Name: biodegradable magnesium stent treatment of saccular aneurysms in a rat model - introduction of the surgical technique
Uploaded: 2017-10-01T13:00:00
Description: Watch this Scientific Journal Video about Biodegradable Magnesium Stent Treatment of Saccular Aneurysms in a Rat Model - Introduction of the Surgical Technique at JoVE.com

We thank Eugen Hofmann and Philine Zumstein for their excellent technical assistance and for sharing their expertise in stent application procedures. We thank Majlinda Kalanderi for the anatomical drawing.

Male Wistar rats with a mean weight of 592 g (&#177;50 SD) and mean age of 20 weeks were housed in animal facilities at a room temperature of 22-24 &#176;C and twelve hour light/dark cycle, with free access to tap water and a pellet diet. The animals received care from humans in accordance with institutional guidelines. The experiments were approved by the Committee for Animal Care of the Canton Bern, Switzerland (BE 102/13). We strictly followed the recommendations for Animal Research: Reporting of In Vivo Expe...

<ol>
	<li>Molyneux, A., 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=International+Subarachnoid+Aneurysm+Trial+(ISAT)+of+neurosurgical+clipping+versus+endovascular+coiling+in+2143+patients+with+ruptured+intracranial+aneurysms:+a+randomised+trial.">International Subarachnoid Aneurysm Trial (ISAT) of neurosurgical clipping versus endovascular coiling in 2143 patients with ruptured intracranial aneurysms: a randomised trial.</a> Lancet. 360 (9342), 1267-1274 (2002).</li><li>Molyneux, A. 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=International+subarachnoid+aneurysm+trial+(ISAT)+of+neurosurgical+clipping+versus+endovascular+coiling+in+2143+patients+with+ruptured+intracranial+aneurysms:+a+randomised+comparison+of+effects+on+survival,+dependency,+seizures,+rebleeding,+subgroups,+and+aneurysm+occlusion.">International subarachnoid aneurysm trial (ISAT) of neurosurgical clipping versus endovascular coiling in 2143 patients with ruptured intracranial aneurysms: a randomised comparison of effects on survival, dependency, seizures, rebleeding, subgroups, and aneurysm occlusion.</a> Lancet. 366 (9488), 809-817 (2005).</li><li>Marbacher, S., 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=Intraluminal+cell+transplantation+prevents+growth+and+rupture+in+a+model+of+rupture-prone+saccular+aneurysms.">Intraluminal cell transplantation prevents growth and rupture in a model of rupture-prone saccular aneurysms.</a> Stroke. 45 (12), 3684-3690 (2014).</li><li>Marbacher, S., Marjamaa, J., Abdelhameed, E., Hernesniemi, J., Niemela, M., Frosen, J. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=The+Helsinki+rat+microsurgical+sidewall+aneurysm+model.">The Helsinki rat microsurgical sidewall aneurysm model.</a> J Vis Exp. (92), e51071 (2014).</li><li>Marbacher, S., 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=Loss+of+mural+cells+leads+to+wall+degeneration,+aneurysm+growth,+and+eventual+rupture+in+a+rat+aneurysm+model.">Loss of mural cells leads to wall degeneration, aneurysm growth, and eventual rupture in a rat aneurysm model.</a> Stroke. 45 (1), 248-254 (2014).</li><li>Pritchett-Corning, K. R., Luo, Y., Mulder, G. B., White, W. J. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Principles+of+rodent+surgery+for+the+new+surgeon.">Principles of rodent surgery for the new surgeon.</a> J Vis Exp. (47), (2011).</li><li>Bernal, 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=Guidelines+for+rodent+survival+surgery.">Guidelines for rodent survival surgery.</a> J Invest Surg. 22 (6), 445-451 (2009).</li><li>Flege, C., 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=Development+and+characterization+of+a+coronary+polylactic+acid+stent+prototype+generated+by+selective+laser+melting.">Development and characterization of a coronary polylactic acid stent prototype generated by selective laser melting.</a> J Mater Sci Mater Med. 24 (1), 241-255 (2013).</li><li>Zartner, P., Cesnjevar, R., Singer, H., Weyand, M. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=First+successful+implantation+of+a+biodegradable+metal+stent+into+the+left+pulmonary+artery+of+a+preterm+baby.">First successful implantation of a biodegradable metal stent into the left pulmonary artery of a preterm baby.</a> Catheter Cardiovasc Interv. 66 (4), 590-594 (2005).</li><li>Hakamata, Y., 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=Green+fluorescent+protein-transgenic+rat:+a+tool+for+organ+transplantation+research.">Green fluorescent protein-transgenic rat: a tool for organ transplantation research.</a> Biochem Biophys Res Commun. 286 (4), 779-785 (2001).</li><li>Bouzeghrane, F., Naggara, O., Kallmes, D. F., Berenstein, A., Raymond, J. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=International+Consortium+of+Neuroendovascular+C.+In+vivo+experimental+intracranial+aneurysm+models:+a+systematic+review.">International Consortium of Neuroendovascular C. In vivo experimental intracranial aneurysm models: a systematic review.</a> AJNR Am J Neuroradiol. 31 (3), 418-423 (2010).</li><li>Aquarius, R., Smits, D., Gounis, M. J., Leenders, W. P., De Vries, J. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Flow+diverter+implantation+in+a+rat+model+of+sidewall+aneurysm:+a+feasibility+study.">Flow diverter implantation in a rat model of sidewall aneurysm: a feasibility study.</a> J Neurointerv Surg. , (2017).</li><li>Indolfi, C., 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=A+new+rat+model+of+small+vessel+stenting.">A new rat model of small vessel stenting.</a> Basic Res Cardiol. 95 (3), 179-185 (2000).</li><li>Oyamada, S., 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=Trans-iliac+rat+aorta+stenting:+a+novel+high+throughput+preclinical+stent+model+for+restenosis+and+thrombosis.">Trans-iliac rat aorta stenting: a novel high throughput preclinical stent model for restenosis and thrombosis.</a> J Surg Res. 166 (1), e91-e95 (2011).</li><li>Kwon, J. S., 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=Comparison+of+bare+metal+stent+and+paclitaxel-eluting+stent+using+a+novel+rat+aorta+stent+model.">Comparison of bare metal stent and paclitaxel-eluting stent using a novel rat aorta stent model.</a> J Vet Sci. 12 (2), 143-149 (2011).</li><li>Rouer, M., Meilhac, O., Delbosc, S., 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=A+new+murine+model+of+endovascular+aortic+aneurysm+repair.">A new murine model of endovascular aortic aneurysm repair.</a> J Vis Exp. (77), e50740 (2013).</li><li>Hardhammar, P. A., 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=Reduction+in+thrombotic+events+with+heparin-coated+Palmaz-Schatz+stents+in+normal+porcine+coronary+arteries.">Reduction in thrombotic events with heparin-coated Palmaz-Schatz stents in normal porcine coronary arteries.</a> Circulation. 93 (3), 423-430 (1996).</li><li>Guerrero, A., 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=Antioxidant+effects+of+a+single+dose+of+acetylsalicylic+acid+and+salicylic+acid+in+rat+brain+slices+subjected+to+oxygen-glucose+deprivation+in+relation+with+its+antiplatelet+effect.">Antioxidant effects of a single dose of acetylsalicylic acid and salicylic acid in rat brain slices subjected to oxygen-glucose deprivation in relation with its antiplatelet effect.</a> Neurosci Lett. 358 (3), 153-156 (2004).</li><li>Niitsu, Y., 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=Repeat+oral+dosing+of+prasugrel,+a+novel+P2Y12+receptor+inhibitor,+results+in+cumulative+and+potent+antiplatelet+and+antithrombotic+activity+in+several+animal+species.">Repeat oral dosing of prasugrel, a novel P2Y12 receptor inhibitor, results in cumulative and potent antiplatelet and antithrombotic activity in several animal species.</a> Eur J Pharmacol. 579 (1-3), 276-282 (2008).</li><li>Kastrati, A., 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=Intracoronary+stenting+and+angiographic+results:+strut+thickness+effect+on+restenosis+outcome+(ISAR-STEREO)+trial.">Intracoronary stenting and angiographic results: strut thickness effect on restenosis outcome (ISAR-STEREO) trial.</a> Circulation. 103 (23), 2816-2821 (2001).</li><li>Lowe, H. C., James, B., Khachigian, L. M. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=A+novel+model+of+in-stent+restenosis:+rat+aortic+stenting.">A novel model of in-stent restenosis: rat aortic stenting.</a> Heart. 91 (3), 393-395 (2005).</li><li>Finn, A. V., 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=A+novel+rat+model+of+carotid+artery+stenting+for+the+understanding+of+restenosis+in+metabolic+diseases.">A novel rat model of carotid artery stenting for the understanding of restenosis in metabolic diseases.</a> J Vasc Res. 39 (5), 414-425 (2002).</li></ol>

