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Triggering genetically-expressed transneuronal tracers by peripheral axotomy reveals convergent and segregated sensory neuron-spinal cord connectivity.

ABSTRACT: To better understand the mechanisms through which non-painful and painful stimuli evoke behavior, new resources to dissect the complex circuits engaged by subsets of primary afferent neurons are required. This is especially true to understand the consequences of injury, when reorganization of central nervous system circuits likely contributes to the persistence of pain. Here we describe a transgenic mouse line (ZWX) in which there is Cre-recombinase-dependent expression of a transneuronal tracer, wheat germ agglutinin (WGA), in primary somatic or visceral afferent neurons, but only after transection of their peripheral axons. The latter requirement allows for both regional and temporal control of tracer expression, even in the adult. Using a variety of Cre lines to target WGA transport to subpopulations of sensory neurons, here we demonstrate the extent to which myelinated and unmyelinated "pain" fibers (nociceptors) engage different spinal cord circuits. We found significant convergence (i.e., manifest as WGA-transneuronal labeling) of unmyelinated afferents, including the TRPV1-expressing subset, and myelinated afferents to NK1-receptor-expressing neurons of lamina I. By contrast, PKCgamma interneurons of inner lamina II only receive a myelinated afferent input. This differential distribution of WGA labeling in the spinal cord indicates that myelinated and unmyelinated sensory neurons target different and spatially segregated populations of postsynaptic neurons. On the other hand, we show that neurons of deeper laminae (III-V) receive direct (i.e., monosynaptic) inputs from myelinated afferents and polysynaptic input from unmyelinated afferents. Taken together, our results indicate that peripheral sensory information is transmitted to the central nervous system both through segregated and convergent pathways.

SUBMITTER: Braz JM

PROVIDER: S-EPMC2760676 | biostudies-literature |

REPOSITORIES: biostudies-literature

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Project description:BackgroundHerpes simplex virus type 1 strain 129 (H129) has represented a promising anterograde neuronal circuit tracing tool, which complements the existing retrograde tracers. However, the current H129 derived tracers are multisynaptic, neither bright enough to label the details of neurons nor capable of determining direct projection targets as monosynaptic tracer.MethodsBased on the bacterial artificial chromosome of H129, we have generated a serial of recombinant viruses for neuronal circuit tracing. Among them, H129-G4 was obtained by inserting binary tandemly connected GFP cassettes into the H129 genome, and H129-ΔTK-tdT was obtained by deleting the thymidine kinase (TK) gene and adding tdTomato coding gene to the H129 genome. Then the obtained viral tracers were tested in vitro and in vivo for the tracing capacity.ResultsH129-G4 is capable of transmitting through multiple synapses, labeling the neurons by green florescent protein, and visualizing the morphological details of the labeled neurons. H129-ΔTK-tdT neither replicates nor spreads in neurons alone, but transmits to and labels the postsynaptic neurons with tdTomato in the presence of complementary expressed TK from a helper virus. H129-ΔTK-tdT is also capable to map the direct projectome of the specific neuron type in the given brain regions in Cre transgenic mice. In the tested brain regions where circuits are well known, the H129-ΔTK-tdT tracing patterns are consistent with the previous results.ConclusionsWith the assistance of the helper virus complimentarily expressing TK, H129-ΔTK-tdT replicates in the initially infected neuron, transmits anterogradely through one synapse, and labeled the postsynaptic neurons with tdTomato. The H129-ΔTK-tdT anterograde monosynaptic tracing system offers a useful tool for mapping the direct output in neuronal circuitry. H129-G4 is an anterograde multisynaptic tracer with a labeling signal strong enough to display the details of neuron morphology.

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Triggering genetically-expressed transneuronal tracers by peripheral axotomy reveals convergent and segregated sensory neuron-spinal cord connectivity.

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