Este estudo foi financiado pelo National Institutes of Health (Grant 1UM1HL112856).

Todos os procedimentos e protocolos foram aprovados pelo Comitê de Cuidados e Uso da Universidade de Illinois, em Chicago animal Institucional. As experiências aqui descritas são experiências agudas não-sobrevivência, e não houve uso de pomada. Manutenção de condições estéreis só ocorre quando os instrumentos cirúrgicos são lavados com etanol 70% em DIH 2 O. Sacrifício de ratos no final da experiência ocorreu através de sobredosagem de IV cetamina / xilazina. 1. Preparação de Instrumentos e Produtos Químicos <ol><li> Preparar a solução de estoque de 0,05 M de 5-HT de HCl em PBS. Depois dilui-se estoque com PBS a uma concentração final de 5-HT de 0,203 mM. </li><li> Diluir dronabinol em óleo de sésamo com uma concentração de 20 ug / uL. </li><li> Corte de 20 cm de comprimento de polietileno (PE) -50 tubagem (ID 0,58 milímetros, 0,965 milímetros OD). Em uma extremidade do tubo, corte uma ponta bisel com uma tesoura, e na outra ponta quadrada, inserir uma agulha de 23 g. Ligue este 23 g agulha para um 1 mlseringa, e encha-o com 0,3 ml de 50 u / ml de heparina. </li><li> Corte quatro pedaços de fio de seda 4-0 trançado e definido para o lado. </li><li> Esterilizar tesouras cirúrgicas, 2 fórceps Graefe curvas, e 3 grampos de micro com etanol a 70% em DIH 2 O. </li></ol> 2. O cateterismo de veia femoral Protocolo para o cateterismo é modificado a partir Jespersen et al 21. <ol><li> Anestesiar uma ratazana Sprague-Dawley com intraperitoneal (IP) de cetamina / xilazina (100 mg / kg: 10 mg / kg). Aperte o dedo do pé do rato e observar qualquer movimento para confirmar um nível adequado de anestesia. Raspar o aspecto ventral da coxa esquerda, e do aspecto ventral do pescoço. Se necessário pelo Comitê de Cuidados e Uso de Animais Institucional do local, aplicar pomada para evitar o ressecamento. </li><li> Rat seguro em decúbito dorsal em uma placa cirúrgica. Utilize uma tesoura cirúrgica para cortar a pele na coxa posterior esquerda. </li><li> Utilizando uma pinça, conduta dissectio contuson do músculo superficial para expor a veia femoral. Utilizar as pinças para separar a veia a partir da artéria femoral, e colocar dois segmentos em torno da veia femoral. </li><li> Use a pinça Graefe curvas para puxar-up da veia para parar o fluxo de sangue. Usando uma seringa de agulha G 22, por punção da veia femoral e, em seguida, insira a ponta chanfrada do tubo PE-50 na veia. </li><li> Verificar se o tubo PE-50 foi inserida correctamente por retracção do êmbolo da seringa. O sangue deve ser visto de entrar no tubo de PE-50. Amarre 2 nós ao redor da veia e PE-50 tubos usando as duas threads. </li></ol> 3. 5-HT induzido por Apnea via IV Infusion <ol><li> Coloque um medidor de tensão piezoelétrico em torno do rato para medir a respiração. </li><li> Altere as configurações para o nível de amplificação do amplificador eletrônico via software amplificador no computador: para amplificar (100x) e filtro passa-banda (10/01 Hz), os sinais respiratórios obtidos a partir do medidor de tensão, ajuste &quot;filtro highpas&quot; at &amp; #8220; AC @ 1 HZ &quot;e&quot; Filter Lowpass &quot;at&quot; 10 Hz &quot;, e definir a amplificação através da introdução de&quot; 10 &quot;para o&quot; ganho inicial &quot;e&quot; 100 &quot;para&quot; Total Gain &quot;.. </li><li> Altere as configurações de taxa de amostragem, abrindo &quot;Entrada Analógica&quot; via software de gravação no computador. Para digitalizar (500 taxa de amostragem Hz) usando um conversor analógico-digital, &quot;sample&quot; at &quot;1.000 Hz&quot; e &quot;pular&quot; todos os outros pontos de gravação por &quot;1&quot; (taxa de amostragem eficaz 500 Hz), e depois gravar o sinal usando o software de gravação. </li><li> Remover a seringa de 1 ml a partir do cateter e inserir uma seringa de vidro de 500 mL cheio com precisão de 0,203 mM de 5-HT no interior do cateter. </li><li> Coloque a 500 ul seringa de vidro de precisão em uma bomba de infusão. Infundir 12,5 ug / kg por 350 ul / kg de uma solução de 5-HT a uma taxa de 63 ml / hr. Executar várias infusões e observar apnéia em rato, que é visto umsa pausa na respiração (≥ 2,5 seg) nos sinais respiratórios gravados no monitor do computador. </li><li> Antes de prosseguir com a cirurgia de pescoço, monitorar o padrão respiratório e verificar se há reflexo dor de toe pitada no rato. Se a respiração for irregular, ou se existe um reflexo dor de aperto do dedo do pé, administrar IP cetamina / xilazina (100 mg / kg: 5 mg / kg) e, em seguida, voltar a confirmar a um nível adequado de anestesia. </li></ol> 4. Neck Surgery para expor Nodoso Ganglia <ol><li> Rat seguro em decúbito dorsal em uma placa cirúrgica. Faça um corte longitudinal mediana usando uma tesoura cirúrgica no pescoço. </li><li> Usando dissecção romba no músculo platisma (usando dois micro grampos para manter esse músculo clara do sítio cirúrgico), expor os músculos sternohyoideus e omohyoideus. Separe esses músculos para expor a artéria carótida interna e um de seus ramos, a artéria pterigopalatina. </li><li> Observe o nervo vago, uma vez que corre ao longo da artéria carótida interna e, em seguida, ao longo th<html> e pterygopalatine artéria. Observe como o nervo vago e artéria pterigopalatina, juntamente com o nervo glossofaríngeo (IX nervo craniano) e do nervo acessório (XI nervo craniano), digite o forame lacerados posteriores na base do crânio. </li><li> Observe que o gânglio nodoso é exibido como um inchaço do nervo vago direito antes de entrar no forame posterior dilacerado. Note também que os ramos da faringe e da laringe nervosas saem os aspectos anterior e posterior, respectivamente, do gânglio nodoso. 5 </li><li> Usando a pinça Graefe, separar o nervo vago a partir das artérias, e coloque um pedaço de fio ao redor do nervo vago. Coloque um grampo na linha para aplicar uma leve tensão no nervo vago, e limpar o gânglio nodoso de qualquer tecido conjuntivo para proporcionar menor resistência ao injetar. </li><li> Repita apnéia 5-HT induzido conforme previsto no ponto 3 para confirmar que nenhum dano foi feito para o gânglio nodoso. </li></ol>itle &quot;&gt; 5. Intranodose Ganglia Injeção de Dronabinol <ol><li> Encha a 10 ul precisão gastight seringa de vidro afixado com um 28 g de meia polegada agulha de seringa feito por encomenda com um 35 ° chanfradas ponta com 5 mL de dronabinol em óleo de gergelim. </li><li> Coloque um grampo micro no segmento de aplicar uma leve tensão no nervo vago, e perfurar o gânglio nodoso tomando cuidado para não perfurar através dele. Pressione a seringa lentamente (≥ 60 seg) para injetar todo o conteúdo da seringa. Note-se que alguns dos conteúdos de seringas vai vazar para fora do gânglio. </li><li> Apneia Repita 5-HT-induzida como indicado na seção 3. </li></ol>

