Die vorliegende Studie wurde vom National Science Council gewährt Dr. Bai-Chuang Shyu (NSC 99-2320-B-001-016-MY3, NSC 100-2311-B-001-003-MY3 und NSC unterstützt 102-2320- B-001-026-MY3). Diese Arbeit wurde am Institut für Biomedizinische Wissenschaften durchgeführt, die Finanzierung von Academia Sinica erhalten.

Das Protokoll in der vorliegenden Studie erhielt die Genehmigung von der Academia Sinica Institutional Animal Care and Utilization Ausschuss in Taiwan. 1. Tierpräparate <ol><li> Erhalten männliche Sprague-Dawley-Ratten (etwa 300-400 g). Pflegen Sie die Ratten in einem klimatisierten Raum (21 - 22 ° C, 50% Luftfeuchtigkeit) unter einem 12 h / 12 h Hell / Dunkel-Zyklus (Licht um 8:00 Uhr auf) mit freiem Zugang zu Nahrung und Wasser. </li></ol> 2. Experimentelles Verfahren <ol><li> Lassen Sie all die Ratten in die Umwelt in ihre Käfige für 1 Woche vor den Experimenten anzupassen. Während der Adaption, führen Sie die von Frey und Plantar-Tests Basislinien zu etablieren. </li><li> von Frey - Test <ol><li> Legen Sie die Ratte in einem Acryl-Gehäuse (30 cm × 30 cm × 80 cm) für 30 min für Wohnzwecke. </li><li> Erhalten von Frey Filamenten (Filament-Nr. 11 - 20), die die gleiche Länge haben, aber Durchmesser variierende einen Bereich zu schaffen,der Kräfte von 2-100 g. </li><li> Um die Pfotenrückzugsschwelle in der Hinterpfote einer Ratte beurteilen zu können, verwenden von Frey Filamenten das Zentrum der hindpaws durch einen netzartigen Anschluss auf der Acrylplatte bei 5 min intertrial Abständen zu stimulieren. Um den maximalen ausgeübten Druck aufnehmen, verwenden Sie die Fäden in aufsteigender Reihenfolge von niedrig bis hoch, bis die maximale angelegte Druck aufgezeichnet wird. 10 </li><li> Bei Ratten, die eine Pfote Rückzug Reaktion auf die Stimulation zeigen, notieren Sie die Glühfaden-Nummer. Bestimmen Sie die Entzugserscheinungen, die in dieser Studie entsprechend dem niedrigsten Reiz. </li><li> Wiederholen der gleichen Prozedur sofort für insgesamt dreimal hintereinander auf der gleichen Ratte. Konvertieren Sie die Glühfaden-Nummer in die entsprechende Kraft (in Gramm) und die Werte im Durchschnitt. </li></ol></li><li> Plantar - Test <ol><li> Legen Sie die Ratte in einem transparenten Plexiglas-Box (unterteilt in vier Rahmen, 80 cm × 30 cm × 15 cm) für 30 min für Wohnzwecke. </li><li> Verwenden Sie ein infrared Strahl die Mitte der Hinterpfote durch eine Glasplatte zu stimulieren. Stellen Sie die Infrarot-Lichtintensität eine durchschnittliche Pfotenrückzugsantwortlatenzzeit von etwa 10 Sekunden zu erhalten. Führen Sie eine Studie durch Drücken einer Taste, die auf der Infrarot-Lichtquelle dreht sich um und startet einen digitalen Solid-State-Timer. Manipulieren die Dauer des Infrarot-Lichtstrahl. </li><li> Notieren Sie sich die Dauer des Infrarotlichts, wenn die Ratten eine Pfotenrückzugsreaktion zeigen. Die längste Dauer sollte nicht mehr als 20 Sekunden in jedem Versuch Gewebeschäden zu vermeiden. Verwenden Sie ein intertrial Abstand von mindestens 5 min sukzessive Stimulation zu vermeiden. </li><li> Den Test mit drei Versuche für die linken und rechten Hinterpfoten, und berechnet den Mittelwert für jede Hinterpfote für jede Ratte. </li></ol></li><li> Kollagenase Lesion Chirurgie <ol><li> Anesthetize die Ratte mit 4% Isofluran bis Verlust der Zehen Prise Reaktion und somatischen Reaktionen auf chirurgische Stimuli auftreten. Pflegen Anästhesie mit 1,5-2% isoflurane für die Dauer der Operation. </li><li> Legen Sie die Ratte in einem stereotaktischen Gerät mit einem einfachen Heizkissen auf Körpertemperatur bei 36,5 aufrechtzuerhalten - 37,5 ° C. Tragen Sie Augencreme auf die Augen Trockenheit während der Narkose zu verhindern. </li><li> Shave das Fell mit sterilisiertem elektrischen Haarschneider und einen glatten Schnitt zu machen (ca. 2-2,5 cm) mit einem Skalpell entlang der Mittellinie der Kopfhaut. Reinigen Sie die Haut und Schädel mit wechselnden Vorsehung Jod und 75% Alkohollösung zur Desinfektion. Während der chirurgischen Phase, verwenden 75% Alkohol alle Geräte, Werkzeuge zu sterilisieren, und die Werkbank sterilen Bedingungen zu halten. Vor, dass alle chirurgischen Materialien müssen in einem Dampfautoklaven sterilisiert werden. </li><li> Verwenden Sie die sterile Handschuhe und Instrumente, und bohren Sie ein kleines Loch (3 mm Durchmesser) in den Schädel mit einer elektrischen Bohrmaschine über die VB (einschließlich ventralen posteromedial Thalamuskern [VPM] und ventralen posterolateral Thalamuskern [VPL]) des Thalamus (3,0 -3,5 mm posterior, 3,0-3,5 mm lateralzu Bregma und von Schädeloberfläche 5,5-5,8 mm Tiefe). </li><li> Microinject 0,5 &amp; mgr; l normaler Salzlösung oder 0.125 U des Typs 4 Kollagenase-Lösung in den Kontroll- und Testgruppen, respectively. </li><li> Halten Sie die Injektionsnadel an Ort und Stelle für weitere 5 min für Arzneimitteldiffusion zu ermöglichen. </li><li> Verwenden Sie Zahnzement das Loch in den Schädel zu füllen, und die Inzision vernähen. Nach Naht der Einschnitt, gelten die lokalen analgetischen Betäubungsmittel (Lidocain-Salbe) auf die Wunde, und bringt sie in ihre Heimatkäfige. </li><li> Nach der Operation Haus einzeln die Ratten in Plastikkäfigen, bis sie Brustlage halten und warm hält von der Anästhesie erholt, bis. </li></ol></li><li> Verhaltenstests nach der chirurgischen Wiederherstellung <ol><li> Nach 7 Tagen der Genesung von der Operation, wiederholen Sie die Prozeduren in den Abschnitten 2.2 und 2.3 für die Testphase. Führen