Neurons can be labeled with unique hues by helper virus-free HSV-1 vectors expressing Brainbow

Abstract

Background-A central problem in neuroscience is elucidating synaptic connections, the connectome. Because mammalian forebrains contain many neurons, labeling specific neurons with unique tags is desirable. A novel technology, Brainbow, creates hundreds of hues by combinatorial expression of multiple fluorescent proteins (FPs).New method-We labeled small numbers of neurons, and their axons, with unique hues, by expressing Brainbow from a helper virus-free Herpes Simplex Virus (HSV-1) vector.Results-The vector expresses a Brainbow cassette containing four FPs from a glutamatergicspecific promoter. Packaging HSV-brainbow produced arrays of seven to eight Brainbow cassettes, and using Cre, each FP gene was in a position to be expressed, in different cassettes. Delivery into rat postrhinal (POR) cortex or hippocampus labeled small numbers of neurons with different, often unique, hues. An area innervated by POR cortex, perirhinal (PER) cortex, contained axons with different hues. Specific axons in PER cortex were matched to specific cell bodies in POR cortex, using hue.Comparison with existing methods-HSV-Brainbow is the only technology for labeling small numbers of neurons with unique hues. In Brainbow mice, many neurons contain the same hue. Brainbow-adeno-associated virus vectors require transduction of the same neuron with multiple vector particles, confounding neuroanatomical studies. Replication-competent Brainbowpseudorabies virus vectors label multiple neurons with the same hue.Conclusions-Attractive properties of HSV-Brainbow include each vector particle contains multiple cassettes, representing numerous hues, recombination products are stabile, and experimental control of the number of labeled neurons. Labeling neurons with unique hues will benefit mapping forebrain circuits.

CRF-Amplified Neuronal TLR4/MCP-1 Signaling Regulates Alcohol Self-Administration

“…This is reflected by the ability of the virus to: (i) establish life-long infection of the peripheral ganglia sensory neurons, known as latency, and (ii) cause encephalitis, which is the most common viral encephalitis and is associated with virus-induced neuronal apoptosis. HSV vectors, including amplicons, retain neurotropism and are therefore recognized as promising vectors for gene therapy of the nervous system and tools to differentially label CNS neurons (Perkins et al, 2003;Taylor et al, 2005;Saeki, 2006;Berges et al, 2007;Suzuki et al, 2008;Manservigi et al, 2010;Cohen et al, 2011;de Silva and Bowers, 2011;Liu et al, 2011;Yang et al, 2011;Fiandaca et al, 2012;Aurelian, 2014;Zhang et al, 2015). The molecular mechanism of this in vivo neuronal lineage-restricted infection/transduction is still unclear.…”

“…; Berges et al, 2007; Suzuki et al, 2008; Manservigi et al, 2010; Cohende Silva and Bowers, 2011;Liu et al, 2011;Yang et al, 2011;Fiandaca et al, 2012;Zhang et al, 2015).…”

Concepts in Light Microscopy of Viruses

“…Multiple fluorogenic genes have also been designed for AAV [ 153 ]. The approach can be used to assess virus replication, competition, spreading, or the arrangement of infected cells in tissue [ 154 ].…”

Use of Adeno‐Associated and Herpes Simplex Viral Vectors for In Vivo Neuronal Expression in Mice

“…Thus, HSV amplicons can accommodate large or multiple transgenes along with their essential regulatory sequences, improving the physiological relevance of gene transfer manipulations (Lim, 2013). Recently, the HSV system has been elegantly coupled with Brainbow technology for circuit tracing (Zhang et al, 2015).…”