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The authors declare that they have no competing financial interests.

Os passos críticos para o sucesso da injecção de neurotransmissores na gânglios nodosos são: 1) a identificação e a limpeza do tecido conjuntivo fora do gânglios nodosos; 2) confirmando a integridade do gânglio nodoso antes da injecção; 3) e usando uma agulha de calibre pequeno para injectar delicadamente em, mas não completamente, através de punção, os gânglios nodosos. O nervo vago inerva muitos órgãos no pescoço e abdômen, e transmite informações importantes, tais co...

Afferent signaling via the vagus nerve (cranial nerve X) transmits important general visceral information to the central nervous system (CNS) from baro-, chemo-, hepatic osmo-, cardiac, pulmonary, and gastric receptors located in the organs of the abdomen and thorax. The vagus nerve communicates information from stimuli such as heart rate, blood pressure, bronchopulmonary irritation, and gastrointestinal distension to the nucleus of solitary tract (NTS) of the medulla. The cell bodies of the pseudounipolar neurons of the vagus nerve are located in the nodose and petrosal ganglia, of which the majority is found in the former. Nodose ganglion cells contain a wealth of receptors for amino acids, monoamines, neuropeptides, and other neurochemicals that when activated, can modify afferent vagus nerve activity.1 Numerous innervations of the afferent vagus nerves coupled with the diversity of receptors located on the nodose ganglia illustrate the biological importance of this cranial nerve, and systemic drugs that do not cross into the CNS targeted at receptors on nodose ganglia can be used to treat various diseases, such as sleep apnea, gastroesophageal reflux disease, or chronic cough.2-4
The ease of access to the nodose ganglia lends itself to experimental manipulation by midline longitudinal incision made at the neck. The vagus nerve emerges from the posterior lacerated foramen at the base of the skull, and immediately displays a swelling of the nerve that is the nodose ganglion. The nodose ganglion is easily recognizable due to two nerve branches that arise from it: anteriorly the pharyngeal branch; and posteriorly superior laryngeal branch.5 Previous experimental manipulations of the nodose ganglia involved electrophysiological recordings,6 injections of immunohistochemical or immunofluorescent compounds for nerve tracings,7-10 superfusion or injections of neuroexcitotoxins,11-13 injections of adeno-associated virus to knockdown receptors,14,15 and injections of receptor-specific neurochemicals to change the activity of the vagus nerve.16,17
Systemic injections of neurochemicals are problematic in that systemic treatment affects both peripheral and central nervous systems. Thus, systemic treatment does not isolate the effect of neurochemicals on afferent vagal nerve activity. This protocol describes a method using readily available equipment of intranodose injections in the Sprague-Dawley rat that modulates vagus nerve activity without affecting the central nervous system. Stimulation of serotonin type 3 (5-HT3) receptors on nodose ganglia by intravenous (IV) infusion of serotonin (5-HT) induces the Bezold-Jarisch reflex, a vagal response trifecta of bradycardia, hypotension, and apnea, which can be abolished by supranodose vagatomy.11,17-19 Apnea is easily measured by placing a respiratory transducer around the abdomen of the rat.17,18 Cannabinoids decrease 5-HT-induced current in nodose ganglia cells,20 and intranodose ganglia injections of dronabinol attenuate 5-HT-induced apnea.

<table><tbody><tr><td>5-HT HCl</td><td>MP Biomedicals</td><td>215376591</td><td>12.5 &amp;micro;g/kg per 350 &amp;micro;l/kg</td></tr><tr><td>Dronabinol (Marinol) 10 mg Capsules (80 &amp;micro;g/&amp;micro;l)</td><td>AbbVie</td><td>NDC 0051-0023-21</td><td>Dilute with sesame oil to 20 &amp;micro;g/&amp;micro;l</td></tr><tr><td>Sesame Oil</td><td>Sigma-Aldrich</td><td>S3547</td><td></td></tr><tr><td>Intramedic Polyethylene-50</td><td>BD</td><td>427411</td><td>Ordered from VWR (Cat. # 63019-047)</td></tr><tr><td>Graefe Forceps</td><td>Roboz</td><td>RS-5138</td><td>Two are needed</td></tr><tr><td>Johns Hopkins Bulldog Clamp</td><td>Roboz</td><td>RS-7441</td><td>Three are needed</td></tr><tr><td>Piezoelectric Strain Gauge</td><td>Ambu</td><td>813255-100</td><td></td></tr><tr><td>Data Acquisition USB Subsystems</td><td>DataWave Technologies</td><td></td><td></td></tr><tr><td>Sciworks Experimenter Software</td><td></td><td></td><td></td></tr><tr><td>CyberAmp</td><td>Axon Instruments</td><td></td><td></td></tr><tr><td>Syringe, 500 &amp;micro;l, Model 1750 TLL</td><td>Hamilton Company</td><td>81220</td><td></td></tr><tr><td>Syringe, 10 &amp;micro;l, Model 1801 RN</td><td>7659-01</td><td></td><td></td></tr><tr><td>Needle, 28 G, Small Hub RN</td><td>7803-02</td><td>Point Style 4, Angle 35, Length 0.5 in</td><td></td></tr></tbody></table>