Sie die Verhaltenstests über 4 Wochen, während die Gesundheitsüberwachung und Entwicklungsstand der Tiere. </li><li&gt; Monitor den Gesundheitszustand der Tiere (zB Körpergewicht, Futtermenge und Freizügigkeit) zwischen der Kontroll- und Versuchsgruppen für die postoperative Phase. </li></ol></li><li> Führen Sie Cannula Implantierung mit PE-50 - Schläuche <ol><li> Anästhesieren die Ratte mit 4% Isofluran und Aufrechterhaltung der Körpertemperatur bei 36,5 bis 37,5 ° C mit einer einfachen Heizkissen. </li><li> Mit einem Skalpell geschnitten, um zwei Löcher (2 cm Durchmesser jeweils) in der Mittellinie des Rückenteils der vorderen Gliedmaßen und Schnitt des ventralen Teil der linken Schulter und Brusthöhle sind. </li><li> Dissoziieren die Haut und Muskel mit einer Schere zwischen den beiden Löchern. </li><li> Ein Ende des PE- 50-Schlauch (20 cm Länge) an die äußere Jugularvene durch das ventrale Loch. Schließen Sie das Anschlussende des gleichen PE- 50 Schlauch mit dem Rückenloch und bringt an der Haut. </li><li> Verwenden Sie eine Spritze Kochsalzlösung in die Halsvene zu injizieren, um sicherzustellen, dass die PE- 50 Schlauchnicht behindert wird. </li><li> Nähen Sie den Schnitt, und injizieren die Ratten mit 6 mg / kg Gentamicin (intraperitoneal). </li><li> Spülen Sie den Schlauch jeden zweiten Tag nach der Operation mit 0,3 ml 0,9% Kochsalzlösung, gefolgt von 0,1 ml Kochsalzlösung mit 20 U / ml Heparin. </li></ol></li><li> Schlussverhaltenstest <ol><li> Eine Woche nach dem Schritt 2.6, wiederholen Sie die Schritte 2,2-2,3 dieses Verhalten zu bestätigen ist stabil im Vergleich zu Schritt 2.5 nach der chirurgischen Wiederherstellung. </li></ol></li><li> Radiotracer - Injection <ol><li> Legen Sie die Ratte in einer Ruhe Käfig 5 - 10 min für die Anpassung. </li><li> Mit einem Splitter verbinden PE- 50 Schlauch mit zwei 1 ml Spritzen. Füllen einer Spritze mit normaler Kochsalzlösung und füllt das andere mit [14 C] -IAP Lösung (125 uCi / kg in einem Volumen von 0,3-0,5 ml). </li><li> Injizieren Sie die Radiotracer in die externe Halsvene, und ersetzen Sie die Spritze mit einer anderen Spritze mit 3 M Kaliumchlorid gefüllt. </li><li> Zehn Sekunden nach dem Radiotracer injection, injizieren 3 M Kaliumchlorid unter einer Überdosierung von Isofluran und opfern , um die Tiere nach Standard Euthanasie Methoden (dh Isofluran aus einem Verdampfer für 50 min) , basierend auf den Richtlinien der Academia Sinica Institutional Animal Care and Utilization Ausschuss in Taiwan. </li><li> Nach 1 min, setzen den Schädel, und schneiden Sie die verbleibenden Muskel ab. Mit rongeurs, ziehen Sie die dorsale Oberfläche des Schädels aus dem Gehirn entfernt. Schneiden Sie die Seiten des Schädels mit rongeurs. Als nächstes wird mit einem Spatel, schneiden Sie die Riechkolben und Nervenverbindungen entlang der ventralen Oberfläche des Gehirns, und das Gehirn zu entfernen. </li><li> Verwenden Sie optimale Schnitttemperatur (OCT) -Verbindung das Gehirn in Trockeneis einzufrieren und Methylbutan (ca. -55 ° C). Lagern Sie das Hirngewebe in einem Gefrierschrank. 11, 12 </li></ol></li><li> Gehirn - Slicing <ol><li> Richten Sie das Gewebe in einem Mikrotom, mit dem Rautenhirns nach unten zeigt. Verwenden Sie ein Kryostat das Gehirn in 20 &amp; zu schneiden# 181; m-dicke Abschnitte. </li><li> Platzieren Sie die Hirnschnitten auf Objektträgern in einem Kryostaten bei -20 ° C, mit einem 240 um Intervall zwischen jeder Scheibe. </li><li> Legen Sie die Objektträger und fünf Standard-Filterpapiere mit abgestuften Radioaktivität in Belichtungskassetten für 3 Tage bei -20 ° C. Entsprechend der Reihenfolge der Gehirnscheiben, die Mikroskop-Objektträger von oben nach unten anordnen. Schließlich legen Sie die Filterpapiere in der Unterseite der Kassetten. 12 </li><li> Entfernen Sie den Phosphorschirm von den Belichtungskassetten und eine Variable-Modus Imager den Phosphorschirm und erzeugen Bilder zu lesen , um zu zeigen , [14 C] -IAP Aufnahme für die Hirnschnitten. </li></ol></li></ol> 3. Datenanalyse <ol><li> Nach dem Schritt 2.9.4, passen Sie die Bilder Statistische Parametric Mapping (SPM) und ImageJ-Software. Rekonstruieren alle Bilder seriellen Koronalschnitte verwenden. Glatte und normalisieren die Bilder gemäß einem Referenzrattenhirnmodell.12, 13 </li><li> Für die quantitative Einschätzungen, messen Sie die Region-of-Interest (ROI) des Gehirns Bilder mit ImageJ Software die Pixelsignalintensität, um zu bestimmen , und statistische Software für die Analyse verwendet werden . 12, 13 </li><li> Um die Verbindungen zwischen verschiedenen Hirnkernen zu untersuchen, verwenden MATLAB Korrelationsanalyse-Software Radioaktivität Verhältnis in einer überregionalen Korrelationsmatrix anzuzeigen, und visualisieren die Matrizen als Farbkarten. Schließlich verwenden für Netzwerkanalyse Pajek Software. 12, 13 </li><li> Verwenden Sie 2 x 5 Zwei-Wege-Mischvarianzanalyse (ANOVA), mit Gruppen- und Wochen als Faktoren, die Dauer der Hitzetoleranz in der Plantar-Test und mechanische Kraft in der von-Frey-Test in den Schein und CPSP Gruppen zu Beginn der Studie zu vergleichen und Woche 1 - 5. Mit einem 2 × 31 zwei-Wege-ANOVA die Radioaktivität Verhältnis gemäß Gruppe und Gehirnbereich zu messen. Wenn es angebracht ist , führen Tukey ehrlich signifikante Differenz (HSD) Post - hoc - Tests. Calculate Pearson Korrelationskoeffizienten Korrelationen zwischen allen ausgewählten Gehirnbereiche in der Sham und CPSP Gruppen zu bewerten. </li></ol>

