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Evaluation of WGA–Cre-dependent topological transgene expression in the rodent brain

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Abstract

Novel neuromodulation techniques in the field of brain research, such as optogenetics, prompt to target specific cell populations. However, not every subpopulation can be distinguished based on brain area or activity of specific promoters, but rather on topology and connectivity. A fascinating tool to detect neuronal circuitry is based on the transsynaptic tracer, wheat germ agglutinin (WGA). When expressed in neurons, it is transported throughout the neuron, secreted, and taken up by synaptically connected neurons. Expression of a WGA and Cre recombinase fusion protein using a viral vector technology in Cre-dependent transgenic animals allows to trace neuronal network connections and to induce topological transgene expression. In this study, we applied and evaluated this technology in specific areas throughout the whole rodent brain, including the hippocampus, striatum, substantia nigra, and the motor cortex. Adeno-associated viral vectors (rAAV) encoding the WGA–Cre fusion protein under control of a CMV promoter were stereotactically injected in Rosa26-STOP-EYFP transgenic mice. After 6 weeks, both the number of transneuronally labeled YFP+/mCherry− cells and the transduced YFP+/mCherry+ cells were quantified in the connected regions. We were able to trace several connections using WGA–Cre transneuronal labeling; however, the labeling efficacy was region-dependent. The observed transneuronal labeling mostly occurred in the anterograde direction without the occurrence of multi-synaptic labeling. Furthermore, we were able to visualize a specific subset of newborn neurons derived from the subventricular zone based on their connectivity.

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Abbreviations

AcbC:

Accumbens nucleus, core

AcbSh:

Accumbens nucleus, shell

AD:

Anterodorsal thalamic nucleus

AM:

Anteromedial thalamic nucleus

APT:

Anterior pretectal nucleus

AV:

Anteroventral thalamic nucleus

AVDM:

Anteroventral thalamic nucleus, dorsomedial part

AVVL:

Anteroventral thalamic nucleus, ventrolateral part

CL:

Centrolateral thalamic nucleus

CPu:

Caudate putamen

HDB:

Nucleus of the horizontal limb of the diagonal band

IAD:

Interanterodorsal thalamic nucleus

F:

Nucleus of the fields of Forel

fi:

Fimbria of the hippocampus

fr:

Fasciculus retroflexus

LGP:

Lateral globus pallidus

LM:

Lateral mammillary nucleus

LDVL:

Laterodorsal thalamic nucleus, ventral part

LM:

Lateral mammillary nucleus

LPMR:

Lateral posterior thalamic nucleus, mediorostral part

LSD:

Lateral septal nucleus, dorsal part

LSI:

Lateral septal nucleus, intermediate part

LSV:

Lateral septal nucleus, ventral part

MD:

Mediodorsal thalamic nucleus

ml:

Medial lemniscus

ML:

Medial mammillary nucleus, lateral part

MM:

Medial mammillary nucleus, medial part

MMn:

Medial mammillary nucleus, median part

MS:

Medial septal nucleus

Rt:

Reticular thalamic nucleus

SFi:

Septofimbrial nucleus

st:

Stria terminalis

Shi:

Septohippocampal nucleus

sm:

Stria medullaris of the thalamus

SNC:

Substantia nigra, compact part

SNR:

Substantia nigra, reticular part

SuML:

Supramammillary nucleus, lateral part

SuMM:

Supramammillary nucleus, medial part

sumx:

Supramammillary decussation

VA–VL:

Ventral anterior—ventrolateral thalamic nucleus

VM:

Ventromedial thalamic nucleus

VPM:

Ventral posteromedial thalamic nucleus

VTA:

Ventral tegmental area

References

  • Aggleton JP, Vann SD, Saunders RC (2005) Projections from the Hippocampal Region to the mammillary bodies in macaque monkeys. Eur J Neurosci 22:2519–2530

    Article  PubMed  Google Scholar 

  • Andersen P, Morris R, Amaral D, Bliss T, O’Keefe J (2006) The hippocampus book. Oxford University Press, Oxford

  • Aschauer DF, Kreuz S, Rumpel S (2013) Analysis of transduction efficiency, tropism and axonal transport of AAV serotypes 1, 2, 5, 6, 8 and 9 in the mouse brain. PLoS One 8(9):1–16

    Article  Google Scholar 

  • Aston-Jones G, Card JP (2000) Use of pseudorabies virus to delineate multisynaptic circuits in brain: opportunities and limitations. J Neurosci Methods 103(1):51–61

