This work was funded by a grant from HEBEI Provincial Department of Human Resources and Social Security (CY201605) and a grant from Natural Science Foundation of Hebei Province (H2017206101), and we are very grateful to Dr. for Guangping Gao, who provided the AAV for this study.

FVB/NJ mice were bred in the animal facility of Key Laboratory of Hebei Neurology. All mouse experiments were approved by the Second Hospital of Hebei Medical University Ethics Committee and carried out according to the regulations of laboratory animal management promulgated by the Ministry of Science and Technology of the People&#39;s Republic of China.
1. Preparation of 20% Lidocaine Hydrochloride Stock Solution
<ol>
	<li>Weigh 2 g of lidocaine hydrochloride. Add 5&#8211;6 mL of sterile wa...

<ol>
	<li>Murlidharan, G., Samulski, R. J., Asokan, A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Biology+of+adeno-associated+viral+vectors+in+the+central+nervous+system.">Biology of adeno-associated viral vectors in the central nervous system.</a> Frontiers in Molecular Neuroscience. 7, 76 (2014).</li><li>Lentz, T. B., Gray, S. J., Samulski, R. J. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Viral+vectors+for+gene+delivery+to+the+central+nervous+system.">Viral vectors for gene delivery to the central nervous system.</a> Neurobiology Disease. 48 (2), 179-188 (2012).</li><li>Yang, B., 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=Global+CNS+transduction+of+adult+mice+by+intravenously+delivered+rAAVrh.8+and+rAAVrh.10+and+nonhuman+primates+by+rAAVrh.10.">Global CNS transduction of adult mice by intravenously delivered rAAVrh.8 and rAAVrh.10 and nonhuman primates by rAAVrh.10.</a> Molecular Therapy. 22 (7), 1299-1309 (2014).</li><li>Guo, 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=A+Single+Injection+of+Recombinant+Adeno-Associated+Virus+into+the+Lumbar+Cistern+Delivers+Transgene+Expression+Throughout+the+Whole+Spinal+Cord.">A Single Injection of Recombinant Adeno-Associated Virus into the Lumbar Cistern Delivers Transgene Expression Throughout the Whole Spinal Cord.</a> Molecular Neurobiology. 53 (5), 3235-3248 (2016).</li><li>Hastie, E., Samulski, R. J. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Adeno-associated+virus+at+50:+a+golden+anniversary+of+discovery,+research,+and+gene+therapy+success--a+personal+perspective.">Adeno-associated virus at 50: a golden anniversary of discovery, research, and gene therapy success--a personal perspective.</a> Human Gene Therapy. 26 (5), 257-265 (2015).</li><li>Hocquemiller, M., Giersch, L., Audrain, M., Parker, S., Cartier, N. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Adeno-Associated+Virus-Based+Gene+Therapy+for+CNS+Diseases.">Adeno-Associated Virus-Based Gene Therapy for CNS Diseases.</a> Human Gene Therapy. 27 (7), 478-496 (2016).</li><li>Tanguy, 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=Systemic+AAVrh10+provides+higher+transgene+expression+than+AAV9+in+the+brain+and+the+spinal+cord+of+neonatal+mice.">Systemic AAVrh10 provides higher transgene expression than AAV9 in the brain and the spinal cord of neonatal mice.</a> Frontiers in Molecular Neuroscience. 8, 36 (2015).</li><li>Federic, T., 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=Robust+spinal+motor+neuron+transduction+following+intrathecal+delivery+of+AAV9+in+pigs.">Robust spinal motor neuron transduction following intrathecal delivery of AAV9 in pigs.</a> Gene Therapy. 19, 852-859 (2012).</li><li>Ayers, J. I., 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=Widespread+and+efficient+transduction+of+spinal+cord+and+brain+following+neonatal+AAV+injection+and+potential+disease+modifying+effect+in+ALS+mice.">Widespread and efficient transduction of spinal cord and brain following neonatal AAV injection and potential disease modifying effect in ALS mice.</a> Molecular Therapy. 23 (1), 53-62 (2015).</li><li>Li, D., 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=Slow+Intrathecal+Injection+of+rAAVrh10+Enhances+its+Transduction+of+Spinal+Cord+and+Therapeutic+Efficacy+in+a+Mutant+SOD1+Model+of+ALS.">Slow Intrathecal Injection of rAAVrh10 Enhances its Transduction of Spinal Cord and Therapeutic Efficacy in a Mutant SOD1 Model of ALS.</a> Neuroscience. 365, 192-205 (2017).</li><li>Borel, F., 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=Therapeutic+rAAVrh10+Mediated+SOD1+Silencing+in+Adult+SOD1(G93A)+Mice+and+Nonhuman+Primates.">Therapeutic rAAVrh10 Mediated SOD1 Silencing in Adult SOD1(G93A) Mice and Nonhuman Primates.</a> Human Gene Therapy. 27 (1), 19-31 (2016).</li><li>Fairbanks, C. A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Spinal+delivery+of+analgesics+in+experimental+models+of+pain+and+analgesia.">Spinal delivery of analgesics in experimental models of pain and analgesia.</a> Advanced Drug Delivery Reviews. 55 (8), 1007-1041 (2003).</li><li>Hylden, J. L., Wilcox, G. L. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Intrathecal+morphine+in+mice:+a+new+technique.">Intrathecal morphine in mice: a new technique.</a> European Journal of Pharmacology. 67, 313-316 (1980).</li><li>Wang, H., 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=Widespread+spinal+cord+transduction+by+intrathecal+injection+of+rAAV+delivers+efficacious+RNAi+therapy+for+amyotrophic+lateral+sclerosis.">Widespread spinal cord transduction by intrathecal injection of rAAV delivers efficacious RNAi therapy for amyotrophic lateral sclerosis.</a> Human Molecular Genetics. 23 (3), 668-681 (2014).</li><li>Wang, 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=scAAV9-VEGF+prolongs+the+survival+of+transgenic+ALS+mice+by+promoting+activation+of+M2+microglia+and+PI3K/Akt+pathway.">scAAV9-VEGF prolongs the survival of transgenic ALS mice by promoting activation of M2 microglia and PI3K/Akt pathway.</a> Brain Research. 1648, 1-10 (2016).</li></ol>