<ol>
	<li>Finnerup, N. B. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=A+review+of+central+neuropathic+pain+states.">A review of central neuropathic pain states.</a> Curr Opin Anaesthesiol. 21 (5), 586-589 (2008).</li><li>Andersen, G., Vestergaard, K., Ingeman-Nielsen, M., Jensen, T. S. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Incidence+of+central+post-stroke+pain.">Incidence of central post-stroke pain.</a> Pain. 61 (2), 187-193 (1995).</li><li>Chen, B., Stitik, T. P., Foye, P. M., Nadler, S. F., DeLisa, J. A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Central+post-stroke+pain+syndrome:+yet+another+use+for+gabapentin?.">Central post-stroke pain syndrome: yet another use for gabapentin?.</a> Am J Phys Med Rehabil. 81 (9), 718-720 (2002).</li><li>Greenspan, J. D., Ohara, S., Sarlani, E., Lenz, F. A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Allodynia+in+patients+with+post-stroke+central+pain+(CPSP)+studied+by+statistical+quantitative+sensory+testing+within+individuals.">Allodynia in patients with post-stroke central pain (CPSP) studied by statistical quantitative sensory testing within individuals.</a> Pain. 109 (3), 357-366 (2004).</li><li>Klit, H., Finnerup, N. B., Jensen, T. S. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Central+post-stroke+pain:+clinical+characteristics,+pathophysiology,+and+management.">Central post-stroke pain: clinical characteristics, pathophysiology, and management.</a> Lancet Neurol. 8 (9), 857-868 (2009).</li><li>Kumar, G., Soni, C. R. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Central+post-stroke+pain:+current+evidence.">Central post-stroke pain: current evidence.</a> J Neurol Sci. 284 (1-2), 10-17 (2009).</li><li>Denk, F., McMahon, S. B., Tracey, I. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Pain+vulnerability:+a+neurobiological+perspective.">Pain vulnerability: a neurobiological perspective.</a> Nat Neurosci. 17 (2), 192-200 (2014).</li><li>Ji, G., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Cognitive+impairment+in+pain+through+amygdala-driven+prefrontal+cortical+deactivation.">Cognitive impairment in pain through amygdala-driven prefrontal cortical deactivation.</a> J Neurosci. 30 (15), 5451-5464 (2010).</li><li>Jungehulsing, G. J., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Levetiracetam+in+patients+with+central+neuropathic+post-stroke+pain:+a+randomized,+double-blind,+placebo-controlled+trial.">Levetiracetam in patients with central neuropathic post-stroke pain: a randomized, double-blind, placebo-controlled trial.</a> Eur J Neurol. 20 (2), 331-337 (2013).</li><li>Chaplan, S. R., Bach, F. W., Pogrel, J. W., Chung, J. M., Yaksh, T. L. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Quantitative+assessment+of+tactile+allodynia+in+the+rat+paw.">Quantitative assessment of tactile allodynia in the rat paw.</a> J Neurosci Methods. 53, 55-63 (1994).</li><li>Jay, T. M., Luciqnani, G., Crane, A. M., Jehle, J., Sokoloff, L. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Measurement+of+local+cerebral+blood+flow+with+[14C]iodoantipyrine+in+the+mouse.">Measurement of local cerebral blood flow with [14C]iodoantipyrine in the mouse.</a> J Cereb Blood Flow Metab. 8 (1), 121-129 (1988).</li><li>Lu, H. C., Chang, W. J., Kuan, Y. H., Huang, A. C., Shyu, B. C. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=A+[14C]iodoantipyrine+study+of+inter-regional+correlations+of+neural+substrates+following+central+post-stroke+pain+in+rats.">A [14C]iodoantipyrine study of inter-regional correlations of neural substrates following central post-stroke pain in rats.</a> Mol Pain. 11 (1), (2015).</li><li>Wang, Z., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Functional+brain+activation+during+retrieval+of+visceral+pain-conditioned+passive+avoidance+in+the+rat.">Functional brain activation during retrieval of visceral pain-conditioned passive avoidance in the rat.</a> Pain. 152, 2746-2756 (2011).</li><li>Wasserman, J. K., Koeberle, P. D. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Development+and+characterization+of+a+hemorrhagic+rat+model+of+central+post-stroke+pain.">Development and characterization of a hemorrhagic rat model of central post-stroke pain.</a> Neuroscience. 161 (1), 173-183 (2009).</li></ol>

Die Autoren haben nichts zu offenbaren.

In den Verhaltenstests zeigten die CPSP Gruppe Reduzierungen des Pfotenrückzugsschwelle bei der thermischen Schmerztest und mechanische Kraft in der Prüfung von Frey zu Beginn der Studie und Wochen . 1 - 5. Die Ergebnisse mit einer früheren Studie waren konsistent 14 Die [14 C] -IAP Verfahren beruht auf der Pixel - Intensität von Gehirnbildern für die quantitative Analyse von Gehirnschnitten. Um die Daten in den Gehirnbilder auszuwerten, wurde die Pixelsignalintens...