    Article  CAS  PubMed  Google Scholar 

  • Boldogköi Z, Sík A, Dénes Á, Reichart A, Toldi J, Gerendai I, Kovács KJ, Palkovits M (2004) Novel tracing paradigms—genetically engineered herpesviruses as tools for mapping functional circuits within the CNS: present status and future prospects. Prog Neurobiol 72(6):417–445

    Article  PubMed  Google Scholar 

  • Borges LF, Sidman RL (1982) Axonal transport of lectins in the peripheral nervous system. J Neurosci 2(5):647–653

    CAS  PubMed  Google Scholar 

  • Braz JM, Rico B, Basbaum AI (2002) Transneuronal tracing of diverse CNS circuits by cre-mediated induction of wheat germ agglutinin in transgenic mice. Proc Natl Acad Sci USA 99(23):15148–15153

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Callaway Edward M (2008) Transneuronal circuit tracing with neurotropic viruses. Curr Opin Neurobiol 18(6):617–623

    Article  CAS  PubMed  Google Scholar 

  • Carretta D, Sbriccoli A, Santarelli M, Pinto F, Granato A, Minciacchi D (1996) Crossed thalamo-cortical and cortico-thalamic projections in adult mice. Neurosci Lett 204(1–2):69–72

    Article  CAS  PubMed  Google Scholar 

  • Castle MJ, Gershenson ZT, Giles AR, Holzbaur ELF, Wolfe JH (2014) Adeno-associated virus serotypes 1, 8, and 9 share conserved mechanisms for anterograde and retrograde axonal transport. Hum Gene Ther 25(8):705–720

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Cearley CN, Wolfe JH (2006) Transduction characteristics of adeno-associated virus vectors expressing cap serotypes 7, 8, 9, and Rh10 in the mouse brain. Mol Ther 13(3):528–537

    Article  CAS  PubMed  Google Scholar 

  • Cenquizca LA, Swanson LW (2007) Spatial organization of direct hippocampal field CA1 axonal projections to the rest of the cerebral cortex. Brain Res Rev 56(1): 1–26. http://www.pubmedcentral.nih.gov/articlerender.fcgi?artid=2171036&tool=pmcentrez&rendertype=abstract. Accessed Nov 11, 2012)

  • Deniau JM (1994) Indirect nucleus accumbens input to the prefrontal cortex via the substantia nigra pars reticulata: a combined anatomical and electrophysiological study in the rat. Neuroscience 61(3):533–545

    Article  CAS  PubMed  Google Scholar 

  • Deshpande A, Bergami M, Ghanem A, Conzelmann K, Lepier A, Götz M, Berninger B (2013) Retrograde monosynaptic tracing reveals the temporal evolution of inputs onto new neurons in the adult dentate gyrus and olfactory bulb. Proc Natl Acad Sci USA 110(12):1152–1161

    Article  Google Scholar 

  • Fabian RH, Coulter JD (1985) Transneuronal transport of lectins. Brain Res 344(1):41–48

    Article  CAS  PubMed  Google Scholar 

  • Ferrari FK, Samulski T, Shenk T, Samulski RJ (1996) Second-strand synthesis is a rate-limiting step for efficient transduction by recombinant adeno-associated virus vectors. J Virol 70(5):3227–3234

    CAS  PubMed  PubMed Central  Google Scholar 

  • Gaykema RPA, Van Der Kuil J, Hersh LB, Luiten PGM (1991) Patterns of direct projections from the hippocampus to the medial septum-diagonal band complex: anterograde tracing with phaseolus vulgaris leucoagglutinin combined with immunohistochemistry of choline acetyltransferase. Neuroscience 43(2/3):349–360

    Article  CAS  PubMed  Google Scholar 

  • Gerfen CR (2004) Basal Ganglia. In: Paxinos G (ed) The rat nervous system. Elsevier academic press, Amsterdam, p 455–508

  • Ginger M, Haberl M, Conzelmann K, Schwarz MK, Frick A (2013) Revealing the secrets of neuronal circuits with recombinant rabies virus technology. Front Neural Circuits 7(January):2

    PubMed  PubMed Central  Google Scholar 

  • Gradinaru V, Zhang F, Ramakrishnan C, Mattis J, Prakash R, Diester I, Goshen I, Thompson KR, Deisseroth K (2010) Molecular and cellular approaches for diversifying and extending optogenetics. Cell 141(1):154–165

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Haber S, McFarland NR (2001) The place of the thalamus in frontal cortical-basal ganglia circuits. The Neuroscientist 7(4):315–324

    Article  CAS  PubMed  Google Scholar 

  • Joel D, Weiner I (2000) Commentary. the connections of the dopaminergic system with the striatum in rats and primates: an analysis with respect to the functional and compartmental organization of the striatum. Neuroscience 96(3):451–474

