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<article dtd-version="3.0" article-type="Invited Methods Article - JoVE Produced Video">
  <front>
    <journal-meta>
      <journal-id journal-id-type="publisher">jove</journal-id>
      <journal-id journal-id-type="email">editorial-office@jove.com</journal-id>
      <journal-title-group>
        <journal-title>Journal of Visualized Experiments</journal-title>
      </journal-title-group>
      <issn pub-type="ppub">1940-087X</issn>
      <issn pub-type="epub"></issn>
    </journal-meta>
    <article-meta>
      <article-id pub-id-type="publisher-id">JoVE59201</article-id>
      <article-id pub-id-type="manuscript">JoVE59201</article-id>
      <article-categories>
        <subj-group subj-group-type="Article Type">
          <subject>Invited Methods Article - JoVE Produced Video</subject>
        </subj-group>
        <subj-group subj-group-type="Category">
          <subject>JoVE Immunology and Infection</subject>
        </subj-group>
        <subj-group subj-group-type="Classification">
          <subject>Central Nervous System Diseases</subject>
          <subject>Brain Diseases</subject>
          <subject>Drug Therapy</subject>
          <subject>Drug Administration Routes</subject>
          <subject>Diseases</subject>
          <subject>Nervous System Diseases</subject>
          <subject>Chemicals and Drugs</subject>
          <subject>Nucleic Acids, Nucleotides, and Nucleosides</subject>
          <subject>Analytical, Diagnostic and Therapeutic Techniques and Equipment</subject>
          <subject>Therapeutics</subject>
        </subj-group>
      </article-categories>
      <title-group>
        <article-title>A positioning device for the placement of mice during intranasal siRNA delivery to the central nervous system</article-title>
      </title-group>
      <contrib-group>
        <contrib contrib-type="author" corresp="yes">
          <role content-type="1" />
          <name>
            <surname>Lee</surname>
            <given-names>Sang-Kyung</given-names>
            <prefix>Dr.</prefix>
          </name>
          <email>sangkyunglee@hanyang.ac.kr</email>
          <xref ref-type="aff" rid="aff1" />
        </contrib>
        <contrib contrib-type="author">
          <role content-type="2" />
          <name>
            <surname>Ullah</surname>
            <given-names>Irfan</given-names>
          </name>
          <email>irfanbiotech.malik@gmail.com</email>
          <xref ref-type="aff" rid="aff2" />
        </contrib>
        <contrib contrib-type="author">
          <role content-type="3" />
          <name>
            <surname>Chung</surname>
            <given-names>Kunho</given-names>
          </name>
          <email>kunho.chung.1207@gmail.com</email>
          <xref ref-type="aff" rid="aff3" />
        </contrib>
        <contrib contrib-type="author">
          <role content-type="4" />
          <name>
            <surname>Beloor</surname>
            <given-names>Jagadish</given-names>
          </name>
          <email>jagadishbeloor@gmail.com</email>
          <xref ref-type="aff" rid="aff4" />
        </contrib>
        <contrib contrib-type="author">
          <role content-type="5" />
          <name>
            <surname>Kumar</surname>
            <given-names>Priti</given-names>
          </name>
          <email>priti.kumar@yale.edu</email>
          <xref ref-type="aff" rid="aff5" />
        </contrib>
        <contrib contrib-type="editor">
          <name>
            <surname>Navani</surname>
            <given-names>Dipesh</given-names>
          </name>
          <degrees>Ph.D.</degrees>
          <email>dipesh.n@jove.com</email>
          <xref ref-type="aff" rid="aff6" />
        </contrib>
        <aff id="aff1">
          <country>KOREA, DEMOCRATIC PEOPLE'S REPUBLIC OF</country>
          <institution>Hanyang University</institution>
        </aff>
        <aff id="aff2">
          <institution>Hanyang University</institution>
          <institution content-type="dept">Chemical Engineering</institution>
          <addr-line content-type="addrline1">ITBT 1018, 222 Wangsimni-ro, Seongdong-gu</addr-line>
          <country>KOREA, REPUBLIC OF</country>
          <addr-line content-type="city">Seoul</addr-line>
          <addr-line content-type="zipcode">04763</addr-line>
          <institution>Hanyang University, Yale University</institution>
        </aff>
        <aff id="aff3">
          <institution>Hanyang University</institution>
          <institution content-type="dept">Chemical Engineering</institution>
          <addr-line content-type="addrline1">ITBT 1018, 222 Wangsimni-ro, Seongdong-gu</addr-line>
          <country>KOREA, REPUBLIC OF</country>
          <addr-line content-type="city">Seoul</addr-line>
          <addr-line content-type="zipcode">04763</addr-line>
          <institution>Hanyang University</institution>
        </aff>
        <aff id="aff4">
          <institution>Yale University</institution>
          <institution content-type="dept">Internal Medicine</institution>
          <addr-line content-type="addrline1">25 York street,</addr-line>
          <country>UNITED STATES</country>
          <addr-line content-type="city">New Haven</addr-line>
          <addr-line content-type="state">Connecticut</addr-line>
          <addr-line content-type="zipcode">06519</addr-line>
          <institution>Yale University</institution>
        </aff>
        <aff id="aff5">
          <institution>Yale University</institution>
          <institution content-type="dept">Internal Medicine</institution>
          <addr-line content-type="addrline1">25 York street,</addr-line>