Schlaganfall Blutung wurde in mehr als 8% der Patienten auftreten gezeigt , die unter neuropathischen Schmerzen leiden, die als zentrale nach Schlaganfall Schmerz (CPSP). 1-3 CPSP von somatosensorischen Dysfunktion führen kann, wodurch die Induktion Überempfindlichkeits und Allodynie. 4 jedoch bleiben die pathophysiologischen Mechanismen der somatosensorischen Dysfunktion in CPSP unsicher. Beispielsweise führt der Verlust von Körperempfindungen von neuronalen Deafferentation im hämorrhagischen Hirnareals. Hyperalgesie kann durch die Übererregbarkeit des zentralen nozizeptiven Neuronen oder zentrale Enthemmung, 5, 6 , aber die neuronalen Substrate verursacht werden , die in CPSP Symptome beteiligt sind , bleiben unbekannt. Einige Studien haben vorgeschlagen , dass die dorsolateralen präfrontalen Kortex (DPFC), rostralen anterioren cingulären Cortex (ACC), der Amygdala, Hippocampus, periaqueductal grau (PAG), rostral ventromedialen Medulla und ihre Verbindungen miteinander nozizeptiven Verarbeitung vermitteln. 7 Zusätzlichein Schmerzen im Zusammenhang mit der Wahrnehmung beteiligt. 8 Daten über die pathophysiologischen Mechanismen von CPSP sind vielfältig, und die Aktivierung der neuronalen Substrate in CPSP muss eine weitere Prüfung ly wurden medialen präfrontalen Kortex (mPFC) -amygdala Schaltungen gezeigt. [14 C] -Iodoantipyrine (IAP) Aufnahme wird verwendet , indirekt zu beobachten , regionale Hirndurchblutung (rCBF), die eine Beziehung zwischen Gehirnaktivität und CBF übernimmt. Obwohl [14 C] -IAP nicht die Gehirnaktivität in Echtzeit, wie zum Beispiel mit der funktionellen Magnetresonanztomographie (fMRI) beurteilen kann, hat es mehrere Vorteile. Zum Beispiel [14 C] -IAP ist zur Messung von spontan auftretenden Gehirn Ereignisse während pathologischen Zuständen geeignet. 9 Außerdem [14 C] -IAP Aufnahme ohne Anästhesie gemessen wird. Es kostet auch weniger als andere Abbildungsverfahren, einschließlich fMRI und Positronen-Emissions-Tomographie (PET). Die [14 C] -IAP Verfahren ist vorgeschlagen worden , für measurin geeignet seing spontane Schmerzen (zB CPSP) , die durch Läsionen des ventralen Nucleus basalis (VB) des Thalamus induziert wird. 9 Das vorliegende Protokoll beschreibt , wie die [14 C] -IAP Verfahren auszuführen , um die Beteiligung von neuronalen Substrate CPSP zu bewerten , die durch Läsionen der VB des Thalamus in einem Tiermodell induziert wird. Die Technik bietet eine Möglichkeit, die pathophysiologischen Mechanismen der Bestimmung, dass CPSP Symptome an den Verhaltens- und neuronale Ebenen zugrunde liegen.