    Article  CAS  PubMed  Google Scholar 

  • Junyent Felix, Kremer Eric J (2015) CAV-2-why a canine virus is a neurobiologist’s best friend. Curr Opin Pharmacol 24:86–93

    Article  CAS  PubMed  Google Scholar 

  • Kelly RM, Strick PL (2003) Rabies as a transneuronal tarcer of circuits in the central nervous system. J Neurosci Meths 103:63–71

    Article  Google Scholar 

  • Kelsch W, Lin C, Mosley CP, Lois C (2009) A critical period for activity-dependent synaptic development during olfactory bulb adult neurogenesis. J Neurosci 29(38):11852–11858

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Kinoshita N, Mizuno T, Yoshihara Y (2002) Adenovirus-mediated WGA gene delivery for transsynaptic labeling of mouse olfactory pathways. Chem Senses 27(3):215–223

    Article  CAS  PubMed  Google Scholar 

  • Kiss J, Csaki H, Bokor H, Kocsis K, Kocsis B (2002) Possible glutamatergic/aspartatergic projections to the supramammillary nucleus and their origins in the rat studied by selective [3H] D-aspartate labelling and immunocytochemistry. Neuroscience 111(3):671–691

    Article  CAS  PubMed  Google Scholar 

  • Lanciego JL, Wouterlood FG (2011) A half century of experimental neuroanatomical tracing. J Chem Neuroanat 42(3):157–183

    Article  PubMed  Google Scholar 

  • Meibach CR, Siegel A (1977) Efferent connections of the septal area in the rat : an analysis utilizing retrograde and anterograde transport methods. Brain Res 119:1–20

    Article  CAS  PubMed  Google Scholar 

  • Ming G, Song H (2011) Adult neurogenesis in the mammalian brain: significant answers and significant questions. Neuron 70(4):687–702

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Nyakas C, Luiten PGM, Spencer DG, Traber J (1987) Detailed projection patterns of septal and diagonal band efferents to the hippocampus in the rat with emphasis on innervation of CA1 and dentate Gyrus. Brain Res Bull 18(4):533–545

    Article  CAS  PubMed  Google Scholar 

  • Paxinos G, Franklin KB (2004) The mouse brain in stereotaxic coordinates. Gulf Professional Publishing

  • Rouiller EM, Welker E (2000) A comparative analysis of the morphology of corticothalamic projections in mammals. Brain Res Bull 53(6):727–741

    Article  CAS  PubMed  Google Scholar 

  • Ruda M, Coulter JD (1982) Axonal and transneuronal transport of wheat germ agglutinin demonstrated by immunocytochemistry. Brain Res 249:237–246

    Article  CAS  PubMed  Google Scholar 

  • Saunders RC, Aggleton JP (2007) Origin and topography of fibers contributing to the fornix in macaque monkeys. Hippocampus 17:396–411

    Article  PubMed  Google Scholar 

  • Sawchenko PE, Gerfen CR (1985) Plant lectins and bacterial toxins as tools for tracing neuronal connections. Trends Neurosci 8:378–384

    Article  CAS  Google Scholar 

  • Schnyder H, Künzle H (1983) Differential labeling in neuronal tracing with wheat germ agglutinin. Neurosci Lett 35(2): 115–20. http://linkinghub.elsevier.com/retrieve/pii/0304394083905372. Accessed 1 May 2015

  • Sherlock DA, Raisman G (1975) A comparison of anterograde and retrograde axonal transport of horseradish peroxidase in the connections of the mammillary nuclei in the rat. Brain Res 85:321–324

    Article  CAS  PubMed  Google Scholar 

  • Srinivas S, Watanabe T, Lin CS, William CM, Tanabe Y, Jessell TM, Costantini F (2001) Cre reporter strains produced by targeted insertion of EYFP and ECFP into the ROSA26 locus. BMC Dev Biol 1:4

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Taymans J, Vandenberghe LH, Van Den Haute C, Thiry I, Deroose CM, Mortelmans L, Wilson JM, Debyser Z, Baekelandt V (2007) Comparative analysis of adeno-associated viral vector serotypes 1, 2, 5, 7, and 8 in mouse brain. Hum Gene Ther 18(3):195–206

    Article  CAS  PubMed  Google Scholar 

  • Van der Perren A, Toelen J, Carlon M, Van den Haute C, Coun F, Heeman B, Reumers V, Vandenberghe LH, Wilson JM, Debyser Z, Baekelandt V (2011) Efficient and stable transduction of dopaminergic neurons in rat substantia nigra by rAAV 2/1, 2/2, 2/5, 2/6.2, 2/7, 2/8 and 2/9. Gene Ther 18(5):517–527

