Name: mouse microsurgery infusion technique for targeted substance delivery into the cns via the internal carotid artery
Uploaded: 2017-01-31T14:00:00
Description: Watch this Scientific Journal Video about Mouse Microsurgery Infusion Technique for Targeted Substance Delivery into the CNS via the Internal Carotid Artery at JoVE.com

We would like to thank Dr. Lei Chen (Icahn School of Medicine at Mount Sinai, NY) who first established the use of this model in our laboratory, and to Dr. Gretchen Wolff (German Cancer Research Center, Heidelberg, Germany) for disseminating the technique in our laboratory. Supported in part by HL126559, DA039576, MH098891, MH63022, MH072567, DA027569, and NSC 2015/17/B/NZ7/02985.

Les procédures impliquées dans le protocole suivant ont été approuvés par l&#39;Université de Miami Institutional Animal Care et utilisation Commission (IACUC). En outre, toutes les procédures sont menées dans des installations agréées par l&#39;Association pour l&#39;évaluation et l&#39;accréditation des animaux de laboratoire Care International (AAALAC). 1. Préparation de la souris pour la chirurgie <ol><li> Anesthésier souris avec de l&#39;isoflurane mélangé avec de l&#3...

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	<li>Chen, L., Swartz, K. R., Toborek, M. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Vessel+microport+technique+for+applications+in+cerebrovascular+research.">Vessel microport technique for applications in cerebrovascular research.</a> J Neurosci Res. 87 (7), 1718-1727 (2009).</li><li>Frisella, W. 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=Intracranial+injection+of+recombinant+adeno-associated+virus+improves+cognitive+function+in+a+murine+model+of+mucopolysaccharidosis+type+VII.">Intracranial injection of recombinant adeno-associated virus improves cognitive function in a murine model of mucopolysaccharidosis type VII.</a> Mol Ther. 3 (3), 351-358 (2001).</li><li>Wei, L., Erinjeri, J. P., Rovainen, C. M., Woolsey, T. A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Collateral+growth+and+angiogenesis+around+cortical+stroke.">Collateral growth and angiogenesis around cortical stroke.</a> Stroke. 32 (9), 2179-2184 (2001).</li><li>Wu, 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=Targeted+delivery+of+methotrexate+to+epidermal+growth+factor+receptor-positive+brain+tumors+by+means+of+cetuximab+(IMC-C225)+dendrimer+bioconjugates.">Targeted delivery of methotrexate to epidermal growth factor receptor-positive brain tumors by means of cetuximab (IMC-C225) dendrimer bioconjugates.</a> Mol Cancer Ther. 5 (1), 52-59 (2006).</li><li>Pignataro, G., Studer, F. E., Wilz, A., Simon, R. P., Boison, D. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Neuroprotection+in+ischemic+mouse+brain+induced+by+stem+cell-derived+brain+implants.">Neuroprotection in ischemic mouse brain induced by stem cell-derived brain implants.</a> J Cereb Blood Flow Metab. 27 (5), 919-927 (2007).</li><li>Sugiyama, Y., Kusuhara, H., Suzuki, H. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Kinetic+and+biochemical+analysis+of+carrier-mediated+efflux+of+drugs+through+the+blood-brain+and+blood-cerebrospinal+fluid+barriers:+importance+in+the+drug+delivery+to+the+brain.">Kinetic and biochemical analysis of carrier-mediated efflux of drugs through the blood-brain and blood-cerebrospinal fluid barriers: importance in the drug delivery to the brain.</a> J Control Release. 62 (1-2), 179-186 (1999).</li><li>Pardridge, W. M. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Drug+and+gene+delivery+to+the+brain:+the+vascular+route.">Drug and gene delivery to the brain: the vascular route.</a> Neuron. 36 (4), 555-558 (2002).</li><li>Vantyghem, S. A., Postenka, C. O., Chambers, A. F. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Estrous+cycle+influences+organ-specific+metastasis+of+B16F10+melanoma+cells.">Estrous cycle influences organ-specific metastasis of B16F10 melanoma cells.</a> Cancer Res. 63 (16), 4763-4765 (2003).</li><li>Huang, R. Q., 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=Efficient+gene+delivery+targeted+to+the+brain+using+a+transferrin-conjugated+polyethyleneglycol-modified+polyamidoamine+dendrimer.">Efficient gene delivery targeted to the brain using a transferrin-conjugated polyethyleneglycol-modified polyamidoamine dendrimer.</a> FASEB J. 21 (4), 1117-1125 (2007).</li><li>Liu, R., Martuza, R. L., Rabkin, S. D. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Intracarotid+delivery+of+oncolytic+HSV+vector+G47Delta+to+metastatic+breast+cancer+in+the+brain.">Intracarotid delivery of oncolytic HSV vector G47Delta to metastatic breast cancer in the brain.</a> Gene Ther. 12 (8), 647-654 (2005).</li><li>Kumar, P., 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=Transvascular+delivery+of+small+interfering+RNA+to+the+central+nervous+system.">Transvascular delivery of small interfering RNA to the central nervous system.</a> Nature. 448 (7149), 39-43 (2007).</li><li>Wrobel, J. K., Wolff, G., Xiao, R., Power, R. F., Toborek, M. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Dietary+Selenium+Supplementation+Modulates+Growth+of+Brain+Metastatic+Tumors+and+Changes+the+Expression+of+Adhesion+Molecules+in+Brain+Microvessels.">Dietary Selenium Supplementation Modulates Growth of Brain Metastatic Tumors and Changes the Expression of Adhesion Molecules in Brain Microvessels.</a> Biol Trace Elem Res. , (2015).</li></ol>