<table border="0" cellpadding="0" cellspacing="0" width="785">
	<tbody>
		<tr height="21">
			<td colspan="4" height="21" style="height:21px;width:785px;">Anesthetic:</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:216px;">Isoflurane</td>
			<td style="width:229px;">Halocarbon Products Corporation&#160;</td>
			<td style="width:173px;">NDC 12164-002-25</td>
			<td style="width:167px;">4%</td>
		</tr>
		<tr height="21">
			<td colspan="4" height="21" style="height:21px;width:785px;">Surgery</td>
		</tr>
		<tr height="63">
			<td height="63" style="height:63px;width:216px;">homeothermic blanket system</td>
			<td style="width:229px;">Harvard Apparatus</td>
			<td style="width:173px;">Model 50&#8211;7079</td>
			<td style="width:167px;">body temperature were maintained at 36.5-37.5&#176;C.</td>
		</tr>
		<tr height="42">
			<td height="42" style="height:42px;width:216px;">10&#181;l micro syringe</td>
			<td style="width:229px;">Hamilton</td>
			<td style="width:173px;">80008, Model 1701SN</td>
			<td style="width:167px;">injected with collagenase</td>
		</tr>
		<tr height="42">
			<td height="42" style="height:42px;width:216px;">polyethylene-50 tubing</td>
			<td style="width:229px;">Becton, Dickinson and Company</td>
			<td style="width:173px;">427411</td>
			<td style="width:167px;">catheterized into external jugular vein</td>
		</tr>
		<tr height="45">
			<td height="45" style="height:45px;width:216px;">1 c.c syringe</td>
			<td style="width:229px;">Terumo Medical Products</td>
			<td style="width:173px;">SS-01T</td>
			<td style="width:167px;">injected with 14C-IAP and saline.</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:216px;">saline (Sodium Chloride 0.9gm)</td>
			<td style="width:229px;">Taiwan Biotech Co., LTD.</td>
			<td style="width:173px;">100-130-0201</td>
			<td style="width:167px;">To flush the tube</td>
		</tr>
		<tr height="21">
			<td colspan="4" height="21" style="height:21px;width:785px;">Drugs:</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:216px;">type 4 collagenase</td>
			<td style="width:229px;">Sigma</td>
			<td style="width:173px;">C5138-500MG</td>
			<td style="width:167px;">0.125 U</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:216px;">Gentamicin</td>
			<td style="width:229px;">Sigma</td>
			<td style="width:173px;">G1264-250MG</td>
			<td style="width:167px;">6 mg/kg</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:216px;">Heparin</td>
			<td style="width:229px;">Sigma</td>
			<td style="width:173px;">H9399</td>
			<td style="width:167px;">20 U/ml; 0.1 ml/day</td>
		</tr>
		<tr height="42">
			<td height="42" style="height:42px;width:216px;">14C-iodoantipyrine (IAP)</td>
			<td style="width:229px;">PerkinElmer</td>
			<td style="width:173px;">NEC712</td>
			<td style="width:167px;">125 mCi/kg in 300 ml of 0.9% saline</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:216px;">Potassium chloride</td>
			<td style="width:229px;">Merck</td>
			<td style="width:173px;">1.04936.1000</td>
			<td style="width:167px;">3 M</td>
		</tr>
		<tr height="21">
			<td colspan="4" height="21" style="height:21px;width:785px;">Behavior system:</td>
		</tr>
		<tr height="126">
			<td height="126" style="height:126px;width:216px;">von Frey esthesiometer</td>
			<td style="width:229px;">Fabrication Enterprises, Inc.</td>
			<td style="width:173px;">Baseline Tactile Monofilaments 12-1666</td>
			<td style="width:167px;">mechanical hyperalgesia was assessed by measuring the withdrawal response to a mechanical stimulus</td>
		</tr>
		<tr height="126">
			<td height="126" style="height:126px;width:216px;">plantar test apparatus</td>
			<td style="width:229px;">IITC Life Science</td>
			<td style="width:173px;">IITC 390G Plantar Test</td>
			<td style="width:167px;">Thermal hyperalgesia was assessed by measuring the hind paw withdrawal latency in response to radiant heat.</td>
		</tr>
		<tr height="21">
			<td colspan="4" height="21" style="height:21px;width:785px;">Brain slice:</td>
		</tr>
		<tr height="42">
			<td height="42" style="height:42px;width:216px;">Optimal Cutting Temperature compound</td>
			<td style="width:229px;">Sakura Fintek Inc</td>
			<td style="width:173px;">4583</td>
			<td style="width:167px;">embedded the brain</td>
		</tr>
		<tr height="63">
			<td height="63" style="height:63px;width:216px;">dry ice</td>
			<td style="width:229px;"></td>
			<td style="width:173px;"></td>
			<td style="width:167px;">frozen in dry ice/methylbutane (approximately &#8722;55&#176;C)</td>
		</tr>
		<tr height="63">
			<td height="63" style="height:63px;width:216px;">methylbutane</td>
			<td style="width:229px;">Sigma</td>
			<td style="width:173px;">M32631-1L</td>
			<td style="width:167px;">frozen in dry ice/methylbutane (approximately &#8722;55&#176;C)</td>
		</tr>
		<tr height="63">
			<td height="63" style="height:63px;width:216px;">Cryostat</td>
			<td style="width:229px;">&#160;Leica Biosystems Nussloch GmbH, Germany</td>
			<td style="width:173px;">Leica CM1850</td>
			<td style="width:167px;">Coronal brain slice were sectioned on this machine.</td>
		</tr>
		<tr height="21">
			<td colspan="4" height="21" style="height:21px;width:785px;">Data analyze</td>
		</tr>
		<tr height="168">
			<td height="168" style="height:168px;width:216px;">exposure cassettes with a phosphor screen</td>
			<td style="width:229px;">Amersham Biosciences&#160;</td>
			<td style="width:173px;">20 cm x 25 cm</td>
			<td style="width:167px;">The slices were dried on glass slides and placed alongside five standard filter papers with graded radioactivity. All of the slides were exposed to the cassettes at &#8722;20&#176;C.</td>
		</tr>
		<tr height="63">
			<td height="63" style="height:63px;width:216px;">&#947;-counter</td>
			<td style="width:229px;">Beckman Coulter</td>
			<td style="width:173px;">Beckman LS 6500 Liquid Scintillation Counter</td>
			<td style="width:167px;">To measure the radioactivity count of the filter papers.</td>
		</tr>
		<tr height="63">
			<td height="63" style="height:63px;width:216px;">Typhoon 9410 Variable Mode Imager</td>
			<td style="width:229px;">&#160;GMI, Inc.</td>
			<td style="width:173px;">WS-S9410</td>
			<td style="width:167px;">To read&#160; phosphor screen which was exposed by brain slice</td>
		</tr>
		<tr height="231">
			<td height="231" style="height:231px;width:216px;">Statistical Parametric Mapping (SPM)</td>
			<td style="width:229px;">Wellcome Centre for Neuroimaging</td>
			<td style="width:173px;">version 8</td>
			<td style="width:167px;">all of the brains were averaged to create the final brain template. To determine significant differences between the images in these two groups, the images were derived by subtracting the sham group from the CPSP group.</td>
		</tr>
		<tr height="294">
			<td height="294" style="height:294px;width:216px;">ImageJ</td>
			<td style="width:229px;">http://imagej.nih.gov/ij</td>
			<td style="width:173px;">version 1.46</td>
			<td style="width:167px;">Adjacent sections were aligned both manually and using Stack- Reg, an automated pixel-based registration algorithm in ImageJ software. All of the original three-dimensionally reconstructed brains were smoothed and normalized to the reference rat brain model.</td>
		</tr>
		<tr height="210">
			<td height="210" style="height:210px;width:216px;">Matlab</td>
			<td style="width:229px;">MathWorks</td>
			<td style="width:173px;">version 2009b</td>
			<td style="width:167px;">used Pearson correlation coefficients to examine the relationships between the CPSP and sham groups. An inter-regional correlation matrix was calculated across animals from each group.</td>
		</tr>
		<tr height="126">
			<td height="126" style="height:126px;width:216px;">Pajek</td>
			<td style="width:229px;">http://Pajek.imfm.si/</td>
			<td style="width:173px;">version 3.06</td>
			<td style="width:167px;">Graphical theoretical analysis was performed on networks defined by the above correlation matrices using Pajek software.</td>
		</tr>
	</tbody>
</table>

autoradiographic measurements of [14c]-iodoantipyrine in rat brain following central post-stroke pain

Approximately 8% of stroke patients present symptoms of central post-stroke pain (CPSP). CPSP is associated with allodynia and hypersensitivity to nociceptive stimuli. Although some studies have shown that neuropathic pain may involve the dorsolateral prefrontal cortex, rostral anterior cingulate cortex, amygdala, hippocampus, periaqueductal gray, rostral ventromedial medulla, and medial thalamus, the neural substrates and their connections that mediate CPSP remain unclear. [14C]-Iodoantipyrine (IAP) uptake can be measured to evaluate spontaneously active pain. It can be used to assess the activation of neural substrates that may be involved in CPSP in an animal model. The [14C]-IAP method in rats is less expensive to perform compared with other brain mapping techniques. The present [14C]-IAP protocol is used to measure the activation of neural substrates that are involved in CPSP that is induced by lesions of the ventral basal nucleus (VB) of the thalamus in a rodent model.