    Article  PubMed  Google Scholar 

  • Vann SD, Aggleton JP (2004) The mammillary bodies: two memory systems in one? Nature reviews. Neuroscience 5:35–44

  • Vertes RP, Hoover WB, Do Valle AC, Sherman A, Rodriguez JJ (2006) Efferent projections of reuniens and rhomboid nuclei of the thalamus in the rat. J Comp Neurol 499:768–796

    Article  PubMed  Google Scholar 

  • Viral (2012) Anterograde trans-synaptic, tracer for, mapping output, pathways of, and genetically marked neurons. NIH Public Access 72(6): 938–50

  • Voorn P, Vanderschuren LJ, Groenewegen HJ, Robbins TW, Pennartz CM (2004) Putting a spin on the dorsal-ventral divide of the striatum. Trends Neurosci 27(8):468–474

    Article  CAS  PubMed  Google Scholar 

  • Whitman MC, Greer CA (2007) Synaptic integration of adult-generated olfactory bulb granule cells: basal axodendritic centrifugal input precedes apical dendrodendritic local circuits. J Neurosci 27(37):9951–9961

    Article  CAS  PubMed  Google Scholar 

  • Witter MP (2010) Connectivity of the Hippocampus (eds) Vassilis Cutsuridis, Bruce Graham, Stuart Cobb, and Imre Vida. Hippocampal Microcircuits: 5–26

  • Witter PM, Amaral GD (2004) Hippocampal formation. In: Paxinos George (ed) The Rat central nervous system. Elsevier Academic press, San Diego, pp 705–727

    Google Scholar 

  • Wyss JM, Swanson LW, Cowan WM (1979) A study of subcortical afferent to the hippocampal formation in the rat. Neuroscience 4:463–476

    Article  CAS  PubMed  Google Scholar 

  • Yoshihara Y (2002) Visualizing selective neural pathways with WGA transgene: combination of neuroanatomy with gene technology. Neurosci Res 44(2):133–140

    Article  CAS  PubMed  Google Scholar 

  • Yoshihara Y, Mizuno T, Nakahira M, Kawasaki M, Watanabe Y, Kagamiyama H, Jishage K, Ueda O, Suzuki H, Tabuchi K, Sawamoto K, Okano H, Noda T, Mori K (1999) A genetic approach to visualization of multisynaptic neural pathways using plant lectin transgene. Neuron 22(1):33–41

    Article  CAS  PubMed  Google Scholar 

  • Zhao H, Otaki JM, Firestein S (1996) Adenovirus-mediated gene transfer in olfactory neurons in vivo. J Neurobiol 30(4):521–530

    Article  CAS  PubMed  Google Scholar 

  • Zhenzhong C, Gerfen CR, Young WS (2013) Hypothalamic and other connections with the dorsal CA2 area of the mouse hippocampus. J Comp Neurol 521(8):1844–1866

    Article  Google Scholar 

Download references

Acknowledgments

The authors would like to thank the Leuven Viral Vector Core for the construction and production of the rAAV vectors and Prof Lieve Moons and Prof Lut Arckens for the use of the confocal laser scanning microscope. The research presented in this paper has received funding from the Fund for Scientific Research-Flanders (FWO—fellowship to SL), the European projects FP7-ICT-2011-C-284801 ENLIGHTENMENT and FP7 HEALTH-F2-2011-278850 (INMiND), the Flemish Agency for Innovation through Science and Technology (IWT—SBO/110068 Optobrain) and the KU Leuven (OT/14/120).

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Authors and Affiliations

  1. Department of Neurosciences, Laboratory for Neurobiology and Gene Therapy, KU Leuven, Leuven, Belgium

    Sarah Libbrecht, Chris Van den Haute, Lina Malinouskaya & Veerle Baekelandt

  2. Department of Pharmaceutical and Pharmacological Sciences, Laboratory for Viral Vector Technology & Gene Therapy, Faculty of Medicine, KU Leuven, Leuven, Belgium

    Rik Gijsbers

  3. Leuven Viral Vector Core, KU Leuven, Leuven, Belgium

    Chris Van den Haute & Rik Gijsbers

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  1. Sarah Libbrecht
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Correspondence to Veerle Baekelandt.

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Libbrecht, S., Van den Haute, C., Malinouskaya, L. et al. Evaluation of WGA–Cre-dependent topological transgene expression in the rodent brain. Brain Struct Funct 222, 717–733 (2017). https://doi.org/10.1007/s00429-016-1241-x

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  • DOI: https://doi.org/10.1007/s00429-016-1241-x

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