Technically, there are several critical steps during the IT injection in awake mice. First, proper gesture and firm control of the mice throughout the entire operation is a prerequisite for successful delivery. Second, the most difficult point is feeling the intervertebral space with the needle tip, as it is necessary not to insert too deeply without resistance or insert forcibly under strong resistance in the case of injuring the animals or bending the needle tip. Third, although the transient paralysis due to lidocaine...

AAV can mediate long-term and widespread gene expression in the CNS transduction with few side effects, and therefore has become one of the most promising vehicles for gene therapy to treat CNS diseases including amyotrophic lateral sclerosis (ALS), Huntington&#39;s disease (HD), Alzheimer&#39;s disease (AD), lysosomal storage diseases (LSD), Gaucher disease (GD), and neuronal ceroid lipofuscinosis (NCL)1. Presently, more than 100 AAV serotypes have been isolated from humans and animals. Among these, at least 12 have been used in preclinical and clinical trials, including the most commonly used gene vectors such as AAV1, 2, 4, 5, 6, 8, 9, rAAVrh.8, and rAAVrh.101,2,3,4,5,6.
Different CNS diseases require different AAV delivery strategies due to the various affected CNS regions and cell types. The CNS regions and cell types that AAV can transduce varies depending on the serotype as well as delivery method. For example, rAAVrh10 has been shown to transduce predominantly astrocytes when delivered by systemic intravenous injection (IV), whereas it transduced both neurons and glia when delivered by intrathecal injection4,7. Additionally, parenchyma injection resulted in local transduction to the vicinity of the injection site, whereas injection into the cerebrospinal fluid (CSF) through intraventricular or intrathecal injection resulted in widespread CNS transduction8. Studies have also demonstrated therapeutic potency of AAV-delivered gene therapy in neurodegenerative disorders by IT administration9,10,11. In diseases that affect broad areas of the CNS such as ALS, intrathecal injection into the CSF has been shown to cover most areas that are afflicted by the disease with a lower dose, compared to a systemic delivery method4,10. Recent studies have also shown that lumbar puncture can be used to inject AAV in mouse models for ALS, which avoids potential injuries associated with laminectomy and intrathecal catheterization4.
Experimental direct lumbar puncture was first used to deliver agents, especially anesthetics, to the spinal cord for analgesia and anesthesia in 188512,13. In this report, we illustrate the lumbar puncture IT injection method in adult mice with the aid of 1% lidocaine hydrochloride, a local amide-derived anesthetic, in the injection solution to evaluate and monitor injection quality. Successful injections were marked by lidocaine-induced transient paralysis, whereas failed injections did not show this behavior. We classified the level of transient weakness as one of five grades to help predict the injection efficiency. Finally, we show that the rAAVrh10 transduction level may be predicted by the grade of paralysis. Therefore, this intrathecal AAV delivery method can be used to enhance AAV-mediated gene-delivery for experimental therapy of CNS diseases.