1A zeigt die experimentelle Zeitleiste. Die Ratten wurden auf den Schein und CPSP Gruppen für die Verhaltenstests (dh von Frey - Test und Plantar - Test) zugeordnet. Am ersten Tag des Experiments diente als Basislinie, und Tests in den Wochen wurden wiederholt, 1 - 5 PE-50-Katheterisierung wurde die Vena jugularis externa in Woche 4 durchgeführt Heparin (20 U / ml, 0,1 ml / Tag) während injiziert wurde Woche 4 und 5 Fünf Minuten nach der Heparin - Injektion,...

Watch this Scientific Journal Video about Autoradiographic Measurements of [14C]-Iodoantipyrine in Rat Brain Following Central Post-Stroke Pain at JoVE.com

Autoradiographic Measurements of [14C]-Iodoantipyrine in Rat Brain Following Central Post-Stroke Pain

We present a protocol to measure [14C]-iodoantipyrine (IAP) uptake and assess the activation of neural substrates that are involved in central post-stroke pain (CPSP) in a rodent model.

Autoradiographiebild Messungen von [<sup&gt; 14</sup&gt; C] -Iodoantipyrine in Rattengehirn Nach Central Post-Stroke Schmerz

Approximately 8% of stroke patients present symptoms of central post-stroke pain (CPSP). CPSP is associated with allodynia and hypersensitivity to ...

autoradiographic-measurements-14c-iodoantipyrine-rat-brain-following

Research

JoVE Journal

Medicine

Autoradiographic Measurements of [14C]-Iodoantipyrine in Rat Brain Following Central Post-Stroke Pain

8.7K Aufrufe.  Fo Guang University. We present a protocol to measure [14C]-iodoantipyrine (IAP) uptake and assess the activation of neural substrates that are involved in central post-stroke pain (CPSP) in a rodent model.

Serie: Autoradiographic Measurements of [14C]-Iodoantipyrine in Rat Brain Following Central Post-Stroke Pain

In vitro Functional Characterization of Mouse Colorectal Afferent Endings

This video demonstrates in detail an in vitro single-fiber electrophysiological recording protocol using a mouse colorectum-nerve preparation. The approach allows unbiased identification and functional characterization of individual colorectal afferents. Extracellular recordings of propagated action potentials (APs) that originate from one or a few afferent (i.e., single-fiber) receptive fields (RFs) in the colorectum are made from teased nerve fiber fascicles. The colorectum is removed with either the pelvic (PN) or lumbar splanchnic (LSN) nerve attached and opened longitudinally. The tissue is placed in a recording chamber, pinned flat and perfused with oxygenated Krebs solution. Focal electrical stimulation is used to locate the colorectal afferent endings, which are further tested by three distinct mechanical stimuli (blunt probing, mucosal stroking and circumferential stretch) to functionally categorize the afferents into five mechanosensitive classes. Endings responding to none of these mechanical stimuli are categorized as mechanically-insensitive afferents (MIAs). Both mechanosensitive and MIAs can be assessed for sensitization (i.e., enhanced response, reduced threshold, and/or acquisition of mechanosensitivity) by localized exposure of RFs to chemicals (e.g., inflammatory soup (IS), capsaicin, adenosine triphosphate (ATP)). We describe the equipment and colorectum&#8211;nerve recording preparation, harvest of colorectum with attached PN or LSN, identification of RFs in the colorectum, single-fiber recording from nerve fascicles, and localized application of chemicals to the RF. In addition, challenges of the preparation and application of standardized mechanical stimulation are also discussed.

In vitro Functional Characterization of Mouse Colorectal Afferent Endings

This video demonstrates a protocol for conducting single-fiber electrophysiological recordings on an in vitro mouse colorectum-nerve preparation.

In-vitro-Funktionelle Charakterisierung von Maus Colorectal Afferente Endings

This video demonstrates in detail an in vitro single-fiber electrophysiological recording protocol using a mouse colorectum-nerve preparation. The ...

Forschung

Medizin

The Monoiodoacetate Model of Osteoarthritis Pain in the Mouse

A major symptom of patients with osteoarthritis (OA) is pain that&#160;is triggered by peripheral as well as central changes within the pain pathways. The current treatments for OA pain such as NSAIDS or opiates are neither sufficiently effective nor devoid of detrimental side effects. Animal models of OA are being developed to improve our understanding of OA-related pain mechanisms and define novel pharmacological targets for therapy. Currently available models of OA in rodents include surgical and chemical interventions into one knee joint. The monoiodoacetate (MIA) model has become a standard for modelling joint disruption in OA in both rats and mice. The model, which is easier to perform in the rat, involves injection of MIA into a knee joint that induces rapid pain-like responses in the ipsilateral limb, the level of which can be controlled by injection of different doses. Intra-articular injection of MIA disrupts chondrocyte glycolysis by inhibiting glyceraldehyde-3-phosphatase dehydrogenase and results in chondrocyte death, neovascularization, subchondral bone necrosis and collapse, as well as inflammation. The morphological changes of the articular cartilage and bone disruption are reflective of some aspects of patient pathology. Along with joint damage, MIA injection induces referred mechanical sensitivity in the ipsilateral hind paw and weight bearing deficits that are measurable and quantifiable. These behavioral changes resemble some of the symptoms reported by the patient population, thereby validating the MIA injection in the knee as a useful and relevant pre-clinical model of OA pain.
The aim of this article is to describe the methodology of intra-articular injections of MIA and the behavioral recordings of the associated development of hypersensitivity with a mind to highlight the necessary steps to give consistent and reliable recordings.

Osteoarthritis (OA), or degenerative joint disease, is a debilitating condition associated with pain that remains only partially controlled by available analgesics. Animal models are being developed to improve our understanding of OA-related pain mechanisms. Here we describe the methodology for the monoiodoacetate model of OA pain in the mouse.