La microchirurgie de perfusion décrit ici a été prouvé être très efficace dans la prestation de substances exogènes de diverses caractéristiques biologiques dans le SNC, ce qui empêche la diffusion non désirée dans tout le corps 1, 12. Rupture de la barrière hémato-encéphalique est une caractéristique pathologique de plusieurs maladies liées au système nerveux central; Par conséquent évaluer la relation entre les substances exogènes à la bar...

Dans les modèles in vivo du système nerveux central (SNC) ont besoin d&#39; une prestation efficace des substances exogènes, tels que les médicaments, les agents pathogènes, ou exosomes, dans le cerveau. Par conséquent, une méthode de distribution idéale devrait provoquer un traumatisme minimal à l&#39;animal, de préserver l&#39;intégrité du réseau neuronal, et d&#39; atteindre des concentrations élevées de la substance dans le cerveau 1. Plusieurs méthodes chirurgicales de distribution de substance locale ont été décrites, y compris intra-gaine, intracérébrale, et les injections intraventriculaire ou implants 2, 3, 4, 5. Ces approches, cependant, sont considérés comme traumatique du système nerveux central, et permettent l&#39;administration des seuls volumes faibles de la substance d&#39;intérêt. En outre, il a été suggéré que les substances exogènes peuvent être éliminées rapidement par le liquide céphalo - 6 </sup&gt;, et une faible gamme de pénétration du parenchyme cérébral a été observée 7 lorsque les techniques mentionnées ci-dessus sont utilisées. Méthodes de libération systémique, comme par voie orale, pulmonaire, sous - cutanée et intraveineuse, sont plus couramment utilisés dans des modèles animaux, bien qu&#39;ils présentent une faible efficacité dans la livraison des substances au système nerveux central, en raison de l&#39; absorption par les autres organes 8, 9. Par conséquent, ces voies d&#39;administration nécessitent des doses élevées de substances administrées, ce qui augmente le risque d&#39;effets secondaires et de toxicité 10, 11. Ici, nous décrivons une technique de microchirurgie d&#39;infusion de souris, ce qui crée effectivement des substances directement dans le cerveau par l&#39;intermédiaire de l&#39;artère carotide interne. En plus de cibler la livraison au CNS, cette technique ne contourne pas les barrières physiologiques normales et est donc très pertinente pour biological processus impliqués dans les passages des agents thérapeutiques ou des agents pathogènes dans le cerveau.