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		<col />
		<col />
		<col />
		<col />
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	<tbody>
		<tr height="32">
			<td height="32" style="height:32px;width:284px;">FVB/NJ mice</td>
			<td style="width:219px;">Charles River Laboratories China</td>
			<td style="width:199px;"></td>
			<td style="width:131px;"></td>
		</tr>
		<tr height="32">
			<td height="32" style="height:32px;">Lidocaine hydrochloride monohydrate</td>
			<td>HEOWNS</td>
			<td>73-78-9</td>
			<td></td>
		</tr>
		<tr height="47">
			<td height="47" style="height:47px;">AAV</td>
			<td style="width:417px;">Viral Vector Core of the Gene Therapy Center at University of Massachusetts Medical School</td>
			<td></td>
			<td></td>
		</tr>
		<tr height="32">
			<td height="32" style="height:32px;">25&#181;L&#160; Hamilton syringe/27-30g needle</td>
			<td>GASTIGHT</td>
			<td>1702</td>
			<td></td>
		</tr>
		<tr height="32">
			<td height="32" style="height:32px;">O.C.T compond</td>
			<td>SAKURA</td>
			<td>4583</td>
			<td></td>
		</tr>
		<tr height="32">
			<td height="32" style="height:32px;">H 2O 2</td>
			<td>SHUI HUAN PAI</td>
			<td>170401</td>
			<td></td>
		</tr>
		<tr height="32">
			<td height="32" style="height:32px;">Goat serum</td>
			<td>Solarbio</td>
			<td>S9070</td>
			<td></td>
		</tr>
		<tr height="32">
			<td height="32" style="height:32px;">Triton X-100</td>
			<td>LIFE SCIENCES</td>
			<td>T8200</td>
			<td></td>
		</tr>
		<tr height="32">
			<td height="32" style="height:32px;">Rabbit anti-GFP</td>
			<td>Life tech</td>
			<td>G10362</td>
			<td>1:333 dilution</td>
		</tr>
		<tr height="32">
			<td height="32" style="height:32px;">The second antibody (goat-anti rabbit)</td>
			<td>Jackson Immuno Research</td>
			<td>111-005-144</td>
			<td>1:1000 dilution</td>
		</tr>
		<tr height="32">
			<td height="32" style="height:32px;">VECTASTAIN ABC REAGENT</td>
			<td>Vector Lab</td>
			<td>PK-6100</td>
			<td></td>
		</tr>
		<tr height="32">
			<td height="32" style="height:32px;">ImmPACT DAB Peroxidase Substrate Kit</td>
			<td>Vector Lab</td>
			<td>SK-4105</td>
			<td></td>
		</tr>
		<tr height="32">
			<td height="32" style="height:32px;">Mounting medium for fluorescence with DAPI</td>
			<td>Vectorshield</td>
			<td>H-1200</td>
			<td></td>
		</tr>
		<tr height="32">
			<td height="32" style="height:32px;width:284px;">NaCl</td>
			<td>Yong Da Chemical</td>
			<td style="width:199px;"></td>
			<td style="width:131px;"></td>
		</tr>
		<tr height="32">
			<td height="32" style="height:32px;width:284px;">NaH2PO4&#183;2H2O</td>
			<td>Yong Da Chemical</td>
			<td style="width:199px;"></td>
			<td style="width:131px;"></td>
		</tr>
		<tr height="32">
			<td height="32" style="height:32px;width:284px;">Na2HPO4&#183;12H2O</td>
