Name: preparation of acute hippocampal slices from rats and transgenic mice for the study of synaptic alterations during aging and amyloid pathology
Uploaded: 2011-03-23T14:15:00
Duration: 14 min 57 s
Description: 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

Work supported by NIH grant AG027297, an award from the Kentucky Spinal Cord and Head Injury Research Trust, and a gift from the Kleberg Foundation.

<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>

The production of this video-article was sponsored by Leica Microsystems.

The steps outlined in this protocol will help insure that brain dissections are carried out at least as rapidly and efficiently in aged, as in young adult rats. We also provide sufficient detail for the beginner to set up their own slice studies on LTP and LTD. If further exploration of aging and AD changes in synaptic function and plasticity is one of your goals, there are at least two other methodological issues, alluded to above, that deserve further consideration. First, several labs have shown that the Ca2+:Mg2+ ratio in recording ACSF can have a marked effect on the induction synaptic plasticity in hippocampal slices2,10,24,25. In mammalian CSF, the Ca2+:Mg2+ ratio is approximately one (e.g. see 26). However, ACSF Ca2+:Mg2+ ratios closer to 2 are commonly used in slice studies of synaptic function and plasticity. In early studies, this practice was probably adapted to optimize induction of LTP, then subsequently became routine for all plasticity studies. However, this practice can be problematic in aging and AD studies because of well-characterized differences in neuronal Ca2+ regulation. Specifically, Ca2+ influx and/or Ca2+-induced Ca2+-release is elevated in aged rats and/or AD model mice during neuronal activation3,27-31. Induction of LTD is particularly sensitive to subtle changes in ACSF Ca2+ levels. Our protocol, which uses 2mM Ca2+ and 2 mM Mg2+, commonly results in LTD for aged, but not young adult animals2, while studies using a Ca2+:Mg2+ ratio closer to two, have observed robust LTD in adults in the absence of an age difference2,10 or in conjunction with reduced LTD in aged rats32. These observations highlight the need to carefully consider ACSF Ca2+ and Mg2+ levels when comparing Ca2+-dependent plasticity in aged and young adult animals.
The second methodological issue concerns the strong dependency of LTP on postsynaptic depolarization33 and possible aging/genotype differences in synaptic strength. In a typical LTP experiment, baseline and LTP stimulation intensity is generally adjusted to produce a half maximal (or three-quarter maximal) EPSP amplitude. The potential problem is that aged rats and APP/PS1 mice usually show reduced synaptic strength relative to their younger and/or wild type counterparts, meaning that baseline EPSP values will also be smaller in aged rats and APP/PS1 mice. Smaller EPSPs may translate to less depolarization during LTP stimulation, resulting in a reduced probability for inducing LTP33. Because of this potential confound, it is difficult to determine whether these animals exhibit a throughput deficit, a plasticity deficit or both. That is, LTP induction mechanisms in aged and/or APP/PS1 mice may be functionally intact (no plasticity deficit), but insufficiently stimulated (throughput deficit) under these conditions. This distinction is critical, as mechanisms for throughput and mechanisms for plasticity may respond very differently to a specific pharmacologic treatment. We try to minimize the impact of reduced throughput on LTP induction by normalizing the EPSP amplitude to the same level (e.g. 1 mV) across all slices prior to LTP stimulation. Other strategies may be effective as well (e.g. use of voltage or current clamp to equalize the membrane potential across groups during LTP stimulation), and should be considered when investigating LTP in these animal models.