<table border="0" cellpadding="0" cellspacing="0" width="635">
	<tbody>
		<tr height="21">
			<td height="21" style="height:21px;width:367px;">Anesthesia instrument</td>
			<td style="width:149px;">Vetequip</td>
			<td align="right" style="width:119px;">901806</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:367px;">Surgical scissors</td>
			<td style="width:149px;">Fine Science Tool</td>
			<td style="width:119px;">14558-09</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:367px;">Surgical forceps straight tip</td>
			<td style="width:149px;">Fine Science Tool</td>
			<td style="width:119px;">00108-11</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:367px;">Surgical forceps angled tip</td>
			<td style="width:149px;">Fine Science Tool</td>
			<td style="width:119px;">00109-11</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:367px;">Spring scissors</td>
			<td style="width:149px;">Fine Science Tool</td>
			<td style="width:119px;">15000-08</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:367px;">Nylon suture</td>
			<td style="width:149px;">Braintree Scientific</td>
			<td style="width:119px;">SUT-S 104</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">Capillary tubing (Micro-Renathane 0.010&#8221; x 0.005&#8221; per ft.)&#160;</td>
			<td style="width:149px;">Braintree Scientific</td>
			<td style="width:119px;">MRE01050</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:367px;">Closing suture</td>
			<td style="width:149px;">VWR</td>
			<td style="width:119px;">95057-036</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:367px;">Isoflurane</td>
			<td style="width:149px;">Piramal</td>
			<td style="width:119px;"></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:367px;">2,3,5-Triphenyltetrazolium chloride</td>
			<td style="width:149px;">FisherScientific</td>
			<td style="width:119px;">50-121-8005</td>
		</tr>
	</tbody>
</table>

mouse microsurgery infusion technique for targeted substance delivery into the cns via the internal carotid artery

Animal models of central nervous system (CNS) diseases and, consequently, blood-brain barrier disruption diseases, require the delivery of exogenous substances into the brain. These exogenous substances may induce injurious impact or constitute therapeutic strategy. The most common delivery methods of exogenous substances into the brain are based on systemic deliveries, such as subcutaneous or intravenous routes. Although commonly used, these approaches have several limitations, including low delivery efficacy into the brain. In contrast, surgical methods that locally deliver substances into the CNS are more specific and prevent the uptake of the exogenous substances by other organs. Several surgical methods for CNS delivery are available; however, they tend to be very traumatic. Here, we describe a mouse infusion microsurgery technique, which effectively delivers substances into the brain via the internal carotid artery, with minimal trauma and no interference with normal CNS functionality.

La technique de microchirurgie d&#39;infusion de souris décrit ici est très polyvalent et a été utilisé pour délivrer différentes substances directement dans le cerveau, y compris la fourniture des cellules tumorales dans un modèle représentatif de la formation de métastases cérébrales 1, 12. Cette technique est également adaptée pour évaluer les aspects...

Watch this Scientific Journal Video about Mouse Microsurgery Infusion Technique for Targeted Substance Delivery into the CNS via the Internal Carotid Artery at JoVE.com

Mouse Microsurgery Infusion Technique for Targeted Substance Delivery into the CNS via the Internal Carotid Artery

The present protocol describes a mouse microsurgery infusion technique, which effectively delivers substances directly into the brain via the internal carotid artery.

Souris microchirurgie Infusion Technique pour ciblée Substance Livraison dans le SNC<i&gt; via</i&gt; L&#39;artère carotide interne

Animal models of central nervous system (CNS) diseases and, consequently, blood-brain barrier disruption diseases, require the delivery of exogenous ...