          <country>UNITED STATES</country>
          <addr-line content-type="city">New Haven</addr-line>
          <addr-line content-type="state">Connecticut</addr-line>
          <addr-line content-type="zipcode">06519</addr-line>
          <institution>Yale University</institution>
        </aff>
        <aff id="aff6">
          <institution>JoVE</institution>
          <addr-line>1 Alewife Center</addr-line>
          <addr-line>Suite 200</addr-line>
          <addr-line>Cambridge, MA  02140</addr-line>
          <country>UNITED STATES</country>
        </aff>
      </contrib-group>
      <history>
        <date date-type="received">
          <day>04</day>
          <month>10</month>
          <year>2018</year>
        </date>
        <date date-type="rev-recd">
          <day>26</day>
          <month>11</month>
          <year>2018</year>
        </date>
        <date date-type="rev-recd">
          <day>20</day>
          <month>11</month>
          <year>2018</year>
        </date>
        <date date-type="accepted">
          <day>30</day>
          <month>11</month>
          <year>2018</year>
        </date>
      </history>
      <abstract>
        <p>Intranasal (IN) drug delivery to the brain has emerged as a promising method to bypass the blood-brain barrier for the delivery of drugs into the CNS. Our recent studies demonstrate the use of a peptide, RVG9R, incorporating the minimal receptor-binding domain of the Rabies virus glycoprotein, in eliciting delivery of siRNA into neurons in the brain. In this protocol, the peptide-siRNA formulation is delivered IN with a pipettor in the dominant hand, while the anesthetized mice are restrained by the scruff, using the non-dominant hand, in a ‘head down-and-forward position’ to avoid drainage into the lung and stomach upon inhalation. This precise gripping of mice can be learned, but is not easy and requires practice and skill to result in effective CNS uptake. Furthermore, the process is long-drawn requiring about 45 minutes for the administration of a total volume ~20-30 μl solution in 1-2 μl droplet volumes per inhalation with 3-4 minute rest periods between each inhalation. The objective of this study is to disclose a “mouse positioning device” that enables appropriate placement of mice for efficient IN administration of the peptide-siRNA formulation. The device was incorporated multiple features in the design of the device such 4 or 8 positioning chairs with adjustable height and tilt to restrain anesthetized mice in the head down-and-forward position enabling easy visualization of the mouse nares and a built-in heating-pad to maintain body temperature during procedure. Importantly, the ability to treat 4 or 8 mice simultaneously with RVG9R-siRNA complexes in this manner enabled studies on a much quicker time scale for the testing of an IN therapeutic siRNA approach. In conclusion, this device allows for appropriate and controlled mouse head positioning for IN application for the CNS delivery of RVG9R-siRNA and other therapeutic molecules such as nanoparticles or antibodies.</p>
      </abstract>
      <kwd-group>
        <kwd>Intranasal administration, brain delivery, siRNA, RVG9R peptide, mouse positioning device, nose-to-brain</kwd>
      </kwd-group>
      <funding-group />
      <counts>
        <fig-count count="0" />
      </counts>
      <custom-meta-group>
        <custom-meta>
          <meta-name>Please indicate whether this article will be Standard Access or Open Access. </meta-name>
          <meta-value>Standard Access (US$2,400);</meta-value>
        </custom-meta>
        <custom-meta>
          <meta-name>Please indicate the &lt;b&gt; city, state/province, and country&lt;/b&gt; where this article will be &lt;b&gt;filmed&lt;/b&gt;. Please do not use abbreviations.</meta-name>
          <meta-value>Yale University, New Haven, Connecticut (06519), USA</meta-value>
        </custom-meta>
        <custom-meta>
          <meta-name>Black and White Image Count</meta-name>
          <meta-value>0</meta-value>
        </custom-meta>
        <custom-meta>
          <meta-name>Color Image Count</meta-name>
          <meta-value>0</meta-value>
        </custom-meta>
        <custom-meta>
          <meta-name>Acquisitions Editor</meta-name>
          <meta-value>Jaydev Upponi</meta-value>
        </custom-meta>
        <custom-meta>
          <meta-name>Assistant who entered lead</meta-name>
          <meta-value>Angela Forgione</meta-value>
        </custom-meta>
        <custom-meta>
          <meta-name>ALA Access Type</meta-name>
          <meta-value>JPV Standard Access ($2400)</meta-value>
        </custom-meta>
      </custom-meta-group>
    </article-meta>
  </front>
  <body>
    <fig>
      <label>Figure 1</label>
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      <label>Figure 4</label>
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    </fig>
    <table-wrap>
      <label>Table of Materials</label>
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    <fig>
      <label>Figure 1</label>
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    <fig>
      <label>Figure 2</label>
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    <fig>
      <label>Figure 3</label>
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    <fig>
      <label>Figure 4</label>
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    </fig>
    <table-wrap>
      <label>Table of Materials</label>
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    <table-wrap>
      <label>Table of Materials</label>
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    </table-wrap>
  </body>
</article>