross-capture using Acute Hippocampal Slices from Rodents

Synaptic tagging and capture (STC) and cross-tagging are two important mechanisms at cellular level that explain how synapse-specificity and associativity is achieved in neurons within a specific time frame. These long-term plasticity-related processes are the leading candidate models to study the basis of memory formation and persistence at the cellular level. Both STC and cross-tagging involve two serial processes: (1) setting of the synaptic tag as triggered by a specific pattern of stimulation, and (2) synaptic capture, whereby the synaptic tag interacts with newly synthesized plasticity-related proteins (PRPs). Much of the understanding about the concepts of STC and cross-tagging arises from the studies done in CA1 region of the hippocampus and because of the technical complexity many of the laboratories are still unable to study these processes. Experimental conditions for the preparation of hippocampal slices and the recording of stable late-LTP/LTD are extremely important to study synaptic tagging/cross-tagging. This video article describes the experimental procedures to study long-term plasticity processes such as STC and cross-tagging in the CA1 pyramidal neurons using stable, long-term field-potential recordings from acute hippocampal slices of rats.

This video article describes experimental procedures to study long-term plasticity and its associative processes such as synaptic tagging, capture and cross-tagging in the CA1 pyramidal neurons using acute hippocampal slices from rodents.

설치류에서 급성 해마 조각을 사용하여 시냅스 태그 / 캡처 및 크로스 캡처 조사

Synaptic tagging and capture (STC) and cross-tagging are two important mechanisms at cellular level that explain how synapse-specificity and associativity ...

Preparation of Acute Brain Slices Using an Optimized N-Methyl-D-glucamine Protective Recovery Method

This protocol is a practical guide to the N-methyl-D-glucamine (NMDG) protective recovery method of brain slice preparation. Numerous recent studies have validated the utility of this method for enhancing neuronal preservation and overall brain slice viability. The implementation of this technique by early adopters has facilitated detailed investigations into brain function using diverse experimental applications and spanning a wide range of animal ages, brain regions, and cell types. Steps are outlined for carrying out the protective recovery brain slice technique using an optimized NMDG artificial cerebrospinal fluid (aCSF) media formulation and enhanced procedure to reliably obtain healthy brain slices for patch clamp electrophysiology. With this updated approach, a substantial improvement is observed in the speed and reliability of gigaohm seal formation during targeted patch clamp recording experiments while maintaining excellent neuronal preservation, thereby facilitating challenging experimental applications. Representative results are provided from multi-neuron patch clamp recording experiments to assay synaptic connectivity in neocortical brain slices prepared from young adult transgenic mice and mature adult human neurosurgical specimens. Furthermore, the optimized NMDG protective recovery method of brain slicing is compatible with both juvenile and adult animals, thus resolving a limitation of the original methodology. In summary, a single media formulation and brain slicing procedure can be implemented across various species and ages to achieve excellent viability and tissue preservation.

Preparation of Acute Brain Slices Using an Optimized N-Methyl-D-glucamine Protective Recovery Method

This protocol demonstrates the implementation of an optimized N-methyl-D-glucamine (NMDG) protective recovery method of brain slice preparation. A single media formulation is used to reliably obtain healthy brain slices from animals of any age and for diverse experimental applications.

최적화 된 N를 사용 하 여 급성 뇌 조각의 준비-메 틸-D-glucamine 보호 복구 방법

This protocol is a practical guide to the N-methyl-D-glucamine (NMDG) protective recovery method of brain slice preparation. Numerous recent studies have ...

Recording Synaptic Plasticity in Acute Hippocampal Slices Maintained in a Small-volume Recycling-, Perfusion-, and Submersion-type Chamber System

Even though experiments on brain slices have been in use since 1951, problems remain that reduce the probability of achieving a stable and successful analysis of synaptic transmission modulation when performing field potential or intracellular recordings. This manuscript describes methodological aspects that might be helpful in improving experimental conditions for the maintenance of acute brain slices and for recording field excitatory postsynaptic potentials in a commercially available submersion chamber with an outflow-carbogenation unit. The outflow-carbogenation helps to stabilize the oxygen level in experiments that rely on the recycling of a small buffer reservoir to enhance the cost-efficiency of drug experiments. In addition, the manuscript presents representative experiments that examine the effects of different carbogenation modes and stimulation paradigms on the activity-dependent synaptic plasticity of synaptic transmission.

This protocol describes the stabilization of the oxygen level in a small volume of recycled buffer and methodological aspects of recording activity-dependent synaptic plasticity in submerged acute hippocampal slices.

