Name: utilizing a cranial window to visualize the middle cerebral artery during endothelin-1 induced middle cerebral artery occlusion
Uploaded: 2013-02-22T12:00:00
Description: Watch this Scientific Journal Video about Utilizing a Cranial Window to Visualize the Middle Cerebral Artery During Endothelin-1 Induced Middle Cerebral Artery Occlusion at JoVE.com

This work was supported by grants from the American Heart Association Greater Southeast Affiliate (09GRNT2060421), the American Medical Association, and from the University of Florida Clinical and Translational Science Institute. Adam Mecca is a NIH/NINDS, NRSA predoctoral fellow (F30 NS-060335). Robert Regenhardt received predoctoral fellowship support from the University of Florida Multidisciplinary Training Program in Hypertension (T32 HL-083810).

This protocol was approved by the Institutional Animal Care and Use Committee (IACUC) at the University of Florida and is in compliance with the "Guide for the Care and Use of Laboratory Animals" (eighth edition, National Academy of Sciences, 2011).
Materials
<ol>
 <li>Animals: Eight-week-old, male, Sprague Dawley rats (Charles River Farms, Wilmington, MA, USA) weighing 250-300 g at the time of surgery. </li>
 <li>Anesthesia
 <ol class="lalpha">
 <li>Inhalation anesthesia system (VetEquip In...

<ol>
	<li>Levasseur, J. E., Wei, E. P., Raper, A. J., Kontos, A. A., Patterson, J. L. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Detailed+description+of+a+cranial+window+technique+for+acute+and+chronic+experiments.">Detailed description of a cranial window technique for acute and chronic experiments.</a> Stroke. 6, 308-317 (1975).</li><li>Baumbach, G. L., Dobrin, P. B., Hart, M. N., Heistad, D. D. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Mechanics+of+cerebral+arterioles+in+hypertensive+rats.">Mechanics of cerebral arterioles in hypertensive rats.</a> Circ. Res. 62, 56-64 (1988).</li><li>Regrigny, O., 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=Effects+of+melatonin+on+rat+pial+arteriolar+diameter+in+vivo.">Effects of melatonin on rat pial arteriolar diameter in vivo.</a> Br. J. Pharmacol. 127, 1666-1670 (1999).</li><li>Mecca, A. P., O'Connor, T. E., Katovich, M. J., Sumners, C. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Candesartan+pretreatment+is+cerebroprotective+in+a+rat+model+of+endothelin-1-induced+middle+cerebral+artery+occlusion.">Candesartan pretreatment is cerebroprotective in a rat model of endothelin-1-induced middle cerebral artery occlusion.</a> Exp. Physiol. 94, 937-946 (2009).</li><li>Mecca, A. 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=Cerebroprotection+by+angiotensin-(1-7)+in+endothelin-1-induced+ischaemic+stroke.">Cerebroprotection by angiotensin-(1-7) in endothelin-1-induced ischaemic stroke.</a> Exp. Physiol. 96, 1084-1096 (2011).</li></ol>

In summary, this cranial window preparation technique is very versatile as it can be altered to meet the needs of many experiments with minor modifications4, 5. For example, we have successfully monitored cerebral blood flow in specific MCA branches using laser doppler flowmetry to focus directly on a cerebral artery visualized through a cranial window (Mecca AP 2009 and 2011). In addition, a similar preparation with the dura incised can be used with topical administration of vasoactive compounds to create an ...