			<td>Yong Da Chemical</td>
			<td style="width:199px;"></td>
			<td style="width:131px;"></td>
		</tr>
		<tr height="32">
			<td height="32" style="height:32px;">Paraformaldehyde</td>
			<td>Yong Da Chemical</td>
			<td>307699</td>
			<td></td>
		</tr>
		<tr height="32">
			<td height="32" style="height:32px;">Adhesion Microscope Slides</td>
			<td>CITOGLAS</td>
			<td>17083</td>
			<td>25*75 mm</td>
		</tr>
		<tr height="32">
			<td height="32" style="height:32px;">SUPER-SLIP MICRO-GLAS</td>
			<td>Electro Microscopy Siences</td>
			<td>72236-60</td>
			<td>24*60 mm</td>
		</tr>
		<tr height="32">
			<td height="32" style="height:32px;">15 ml Centrifuge tube</td>
			<td>CORNING</td>
			<td>430790</td>
			<td></td>
		</tr>
		<tr height="32">
			<td height="32" style="height:32px;">96 well cell culture cluster</td>
			<td>Coster</td>
			<td>3599</td>
			<td></td>
		</tr>
		<tr height="32">
			<td height="32" style="height:32px;">24 well cell culture cluster</td>
			<td>Coster</td>
			<td>3524</td>
			<td></td>
		</tr>
		<tr height="32">
			<td height="32" style="height:32px;">70% Ethanol</td>
			<td>WEN ZHI</td>
			<td></td>
			<td></td>
		</tr>
		<tr height="32">
			<td height="32" style="height:32px;">Gauze</td>
			<td>Wei AN</td>
			<td>05171112</td>
			<td>8cm*10cm*12cm</td>
		</tr>
		<tr height="32">
			<td height="32" style="height:32px;">1mL syringe</td>
			<td>Hong Da</td>
			<td></td>
			<td></td>
		</tr>
		<tr height="32">
			<td height="32" style="height:32px;width:284px;">Microtubes</td>
			<td>Plasmed</td>
			<td></td>
			<td></td>
		</tr>
		<tr height="32">
			<td height="32" style="height:32px;">Micropipet&#160;</td>
			<td>eppendorf</td>
			<td></td>
			<td></td>
		</tr>
		<tr height="32">
			<td height="32" style="height:32px;">Peppet tips</td>
			<td>Rainin</td>
			<td></td>
			<td></td>
		</tr>
		<tr height="32">
			<td height="32" style="height:32px;">Centirifuge</td>
			<td>eppendorf</td>
			<td>5427R</td>
			<td></td>
		</tr>
		<tr height="32">
			<td height="32" style="height:32px;">Regerator</td>
			<td>Haier</td>
			<td>BCD-539WT</td>
			<td></td>
		</tr>
		<tr height="32">
			<td height="32" style="height:32px;">Filter</td>
			<td>MILLEX GP</td>
			<td>R4PA42342</td>
			<td></td>
		</tr>
		<tr height="32">
			<td height="32" style="height:32px;width:284px;">Pump</td>
			<td style="width:219px;">LongerPump</td>
			<td style="width:199px;">BT-100-2J/YZ1515X</td>
			<td style="width:131px;"></td>
		</tr>
		<tr height="32">
			<td height="32" style="height:32px;">Microscope</td>
			<td>Olympus</td>
			<td>BX53</td>
			<td></td>
		</tr>
		<tr height="32">
			<td height="32" style="height:32px;width:284px;">Freezing-microtome</td>
			<td style="width:219px;">Leica</td>
			<td style="width:199px;">CM1520</td>
			<td style="width:131px;"></td>
		</tr>
	</tbody>
</table>