급성 Hippocampal 조각에 작은 볼륨 재활용-, 관류-침수 형 챔버 시스템 유지에 시 냅 스가 소성을 기록

Even though experiments on brain slices have been in use since 1951, problems remain that reduce the probability of achieving a stable and successful ...

Single Synapse Indicators of Glutamate Release and Uptake in Acute Brain Slices from Normal and Huntington Mice

Synapses are highly compartmentalized functional units that operate independently on each other. In Huntington&#39;s disease (HD) and other neurodegenerative disorders, this independence might be compromised due to insufficient glutamate clearance and the resulting spill-in and spill-out effects. Altered astrocytic coverage of the presynaptic terminals and/or dendritic spines as well as a reduced size of glutamate transporter clusters at glutamate release sites have been implicated in the pathogenesis of diseases resulting in symptoms of dys-/hyperkinesia. However, the mechanisms leading to the dysfunction of glutamatergic synapses in HD are not well understood. Improving and applying synapse imaging we have obtained data shedding new light on the mechanisms impeding the initiation of movements. Here, we describe the principle elements of a relatively inexpensive approach to achieve single synapse resolution by using the new genetically encoded ultrafast glutamate sensor iGluu, wide-field optics, a scientific CMOS (sCMOS) camera, a 473 nm laser and a laser positioning system to evaluate the state of corticostriatal synapses in acute slices from age appropriate healthy or diseased mice. Glutamate transients were constructed from single or multiple pixels to obtain estimates of i) glutamate release based on the maximal elevation of the glutamate concentration [Glu] next to the active zone and ii) glutamate uptake as reflected in the time constant of decay (TauD) of the perisynaptic [Glu]. Differences in the resting bouton size and contrasting patterns of short-term plasticity served as criteria for the identification of corticostriatal terminals as belonging to the intratelencephalic (IT) or the pyramidal tract (PT) pathway. Using these methods, we discovered that in symptomatic HD mice ~40% of PT-type corticostriatal synapses exhibited insufficient glutamate clearance, suggesting that these synapses might be at risk to excitotoxic damage. The results underline the usefulness of TauD as a biomarker of dysfunctional synapses in Huntington mice with a hypokinetic phenotype.

We present a protocol to evaluate the balance between glutamate release and clearance at single corticostriatal glutamatergic synapses in acute slices from adult mice. This protocol uses the fluorescent sensor iGluu for glutamate detection, a sCMOS camera for signal acquisition and a device for focal laser illumination.

정상 및 헌팅턴 마우스의 급성 뇌 슬라이스에서 글루타메이트 방출 및 섭취량의 단일 시냅스 지표

Synapses are highly compartmentalized functional units that operate independently on each other. In Huntington&#39;s disease (HD) and other ...

Preparation of Acute Human Hippocampal Slices for Electrophysiological Recordings

Epilepsy affects about 1% of the world population and leads to a severe decrease in quality of life due to ongoing seizures as well as high risk for sudden death. Despite an abundance of available treatment options, about 30% of patients are drug-resistant. Several novel therapeutics have been developed using animal models, though the rate of drug-resistant patients remains unaltered. One of probable reasons is the lack of translation between rodent models and humans, such as a weak representation of human pharmacoresistance in animal models. Resected human brain tissue as a preclinical evaluation tool has the advantage to bridge this translational gap. Described here is a method for high quality preparation of human hippocampal brain slices and subsequent stable induction of epileptiform activity. The protocol describes the induction of burst activity during application of 8 mM KCl and 4-aminopyridin. This activity is sensitive to established AED lacosamide or novel antiepileptic candidates, such as dimethylethanolamine (DMEA). In addition, the method describes induction of seizure-like events in CA1 of human hippocampal brain slices by reduction of extracellular Mg2+ and application of bicuculline, a GABAA receptor blocker. The experimental set-up can be used to screen potential antiepileptic substances for their effects on epileptiform activity. Furthermore, mechanisms of action postulated for specific compounds can be validated using this approach in human tissue (e.g., using patch-clamp recordings). To conclude, investigation of vital human brain tissue ex vivo (here, resected hippocampus from patients suffering from temporal lobe epilepsy) will improve the current knowledge of physiological and pathological mechanisms in the human brain.

The presented protocol describes the transport and preparation of resected human hippocampal tissue with the ultimate goal to use vital brain slices as a preclinical evaluation tool for potential antiepileptic substances.

전기 생리학적 기록을 위한 급성 인간 해마 슬라이스의 준비

Epilepsy affects about 1% of the world population and leads to a severe decrease in quality of life due to ongoing seizures as well as high risk for ...