Givaudan-Roure Lecture - Association for Chemoreception Sciences

139 Poster [ ] Olfactory Development
NO-MEDIATED SIGNALING FROM OLFACTORY
RECEPTORS TO PERIPHERAL NERVE GLIA IN THE MOTH
OLFACTORY PATHWAY
Oland L.A. 1, Gibson N.J. 2, Tolbert L.P. 2 1A.R.L. Division of
Neurobiology, University of Arizona, Tucson, AZ; 2Neurobiology,
University of Arizona, Tucson, AZ
In the developing olfactory (antennal) nerve of the moth Manduca
sexta, the glia that populate the nerve arise in the antenna and migrate
along the nerve. They reach the CNS/PNS interface region of the nerve
several stages after the first axons have arrived and begin to ensheathe
bundles of axons in the nerve. In recent in vitro studies, antennal nerve
(AN) glia were found to advance readily along glial processes, and
ORN axons were found to induce the AN glia to form multi-cellular
arrays (Tucker and Tolbert, 2003). Array formation did not require
direct ORN-AN glia contact, but occurred only in glia in close
proximity to the axons, suggesting that the axons release a short-range
diffusible signal. Evidence from previous studies of nitric oxide (NO) in
developing Manduca indicated that a Ca ++ -dependent NO synthase is
strongly expressed in the antenna (Nighorn et al., 1998), that the ORN
axons express NO synthase during stages of axon ingrowth when the
AN glia are populating the nerve (Gibson and Nighorn, 2000), and that
NO regulates AN glial migration (Gibson et al., 2001). Current
experiments test whether the formation of arrays by AN glia in vitro is a
consequence of NO release by ORN axons. In co-cultures of ORN
neurons and PN glial cells, glial arrays can by reduced by a NOS
inhibitor. In cultures of AN glia only, NO donors can induce glial
chaining. Our initial results suggest that NO signaling from the ORN
axons may be involved in the normal developmental processes of AN
glia elongation and migration. Supported by NIH-DC04598 to LPT.
140 Poster [ ] Olfactory Development
EXPRESSION PROFILES OF GENES REGULATED BY
THYROID HORMONE IN THE NOSE OF XENOPUS LAEVIS
Walworth E. 1, Burd G. 1 1Molecular and Cellular Biology, University of
Arizona, Tucson, AZ
In Xenopus, metamorphosis is initiated by rising levels of
endogenous thyroid hormone (T ). During metamorphosis from tadpole
3
to frog, essentially every organ/tissue is remodeled and many new
structures must be formed. The nose of Xenopus tadpoles contains two
areas of sensory epithelium. At metamorphosis, one area is completely
remodeled and a new, third area of sensory epithelium is formed de
novo. Apoptosis, cell proliferation, remodeling of the extracellular
matrix and cell differentiation all take place during this transformation.
Previously by our laboratory, more than 125 gene fragments associated
with metamorphosis in the frog nose were isolated and identified using
representational difference analysis, including gene fragments known to
be associated with all of the above cellular processes. In this study, 56
of those genes were subjected to real time quantitative RT-PCR at 5
different stages of development throughout metamorphosis. We found
the expression levels of several of the T early response genes,
3
including Gene 12, transcription factors (nuclear factor I-B, TH/bZIP,
TRβ) and an extracellular matrix remodeling enzyme (stromelysin 3),
were elevated 3 to 57 fold in response to 48 hours of T treatment, and
3
expression remained elevated throughout metamorphic climax. The
elevated levels of expression were visualized by in situ hybridization.
Therefore, these genes which are known to participate in remodeling
other tissues undergoing metamorphosis, also aid in remodeling the
nasal capsule. Support: NIDCD #DC03905
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141 Poster [ ] Olfactory Development
DE NOVO DNA METHYL TRANSFERASES AND METHYL
DNA BINDING DOMAIN PROTEINS IN OLFACTORY
NEUROGENESIS
Macdonald J. 1, Gin C. 1, Roskams A.J. 1 1Zoology, University of British
Columbia, Vancouver, British Columbia, Canada
Epigenetic Regulation (Silencing) of mammalian genomic DNA
occurs when de novo methyltransferases (DNMTs) methylate sites in
the mammalian genome to form targets for methyl-CpG binding
domain proteins (MBDs). MBDs form repressor complexes that can
directly repress promoters or modify chromatin structure by recruitment
of histone deacetylases (HDACs). In the mouse olfactory epithelium,
DNMT1, 2, 3a, and 3b are expressed in the OE from embryonic
development through adulthood. DNMT3a and 3b expression is
maximal in developing ORNs at peaks of neurogenesis. DNMT3b is
expressed in presumptive globose basal cells, sustentacular cells, and
cells migrating from the basal layer to the sustentacular layer in the
E16.5 OE. DNMT3b is primarily expressed (with PCNA) in actively
proliferating cells and may thus play a role in ORN progenitor cell
cycle exit or neuronal fate choice. DNMT3b+/PCNA+ cells migrating
from the basal layer are NST negative and likely represent an
alternative lineage to the neuronal lineage. DNMT3a is expressed in
post-mitotic immature ORNs and a subset of mature ORNs and may
play a role in lineage restriction and induction of terminal ORN
differentiation in non-proliferating neuro-precursors. Expression of
HDAC2 is induced at the earliest stages of neuronal differentiation and
is down-regulated at terminal differentiation of ORNs. A small
percentage of DNMT3b positive cells begin to express HDAC2, while
the majority of DNMT3a positive cells express HDAC2. This
continuum of HDAC2 expression suggests that it may be appropriately
expressed to mediate gene silencing responses to DNA methylation
catalyzed by both DNMT3b and DNMT3a.
142 Poster [ ] Olfactory Development
MECHANISMS BY WHICH BDNF AND NGF ACT AND
INTERACT WITH ATRIAL NATRIURETIC PEPTIDE TYPE-C
TO PROMOTE PROLIFERATION OR DIFFERENTIATION OF
OLFACTORY NEURONAL PRECURSORS.
Simpson P.J. 1, Moon C. 1, Ronnett G.V. 1 1Neuroscience, Johns Hopkins
University, Baltimore, MD
Both BDNF and NGF promote the proliferation of a subset of
olfactory neuronal precursor cells. Concurrent exposure to atrial
natriuretic peptide type-C (CNP) inhibits this proliferation and
promotes differentiation. Immunocytochemical analysis of cell cycle
regulatory proteins shows that both BDNF and NGF increase levels of
cyclin D1 and cdk4 although the mechanisms behind these increases
differ. NGF also increases p27. Addition of CNP with neurotrophin
inhibits increases in cyclin D1 and p27 while alternatively increasing
levels of a variety of cell cycle inhibitory proteins, including p21. The
profile and time course of this inhibitor expression varies between
BDNF and NGF. Analysis of the mechanisms behind changes in cyclin
D1 and p21 shows that inhibition of the MAPK pathway blocks
alterations in cyclin D1 levels in response to either neurotrophins or
neurotrophins with CNP. CNP however does not promote Erk1/2
phosphorylation, suggesting that it may alter the specificity of MEK1/2
signaling. Other signal transduction pathways are responsible for
alterations in p21. Inclusion of cycloheximide results in an increase in
p21 levels in response to neurotrophins and an increase in cyclin D1
levels in response to neurotrophins with CNP. These results are
mimicked by application of 26S proteosome inhibitors. This suggests
that the switch between proliferation and differentiation is primarily
regulated through reciprocal degradation of inhibitory or progressive
cell cycle proteins with a requirement for protein synthesis. Supported
by NIDCD 5RO3DC005704-02 (PJS).