a method of nodose ganglia injection in sprague-dawley rat

Afferent signaling via the vagus nerve transmits important general visceral information to the central nervous system from many diverse receptors located in the organs of the abdomen and thorax. The vagus nerve communicates information from stimuli such as heart rate, blood pressure, bronchopulmonary irritation, and gastrointestinal distension to the nucleus of solitary tract of the medulla. The cell bodies of the vagus nerve are located in the nodose and petrosal ganglia, of which the majority are located in the former. The nodose ganglia contain a wealth of receptors for amino acids, monoamines, neuropeptides, and other neurochemicals that can modify afferent vagus nerve activity. Modifying vagal afferents through systemic peripheral drug treatments targeted at the receptors on nodose ganglia has the potential of treating diseases such as sleep apnea, gastroesophageal reflux disease, or chronic cough. The protocol here describes a method of injection neurochemicals directly into the nodose ganglion. Injecting neurochemicals directly into the nodose ganglia allows study of effects solely on cell bodies that modulate afferent nerve activity, and prevents the complication of involving the central nervous system as seen in systemic neurochemical treatment. Using readily available and inexpensive equipment, intranodose ganglia injections are easily done in anesthetized Sprague-Dawley rats.

A Figura 1 representa gravação respiração amostra em ratos que tinham infusão de 5-HT para provocar apneia antes e depois da injecção dos gânglios intranodose de dronabinol. 5-HT activa 5-HT3 sobre os gânglios nodosos que contribuem para o reflexo de Bezold-Jarisch de bradicardia, hipotensão, e apneia. 11,17-19 injecções gânglios Intranodose dronabinol de activar os receptores CB inibitórios, ou modula alostericamente 5-HT 3 receptores que inibem a excitaç...

Watch this Scientific Journal Video about A Method of Nodose Ganglia Injection in Sprague-Dawley Rat at JoVE.com

A Method of Nodose Ganglia Injection in Sprague-Dawley Rat

Afferent vagal signaling transmits important information to central nervous system from receptors located in organs of the abdomen and thorax. The nodose ganglia of vagus nerves contain many types of receptors that modulate vagal activity. This protocol describes a method of local injections of neurochemicals into the nodose ganglia.

Um método de Nodoso Gânglios Injecção em ratos Sprague-Dawley

Afferent signaling via the vagus nerve transmits important general visceral information to the central nervous system from many diverse receptors located ...

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18.7K visualizações.  University of Illinois at Chicago. Afferent vagal signaling transmits important information to central nervous system from receptors located in organs of the abdomen and thorax. The nodose ganglia of vagus nerves contain many types of receptors that modulate vagal activity. This protocol describes a method of local injections of neurochemicals into the nodose ganglia.

Vídeo: A Method of Nodose Ganglia Injection in Sprague-Dawley Rat

Minimally-invasive Technique for Injection into Rat Optic Nerve

The rat optic nerve is a useful model for stem cell regeneration research. Direct injection into the rat optic nerve allows delivery into the central nervous system in a minimally-invasive surgery without bone removal. This technique describes an approach to visualization and direct injection of the optic nerve following minor fascial dissection from the orbital ridge, using a conjunctival traction suture to gently pull the eye down and out. Representative examples of an injected optic nerve show successful injection of dyed beads.

Direct injection into the rat optic nerve is useful for regenerative research. We demonstrate a minimally-invasive technique for direct injection into a rat optic nerve that does not involve opening the skull. Using this method, surgical complications are minimized and recovery is more rapid.

Técnica minimamente invasiva para a injeção em Rat Optic Nerve

The rat optic nerve is a useful model for stem cell regeneration research. Direct injection into the rat optic nerve allows delivery into the central ...