Bioabsorbable stents and animal models 
In recent years the general trend in medicine has been away from permanent implants (which remain in the patient&#39;s body for the rest of their life) to bio-absorbable materials. Magnesium stents, in particular, are already quite established in cardiology8,9. Unfortunately these stents have not yet been tested for other applications, such as cerebrovascular diseases. For this reason we decided to stu...

Subarachnoid hemorrhage due to a ruptured intracranial aneurysm is associated with a high mortality rate and poor neurological outcome in many survivors. There are currently two general approaches to occlude IA: either microsurgical clipping (which requires operative exposure of the aneurysm), or endovascular occlusion. As the less invasive endovascular coil treatment of narrow-necked IA has been shown to be associated with slightly lower morbidity (especially in the posterior circulation1,2), endovascular treatment options have become the preferred modality of many neurosurgical centers. Numerous devices have been developed in order to extend the indications of endovascular treatment and overcome the main limitation of IA recurrence after coiling. Intracranial stents are especially promising to overcome these limitations, as they serve as a scaffold for neo-endothelization and coil herniation prevention, as well as protect the parent artery and improve intraluminal intraaneurysmal thrombosis caused by reduction of blood inflow. There is a need to study novel intracranial stents in a low cost animal model; at both macroscopic and molecular levels.
The aim of this study was to design a safe, fast, and standardized surgical technique for stent application in an already established saccular aneurysm model in rats3,4,5. In the present project, we evaluated the role of a biodegradable magnesium stent.

