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Directed stepwise tracing of polysynaptic neuronal circuits with replication-deficient pseudorabies virus.

ABSTRACT: Brain functions are accomplished by polysynaptic circuits formed by neurons wired together through multiple orders of synaptic connections. Polysynaptic connectivity has been difficult to examine due to a lack of methods of continuously tracing the pathways in a controlled manner. Here, we demonstrate directed, stepwise retrograde polysynaptic tracing by inducible reconstitution of replication-deficient trans-neuronal pseudorabies virus (PRV^ΔIE) in the brain. Furthermore, PRV^ΔIE replication can be temporally restricted to minimize its neurotoxicity. With this tool, we delineate a wiring diagram between the hippocampus and striatum-two major brain systems for learning, memory, and navigation-that consists of projections from specific hippocampal domains to specific striatal areas via distinct intermediate brain regions. Therefore, this inducible PRV^ΔIE system provides a tool for dissecting polysynaptic circuits underlying complex brain functions.

SUBMITTER: Du W

PROVIDER: S-EPMC10326449 | biostudies-literature | 2023 Jun

REPOSITORIES: biostudies-literature

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Directed stepwise tracing of polysynaptic neuronal circuits with replication-deficient pseudorabies virus.

Du Wenqin W Li Elizabeth E Guo Jun J Arano Rachel R Kim Yerim Y Chen Yuh-Tarng YT Thompson Alyssa A Oh So Jung SJ Samuel Aspen A Li Ying Y Oyibo Hassana K HK Xu Wei W

Cell reports methods 20230616 6

Brain functions are accomplished by polysynaptic circuits formed by neurons wired together through multiple orders of synaptic connections. Polysynaptic connectivity has been difficult to examine due to a lack of methods of continuously tracing the pathways in a controlled manner. Here, we demonstrate directed, stepwise retrograde polysynaptic tracing by inducible reconstitution of replication-deficient <i>trans</i>-neuronal pseudorabies virus (PRV<sup>ΔIE</sup>) in the brain. Furthermore, PRV<s ...[more]

PMID: 37426757

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Project description:Alpha-herpesviruses, including human herpes simplex virus 1 & 2, varicella zoster virus and the swine pseudorabies virus (PRV), infect the peripheral nervous system of their hosts. Symptoms of infection often include itching, numbness, or pain indicative of altered neurological function. To determine if there is an in vitro electrophysiological correlate to these characteristic in vivo symptoms, we infected cultured rat sympathetic neurons with well-characterized strains of PRV known to produce virulent or attenuated symptoms in animals. Whole-cell patch clamp recordings were made at various times after infection. By 8 hours of infection with virulent PRV, action potential (AP) firing rates increased substantially and were accompanied by hyperpolarized resting membrane potentials and spikelet-like events. Coincident with the increase in AP firing rate, adjacent neurons exhibited coupled firing events, first with AP-spikelets and later with near identical resting membrane potentials and AP firing. Small fusion pores between adjacent cell bodies formed early after infection as demonstrated by transfer of the low molecular weight dye, Lucifer Yellow. Later, larger pores formed as demonstrated by transfer of high molecular weight Texas red-dextran conjugates between infected cells. Further evidence for viral-induced fusion pores was obtained by infecting neurons with a viral mutant defective for glycoprotein B, a component of the viral membrane fusion complex. These infected neurons were essentially identical to mock infected neurons: no increased AP firing, no spikelet-like events, and no electrical or dye transfer. Infection with PRV Bartha, an attenuated circuit-tracing strain delayed, but did not eliminate the increased neuronal activity and coupling events. We suggest that formation of fusion pores between infected neurons results in electrical coupling and elevated firing rates, and that these processes may contribute to the altered neural function seen in PRV-infected animals.

Dataset Information

Directed stepwise tracing of polysynaptic neuronal circuits with replication-deficient pseudorabies virus.

Publications

Directed stepwise tracing of polysynaptic neuronal circuits with replication-deficient pseudorabies virus.

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