direct intrathecal injection of recombinant adeno-associated viruses in adult mice

Intrathecal (IT) injection of adeno-associated virus (AAV) has drawn considerable interest in CNS gene therapy by virtue of its safety, noninvasiveness, and excellent transduction efficacy in the CNS.&#160;Previous studies have demonstrated the therapeutic potency of AAV-delivered gene therapy in neurodegenerative disorders by IT administration. However, high rates of unpredictable failure due to the technical limitation of IT administration in small animals have been reported. Here, we established a scoring system to indicate the success extent of lumbar puncture in small animals by adding 1% lidocaine hydrochloride into the injection solution. We further show that the extent of transient weakness following injection can predict the transduction efficiency of AAV. Thus, this IT injection method can be used to optimize therapeutic trials in mouse models of CNS diseases that afflict wide regions of the CNS.

Mice showed different degrees of transient weakness right after IT injection of AAV solution in 1% lidocaine hydrochloride due to various quality of intrathecal injection. According to the semi-quantitative 5-grade scoring system we have established, we tested the transduction patterns of AAV in mice with different degrees of lidocaine-induced limb weakness (score 0, n = 2; score 1, n = 1; score 4, n = 4; score 5, n = 3). EGFP immunostaining of spinal cords showed either no or little tran...

Watch this Scientific Journal Video about Direct Intrathecal Injection of Recombinant Adeno-associated Viruses in Adult Mice at JoVE.com

Direct Intrathecal Injection of Recombinant Adeno-associated Viruses in Adult Mice

Here we present a direct intrathecal injection technique using 1% lidocaine hydrochloride in a viral solution to ensure efficient adeno-associated virus delivery to small animals and establish a scoring system to predict transduction efficiency in the central nervous system according to the degree of transient weakness induced by lidocaine.

Intrathecal (IT) injection of adeno-associated virus (AAV) has drawn considerable interest in CNS gene therapy by virtue of its safety, noninvasiveness, ...

