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Differences in neurotropism and neurotoxicity among retrograde viral tracers.

ABSTRACT: BACKGROUND:Neurotropic virus-based tracers have been extensively applied in mapping and manipulation of neural circuits. However, their neurotropic and neurotoxic properties remain to be fully characterized. METHODS:Through neural circuit tracing, we systematically compared the neurotropism discrepancy among different multi-trans-synaptic and mono-synaptic retrograde viral tracers including pseudorabies virus (PRV), rabies virus (RV), and the newly engineered retro adeno-associated virus (rAAV2-retro) tracers. The (single-cell) RNA sequencing analysis was utilized for seeking possible attribution to neurotropism discrepancy and comparing cell toxicity caused by viral infection between glycoprotein-deleted RV (RV-?G) and rAAV2-retro. Viral toxicity induced microglia activation and neuronal protein change were evaluated by immunohistochemistry. RESULTS:Multi-trans-synaptic retrograde viral tracers, PRV and RV, exhibit differential neurotropism when they were used for central neural circuit tracing from popliteal lymph nodes. Mono-synaptic retrograde tracers, including RV-?G and rAAV2-retro, displayed discrepant neurotropic property, when they were applied to trace the inputs of lateral hypothalamic area and medial preoptic nucleus. rAAV2-retro demonstrated preference in cerebral cortex, whereas RV-?G prefers to label basal ganglia and hypothalamus. Remarkably, we detected a distinct preference for specific cortical layer of rAAV2-retro in layer 5 and RV-?G in layer 6 when they were injected into dorsal lateral geniculate nucleus to label corticothalamic neurons in primary visual cortex. Complementation of TVA receptor gene in RV-resistant neurons enabled EnvA-pseudotyped RV infection, supporting receptors attribution to viral neurotropism. Furthermore, both RV-?G and rAAV2-retro exerted neurotoxic influence at the injection sites and retrogradely labeled sites, while the changes were more profound for RV-?G infection. Finally, we demonstrated a proof-of-concept strategy for more comprehensive high-order circuit tracing of a specific target nucleus by combining rAAV2-retro, RV, and rAAV tracers. CONCLUSIONS:Different multi-trans-synaptic and mono-synaptic retrograde viral tracers exhibited discrepant neurotropism within certain brain regions, even cortical layer preference. More neurotoxicity was observed under RV-?G infection as compared with rAAV2-retro. By combining rAAV2-retro, RV, and rAAV tracers, high-order circuit tracing can be achieved. Our findings provide important reference for appropriate application of viral tracers to delineate the landscape and dissect the function of neural network.

SUBMITTER: Sun L

PROVIDER: S-EPMC6368820 | biostudies-literature | 2019 Feb

REPOSITORIES: biostudies-literature

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Differences in neurotropism and neurotoxicity among retrograde viral tracers.

Sun Leqiang L Tang Yajie Y Yan Keji K Yu Jinsong J Zou Yanyan Y Xu Weize W Xiao Ke K Zhang Zhihui Z Li Weiming W Wu Beili B Hu Zhe Z Chen Kening K Fu Zhen F ZF Dai Jinxia J Cao Gang G

Molecular neurodegeneration 20190208 1

<h4>Background</h4>Neurotropic virus-based tracers have been extensively applied in mapping and manipulation of neural circuits. However, their neurotropic and neurotoxic properties remain to be fully characterized.<h4>Methods</h4>Through neural circuit tracing, we systematically compared the neurotropism discrepancy among different multi-trans-synaptic and mono-synaptic retrograde viral tracers including pseudorabies virus (PRV), rabies virus (RV), and the newly engineered retro adeno-associate ...[more]

PMID: 30736827

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Project description:Viral capsid components that bind cellular receptors mediate critical functions in viral tropism and disease pathogenesis. Mammalian orthoreoviruses (reoviruses) spread systemically in newborn mice to cause serotype-specific disease in the central nervous system (CNS). Serotype 1 (T1) reovirus infects ependymal cells to cause nonlethal hydrocephalus, whereas serotype 3 (T3) reovirus infects neurons to cause fulminant and lethal encephalitis. This serotype-dependent difference in tropism and concomitant disease is attributed to the ?1 viral attachment protein, which is composed of head, body, and tail domains. To identify ?1 sequences that contribute to tropism for specific cell types in the CNS, we engineered a panel of viruses expressing chimeric ?1 proteins in which discrete ?1 domains have been reciprocally exchanged. Parental and chimeric ?1 viruses were compared for replication, tropism, and disease induction following intracranial inoculation of newborn mice. Viruses expressing T1 ?1 head sequences infect the ependyma, produce relatively lower titers in the brain, and do not cause significant disease. In contrast, viruses expressing T3 ?1 head sequences efficiently infect neurons, replicate to relatively higher titers in the brain, and cause a lethal encephalitis. Additionally, T3 ?1 head-expressing viruses display enhanced infectivity of cultured primary cortical neurons compared with T1 ?1 head-expressing viruses. These results indicate that T3 ?1 head domain sequences promote infection of neurons, likely by interaction with a neuron-specific receptor, and dictate tropism in the CNS and induction of encephalitis.IMPORTANCE Viral encephalitis is a serious and often life-threatening inflammation of the brain. Mammalian orthoreoviruses are promising oncolytic therapeutics for humans but establish virulent, serotype-dependent disease in the central nervous system (CNS) of many young mammals. Serotype 1 reoviruses infect ependymal cells and produce hydrocephalus, whereas serotype 3 reoviruses infect neurons and cause encephalitis. Reovirus neurotropism is hypothesized to be dictated by the filamentous ?1 viral attachment protein. However, it is not apparent how this protein mediates disease. We discovered that sequences forming the most virion-distal domain of T1 and T3 ?1 coordinate infection of either ependyma or neurons, respectively, leading to mutually exclusive patterns of tropism and disease in the CNS. These studies contribute new knowledge about how reoviruses target cells for infection in the brain and inform the rational design of improved oncolytic therapies to mitigate difficult-to-treat tumors of the CNS.

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Differences in neurotropism and neurotoxicity among retrograde viral tracers.

Publications

Differences in neurotropism and neurotoxicity among retrograde viral tracers.

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