<table border="1">
<tbody>
 <tr>
 <td>Name of the reagent</td>
 <td>Company</td>
 <td>Catalogue number</td>
 </tr>
 <tr>
 <td>NaCl</td>
 <td>Fisher</td>
 <td>BP358-1</td>
 </tr>
 <tr>
 <td>KCl</td>
 <td>Fisher</td>
 <td>BP366-500</td>
 </tr>
 <tr>
 <td>KH2PO4 (monobasic)</td>
 <td>Sigma</td>
 <td>P5379-100G</td>
 </tr>
 <tr>
 <td>MgSO4</td>
 <td>Sigma</td>
 <td>M2643-500G</td>
 </tr>
 <tr>
 <td>CaCl2 (dihydrate)</td>
 <td>Sigma </td>
 <td>C3306-250G</td>
 </tr>
 <tr>
 <td>NaHCO3</td>
 <td>Fisher </td>
 <td>S233-500</td>
 </tr>
 <tr>
 <td>C6H12O6 (dextrose)</td>
 <td>Fisher </td>
 <td>BP350-1</td>
 </tr>
</tbody>
</table>
Table 1. Reagents required
<table border="1">
<tbody>
 <tr>
 <td>Name of the equipment</td>
 <td>Company</td>
 <td>Catalogue number</td>
 <td>Comments </td>
 </tr>
 <tr>
 <td>Erlenmeyer Flasks</td>
 <td>Fisher</td>
 <td>FB-500-2000 
 FB-500-1000</td>
 <td>&nbsp;</td>
 </tr>
 <tr>
 <td>Aquarium Bubbler</td>
 <td>&nbsp;</td>
 <td>&nbsp;</td>
 <td>Used for oxygenating media. Available at most pet stores</td>
 </tr>
 <tr>
 <td>50 mL Glass beaker</td>
 <td>Fisher</td>
 <td>02-540G</td>
 <td>For brain storarge in ACSF</td>
 </tr>
 <tr>
 <td>Parafilm</td>
 <td>Fisher</td>
 <td>13-374-10</td>
 <td>&nbsp;</td>
 </tr>
 <tr>
 <td>Small Animal Guillotine</td>
 <td>World Precision Instruments (WPI)</td>
 <td>DCAP-M</td>
 <td>&nbsp;</td>
 </tr>
 <tr>
 <td>Flat paper towel</td>
 <td>&nbsp;</td>
 <td>&nbsp;</td>
 <td>&nbsp;</td>
 </tr>
 <tr>
 <td>#11 Feather surgical blade</td>
 <td>Fisher</td>
 <td>08-916-5B</td>
 <td>&nbsp;</td>
 </tr>
 <tr>
 <td>Beebee bone scissors</td>
 <td>Fine Science Tools (FST)</td>
 <td>16044-10</td>
 <td>&nbsp;</td>
 </tr>
 <tr>
 <td>Lempert Rongeurs</td>
 <td>Roboz</td>
 <td>RS-8321</td>
 <td>Use for rats</td>
 </tr>
 <tr>
 <td>Friedman-Pearson Rongeurs</td>
 <td>FST</td>
 <td>16020-14</td>
 <td>Use for mice or rats</td>
 </tr>
 <tr>
 <td>Hippocampus tool</td>
 <td>FST</td>
 <td>10099-15</td>
 <td>&nbsp;</td>
 </tr>
 <tr>
 <td>Spoon</td>
 <td>&nbsp;</td>
 <td>&nbsp;</td>
 <td>A plastic teaspoon will do</td>
 </tr>
 <tr>
 <td>Spatula</td>
 <td>Fisher</td>
 <td>21-401-25A </td>
 <td>Spatula</td>
 </tr>
 <tr>
 <td>Surgical iris scissors</td>
 <td>FST</td>
 <td>14058-09</td>
 <td>&nbsp;</td>
 </tr>
 <tr>
 <td>plastic transfer pipets</td>
 <td>Fisher</td>
 <td>13-711-43 </td>
 <td>&nbsp;</td>
 </tr>
 <tr>
 <td>110mm Whatman filter paper</td>
 <td>Fisher</td>
 <td>09-805E</td>
 <td>Whatman cat. 1001-110</td>
 </tr>
 <tr>
 <td>Glass petri dish</td>
 <td>Fisher</td>
 <td>&nbsp;</td>
 <td>&nbsp;</td>
 </tr>
 <tr>
 <td><a href="http://www.leica-microsystems.com/products/total-histology/sectioning/vibrating-blade-microtomes/">Leica VT1000P Manual Vibrating Microtome</a></td>
 <td>Vibratome</td>
 <td>VT1000P</td>
 <td>&nbsp;</td>
 </tr>
 <tr>
 <td>0.1mm FA-10 Feather S blade</td>
 <td>Ted Pella</td>
 <td>121-9</td>
 <td>0.1mm FA-10 Feather S blade</td>
 </tr>
 <tr>
 <td>Borosilicate Glass Pasteur Pipet (with rubber bulb)</td>
 <td>Fisher</td>
 <td>13-678-20A</td>
 <td>For transferring slices: Tip is broken off and heat-polished for larger opening</td>
 </tr>
 <tr>
 <td>35 mm Polysterine Culture dish</td>
 <td>Corning</td>
 <td>430588</td>
 <td>Used for collecting slices after dissection</td>
 </tr>
</tbody>
</table>
Table 2. Tools and materials for dissection
<table border="1">
<tbody>
 <tr>
 <td>Equipment/Materials</td>
 <td>Company</td>
 <td>Comments (optional)</td>
 </tr>
 <tr>
 <td>Holding chamber</td>
 <td>Custom Built</td>
 <td>&nbsp;</td>
 </tr>
 <tr>
 <td>P-97 Horizontal Pipette Puller</td>
 <td>Sutter Instrument Co.</td>
 <td>&nbsp;</td>
 </tr>
 <tr>
 <td>Vibration isolation table</td>
 <td>Technical Manufacturing Corporation (TMC)</td>
 <td>&nbsp;</td>
 </tr>
 <tr>
 <td>Faraday cage</td>
 <td>Custom built</td>
 <td>&nbsp;</td>
 </tr>
 <tr>
 <td>Pyrex Aspirator Bottle with Bottom Sidearm (Product #1220-1L)</td>
 <td>Corning</td>
 <td>&nbsp;</td>
 </tr>
 <tr>
 <td>Gravity-contolled IV set with regulator (Product # 2C8891)</td>
 <td>Baxter</td>
 <td>&nbsp;</td>
 </tr>
 <tr>
 <td>Central Vacuum Line</td>
 <td>&nbsp;</td>
 <td>Available in most modern labs</td>
 </tr>
 <tr>
 <td>95% O2 / 5% CO2 Gas Mix </td>
 <td>Scott-Gross Co.</td>
 <td>&nbsp;</td>
 </tr>
 <tr>
 <td>TygonTM Lab tubing 
 For O2/CO2 delivery</td>
 <td>Fisher Scientific</td>
 <td>Non-toxic, non-oxidizing, comes in a variety of sizes.</td>
 </tr>
 <tr>
 <td>Eclipse E600FN Microscope</td>
 <td>Nikon</td>
 <td>with 10x and 40x objectives, near infared filter, and GFP,DS-Red2 filters</td>
 </tr>
 <tr>
 <td>Cool Snap ES Digital Camera</td>
 <td>Photometrics</td>
 <td>Cool Snap ES Digital Camera</td>
 </tr>
 <tr>
 <td>X-Cite Fluorescent Illuminator</td>
 <td>EXFO</td>
 <td>X-Cite Fluorescent Illuminator</td>
 </tr>
 <tr>
 <td>Microscope Platform</td>
 <td>Siskiyou</td>
 <td>Custom assembled</td>
 </tr>
 <tr>
 <td>RC-22 Submersible recording chamber (Product # 64-0228)</td>
 <td>Warner Instruments (WI)</td>
 <td>Requires P-1 platform and stage adaptor (Product # 64-0277 From Warner)</td>
 </tr>
 <tr>
 <td>TC2BIP 2/3Ch Temperature controller </td>
 <td>Cell Microcontrols</td>
 <td>TC2BIP 2/3Ch Temperature controller </td>
 </tr>
 <tr>
 <td>4 Axis Manual Miniature manipulator</td>
 <td>Siskiyou</td>
 <td>&nbsp;</td>
 </tr>
 <tr>
 <td>Platinum Iridium Wire (0.002 in) 
 (item # PTT0203)</td>
 <td>WPI</td>
 <td>&nbsp;</td>
 </tr>
 <tr>
 <td>A365 Stimulus Isolator</td>
 <td>WPI</td>
 <td>A365 Stimulus Isolator</td>
 </tr>
 <tr>
 <td>Multiclamp 700b Amplifier</td>
 <td>Axon Instruments</td>
 <td>&nbsp;</td>
 </tr>
 <tr>
 <td>Digidata 1322A A/D converter</td>
 <td>Axon Instruments</td>
 <td>&nbsp;</td>
 </tr>
 <tr>
 <td>PClamp software</td>
 <td>Axon Instruments</td>
 <td>&nbsp;</td>
 </tr>
 <tr>
 <td>Personal Computer (Pentium 4)</td>
 <td>Dell</td>
 <td>&nbsp;</td>
 </tr>
</tbody>
</table>
Table 3. Electrophysiology equipment and materials