Neurociência

Dorsal Root Ganglion Injection and Dorsal Root Crush Injury as a Model for Sensory Axon Regeneration

Achieving axon regeneration after nervous system injury is a challenging task. As different parts of the central nervous system (CNS) differ from each other anatomically, it is important to identify an appropriate model to use for the study of axon regeneration. By using a suitable model, we can formulate a specific treatment based on the severity of injury, the neuronal cell type of interest, and the desired spinal tract for assessing regeneration. Within the sensory pathway, DRG neurons are responsible for relaying sensory information from the periphery to the CNS. We present here a protocol that uses a DRG injection with a viral vector and a concurrent dorsal root crush injury in the lower cervical spinal cord of an adult rat as a model to study sensory axon regeneration. As demonstrated using a control virus, AAV5-GFP, we show the effectiveness of a direct DRG injection in transducing DRG neurons and tracing sensory axons into the spinal cord. We also show the effectiveness of the dorsal root crush injury in denervating the forepaw as an injury model for evaluating axon regeneration. Despite the requirement for specialized training to perform this invasive surgical procedure, the protocol is flexible, and potential users can modify many parts to accommodate their experimental requirements. Importantly, it can serve as a foundation for those in search of a suitable animal model for their studies. We believe that this article will help new users to learn the procedure in a very efficient and effective manner.

This protocol presents the use of a dorsal root ganglion (DRG) injection with a viral vector and a concurrent dorsal root crush injury in an adult rat as a model to study sensory axon regeneration. This model is suitable for investigating the use of gene therapy to promote sensory axon regeneration.

Dorsal Root Injection gânglio da raiz dorsal e lesão por esmagamento como um modelo para Sensorial Axon Regeneração

Achieving axon regeneration after nervous system injury is a challenging task. As different parts of the central nervous system (CNS) differ from each ...

Laminotomy for Lumbar Dorsal Root Ganglion Access and Injection in Swine

Dorsal root ganglia (DRG) are anatomically well defined structures that contain all primary sensory neurons below the head. This fact makes DRG attractive targets for injection of novel therapeutics aimed at treating chronic pain. In small animal models, laminectomy has been used to facilitate DRG injection because it involves surgical removal of the vertebral bone surrounding each DRG. We demonstrate a technique for intraganglionic injection of lumbar DRG in a large animal species, namely, swine. Laminotomy is performed to allow direct access to DRG using standard neurosurgical techniques, instruments, and materials. Compared with more extensive bone removal via laminectomy, we implement laminotomy to conserve spinal anatomy while achieving sufficient DRG access. Intraoperative progress of DRG injection is monitored using a non-toxic dye. Following euthanasia on post-operative day 21, the success of injection is determined by histology for intraganglionic distribution of 4&#39;,6-diamidino-2-phenylindole (DAPI). We inject a biologically inactive solution to demonstrate the protocol. This method could be applied in future preclinical studies to target therapeutic solutions to DRG. Our methodology should facilitate testing the translatability of intraganglionic small animal paradigms in a large animal species. Additionally, this protocol may serve as a key resource for those planning preclinical studies of DRG injection in swine.

We describe a method for laminotomy in swine that provides access to lumbar dorsal root ganglia (DRG) for intraganglionic injection. Injection progress is monitored intraoperatively and histologically confirmed up to 21 days after surgery. This protocol could be used for future preclinical studies involving DRG injection.

Laminotomy para acesso de gânglio lombar Dorsal raiz e injeção em suínos

Dorsal root ganglia (DRG) are anatomically well defined structures that contain all primary sensory neurons below the head. This fact makes DRG attractive ...

Adeno-associated Virus-mediated Transgene Expression in Genetically Defined Neurons of the Spinal Cord

