Abstracts - Association for Chemoreception Sciences

epressive sequence contains binding sites for the transcriptional
repressor zbtb7b, which is abundantly expressed in the peripheral
zebrafish olfactory system. Preliminary results suggest that
negative regulation confers a higher specificity of transgene
expression to ciliated OSNs. A major distinction within the OR
repertoire can be made between evolutionary ancient class I ORs
and the group of class II receptors, which massively expanded in
the tetrapod lineage. OR101-1 appears to be the only class II OR
in zebrafish. OSNs that fail to express a functional OR usually
undergo a second OR gene choice resulting in heterogeneity of
OR expression and widespread axonal projection of transgenic
OSN axons to olfactory bulb glomeruli. Transgenic OSNs
expressing a deletion allele of the OR101-1 gene in contrast
target a single specific glomerulus, in the zebrafish olfactory
bulb suggesting that second OR gene choice is severely restricted
in those OSNs. The results imply that distinct regulatory
mechanisms evolved concomitantly with the appearance of
the first class II OR in the teleost lineage. Acknowledgements:
TÜBITAK, The Scientific and Technological Research Council
of Turkey, Grant Awards: 107T760, 112T168
#P252 POSTER SESSION V:
HUMAN TASTE PSYCHOPHYSICS;
OLFACTION RECEPTORS; TASTE DEVELOPMENT
Olfactory Receptor (OR) switching is influenced by genome
position in olfactory-placode (OP)-derived cells.
Robert P. Lane, Seda Kilinc
Wesleyan University Middletown, CT, USA
We previously characterized olfactory receptor (OR) expression
in the OP6 and OP27 cell lines and made two general
observations: OR choice is not a heritable property; and the
range of OR representation in OP populations appears biased
for a subset of the full OR repertoire. We used custom arrays
and deep sequencing to analyze the complete expressed OR
repertoire in OP cultures. OP6 and OP27 cell lines have
significant overlap in OR representation, consistent with similar
pre-specification of the two founder cells, which had been
isolated from a common developmental milieu. However, OR
representation in OP cultures is not constrained by presumptive
expression zones within mouse olfactory epithelium, as
might be predicted if the range of OR choices had been prespecified
by developmental (i.e., spatial) cues. Instead, we find
strong evidence for ‘position-effects’: neighboring OR genes
are significantly over-represented in divergent populations,
suggesting a tendency to switch within versus across OR
clusters. Surprisingly, we do not observe differences in common
epigenetic marks between “active” versus “inactive” ORs, nor
does locus positioning relative to nuclear chromocenters appear
to be predictive of selection probability. We suggest a switching
model in which the epigenetic microenvironment established by
previous OR transcription increases the likelihood of selection/
re-selection within that genome region. We hypothesize that
the OP6 and OP27 founder cells had been similarly specified,
initially leading to a similar bias for OR switching, but that in
the absence of further developmental cues during subsequent
culturing, probabilistic switching histories have resulted in
slow evolution from initial biases, thus independently evolving
OR subrepertoires. Acknowledgements: NIH R01-DC006267
NSF 0842868
#P253 POSTER SESSION V:
HUMAN TASTE PSYCHOPHYSICS;
OLFACTION RECEPTORS; TASTE DEVELOPMENT
Blockade of Insect Odorant Receptor Currents by
Amiloride Derivatives
Gregory M Pask 1 , Yuriy V Bobkov 2 , Elizabeth A Corey 2 ,
Barry W Ache 2,3 , Laurence J Zwiebel 1,4
1
Department of Biological Sciences, Vanderbilt University Nashville,
TN, USA, 2 Whitney Laboratory, Center for Smell and Taste, and
McKnight Brain Institute, University of Florida Gainesville, FL, USA,
3
Departments of Biology and Neuroscience, University of Florida
Gainesville, FL, USA, 4 Department of Pharmacology, Vanderbilt Brain
Institute and Center for Human Genetics, Institutes of Chemical Biology
and Global Health and Program in Developmental Biology, Vanderbilt
University Medical Center Nashville, TN, USA
Insect odorant receptors (ORs) function as heteromeric odorantgated
ion channels consisting of a conserved coreceptor, Orco,
and an odorant-sensitive tuning subunit. Although some
OR modulators have been identified, an extended library of
pharmacological tools is currently lacking and would aid in
furthering our understanding of insect OR complexes. Using
whole-cell patch clamp techniques, we demonstrate that
amiloride and several derivatives, which have been extensively
used as blockers for various ion channels and transporters, also
block odorant-gated currents from two OR complexes from
the malaria vector mosquito Anopheles gambiae, AgOr48 +
AgOrco and AgOr65 + AgOrco. In addition, currents from
both heteromeric OR complexes were susceptible to HMA
blockade when activated by VUAA1, an agonist that targets
the Orco channel subunit. HMA was also capable of blocking
VUAA1-evoked currents of Orco homomers from 4 different
insect orders, demonstrating that HMA blockade is not unique
to AgOR complexes. Additionally, amiloride derivatives have
provided insights into the properties of both the channel pore
and spontaneous gating across different insect OR complexes.
Amiloride derivatives therefore represent a valuable class of
channel blockers that can be used to further investigate the
pharmacological and biophysical properties of insect OR
function. Acknowledgements: This work was supported by the
National Institute of Allergy and Infectious Disease [AI056402]
to L.J.Z and the National Institute on Deafness and Other
Communication Disorders (NIDCD) [DC001655] to B.W.A. at
the National Institutes of Health, and the Foundation for the
National Institutes of Health through the Grand Challenges
in Global Health Initiative [VCTR121] to L.J.Z. G.M.P was
supported by the NIDCD through an NRSA F31 [DC011989].
POSTER PRESENTATIONS
Abstracts are printed as submitted by the author(s).
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