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Abstract:
Functional logic employed by the nervous system for
information processing resides mainly in the wiring patterns among
specific types of neurons. Therefore, detailed knowledge on
neuronal networks is essential for understanding a variety of brain
functions. A powerful and long-awaited method for analyzing the
neuronal connectivity patterns would be to deliver tracers
selectively to specific types of neurons and at the same time to
label trans-synaptically their axonal target neurons. For this
purpose, we took advantage of a unique property of plant lectin,
wheat germ agglutinin (WGA), which has been used as a
trans-synaptic tracer in classical neuroanatomical studies. Here,
we report the development of a novel genetic strategy which employs
WGA cDNA as a transgene, for the visualization of selective and
functional neural pathways in the nervous system.
As a first example, we employed the L7 (Pcp2) promoter elements
to direct the expression of WGA in cerebellar Purkinje cells and
generated transgenic mouse lines. WGA mRNA was abundantly produced
in Purkinje cells. Its protein product was detected not only in the
Purkinje cells, but also in neurons in the deep cerebellar nuclei,
suggesting that WGA produced in the Purkinje cells (1st-order
neurons) were transported to their nerve terminals and then
trans-synaptically conveyed to the deep cerebellar nuclei neurons
(2nd-order neurons). Furthermore, WGA protein was detected in the
red nucleus, the thalamic ventrolateral nucleus, and several other
areas, all of which receive massive projection from the deep
cerebellar nuclei.
As a second example, we applied this WGA transgene technique to
the olfactory system. Recently, there has been a great progress in
the understanding of the olfactory system by employing
electrophisiological and molecular biological techniques. In
particular, the findings of two basic principles in projection
patterns of primary olfactory axons (glomerular convergence and
zone-to-zone projection) and molecular receptive ranges of the
olfactory bulb neurons have shed light on molecular mechanisms of
the information processing in the primary olfactory system.
However, the functional and molecular analyses of the central
connections from the olfactory bulb to the olfactory cortex remain
to elucidated. WGA mRNA and protein were robustly expressed in the
olfactory sensory neurons under the control of the OMP (olfactory
marker protein) promoter elements in transgenic mice. WGA protein
was anterogradely transported from the olfactory epithelium to the
olfactory bulb, and trans-synaptically transferred in glomeruli to
the dendrites of mitral/tufted cells (the second-order neurons).
Furthermore, WGA protein was detected in target regions of
mitral/tufted cells such as anterior olfactory nucleus, the
olfactory tubercle,and the piriform cortex. Thus, this technique
for visualization of specific multi-synaptic neural pathways will
provide extremely valuable tool for the studies of formation,
refinement, maintenance, and remodelling of neural networks in the
brain.
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