<table style="border-collapse:collapse;width:510pt" width="680">
	<colgroup>
		<col style="width:220pt" width="293" />
		<col style="width:115pt" width="153" />
		<col span="2" style="width:88pt" width="117" />
	</colgroup>
	<tbody>
		<tr height="20" style="height:15.0pt">
			<td class="xl17" height="20" style="width:220pt;height:15.0pt" width="293">Medetomidine</td>
			<td class="xl17" style="width:115pt" width="153"></td>
			<td class="xl17" style="width:88pt" width="117"></td>
			<td class="xl17" style="width:88pt" width="117">any generic</td>
		</tr>
		<tr height="20" style="height:15.0pt">
			<td class="xl17" height="20" style="height:15.0pt">Ketamin</td>
			<td class="xl17"></td>
			<td class="xl17"></td>
			<td class="xl17">any generic</td>
		</tr>
		<tr height="20" style="height:15.0pt">
			<td class="xl17" height="20" style="height:15.0pt">Buprenorphine</td>
			<td class="xl17"></td>
			<td class="xl17"></td>
			<td class="xl17">any generic</td>
		</tr>
		<tr height="20" style="height:15.0pt">
			<td class="xl17" height="20" style="height:15.0pt">Phosphate buffered saline</td>
			<td class="xl17"></td>
			<td class="xl17"></td>
			<td class="xl17"></td>
		</tr>
		<tr height="20" style="height:15.0pt">
			<td class="xl17" height="20" style="height:15.0pt">Sodium dodecyl sulfate (0.1%)</td>
			<td class="xl17"></td>
			<td class="xl17"></td>
			<td class="xl17"></td>
		</tr>
		<tr height="20" style="height:15.0pt">
			<td class="xl17" height="20" style="height:15.0pt">3-0 resorbable suture</td>
			<td class="xl17">Ethicon Inc., USA</td>
			<td class="xl17">VCP428G</td>
			<td class="xl17"></td>
		</tr>
		<tr height="20" style="height:15.0pt">
			<td class="xl17" height="20" style="height:15.0pt">5-0 non absorbable suture</td>
			<td class="xl17">Ethicon Inc., USA</td>
			<td class="xl17">8618G</td>
			<td class="xl17"></td>
		</tr>
		<tr height="20" style="height:15.0pt">
			<td class="xl17" height="20" style="height:15.0pt">6-0 non-absorbable suture</td>
			<td class="xl17">B. Braun, Germany</td>
			<td class="xl17">C0766070</td>
			<td class="xl17"></td>
		</tr>
		<tr height="20" style="height:15.0pt">
			<td class="xl17" height="20" style="height:15.0pt">9-0 non-absorbable suture</td>
			<td class="xl17">B. Braun, Germany</td>
			<td class="xl17">G1111140</td>
			<td class="xl17"></td>
		</tr>
		<tr height="20" style="height:15.0pt">
			<td class="xl17" height="20" style="height:15.0pt">10-0 non-absorbable suture</td>
			<td class="xl17">Covidien, USA</td>
			<td class="xl17">N2530 Monosof</td>
			<td class="xl17"></td>
		</tr>
		<tr height="20" style="height:15.0pt">
			<td class="xl17" height="20" style="height:15.0pt">Operation microscope</td>
			<td class="xl17">Zeiss, Germany</td>
			<td class="xl17"></td>
			<td class="xl17"></td>
		</tr>
		<tr height="20" style="height:15.0pt">
			<td class="xl17" height="20" style="height:15.0pt">Digital microscope camera</td>
			<td class="xl17">Sony, Japan</td>
			<td class="xl17">HXR-MC1P</td>
			<td class="xl17"></td>
		</tr>
		<tr height="20" style="height:15.0pt">
			<td class="xl17" height="20" style="height:15.0pt">Standard surgical instruments</td>
			<td class="xl17"></td>
			<td class="xl17">multiple</td>
			<td class="xl17">see protocol 7.a</td>
		</tr>
		<tr height="20" style="height:15.0pt">
			<td class="xl17" height="20" style="height:15.0pt">Microsurgical instruments</td>
			<td class="xl17"></td>
			<td class="xl17">multiple</td>
			<td class="xl17">see protocol 7.b</td>
		</tr>
		<tr height="20" style="height:15.0pt">
			<td class="xl17" height="20" style="height:15.0pt">Vascular clip applicator</td>
			<td class="xl17">B. Braun, Germany</td>
			<td class="xl17">FT495T</td>
			<td class="xl17"></td>
		</tr>
		<tr height="20" style="height:15.0pt">
			<td class="xl17" height="20" style="height:15.0pt">Temporary vascular clamps</td>
			<td class="xl17">B. Braun, Germany</td>
			<td class="xl17"></td>
			<td class="xl17"></td>
		</tr>
		<tr height="20" style="height:15.0pt">
			<td class="xl17" height="20" style="height:15.0pt">19G Puncture needle&#160;</td>
			<td class="xl17">Angiomed GmbH, Germany</td>
			<td align="right" class="xl17">15820010</td>
			<td class="xl17"></td>
		</tr>
		<tr height="20" style="height:15.0pt">
			<td class="xl17" height="20" style="height:15.0pt">Hydrophobic guide wire</td>
			<td class="xl17">Cook Medical, USA</td>
			<td class="xl17">G00650</td>
			<td class="xl17"></td>
		</tr>
		<tr height="20" style="height:15.0pt">
			<td class="xl17" height="20" style="height:15.0pt">4F sheat</td>
			<td class="xl17">Cordis Corporation, USA</td>
			<td class="xl17">504-604A</td>
			<td class="xl17"></td>
		</tr>
		<tr height="20" style="height:15.0pt">
			<td class="xl17" height="20" style="height:15.0pt">Inflation syringe</td>
			<td class="xl17"></td>
			<td class="xl17"></td>
			<td class="xl17"></td>
		</tr>
		<tr height="20" style="height:15.0pt">
			<td class="xl17" height="20" style="height:15.0pt">Laboratory shaker</td>
			<td class="xl17">Stuart</td>
			<td class="xl17">SRT6</td>
			<td class="xl17"></td>
		</tr>
		<tr height="20" style="height:15.0pt">
			<td class="xl17" height="20" style="height:15.0pt">Magnesium Stent 2.5/6 AMS with Polymer coating</td>
			<td class="xl17">Biotronik, Switzerland</td>
			<td class="xl17"></td>
			<td class="xl17"></td>
		</tr>
		<tr height="21" style="height:15.75pt">
			<td class="xl18" height="21" style="width:220pt;height:15.75pt" width="293">Surgery drape&#160;</td>
			<td class="xl17"></td>
			<td class="xl17"></td>
			<td class="xl17"></td>
		</tr>
		<tr height="20" style="height:15.0pt">
			<td class="xl17" height="20" style="height:15.0pt">Sterile cellulose swabs</td>
			<td class="xl17"></td>
			<td class="xl17"></td>
			<td class="xl17"></td>
		</tr>
		<tr height="20" style="height:15.0pt">
			<td class="xl17" height="20" style="height:15.0pt">Syringes 1 ml and 2 ml</td>
			<td class="xl17"></td>
			<td class="xl17"></td>
			<td class="xl17"></td>
		</tr>
		<tr height="20" style="height:15.0pt">
			<td class="xl17" height="20" style="height:15.0pt">Hollow needles 18G and 26G</td>
			<td class="xl17"></td>
			<td class="xl17"></td>
			<td class="xl17"></td>
		</tr>
		<tr height="20" style="height:15.0pt">
			<td class="xl17" height="20" style="height:15.0pt">Isotonic sodium chloride</td>
			<td class="xl17"></td>
			<td class="xl17"></td>
			<td class="xl17"></td>
		</tr>
		<tr height="20" style="height:15.0pt">
			<td class="xl17" height="20" style="height:15.0pt">Microtubes</td>
			<td class="xl17"></td>
			<td class="xl17"></td>
			<td class="xl17"></td>
		</tr>
		<tr height="20" style="height:15.0pt">
			<td class="xl17" height="20" style="height:15.0pt">Eye ointment</td>
			<td class="xl17">Bausch + Lomb Inc, USA</td>
			<td class="xl17">Lacrinorm</td>
			<td class="xl17">any generic</td>
		</tr>
		<tr height="20" style="height:15.0pt">
			<td class="xl17" height="20" style="height:15.0pt">Small animal shaver</td>
			<td class="xl19"></td>
			<td class="xl19"></td>
			<td class="xl19"></td>
		</tr>
	</tbody>
</table>