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

The rodent hippocampal slice preparation is perhaps the most broadly used tool for investigating mammalian synaptic function and plasticity. The hippocampus can be extracted quickly and easily from rats and mice and slices remain viable for hours in oxygenated artificial cerebrospinal fluid. Moreover, basic electrophysisologic techniques are easily applied to the investigation of synaptic function in hippocampal slices and have provided some of the best biomarkers for cognitive impairments. The hippocampal slice is especially popular for the study of synaptic plasticity mechanisms involved in learning and memory. Changes in the induction of long-term potentiation and depression (LTP and LTD) of synaptic efficacy in hippocampal slices (or lack thereof) are frequently used to describe the neurologic phenotype of cognitively-impaired animals and/or to evaluate the mechanism of action of nootropic compounds. This article outlines the procedures we use for preparing hippocampal slices from rats and transgenic mice for the study of synaptic alterations associated with brain aging and Alzheimer's disease (AD)1-3. Use of aged rats and AD model mice can present a unique set of challenges to researchers accustomed to using younger rats and/or mice in their research. Aged rats have thicker skulls and tougher connective tissue than younger rats and mice, which can delay brain extraction and/or dissection and consequently negate or exaggerate real age-differences in synaptic function and plasticity. Aging and amyloid pathology may also exacerbate hippocampal damage sustained during the dissection procedure, again complicating any inferences drawn from physiologic assessment. Here, we discuss the steps taken during the dissection procedure to minimize these problems. Examples of synaptic responses acquired in "healthy" and "unhealthy" slices from rats and mice are provided, as well as representative synaptic plasticity experiments. The possible impact of other methodological factors on synaptic function in these animal models (e.g. recording solution components, stimulation parameters) are also discussed. While the focus of this article is on the use of aged rats and transgenic mice, novices to slice physiology should find enough detail here to get started on their own studies, using a variety of rodent models.

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

This article outlines procedures for preparing hippocampal slices from rats and transgenic mice for the study of synaptic alterations associated with brain aging and age-related neurodegenerative diseases, such as Alzheimer&#x2019;s disease.

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

Research

JoVE Journal

Neuroscience

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

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

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

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Preparation of Acute Subventricular Zone Slices for Calcium Imaging

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

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Stereotaxic Infusion of Oligomeric Amyloid-beta into the Mouse Hippocampus

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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 ...

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

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 ...

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 ...

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 ...

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