Intracerebroventricular and Intravascular Injection of Viral Particles and Fluorescent Microbeads into the Neonatal Brain
| Autor: | Isao Kosugi, Yoshihiro Tsutsui, Makiko Sakao-Suzuki, Toshihide Iwashita, Shiori Meguro, Hideya Kawasaki |
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| Rok vydání: | 2016 |
| Předmět: |
0301 basic medicine
Muromegalovirus Pathology medicine.medical_specialty General Chemical Engineering Central nervous system Viremia In situ hybridization General Biochemistry Genetics and Molecular Biology Green fluorescent protein Mice 03 medical and health sciences 0302 clinical medicine Cerebrospinal fluid Animals Medicine Injections Intraventricular General Immunology and Microbiology business.industry General Neuroscience Viral encephalitis Meninges Brain medicine.disease Microspheres 030104 developmental biology medicine.anatomical_structure Cytomegalovirus Infections Choroid plexus business 030217 neurology & neurosurgery Neuroscience |
| Zdroj: | Journal of Visualized Experiments. |
| ISSN: | 1940-087X |
| DOI: | 10.3791/54164 |
| Popis: | In the study on the pathogenesis of viral encephalitis, the infection method is critical. The first of the two main infectious routes to the brain is the hematogenous route, which involves infection of the endothelial cells and pericytes of the brain. The second is the intracerebroventricular (ICV) route. Once within the central nervous system (CNS), viruses may spread to the subarachnoid space, meninges, and choroid plexus via the cerebrospinal fluid. In experimental models, the earliest stages of CNS viral distribution are not well characterized, and it is unclear whether only certain cells are initially infected. Here, we have analyzed the distribution of cytomegalovirus (CMV) particles during the acute phase of infection, termed primary viremia, following ICV or intravascular (IV) injection into the neonatal mouse brain. In the ICV injection model, 5 µl of murine CMV (MCMV) or fluorescent microbeads were injected into the lateral ventricle at the midpoint between the ear and eye using a 10-µl syringe with a 27 G needle. In the IV injection model, a 1-ml syringe with a 35 G needle was used. A transilluminator was used to visualize the superficial temporal (facial) vein of the neonatal mouse. We infused 50 µl of MCMV or fluorescent microbeads into the superficial temporal vein. Brains were harvested at different time points post-injection. MCMV genomes were detected using the in situ hybridization method. Fluorescent microbeads or green fluorescent protein expressing recombinant MCMV particles were observed by fluorescent microscopy. These techniques can be applied to many other pathogens to investigate the pathogenesis of encephalitis. |
| Databáze: | OpenAIRE |
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