biodegradable magnesium stent treatment of saccular aneurysms in a rat model - introduction of the surgical technique

The steady progess in the armamentarium of techniques available for endovascular treatment of intracranial aneurysms requires affordable and reproducable experimental animal models to test novel embolization materials such as stents and flow diverters. The aim of the present project was to design a safe, fast, and standardized surgical technique for stent assisted embolization of saccular aneurysms in a rat animal model.
Saccular aneurysms were created from an arterial graft from the descending aorta.The aneurysms were microsurgically transplanted through end-to-side anastomosis to the infrarenal abdominal aorta of a syngenic male Wistar rat weighing &#62;500 g. Following aneurysm anastomosis, aneurysm embolization was performed using balloon expandable magnesium stents (2.5 mm x 6 mm). The stent system was retrograde introduced from the lower abdominal aorta using a modified Seldinger technique.
Following a pilot series of 6 animals, a total of 67 rats were operated according to established standard operating procedures. Mean surgery time, mean anastomosis time, and mean suturing time of the artery puncture site were 167 &#177; 22 min, 26 &#177; 6 min and 11 &#177; 5 min, respectively. The mortality rate was 6% (n=4). The morbidity rate was 7.5% (n=5), and in-stent thrombosis was found in 4 cases (n=2 early, n=2 late in stent thrombosis).
The results demonstrate the feasibility of standardized stent occlusion of saccular sidewall aneurysms in rats - with low rates of morbidity and mortality. This stent embolization procedure combines the opportunity to study novel concepts of stent or flow diverter based devices as well as the molecular aspects of healing.

The average duration of surgery was 167 (&#177; 22) min, 26 (&#177; 6) min of which were needed for aneurysm creation and a further 23 (&#177; 7) min required for stent application and reconstruction of the arteriotomy (Figure 3).
Mortality, morbidity, and macroscopic in-stent thrombosis were the primary endpoints of the study. The regular follow-up periods were 7 days (n=28), and 21 days (n=32) res...

Watch this Scientific Journal Video about Biodegradable Magnesium Stent Treatment of Saccular Aneurysms in a Rat Model - Introduction of the Surgical Technique at JoVE.com

Biodegradable Magnesium Stent Treatment of Saccular Aneurysms in a Rat Model - Introduction of the Surgical Technique

Reproducable experimental animal models are needed for the testing of novel embolization materials, which have been designed to treat endovascular occlusion of intracranial aneurysms (IA). The present study aims to develop a safe and standardized surgical technique for stent assisted embolization of saccular aneurysms in a rat animal model.

The steady progess in the armamentarium of techniques available for endovascular treatment of intracranial aneurysms requires affordable and reproducable ...