Selective manipulation of spinal neuronal subpopulations has mainly been achieved by two different methods: 1) Intersectional genetics, whereby double or triple transgenic mice are generated in order to achieve selective expression of a reporter or effector gene (e.g., from the Rosa26 locus) in the desired spinal population. 2) Intraspinal injection of Cre-dependent recombinant adeno-associated virus (rAAV); here Cre-dependent AAV vectors coding for the reporter or effector gene of choice are injected into the spinal cord of mice expressing Cre recombinase in the desired neuronal subpopulation. This protocol describes how to generate Cre-dependent rAAV vectors and how to transduce neurons in the dorsal horn of the lumbar spinal cord segments L3-L5 with rAAVs. As the lumbar spinal segments L3-L5 are innervated by those peripheral sensory neurons that transmit sensory information from the hindlimbs, spontaneous behavior and responses to sensory tests applied to the hindlimb ipsilateral to the injection side can be analyzed in order to interrogate the function of the manipulated neurons in sensory processing. We provide examples of how this technique can be used to analyze genetically defined subsets of spinal neurons. The main advantages of virus-mediated transgene expression in Cre transgenic mice compared to classical reporter mouse-induced transgene expression are the following: 1) Different Cre-dependent rAAVs encoding various reporter or effector proteins can be injected into a single Cre transgenic line, thus overcoming the need to create several multiple transgenic mouse lines. 2) Intraspinal injection limits manipulation of Cre-expressing cells to the injection site and to the time after injection. The main disadvantages are: 1) Reporter gene expression from rAAVs is more variable. 2) Surgery is required to transduce the spinal neurons of interest. Which of the two methods is more appropriate depends on the neuron population and research question to be addressed.

Intraspinal injection of recombinase dependent recombinant adeno-associated virus (rAAV) can be used to manipulate any genetically labelled cell type in the spinal cord. Here we describe how to transduce neurons in the dorsal horn of the lumbar spinal cord. This technique enables functional interrogation of the manipulated neuron subtype.

Expressão do Transgene de mediada por vírus adeno-associado em geneticamente definidos neurônios da medula espinhal

Selective manipulation of spinal neuronal subpopulations has mainly been achieved by two different methods: 1) Intersectional genetics, whereby double or ...

Intrathecal Delivery of Antisense Oligonucleotides in the Rat Central Nervous System

The blood brain barrier (BBB) is an important defense against the entrance of potentially toxic or pathogenic agents from the blood into the central nervous system (CNS). However, its existence also dramatically lowers the accessibility of systemically administered therapeutic agents to the CNS. One method to overcome this, is to inject those agents directly into the cerebrospinal fluid (CSF), thus bypassing the BBB. This can be done via implantation of a catheter for either continuous infusion using an osmotic pump, or for single bolus delivery. In this article, we describe a surgical protocol for delivery of CNS-targeting antisense oligonucleotides (ASOs) via a catheter implanted directly into the cauda equina space of the adult rat spine. As representative results, we show the efficacy of a single bolus ASO intrathecal (IT) injection via this catheterization system in knocking down the target RNA in different regions of the rat CNS. The procedure is safe, effective and does not require expensive equipment or surgical tools. The technique described here can be adapted to deliver drugs in other modalities as well.

Here, we describe a method for delivering drugs to the rat central nervous system by implanting a catheter into the lumbar intrathecal space of the spine. We focus on the delivery of antisense oligonucleotides, though this method is suitable for delivery of other therapeutic modalities as well.

Entrega intrathecal de oligonucleotides do antisense no sistema nervoso central do rato

The blood brain barrier (BBB) is an important defense against the entrance of potentially toxic or pathogenic agents from the blood into the central ...

Entrega de punção lombar de AAV9 em ratos juvenis para terapia gênica do SNC

Intrathecal Vector Delivery in Juvenile Rats via Lumbar Cistern Injection

Gene therapy is a powerful technology to deliver new genes to a patient for the treatment of disease, be it to introduce a functional gene, inactivate a toxic gene, or provide a gene whose product can modulate the biology of the disease. The delivery method for the therapeutic vector can take many forms, ranging from intravenous infusion for systemic delivery to direct injection into the target tissue. For neurodegenerative disorders, it is often desirable to skew transduction towards the brain and/or spinal cord. The least invasive approach to target the entire central nervous system involves injection into the cerebrospinal fluid (CSF), allowing the therapeutic to reach a large fraction of the central nervous system. The safest approach to deliver a vector into the CSF is the lumbar intrathecal injection, where a needle is introduced into the lumbar cistern of the spinal cord. This technique, also known as a lumbar puncture, has been widely used in neonatal and adult rodents and in large animal models. While the technique is similar across species and developmental stages, subtle differences in size, structure, and elasticity of tissues surrounding the intrathecal space require accommodations in the approach. This article describes a method for performing lumbar puncture in juvenile rats to deliver an adeno-associated serotype 9 vector. Here, 25-35 &#181;L of vector were injected into the lumbar cistern, and a green fluorescent protein (GFP) reporter was used to evaluate the transduction profile resulting from each injection. The benefits and challenges of this approach are discussed.

