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Use of rabies virus as a transneuronal tracer of
neuronal connections implications for the
understanding of rabies pathogenesis Gabriella
Ugolini NBCM, CNRS Gif-sur-Yvette
Kuypers Ugolini, Trends Neurosci. 1990
Amplification of the signal self-amplifying
marker
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Rabies Virus
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Transneuronal tracing with rabies virus 1 -
Amplification of the signal self-amplifying
marker. 2 - Exclusive tropism for neurones in
vivo. 3 - Absence of degeneration of infected
neurones possibility of combined visualisation
of neurotransmitters other tracers. 4 -
Specificity propagation exclusively by
transneuronal transfer between connected neurones
at chemical synapses. 5 - Intracellular
transport is preferentially addressed to
dendrites transneuronal transfer occurs only in
the retrograde direction. 6 - Ubiquitous
distribution of rabies receptors in the CNS, but
not in the peripheral nervous system. 7 - The
only technique allowing the identification of
neuronal connections across a practically
unlimited number of synapses.
CVS strain (1010 PFU/ml) Asymptomatic period
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2 - No propagation via gap junctions (DM MNs) 3
- Sequential infection of 2, 3 and 4 order
neurons 4 - Centrifugal transfer to sensory and
autonomic neurons at long time points, during the
asymptomatic period
1 - Peripheral uptake is restricted to
motoneurons no uptake via sensory and autonomic
neurons
RAT
Tang, Rampin, Giuliano Ugolini (1999) J. Comp.
Neurol. 414167-192.
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PRIMATES (macaque monkeys) injection into the
lateral rectus (LR) muscle 1 - Peripheral
uptake is restricted to motoneurons2 -
Ubiquitous distribution of rabies receptors
within the CNS3 - Centrifugal transfer to the
vestibular (Scarpas) ganglionat 3 days p.i.
(during the asymptomatic incubation period)
Ugolini et al. J. Comp. Neurol. 2006
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PRIMATES differences in monosynaptic pathways to
motoneurons of the lateral rectus muscle (LR)
which innervate slow and fast muscle fibers

• Muscle defective replication in fibrocytes, no
  virus in myocites.
• No spread within the muscle uptake occurs only
  at the site of inoculation.
• - Combined visualisation of rabies virus and
  choline acetyltransferase (CAT) infected
  motoneurons remain viable.

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Primates Injection of rabies virus into the CNS
(Posterior parietal cortex, areas VIP, MIP)

• Same pattern of propagation of rabies virus after
  central and peripheral inoculations
• 1 - Infection of first-order neurons at 2 days.
• 2 - Transneuronal transfer occurs only
  retrogradely (no anterograde transfer to the
  pontine nuclei).
• 3 - Transfer to connected neurons at sequential
  intervals of 12 hrs.
• 4 - No local spread or cell death.

3 order at 3 days p.i. centrifugal transfer to
the vestibular (Scarpas) ganglion
2 order at 2.5 days p.i. vestibular nuclei
Co-injection of rabies virus a conventional
tracer (Cholera toxin B fragment, CTB) no
interference between uptake of rabies CTB
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Scarpas ganglion
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• Rabies pathogenesis
•  
• In primates rodents (rats, guinea pigs),
  peripheral uptake is restricted to motor
  endplates/axons in keeping with the presynaptic
  location of NCAM and with a role of Ach nicotinic
  receptors (despite their mainly post-synaptic
  location).
• Rabies virus does not spread within the muscle
  uptake occurs only at the site of inoculation
  importance of complete wound infiltration with
  rabies antibodies as soon as possible, to prevent
  virus entry!
• Motoneurons are the only gateway for propagation
  of rabies virus to the CNS.
• Rabies virus propagates exclusively by retrograde
  transneuronal transfer at chemical synapses - not
  via gap junctions or local spread.
• Transneuronal transfer occurs only retrogradely
  due to the fact that, after replication,
  centrifugal intracellular transport of rabies
  targets only dendrites, and not axons.
• Retrograde transport and transneuronal transfer
  occur at high speed, by active axonal transport
  (P/LC8 interactions and microtubule-based
  transport)

• Ubiquitous distribution of rabies receptor(s)
  within the CNS transneuronal transfer involves
  all known populations connected to first-order
  neurons, regardless of their transmitters.
• Extensive propagation of rabies virus within the
  CNS during the asymptomatic period! Each
  successive step of transfer (to 2, 3, 4 order
  neurons) requires only 12 hrs, regardless of the
  distance.
• Infection of sensory and autonomic neurons
  requires longer incubation times because it
  reflects centrifugal propagation from the CNS to
  the periphery.

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Acknowledgments
Supported by EU grants BIO4-CT98-0546
(TransVirus) QLK6-CT-2002-0015 (EUROKINESIS)