The overall goal of this standardized method for occluding saccular sidewall aneurysms in rats using biodegradable magnesium stents is to study novel concepts in stent devices and the molecular aspects of aneurysm healing. This method can help answer key questions in the neurosurgical and neuroradiological field related to the function of new stent or flow diverter devices. The main advantage of this technique is that it represents a standardized and safe procedure with low morbidity and mortality rates.It is also highly reproducible and thus improves the reliability of associated studies. Under general anesthesia, open the abdomen of the donor rat below the xiphoid process. First incise along the midventral line and separate the skin from the musculature.Then dissect the abdominal musculature and identify the diaphragm above the liver at the insertion point of the thoracic cavity. Next access the thoracic cavity and euthanize the rat with an intracardiac ketamine injection. Then mobilize the left lung and fix it to the right side using a cellulose swab.Next expose the aortic arc and above the aorta, clamp the azygos and left cranial caval veins using two mosquito clamps. Then cut between the clamps and leave the clamps attached. Now increase the access to the aortic arc and descending aorta using micro scissors and micro forceps.Along the aorta, identify the left subclavian artery and first intercostal artery. Now place a nonabsorbable 6-0 suture just cranial of the first intercostal artery and tie it up. At the proximal end, cut the aorta right after the origin of the left subclavian artery.Finally remove the newly created aneurysm with a cut just below the ligature. After measuring the aneurysm, immediately transplant the graft into the recipient rat. Start the surgery on the recipient rat with a mid ventral incision distal of the xiphoid process and open the abdominal muscles along the linea alba.Then move aside the small intestines, the cecum, and the colon and attach a self-holding retractor to keep the abdominal cavity open. Place a swab beneath the liver to improve the retroperitoneal exposure. Fat masses can be moved aside as well.Now open the parietal peritoneum to expose the abdominal aorta. First bluntly dissect the surrounding retroperitoneal fat to expose a longer length of the vessel. Be careful when handling the ureters, testicular vessels, and the superior mesenteric artery.They are often covered by a thin layer of adipose tissue. When proceeding to a sharp dissection, only grasp the adventitia to avoid damaging the vessel wall. Once exposed, dissect the abdominal aorta.Start at the distal segment which is the most technically challenging segment to dissect. Use micro scissors and micro forceps starting at the vena cava alongside the arterial adventitial layer between the vessels. Small lumbar arteries may arise as segmental vessels from the dorsal surface of the aorta.Coagulate and cut these as needed. Also try to identify the right iliolumbar artery. Its origin is most frequently caudal to the left counterpart but this varies.Once the dissection is finished, without stretching the vessel, place a small, colored rubber pad underneath it to indicate where the arterial puncture will be performed. Now proceed with the dissection of the abdominal aorta from the vena cava in the middle segment. Identify the superior mesenteric artery at its origin from the abdominal aorta and keep it intact.Also do not damage the lumbosacral plexus. Use lots of flushing and be mindful of the short coagulation time. Ultimately establish the location of the stent in the middle segment and be sure to leave enough room for the abdominal vessels around the stent.Now place a small piece of gauze and a colored rubber pad under the aorta for better exposure. Then dissect the proximal segment of the abdominal aorta located proximal to the renal vein. After opening the parietal peritoneum, separate the vessels with two blunt forceps.Then after a dorsal circumnavigation of the aorta, place a small piece of a colored rubber beneath it to facilitate the future application of a proximal clip. Clamp the aorta in the middle segment. First apply the distal clamp then the proximal clamp.Next remove the adventitia at the planned anastomosis site using micro forceps and micro scissors. Do not injure the vessel wall. Next lift a small piece of vessel wall using micro forceps and perform a linear incision with micro scissors to make a hole with the same size as the aneurysm base.Then flush the artery with saline in both directions using a blunted needle. Now place the first two stitches of the end-to-side anastomosis at the proximal and distal end of the arteriotomy. Grasp the vessel as little as possible while placing the sutures.Then close the incision with interrupted sutures to complete the anastomosis. To proceed, remove the distal clamp first. Place an extra suture in case of major bleeding from backflow.Then remove both clamps and rinse the anastomosis site. Check the patency of the distal abdominal aorta using a direct milking test. Then remove the gauze swab under the colored rubber pad and wait 10 minutes before proceeding with the stent implantation.To start the stent implantation, first clip the artery puncture site in the distal segment at the distal clamp and then the proximal clamp. Then remove the loose connective tissue and some of the adventitia at the puncture site. Next place a large temporary clamp in the proximal segment to avoid major hemorrhaging during the arterial puncture.Now lift the vessel wall and puncture the distal arterial segment using an 18-gauge puncture needle and thoroughly flush the artery with saline. Next insert a hydrophobic guidewire into the puncture site and remove the proximal clamp from the distal segment. Advance the guidewire to the clamp in the proximal segment.Then insert the Teflon introducer using the Seldinger technique. Advance the dilater to be proximal to the anastomosis site. Then check that the three-way stopcock is turned off to avoid unnecessary loss of blood.Next remove the guidewire and dilater. After vacuuming the balloon system, set the system to one bar of pressure, and then insert the observable magnesium stent using the size four Fogarty introducer. When the stent is finally in place, carefully inflate the balloon to eight or nine bar while continuously checking the anastomosis sutures for ripping.Then deflate and remove the balloon. While removing the introducer, be ready with a temporary clip as retrograde oozing from the proximal patent arteries may be significant. Now place a clip in the proximal distal segment and remove the temporary clamp in the proximal segment.Next thoroughly flush the puncture site with saline and close it with interrupted, nonabsorbable 10-0 sutures. If possible avoid grasping the vessel wall. Check for distal artery patency with the direct milking test.Remove the colored rubber pad. Finish by closing the surgical site as described in the tech's protocol. The average duration of surgery was about three hours of which about 1/2 hour was needed for aneurysm creation and another half hour was needed for stent application and reconstruction of the arteriotomy.There were no deaths during surgery. In 87%of surgeries, the desired endpoint of a patent artery was achieved. The other animals included five that experienced complications and four that died within three days due to early in stent thrombosis.In two animals, late in stent thrombosis occurred 21 days postoperatively with no accompanying neurological deficits. One animal suffered wound dehiscence on the second postoperative day with a consecutive loss of volume. We first had the idea for this method when we recognized that biodegradable stent devices gained increasing popularity in the treatment of coronary heart disease and realized that today's techniques of stent insertion are associated with high rates of morbidity and mortality.Once mastered, this technique can be performed in an average time of two hours and 15 minutes if it is performed properly. Following this procedure, other methods like intra aneurysm coil embolization can be performed in order to answer additional questions about aneurysm healing.