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<tbody>
 <tr>
 <td>Name of the reagent</td>
 <td>Company</td>
 <td>Catalogue number</td>
 <td>Comments (optional)</td>
 </tr>
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 <td>Inhalation anesthesia system</td>
 <td> VetEquip Inc., Pleasanton, CA, USA</td>
 <td>901806</td>
 <td></td>
 </tr>
 <tr>
 <td>Isoflurane anesthetic</td>
 <td>Baxter Pharmaceutics, Deerfield, IL, USA</td>
 <td>1001936060</td>
 <td></td>
 </tr>
 <tr>
 <td>Small animal stereotaxic system </td>
 <td>David Kopf Instruments, Tujunga, CA, USA</td>
 <td>900</td>
 <td></td>
 </tr>
 <tr>
 <td>Non-rupture ear bars, rat</td>
 <td>David Kopf Instruments, Tujunga, CA, USA</td>
 <td>957</td>
 <td></td>
 </tr>
 <tr>
 <td>Rat gas anesthesia head holder </td>
 <td>David Kopf Instruments, Tujunga, CA, USA</td>
 <td>1929</td>
 <td></td>
 </tr>
 <tr>
 <td>BAT-12 microprobe thermometer</td>
 <td>World Precision Instruments, Inc., Sarasota, FL, USA</td>
 <td>BAT-12</td>
 <td></td>
 </tr>
 <tr>
 <td>T/PUMP, Thermal blanket</td>
 <td>Gaymar Industries, Inc., Orchard Park, NY, USA</td>
 <td>T/PUMP, TP600</td>
 <td></td>
 </tr>
 <tr>
 <td>Metzenbaum Scissors</td>
 <td>World Precision Instruments, Inc., Sarasota, FL, USA</td>
 <td>501254</td>
 <td></td>
 </tr>
 <tr>
 <td>Iris forceps</td>
 <td>World Precision Instruments, Inc., Sarasota, FL, USA</td>
 <td>15915</td>
 <td></td>
 </tr>
 <tr>
 <td>Bulldog clamp retractors</td>
 <td>World Precision Instruments, Inc., Sarasota, FL, USA</td>
 <td>14119-G</td>
 <td></td>
 </tr>
 <tr>
 <td>10 &mu;l syringe 26-gaugue</td>
 <td>World Precision Instruments, Inc., Sarasota, FL, USA</td>
 <td>SGE010RNS</td>
 <td></td>
 </tr>
 <tr>
 <td>Bovie, high temperature cautery kit</td>
 <td>World Precision Instruments, Inc., Sarasota, FL, USA</td>
 <td>500392</td>
 <td></td>
 </tr>
 <tr>
 <td>Rat tooth forceps 0.12</td>
 <td>Stotz</td>
 <td>E1811</td>
 <td></td>
 </tr>
 <tr>
 <td>Micromotor drill</td>
 <td>Stoelting, Wood Dale, IL, USA</td>
 <td>51449</td>
 <td></td>
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 <td>0.8 mm round drill bur</td>
 <td>Roboz Surgical Instrument Co., Inc., Gaithersburg, MD, USA</td>
 <td>RS-6280C-1</td>
 <td></td>
 </tr>
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 <td>STORZ Bonn suturing forceps</td>
 <td>Bausch and Lomb, Inc., Rochester, NY, USA</td>
 <td></td>
 <td></td>
 </tr>
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 <td>Nylon Suture, size 3.0</td>
 <td>Oasis, Mettawa, IL, USA</td>
 <td>MV-663</td>
 <td></td>
 </tr>
 <tr>
 <td>Cotton swabs</td>
 <td>Fisher Scientific, Pittsburg, PA, USA</td>
 <td>22-029-488</td>
 <td></td>
 </tr>
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 <td>Puralube eye ointment</td>
 <td>Fisher Scientific, Pittsburg, PA, USA</td>
 <td>NC0138063</td>
 <td></td>
 </tr>
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 <td>Electric hair clippers </td>
 <td>Oster, Providence, RI, USA</td>
 <td>78005-301</td>
 <td></td>
 </tr>
 <tr>
 <td>ET-1 diluted to 80 &mu;M concentration in PBS </td>
 <td>American Peptide, Sunnyvale, CA, USA</td>
 <td>88-1-10A</td>
 <td></td>
 </tr>
 <tr>
 <td>Chlorhexidine, 2% </td>
 <td>Agrilabs, St. Joseph, MO, USA</td>
 <td>1040, Rev. 6-06, NAC No.: 10580322</td>
 <td></td>
 </tr>
 <tr>
 <td>Surgical microscope </td>
 <td>Seiler Instrument and Manufacturing, St. Louis, MO, USA</td>
 <td>Evolution xR6</td>
 <td></td>
 </tr>
 <tr>
 <td>Sony Handycam</td>
 <td>Sony, Minato, Tokyo, Japan</td>
 <td>HDR-SR12</td>
 <td></td>
 </tr>
 <tr>
 <td>Fiber optic illuminator </td>
 <td>TechniQuip Corp., Livermore, CA, USA</td>
 <td>FO1&#x2013;150</td>
 <td></td>
 </tr>
 <tr>
 <td>VLC media Player</td>
 <td>(Paris, France)</td>
 <td></td>
 <td></td>
 </tr>
 <tr>
 <td>Image J software</td>
 <td>U.S. National Institutes of Health, Bethesda, MA, USA</td>
 <td></td>
 <td></td>
 </tr>
 </tbody>
</table>

utilizing a cranial window to visualize the middle cerebral artery during endothelin-1 induced middle cerebral artery occlusion

Creation of a cranial window is a method that allows direct visualization of structures on the cortical surface of the brain1-3. This technique can be performed in many locations overlying the rat cerebrum, but is most easily carried out by creating a craniectomy over the readily accessible frontal or parietal bones. Most frequently, we have used this technique in combination with the endothelin-1 middle cerebral artery occlusion model of ischemic stroke to quantify the changes in middle cerebral artery vessel diameter that occur with injection of endothelin-1 into the brain parenchyma adjacent to the proximal MCA4, 5. In order to visualize the proximal portion of the MCA during endothelin -1 induced MCAO, we use a technique to create a cranial window through the temporal bone on the lateral aspect of the rat skull (Figure 1). Cerebral arteries can be visualized either with the dura intact or with the dura incised and retracted. Most commonly, we leave the dura intact during visualization since endothelin-1 induced MCAO involves delivery of the vasoconstricting peptide into the brain parenchyma. This bypasses the need to incise the dura directly over the visualized vessels for drug delivery. This protocol will describe how to create a cranial window to visualize cerebral arteries in a step-wise fashion, as well as how to avoid many of the potential pitfalls pertaining to this method.

Still images taken from the captured video show that the change in cerebral artery diameter after ET-1 injection can be readily appreciated using this cranial window technique (Figure 2). Within minutes of ET-1 injection, the vessel will begin to constrict. Eventually the vessels will be difficult to visualize and the brain tissue will become pale. After about 20 min the effects of ET-1 will diminish and the vessels will begin to dilate, gradually returning to baseline diameter after about 45 min. In add...

Watch this Scientific Journal Video about Utilizing a Cranial Window to Visualize the Middle Cerebral Artery During Endothelin-1 Induced Middle Cerebral Artery Occlusion at JoVE.com

Utilizing a Cranial Window to Visualize the Middle Cerebral Artery During Endothelin-1 Induced Middle Cerebral Artery Occlusion

This article describes a method for visualizing rat cerebral arteries through a cranial window using temporal craniectomy in order to view proximal portions of the middle cerebral artery (Figure 1). This versatile method can be combined with various techniques of drug delivery to measure cerebral artery reactivity in vivo.

Creation of a cranial window is a method that allows direct visualization of structures on the cortical surface of the brain1-3. This technique can be ...

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