This method facilitates the efficient intrathecal delivery of adeno-associated viral transgenes in awake small animals for the experimental treatment of central nervous system diseases such as ALS. The main advantage of this method is that the viral solution includes 1%lidocaine hydrochloride which induces a transient paralysis after a successful injection. Although this method provide a visual indicator for judging the success of the intrathecal automated situation, sufficient practice is essential for improving the success rate of the delivery.Before beginning the procedure, attach a 27 gauge needle to a 25 microliter Hamilton syringe and align the beveled tip of the needle with the volumetric scale on the syringe. Then carefully load 8 microliters of 4 x 10 to the 10th genome copies of the virus solution into the syringe, taking care to avoid bubbles. Next, place an awake 30 to 70 day old 12 to 30 gram mouse on a bed piece in the prone position in a biosafety hood and cover the upper body with sterile gauze to calm the mouse and to avoid being bitten.Firmly grip the mouse on its pelvic girdle with the thumb on one side and the forefinger and middle finger on the other side keeping the skin between the bilateral pelvic girdles taut with a thumb and forefinger. Then shave the fur between the bilateral pelvic girdles and sterilize the exposed skin with an iodine based scrub and 70%ethanol. For direct intrathecal delivery of the adeno-associated virus solution, palpate the intervertebral space along the midline between the bilateral pelvic girdles and use a fingernail to depress an indentation into the skin to indicate the L5 to L6 intervertebral space.Rotate the base of the tail slightly and gently to reveal the midline of the spine and adjust the bevel of the needle toward the head of the animal. With the mouse fixed firmly into position, align the needle along the midline of the spine and insert the needle gently and vertically into the center of the indentation keeping the syringe in a central sagittal plane. When the needle comes into contact with the bone, slowly decrease the angle to approximately 30 degrees and slip the needle into the intervertebral space.A sudden tail flick is a sign of a successful entry into the intradural space. When the needle enters the intervertebral space, the tip will feel firmly clamped. Inject the vector solution, and maintain the needle within the intradural space for one minute.Then slowly withdraw the needle with rotation to minimize leakage. Immediately score the transient weakness of the mouse limbs to evaluate the injection quality and return the mouse to its cage for recovery from the paralysis. At the appropriate experimental endpoint, fix the limbs and head of the injected mouse in the prone position on a foam box cover and use scissors to strip and remove the skin from the head to the sacrum.Clip the skull between the eyes, cut along the midline of the skull and the horizontal line above the cerebellum and open the skull on each side. Use tweezers to remove the occipital bone and use ophthalmic scissors to open the spinal canal bilaterally. Cut the ribs on both sides and carefully remove the upper section of vertebrae.Use curved tweezers to lift the brain and sever the nerves of the skull base. Then carefully extract the whole brain and spinal cord and fix the tissues in 4%paraformaldehyde for 24 hours. At the end of the fixation period, cryoprotect the brain and cervical and lumbar spinal cord in 30%sucrose solution overnight at 4 degrees Celsius followed by embedding in optimum cutting temperature compound before snap freezing in liquid nitrogen.Then use a cryostat to obtain 25 micrometer thick sections of each tissue, storing the frozen sections in 0.01 molar PBS at 4 degrees Celsius as they are obtained. For immunohistochemical analysis of the tissues, pretreat the free floating sections with 1%hydrogen peroxide for 10 minutes followed by a 10 minute wash in PBS. Next incubate the samples in blocking solution containing 5%serum and 0.3%non-ionic detergent in PBS for 1 hour followed by overnight labeling at 4 degrees Celsius with the appropriate primary antibodies of interest.The next day, wash the sections with 3 10 minute washes in PBS plus Tween and incubate the slides with the appropriate corresponding biotinylated secondary antibodies at room temperature for 1 hour after the last wash. At the end of the incubation, wash the sections 3 times in fresh PBST as just demonstrated and incubate the samples with infinity biotin peroxidase complex for 40 minutes followed by staining with an appropriate chromogenic agent. Mount the labeled sections onto glass microscope slides and soak the slides in anhydrous ethanol for 5 minutes once the mounted sections have fully dried.Next soak the slides in xylene for 10 minutes and seal them with an appropriate mounting medium. Then image the slides with a light microscope equipped with a charge coupled device at 100, 200, and 400 times magnifications. eGFP immunostaining of cervical and lumbar spinal cord tissue sections reveals little or no transduction in the lumbar spinal cord of mice with a weakness score of 0, slightly enhanced transduction in mice with a weakness score of 1, and strong and widespread transductions in mice with a weakness score of 4 or 5.Quantification of the GFP staining intensity of spinal cord tissue sections from mice displaying various degrees of transient limb weakness indicates that the severity of the weakness after the virus solution injection closely correlates with the extent of the spinal cord transduction. In the brain, robust eGFP signals are detected in the olfactory bulb, dorsolateral prefrontal cortex, dentate gyrus, and the CA3 zone of the hippocampus, cerebellar cortex, and marginal areas of the brainstem, including the facial nucleus, choroid plexus, and the ependymal epithelial cells. Motor neurons in the anterior horns are also strongly transduced in different levels of the spinal cord.Moreover, in the cortex, GFP positive neurons including pyramidal cells are detected as well as various glial cell types including microglia, astrocytes, and oligodendrocytes. This technique enables researchers in the neurology field to explore the therapeutic effects of direct intrathecal delivery in small awake animals on central nervous system diseases.

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35.2K 조회수.  Second Hospital of Hebei Medical University. 이 방법은 ALS와 같은 중추 신경계 질환의 실험 적 치료를 위해 작은 동물을 깨우는 아데노 관련 바이러스 성 바이러스 성 트랜스게놈의 효율적인 장내 전달을 용이하게한다. 이 방법의 주요 장점은 바이러스 용액이 성공적인 주입 후 일시적인 마비를 유도하는 1 % 리도카인 염산염을 포함한다는 것입니다. 이 방법은 인트라더칼 자동화 상황의 성공을 판단하기위한 시각적 지?...