biodegradable-magnesium-stent-treatment-saccular-aneurysms-rat-model

Research

JoVE Journal

Neuroscience

Application of a NMDA Receptor Conductance in Rat Midbrain Dopaminergic Neurons Using the Dynamic Clamp Technique

Neuroscientists study the function of the brain by investigating how neurons in the brain communicate. Many investigators look at changes in the electrical activity of one or more neurons in response to an experimentally-controlled input. The electrical activity of neurons can be recorded in isolated brain slices using patch clamp techniques with glass micropipettes. Traditionally, experimenters can mimic neuronal input by direct injection of current through the pipette, electrical stimulation of the other cells or remaining axonal connections in the slice, or pharmacological manipulation by receptors located on the neuronal membrane of the recorded cell.
Direct current injection has the advantages of passing a predetermined current waveform with high temporal precision at the site of the recording (usually the soma). However, it does not change the resistance of the neuronal membrane as no ion channels are physically opened. Current injection usually employs rectangular pulses and thus does not model the kinetics of ion channels. Finally, current injection cannot mimic the chemical changes in the cell that occurs with the opening of ion channels.
Receptors can be physically activated by electrical or pharmacological stimulation. The experimenter has good temporal precision of receptor activation with electrical stimulation of the slice. However, there is limited spatial precision of receptor activation and the exact nature of what is activated upon stimulation is unknown. This latter problem can be partially alleviated by specific pharmacological agents. Unfortunately, the time course of activation of pharmacological agents is typically slow and the spatial precision of inputs onto the recorded cell is unknown.
The dynamic clamp technique allows an experimenter to change the current passed directly into the cell based on real-time feedback of the membrane potential of the cell (Robinson and Kawai 1993, Sharp et al., 1993a,b; for review, see Prinz et al. 2004). This allows an experimenter to mimic the electrical changes that occur at the site of the recording in response to activation of a receptor. Real-time changes in applied current are determined by a mathematical equation implemented in hardware.
We have recently used the dynamic clamp technique to investigate the generation of bursts of action potentials by phasic activation of NMDA receptors in dopaminergic neurons of the substantia nigra pars compacta (Deister et al., 2009; Lobb et al., 2010). In this video, we demonstrate the procedures needed to apply a NMDA receptor conductance into a dopaminergic neuron.

In this video, we demonstrate how to apply a conductance into a dopaminergic neuron recorded in the whole cell configuration in rat brain slices. This technique is called the dynamic clamp.

Neuroscientists study the function of the brain by investigating how neurons in the brain communicate. Many investigators look at changes in the ...

Visual Evoked Potential Recording in a Rat Model of Experimental Optic Nerve Demyelination

The visual evoked potential (VEP) recording is widely used in clinical practice to assess the severity of optic neuritis in its acute phase, and to monitor the disease course in the follow-up period. Changes in the VEP parameters closely correlate with pathological damage in the optic nerve. This protocol provides a detailed description about the rodent model of optic nerve microinjection, in which a partial demyelination lesion is produced in the optic nerve. VEP recording techniques are also discussed. Using skull implanted electrodes, we are able to acquire reproducible intra-session and between-session VEP traces. VEPs can be recorded on individual animals over a period of time to assess the functional changes in the optic nerve longitudinally. The optic nerve demyelination model, in conjunction with the VEP recording protocol, provides a tool to investigate the disease processes associated with demyelination and remyelination, and can potentially be employed to evaluate the effects of new remyelinating drugs or neuroprotective therapies.

Focal demyelination is induced in the optic nerve using lysolecithin microinjection. Visual evoked potentials are recorded via skull electrodes implanted over the visual cortex to examine the signal conduction along the visual pathway in vivo. This protocol details the surgical procedures underlying electrode implantation and optic nerve microinjection.

The visual evoked potential (VEP) recording is widely used in clinical practice to assess the severity of optic neuritis in its acute phase, and to ...

The Use of Trace Eyeblink Classical Conditioning to Assess Hippocampal Dysfunction in a Rat Model of Fetal Alcohol Spectrum Disorders

Neonatal rats were administered a relatively high concentration of ethyl alcohol (11.9% v/v) during postnatal days 4-9, a time when the fetal brain undergoes rapid organizational change and is similar to accelerated brain changes that occur during the third trimester in humans. This model of fetal alcohol spectrum disorders (FASDs) produces severe brain damage, mimicking the amount and pattern of binge-drinking that occurs in some pregnant alcoholic mothers. We describe the use of trace eyeblink classical conditioning (ECC), a higher-order variant of associative learning, to assess long-term hippocampal dysfunction that is typically seen in alcohol-exposed adult offspring. At 90 days of age, rodents were surgically prepared with recording and stimulating electrodes, which measured electromyographic (EMG) blink activity from the left eyelid muscle and delivered mild shock posterior to the left eye, respectively. After a 5 day recovery period, they underwent 6 sessions of trace ECC to determine associative learning differences between alcohol-exposed and control rats. Trace ECC is one of many possible ECC procedures that can be easily modified using the same equipment and software, so that different neural systems can be assessed. ECC procedures in general, can be used as diagnostic tools for detecting neural pathology in different brain systems and different conditions that insult the brain.

Trace eyeblink classical conditioning (ECC) was used to assess hippocampal-dependent associative learning in adult rats that were administered a high concentration (11.9% v/v) of alcohol during early neonatal brain development. In general, ECC procedures are sound diagnostic tools for detecting brain dysfunction across many psychological and biomedical settings.

Neonatal rats were administered a relatively high concentration of ethyl alcohol (11.9% v/v) during postnatal days 4-9, a time when the fetal brain ...