Autor em destaque: Avaliando a eficácia da terapia gênica intratecal em ratos juvenis

A surgical procedure is described to perform injections into the lumbar cistern of the juvenile rat. This approach has been used for the intrathecal delivery of gene therapy vectors, but it is anticipated that this approach can be used for a variety of therapeutics, including cells and drugs.

Entrega de vetor intratecal em ratos juvenis por injeção de cisterna lombar

Gene therapy is a powerful technology to deliver new genes to a patient for the treatment of disease, be it to introduce a functional gene, inactivate a ...

Direct Vagus Nerve Injection Protocol For Rats

There is a relative abundance of strategies and methodologies to facilitate drug delivery to the central nervous system. However, drug delivery directly to the peripheral nervous system is less common, with fewer detailed methods publications available to aid researchers. Here, we describe a direct nerve injection method for peripheral nervous system drug delivery, using the vagus nerve as a model nerve. This method can be used in the treatment of autonomic nervous system disorders through targeting of the left vagus nerve, although this general injection method can be extrapolated to injection of other nerves with minor modification. This method explains all critical steps involved in the procedure involving microsurgery in anesthetized adult rats under a dissecting microscope. The use of a tracking dye is described to facilitate the monitoring of injection fidelity in real time. Illustrations of successful and failed injections are provided. If carried out properly, direct vagus nerve injections can be conducted in a safe manner that is well-tolerated by the rat without post-delivery complications. For example, once the surgeons were trained in this method, six out of six rats were successfully injected without any complications. This method of direct nerve injection for preclinical rat studies is capable of delivering agents (including but not limited to gene therapy) to peripheral nerves.

We present a protocol for direct vagus nerve injection in rats, enabling drug delivery directly into the nerve without post-injection complications. This method applies to preclinical neurological studies involving autonomic nervous system manipulation. It can be used for direct nerve injection for other nerves in rats and other species, with necessary modifications.

Protocolo de injeção direta do nervo vago para ratos

There is a relative abundance of strategies and methodologies to facilitate drug delivery to the central nervous system. However, drug delivery directly ...

Lumbar Intrathecal Injection of Gene Therapy Vectors for Central Nervous System Targeting in Mice and Rats

One method to target the central nervous system for treating neurological diseases involves utilizing the lumbar intrathecal route of administration. This approach bypasses the blood-brain barrier to directly access the cerebrospinal fluid and preferentially target cells within the central nervous system. Multiple published preclinical studies employing the lumbar intrathecal injection route have contributed to the development of gene therapy clinical trials; however, the described protocols are variable and dispersed across multiple resources. Here, a comprehensive set of protocols for lumbar intrathecal injections in neonatal, juvenile, and adult mice and rats for preclinical gene therapy studies is presented. With proper training, this injection technique can be performed quickly and reliably. In addition to detailing the injection protocol at each developmental stage, associated parameters, such as injection volume, that can influence study outcomes are discussed. To demonstrate the application of lumbar intrathecal injections for targeting the central nervous system, the expression of adeno-associated virus serotype 9 in the brain, spinal cord, and peripheral tissues is presented following a successful or unsuccessful injection.

A lumbar intrathecal injection represents a translationally relevant route of administration for delivering gene therapy to the central nervous system. This comprehensive standardized protocol for lumbar intrathecal injections in neonatal, juvenile, and adult mice and rats aims to guide researchers in adopting this technique for preclinical gene therapy studies.

Injeção intratecal lombar de vetores de terapia gênica para direcionamento do sistema nervoso central em camundongos e ratos

One method to target the central nervous system for treating neurological diseases involves utilizing the lumbar intrathecal route of administration. This ...

Um método de Nodoso Gânglios Injecção em ratos Sprague-Dawley

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