The overall goal of this surgical technique is to deliver exogenous substances into the mouse brain without penetrating the cranium. This method can help answer key questions in neuroscience regarding the action of drugs, pathogens, and exosomes in the brain. The main advantage of this technique is that it causes minimal trauma to the animal, preserves the integrity of the neuronal network, and achieves high concentrations of the substance in the brain.Begin by administering isoflurane mixed with oxygen to anesthetize the mouse in an induction chamber. After induction of anesthesia, transfer the animal to the surgical surface under a stereo microscope, and place the nose cone to maintain a surgical plane of anesthesia. Ensure that the respiratory rate is around one to two respirations per second without gasping.In addition, ensure that the animal does not react to whisker stimulation or a toe pinch. Use a sterile swab to apply a drop of ophthalmic lubricant on each eye to prevent dryness. After shaving the surgical site with a razor, prep the area with 70%ethanol and chlorhexidine.While looking through the stereo microscope, use surgical scissors and forceps to create a midline incision in the neck from above the breastbone to below the jaw. Use forceps to carefully separate fatty and connective tissue to expose the trachea. Then place a soft, round object of about zero point five centimeters in diameter under the back of the neck to extend the neck and further expose the area.Next, separate the tissue around the trachea using hooks or a tissue retractor. On the left side of the trachea, carefully tweeze apart the connective tissue to expose the left common carotid artery, or CCA. Using forceps, carefully remove all connective tissue to expose the CCA bifurcation in the beginning of both the external and internal carotid arteries.Once the ECA and ICA are exposed, use forceps to place a segment of one-centimeter nylon suture under the external carotid artery. Then place a second one-centimeter segment. Then tie a permanent knot at the highest point possible of the ECA.At the lowest point possible of the ECA, immediately above the CCA bifurcation, tie a looser, removable knot. Then use vessel clips to close the CCA at the lowest possible point, and to close the ICA. Properly closing the internal carotid artery before making a small cut in the external carotid artery will prevent bleeding.Now, using microdissection spring scissors, make a two-millimeter cut in the ECA between the two knots. It is important to keep the cuts small, as a large cut will cause leaking of the substance to be infused. Assemble the infusion system by attaching six inches of capillary tubing to a tuberculin syringe containing 250 microliters of the substance to be infused.Then gently insert the tip of the capillary tubing into the incision in the ECA. Keep advancing the tip until it reaches midway between the bifurcation and the clip on the CCA. Tie down the lower ECA knot.Verify that the knot is loose enough to allow capillary fluidity, and tight enough to prevent leaking. Then, remove the clip from the ICA. Gently apply pressure to the syringe piston, allowing substance infusion at approximately 10 microliters per second.Following infusion of the test substance, place the clip back on the ICA. Gently remove the capillary tubing. Then, tie down the lower ECA knot completely.Remove the clip from the ICA and the CCA. Then remove the retractor or tissue hooks and pillow. Clean the area to be sutured with sterile saline, and then close the incision using nylon sutures.After administering an approved anti-inflammatory agent and analgesic to alleviate post-operative discomfort, place the animal in a cage on a heating pad for at least one hour, and monitor recovery. This histogram demonstrates HIV detection by real-time PCRs seven days after viral infusion via the left internal carotid artery. As expected, levels of the virus are higher in the left hemisphere.This immunofluorescence image shows a mouse brain section depicting GFP tag CD63, an exosome marker, after extracellular vesicle infusion via the internal carotid artery. The CD63 expressing extracellular vesicles, shown in green, are associated with the mouse brain microvessel. Once mastered, this technique can be done in 20 minutes if it is performed properly.After its development, this technique paved the way for researchers in the field of neurodegenerative diseases to explore drug, pathogens, and exosome interactions in mice.

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Neuroscience

Mouse Microsurgery Infusion Technique for Targeted Substance Delivery into the CNS via the Internal Carotid Artery

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