The Optical Fractionator Technique to Estimate Cell Numbers in a Rat Model of Electroconvulsive Therapy

Stereological methods are designed to describe quantitative parameters without making assumptions about size, shape, orientation and distribution of cells or structures. These methods have been revolutionary for quantitative analysis of the mammalian brain, in which volumetric cell populations are too high to count manually, and stereology is now the technique of choice whenever estimates of three-dimensional quantities need to be extracted from measurements on two-dimensional sections. All stereological methods are in principle unbiased; however, they rely on proper knowledge about the structure of interest and the characteristics of the tissue. Stereology is based on Systematic Uniformly Random Sampling (SURS), with adjustment of sampling to the most efficient level in respect to precision, providing reliable, quantitative information about the whole structure of interest. Here we present the optical fractionator in conjunction with BrdU immunohistochemistry to estimate the production and survival of newly-formed neurons in the granule cell layer (including the sub-granular zone) of the rat hippocampus following electroconvulsive stimulation, which is among the most potent stimulators of neurogenesis. The optical fractionator technique is designed to provide estimates of the total number of cells from thick sections sampled from the full structure. Thick sections provide the opportunity to observe cells in their full 3-D extent and thus, allow for easy and robust cell classification based on morphological criteria. When correctly implemented, the sensitivity and efficiency of the optical fractionator provides accurate estimates with a fixed and predetermined precision.

Here, we present a stereological method, the optical fractionator, used to quantify the formation of new neurons, and their survival, in the rat hippocampus following electroconvulsive stimulation. When correctly implemented, the sensitivity and efficiency of stereological methods ensures accurate estimates with a fixed and predetermined precision.

Stereological methods are designed to describe quantitative parameters without making assumptions about size, shape, orientation and distribution of cells ...

A Protocol for Measuring Cue Reactivity in a Rat Model of Cocaine Use Disorder

Cocaine use disorder (CUD) follows a trajectory of repetitive self-administration during which previously neutral stimuli gain incentive value. Cue reactivity, the sensitivity to cues previously linked with the drug-taking experience, plays a prominent role in human craving during abstinence. Cue reactivity can be assessed as the attentional orientation toward drug-associated cues that is measurable as appetitive approach behavior in both preclinical and human studies. Herein describes an assessment of cue reactivity in rats trained to self-administer cocaine. Cocaine self-administration is paired with the presentation of discrete cues that act as conditioned reinforcers (i.e., house light, stimulus light, infusion pump sounds). Following a period of abstinence, lever presses in the cocaine self-administration context accompanied by the discrete cues previously paired with cocaine infusion are measured as cue reactivity. This model is useful to explore neurobiological mechanisms underlying cue reactivity processes as well as to assess pharmacotherapies to suppress cue reactivity and therefore, modify relapse vulnerability. Advantages of the model include its translational relevance, and its face and predictive validities.&#160;The primary limitation of the model is that the cue reactivity task can only be performed infrequently and must only be used in short duration (e.g., 1 hour), otherwise rats will begin to extinguish the pairing of the discrete cues with the cocaine stimulus. The model is extendable to any positively reinforcing stimulus paired with discrete cues; though particularly applicable to drugs of abuse, this model may hold future applications in fields such as obesity, where palatable food rewards can act as positively reinforcing stimuli.

Cue reactivity is conceptualized as sensitivity to cues linked with drug-taking experiences that contribute to craving and relapse in abstinent humans. Cue reactivity is modeled in rats by measuring attentional orientation toward drug-associated cues that results in appetitive approach behavior in a cue reactivity test following self-administration and forced abstinence.

Cocaine use disorder (CUD) follows a trajectory of repetitive self-administration during which previously neutral stimuli gain incentive value. Cue ...

Simple Surgical Induction of Conductive Hearing Loss with Verification Using Otoscope Visualization and Behavioral Clap Startle Response in Rat

Conductive hearing loss (CHL) is a prevalent hearing impairment in humans. The goal of the protocol is to describe a simple surgical procedure for inducing CHL in rodents. The protocol demonstrates CHL by tympanic membrane puncture. Verification of CHL surgery was by otoscope examination and behavioral assessment by clap startle response, both replicable and reliable, and are simple methods to demonstrate hearing loss has occurred. The simple CHL procedure is advantageous due to its reproducibility and flexibility to different pursuits in hearing loss research. The limitations of inducing CHL by a surgical approach are associated with the learning curve to perform the surgical procedure and confidence in audiological examination. Inducing a hearing impairment by CHL allows one to readily study the neural manifestations and behavioral outcomes of hearing loss.

Here, we present a protocol to establish a replicable conductive hearing loss induction via surgical tympanic membrane puncture and verification by otoscope visualization and behavioral assessment by clap startle.

Conductive hearing loss (CHL) is a prevalent hearing impairment in humans. The goal of the protocol is to describe a simple surgical procedure for ...

Facial Nerve Surgery in the Rat Model to Study Axonal Inhibition and Regeneration

This protocol describes consistent and reproducible methods to study axonal regeneration and inhibition in a rat facial nerve injury model. The facial nerve can be manipulated along its entire length, from its intracranial segment to its extratemporal course. There are three primary types of nerve injury used for the experimental study of regenerative properties: nerve crush, transection, and nerve gap. The range of possible interventions is vast, including surgical manipulation of the nerve, delivery of neuroactive reagents or cells, and either central or end-organ manipulations. Advantages of this model for studying nerve regeneration include simplicity, reproducibility, interspecies consistency, reliable survival rates of the rat, and an increased anatomic size relative to murine models. Its limitations involve a more limited genetic manipulation versus the mouse model and the superlative regenerative capability of the rat, such that the facial nerve scientist must carefully assess time points for recovery and whether to translate results to higher animals and human studies. The rat model for facial nerve injury allows for functional, electrophysiological, and histomorphometric parameters for the interpretation and comparison of nerve regeneration. It thereby boasts tremendous potential toward furthering the understanding and treatment of the devastating consequences of facial nerve injury in human patients.