Das Monoiodoacetate Modell von Osteoarthritis Schmerzen in der Maus

A major symptom of patients with osteoarthritis (OA) is pain that&#160;is triggered by peripheral as well as central changes within the pain pathways. The ...

Patch Angioplasty in the Rat Aorta or Inferior Vena Cava

Pericardial patches are commonly used in vascular surgery to close vessels. To facilitate studies of the neointimal hyperplasia that forms on the patch, we developed a rat model of patch angioplasty that can be used in either a vein or an artery, creating a patch venoplasty or a patch arterioplasty, respectively. Technical aspects of this model are discussed. The infra-renal IVC or aorta are dissected and then clamped proximally and distally. A 3 mm venotomy or arteriotomy is performed in the infrarenal inferior vena cava or aorta of 6 to 8 week-old Wistar rats. A bovine pericardial patch (3 mm x 1.5 mm x 0.6 mm) is then used to close the site using a 10-0 nylon suture. Compared to arterial patches, venous patches show increased neointimal thickness on postoperative day 7. This novel model of pericardial patch angioplasty can be used to examine neointimal hyperplasia on vascular biomaterials, as well as to compare the differences between the arterial and venous environments.

We have established a model of pericardial patch angioplasty that can be used in either small-diameter veins or arteries. This model can be used to compare venous and arterial neointimal hyperplasia formation.

Patch-Angioplastie in der Ratte Aorta oder Vena Cava

Pericardial patches are commonly used in vascular surgery to close vessels. To facilitate studies of the neointimal hyperplasia that forms on the patch, ...

A Model of Free Tissue Transfer: The Rat Epigastric Free Flap

Free tissue transfer has been increasingly used in clinical practice since the 1970s, allowing reconstruction of complex and otherwise untreatable defects resulting from tumor extirpation, trauma, infections, malformations or burns. Free flaps are particularly useful for reconstructing highly complex anatomical regions, like those of the head and neck, the hand, the foot and the perineum. Moreover, basic and translational research in the area of free tissue transfer is of great clinical potential. Notwithstanding, surgical trainees and researchers are frequently deterred from using microsurgical models of tissue transfer, due to lack of information regarding the technical aspects involved in the operative procedures. The aim of this paper is to present the steps required to transfer a fasciocutaneous epigastric free flap to the neck in the rat.
This flap is based on the superficial epigastric artery and vein, which originates from and drain into the femoral artery and vein, respectively. On average the caliber of the superficial epigastric vein is 0.6 to 0.8 mm, contrasting with the 0.3 to 0.5 mm of the superficial epigastric artery. Histologically, the flap is a composite block of tissues, containing skin (epidermis and dermis), a layer of fat tissue (panniculus adiposus), a layer of striated muscle (panniculus carnosus), and a layer of loose areolar tissue.
Succinctly, the epigastric flap is raised on its pedicle vessels that are then anastomosed to the external jugular vein and to the carotid artery on the ventral surface of the rat's neck. According to our experience, this model guarantees the complete survival of approximately 70 to 80% of epigastric flaps transferred to the neck region. The flap can be evaluated whenever needed by visual inspection. Hence, the authors believe this is a good experimental model for microsurgical research and training.

This paper describes the steps required to raise a fasciocutaneous epigastric free flap and transfer it to the neck in the rat.

Ein Modell der freien Gewebetransfer: Die Ratte Epigastrische Freie Flap

Free tissue transfer has been increasingly used in clinical practice since the 1970s, allowing reconstruction of complex and otherwise untreatable defects ...

Reproducible Motor Deficit Following Aortic Occlusion in a Rat Model Of Spinal Cord Ischemia

Spinal cord ischemia is a fatal complication following thoracoabdominal aortic aneurysm surgery. Researchers can investigate the strategies for preventing and treating this complication using experimental models of spinal cord ischemia. The model described here demonstrates varying degrees of paraplegia that relate to the length of occlusion following thoracic aortic occlusion in a rat spinal cord ischemia model.
A 2-Fr. balloon-tipped catheter was advanced through the femoral artery into the descending thoracic aorta until the catheter tip was placed at the left subclavian artery in anesthetized male Sprague-Dawley rats. Spinal cord ischemia was induced by inflating the catheter balloon. After a set period of occlusion (9, 10, or 11 min), the balloon was deflated. Neurologic assessment was performed using the motor deficit index at 24 h after surgery, and the spinal cord was harvested for histopathological examination.
Rats that underwent 9 min of aortic occlusion showed mild and reversible motor impairment in the hind limb. Rats subjected to 10 min of aortic occlusion presented with moderate but reversible motor impairment. Rats subjected to 11 min of aortic occlusion displayed complete and persistent paralysis. The motor neurons in the spinal cord sections were more preserved in rats subjected to shorter duration of aortic occlusion.
Researchers can achieve a reproducible hind limb motor deficit following thoracic aortic occlusion using this spinal cord ischemia model.

This study demonstrates the technique to make a minimally invasive and easily reproducible model of spinal cord ischemia in rats. Various degrees of hind limb motor deficit can be produced by controlling the aortic occlusion time.

Reproduzierbare motorische Defizit nach Aortengelenk in einem Rattenmodell der Rückenmark Ischämie

Spinal cord ischemia is a fatal complication following thoracoabdominal aortic aneurysm surgery. Researchers can investigate the strategies for preventing ...

Spontaneous and Evoked Measures of Pain in Murine Models of Monoarticular Knee Pain

Pain is the main cause of disability from arthritis. There is currently an unmet need for adequate treatments for arthritis pain. Pre-clinical models are necessary and useful for studying the mechanisms of pain and for evaluating efficacy of arthritis therapies. Measuring pain in animal models of arthritis is challenging. We have developed methods for measuring evoked and spontaneous pain in three models of murine arthritis. We quantitate the evoked pain responses of mice subjected to firm palpation of a painful knee. We also evaluate spontaneous pain by the proportion of weight and the amount of time placed on each of their 4 limbs after induction of arthritis pain in one knee. Joint pain in these mouse models produces a significant increase in evoked pain responses and an alteration in weight bearing. Since mice are quadrupeds, they offload the painful limb to the contralateral limb, to the forelimbs, or some combination. These methods are simple, require minimal equipment, and are reproducible and sensitive for detecting pain. They are useful for studying both disease-modifying arthritis treatments and analgesics in mice.