This protocol describes a reproducible approach to facial nerve surgery in the rat model, including descriptions of various inducible patterns of injury.

This protocol describes consistent and reproducible methods to study axonal regeneration and inhibition in a rat facial nerve injury model. The facial ...

A Revised Surgical Approach to Induce Endolymphatic Hydrops in the Guinea Pig

Endolymphatic hydrops is an enlargement of scala media that is most often associated with Meniere&#39;s disease, though the pathophysiologic mechanism(s) remain unclear. In order to adequately study the attributes of endolymphatic hydrops, such as the origins of low-frequency hearing loss, a reliable model is needed. The guinea pig is a&#160;good model because it hears in the low-frequency regions that are putatively affected by endolymphatic hydrops. Previous research has demonstrated that endolymphatic hydrops can be induced surgically via intradural or extradural approaches that involve drilling on the endolymphatic duct and sac. However, whether it was possible to create an endolymphatic hydrops model using an extradural approach that avoided dangerous drilling on the endolymphatic duct and sac was unknown. The objective of this study was to demonstrate a revised extradural approach to induce experimental endolymphatic hydrops at 30 days post-operatively by obliterating the endolymphatic sac and injuring the endolymphatic duct with a fine pick. The sample size consisted of seven guinea pigs. Functional measurements of hearing were made and temporal bones were subsequently harvested for histologic analysis. The approach had a success rate of 86% in achieving endolymphatic hydrops. The risk of cerebrospinal fluid leak was minimal. No perioperative deaths or injuries to the posterior semicircular canal occurred in the sample. The presented method demonstrates a safe and reliable way to induce endolymphatic hydrops at a relatively quick time point of 30 days. The clinical implications are that the presented method provides a reliable model to further explore the origins of low-frequency hearing loss that can be associated endolymphatic hydrops.

This article demonstrates an extradural approach to obliterate the guinea pig endolymphatic sac and injure the endolymphatic duct with a fine pick in order to induce experimental endolymphatic hydrops.

Endolymphatic hydrops is an enlargement of scala media that is most often associated with Meniere&#39;s disease, though the pathophysiologic mechanism(s) ...

Applying the RatWalker System for Gait Analysis in a Genetic Rat Model of Parkinson's Disease

Parkinson's disease (PD) is a progressive neurodegenerative disorder caused by the loss of dopaminergic (DA) neurons in the substantia nigra pars compacta. Gait abnormalities, including decreased arm swing, slower walking speed, and shorter steps are common in PD patients and appear early in the course of disease. Thus, the quantification of motor patterns in animal models of PD will be important for phenotypic characterization during disease course and upon therapeutic treatment. Most cases of PD are idiopathic; however, the identification of hereditary forms of PD uncovered gene mutations and variants, such as loss-of-function mutations in Pink1 and Parkin, two proteins involved in mitochondrial quality control that could be harnessed to create animal models. While mice are resistant to neurodegeneration upon loss of Pink1 and Parkin (single and combined deletion), in rats, Pink1 but not Parkin deficiency leads to nigral DA neuron loss and motor impairment. Here, we report the utility of FTIR imaging to uncover gait changes in freely walking young (2 months of age) male rats with combined loss of Pink1 and Parkin prior to the development of gross visually apparent motor abnormality as these rats age (observed at 4-6 months), characterized by hindlimb dragging as previously reported in Pink1 knockout (KO) rats.

Applying the RatWalker System for Gait Analysis in a Genetic Rat Model of Parkinson&#039;s Disease

Here we describe the RatWalker system, built by redesigning the MouseWalker apparatus to accommodate for the increased size and weight of rats. This system uses frustrated total internal reflection (FTIR), high-speed video capture, and open-access analysis software to track and quantify gait parameters.

Parkinson's disease (PD) is a progressive neurodegenerative disorder caused by the loss of dopaminergic (DA) neurons in the substantia nigra pars ...

Establishment of a Rat Model of Superior Sagittal-Sinus Occlusion via a Thread-Embolism Method

The mechanisms contributing to the natural onset of cerebral venous sinus thrombosis (CVST) are mostly unknown, and a variety of uncontrollable factors are involved in the course of the disease, resulting in great limitations in clinical research. Therefore, the establishment of stable CVST animal models that can standardize a variety of uncontrollable confounding factors have helped to circumvent shortcomings in clinical research. In recent decades, a variety of CVST animal models have been constructed, but the results based on these models have been inconsistent and incomplete. Hence, in order to further explore the pathophysiological mechanisms of CVST, it is necessary to establish a novel and highly compatible animal model, which has important practical value and scientific significance for the diagnosis and treatment of CVST. In the present study, a novel Sprague-Dawley (SD) rat model of superior sagittal sinus (SSS) thrombosis was established via a thread-embolization method, and the stability and reliability of the model were verified. Additionally, we evaluated changes in cerebral venous blood flow in rats after the formation of CVST. Collectively, the SD-rat SSS-thrombosis model represents a novel CVST animal model that is easily established, minimizes trauma, yields good stability, and allows for accurately controlling ischemic timing and location.

Here, we establish a novel Sprague-Dawley (SD) rat model of superior sagittal sinus (SSS) thrombosis via a thread-embolization method, and the stability and reliability of the model were verified.

The mechanisms contributing to the natural onset of cerebral venous sinus thrombosis (CVST) are mostly unknown, and a variety of uncontrollable factors ...