We have developed an evoked measure of arthritis pain and coupled it with a standardized method for measuring spontaneous pain in different murine models of chemically induced arthritis. These measures are sensitive and reproducible for different types of joint pain.

Spontan und evozierten Maßnahmen des Schmerzes in murinen Modellen der monoartikulär Knieschmerzen

Pain is the main cause of disability from arthritis. There is currently an unmet need for adequate treatments for arthritis pain. Pre-clinical models are ...

A New Best Practice for Validating Tail Vein Injections in Rat with Near-infrared-Labeled Agents

Intravenous (IV) administration of agents into the tail vein of rats can be both difficult and inconsistent. Optimizing tail vein injections is a key part of many experimental procedures where reagents need to be introduced directly into the bloodstream. Unwittingly, the injection can be subcutaneous, possibly altering the scientific outcomes. Utilizing a nanoemulsion-based biological probe with an incorporated near-infrared fluorescent (NIRF) dye, this method offers the capability of imaging a successful tail vein injection in vivo. With the use of a NIRF imager, images are taken before and after the injection of the agent. An acceptable IV injection is then qualitatively or quantitatively determined based on the intensity of the NIRF signal at the site of injection.

Here we present a method to validate tail vein injections in rats by utilizing near-infrared fluorescence imaging data from dyes incorporated into agents or biological probes. The tail is imaged before and after the injection, the fluorescent signal is quantified, and an assessment of the injection quality is made.

Neue bewährte Methode zur Validierung von Tail Vene Injektionen in Ratte mit nahe-Infrarot-Label Agenten

Intravenous (IV) administration of agents into the tail vein of rats can be both difficult and inconsistent. Optimizing tail vein injections is a key part ...

Blood Collection Through Subclavian Vein Puncture in Mice

The mouse is the foremost mammalian model for studying human disease and human health. However, blood sample collection from mice is challenging in research work. Tail blood collection is a popular method when a small amount of blood sample is needed. Orbital artery could be considered if a large amount of blood is needed but this blood collection method has ethical issues. Formerly, we demonstrated the feasibility and safety of blood sample collection through subclavian vein puncture in rats, and here we investigate whether this method could be used in mice. We report that this method is safe and practical for blood collection in mice. Blood collection through the subclavian vein puncture in mice can be a convenient method in daily research works.

Here, we present a protocol to take blood samples from the subclavian vein of mice.

Blutentnahme durch Subclavian-Vein-Punktion in Mäusen

The mouse is the foremost mammalian model for studying human disease and human health. However, blood sample collection from mice is challenging in ...

Functional and Physiological Methods of Evaluating Median Nerve Regeneration in the Rat

The main goal of this investigation is to show how to create and repair different types of median nerve (MN) lesions in the rat. Moreover, different methods of simulating postoperative physiotherapy are presented. Multiple standardized strategies are used to assess motor and sensory recovery using an MN model of peripheral nerve lesion and repair, thus permitting easy comparison of the results. Several options are included for providing a postoperative physiotherapy-like environment to rats that have undergone MN injuries. Finally, the paper provides a method to evaluate the recovery of the MN using several noninvasive tests (i.e., grasping test, pin prick test, ladder rung walking test, rope climbing test, and walking track analysis), and physiological measurements (infrared thermography, electroneuromyography, flexion strength evaluation, and flexor carpi radialis muscle weight determination). Hence, this model seems particularly appropriate to replicate a clinical scenario, facilitating extrapolation of results to the human species.
Although the sciatic nerve is the most studied nerve in peripheral nerve research, analysis of the rat MN presents various advantages. For example, there is a reduced incidence of joint contractures and automutilation of the affected limb in MN lesion studies. Furthermore, the MN is not covered by muscle masses, making its dissection easier than that of the sciatic nerve. In addition, MN recovery is observed sooner, because the MN is shorter than the sciatic nerve. Also, the MN has a parallel path to the ulnar nerve in the arm. Hence, the ulnar nerve can be easily used as the nerve graft for repairing MN injuries. Finally, the MN in rats is located in the forelimb, akin to the human upper limb; in humans, the upper limb is the site of most peripheral nerve lesions.

Presented is a protocol to produce different types of median nerve (MN) lesions and repair in the rat. Additionally, the protocol shows how to evaluate the functional recovery of the nerve using several noninvasive behavioral tests and physiological measurements.

Funktionelle und physiologische Methoden zur Beurteilung der medianen Nervenregeneration in der Ratte

The main goal of this investigation is to show how to create and repair different types of median nerve (MN) lesions in the rat. Moreover, different ...

Protocol for Developing a Femur Osteotomy Model in Wistar Albino Rats

Fracture healing is a physiological process resulting in the regeneration of bone defects by the coordinated action of osteoblasts and osteoclasts. Osteoanabolic drugs have the potential to augment the repair of fractures but have constraints like high costs or undesirable side effects. The bone healing potential of a drug can initially be determined by in vitro studies, but in vivo studies are needed for the final proof of concept. Our objective was to develop a femur osteotomy rodent model that could help researchers understand the development of callus formation following fracture of the shaft of the femur and that could help establish whether a potential drug has bone healing properties. Adult male Wistar albino rats were used after Institutional Animal Ethics Committee clearance. The rodents were anesthetized, and under aseptic conditions, complete transverse fractures at the middle one-third of the shafts of the femurs were created using open osteotomy. The fractures were reduced and internally fixed using intramedullary K-wires, and secondary fracture healing was allowed to take place. After surgery, intraperitoneal analgesics and antibiotics were given for 5 days. Sequential weekly x-rays assessed callus formation. The rats were sacrificed based on radiologically pre-determined time points, and the development of the fracture callus was analyzed radiologically and using immunohistochemistry.

Here, we present a protocol to iatrogenically fracture the shaft of the femur of Wistar albino rats and follow up on the development of the callus. This femur osteotomy model can help researchers evaluate the process of fracture healing and to study how a drug could influence fracture healing.

Protokoll zur Entwicklung eines Femurosteotomiemodells bei Wistar Albino-Ratten

Fracture healing is a physiological process resulting in the regeneration of bone defects by the coordinated action of osteoblasts and osteoclasts. ...