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#P178
POSTER SESSION V:
TRIGEMINAL SYSTEM; BEHAVIOR
AND PSYCHOPHYSICS; ODORANT
RECEPTORS & OLFACTION PERIPHERY
Methyl syringate, a low-molecular-weight phenolic ester,
activates the chemosensory cation channel TRPA1
Mee-Ra Rhyu
1
, Hee-Jin Son
1
, Yong-Ho Kim
2
, Seok-Bae Oh
2
,
Sho Ishii
3
, Takumi Misaka
3
1
Korea Food Research Institute Seoul, Korea,
2
Seoul National
University Seoul, Korea,
3
The University of Tokyo Tokyo, Japan
We have previously shown that methyl syringate, a component of
the
Kalopanax pictus
Nakai (Araliaceae), a wild vegetable widely
used in Korea for its unique somatosensory properties interact with
TRPA1 in calcium imaging analyses using a human TRPA1
expressing cells. We now further analyzed the action of methyl
syringate in cell-based assay and whole-cell patch-clamp
recordings in the extracellular Ca
2+
-free condition at a holding
potential of -60 mV in hTRPA1 stably expressing cells. In cell-
based assay, methyl syringate (1mM) significantly increased
intracellular Ca
2+
in a dose-dependent manner (EC
50
= 565.2 µM),
which was completely blocked by HC-030031, a specific TRPA1
antagonist. This was 90- or 10-fold low in comparison to AITC
(EC
50
= 6.246 µM), the most potent TRPA1 agonist among all
natural products, or CALD (EC
50
= 64.01 µM), a pungent
compound present in cinnamon oil, respectively. When methyl
syringate (1-3 mM) was applied to AITC-sensitive cells for 15 s,
inward currents were clearly evoked in a dose-dependent manner.
Further, the
I-V
relationship indicated that methyl syringate
(3 mM)-induced currents have the characteristics of TRPA1-
mediated currents with a reversal potential of ~0 mV and a slight
outward rectification. These results demonstrate that methyl
syringate activates human TRPA1, providing a mechanistic
rationale for the unique somatosensory properties of
K. pictus
.
Acknowledgements: Supported by a Korea Food Research Institute
(KFRI) grant E0105101
#P179
POSTER SESSION V:
TRIGEMINAL SYSTEM; BEHAVIOR
AND PSYCHOPHYSICS; ODORANT
RECEPTORS & OLFACTION PERIPHERY
Comparison of functional characteristics between the
TRPM8 ionchannel of
Mus musculus
and
Gallus gallus
Sven Zielke
1,2
, Jonas Petersen
1
, Christian H. Wetzel
1
1
Ruhr-University Bochum/Department of Cellphysiology Bochum,
Germany,
2
International Max Planck Research School in Chemical
Biology (IMPRS-CB) Dortmund, Germany
Transient receptor potential (TRP) channels can be activated by a
huge variety of endogenous and exogenous chemical compounds
and by physical stimuli like touch and temperature. In the present
study, we focused on the TRPM8 receptor of
Gallus gallus
(cTRPM8) and
Mus musculus
(mTRPM8) and compare the
functional characteristics of these two ortholog receptors.
The mammalian TRPM8 channels are well characterized. It is
known, that the mammalian TRPM8 receptor can be activated by
temperatures below 25°C, and by natural as well as synthetic
chemical compounds like menthol, eucalyptol, icilin or the second
messenger PIP2. We measured temperature and menthol
responsiveness of sensory neurons from dissociated dorsal root
ganglia and transfected HEK-293 cells. It is known, that avians and
mammals have different basal body temperatures (39°C and 37°C,
respectively). Furthermore, variations in the amino acid sequence
of both species, e.g. in the coiled-coil domain which is necessary
for the activation of the receptor, could be responsible for
functional differences. We aimed to investigate functional
properties of the ortholog receptors and compared the differences in
detection of cool temperatures or after menthol stimulion between
the two species. Therefore, we used the patch-clamp and Ca
2+
-
imaging technique to characterize the activation temperature and
the response behavior after stimulation with menthol. We found
that the cTRPM8 has an approximately 3°C higher activation
Temperature than the mouse ortholog. Furthermore the cTRPM8
leads to a lower intracellular Ca
2+
increase by stimulation with
menthol and cooling than the mTRPM8. Acknowledgements:
DFG (WE2298/4-1), International Max Planck Research School in
Chemical Biology (IMPRS-CB)
#P180
POSTER SESSION V:
TRIGEMINAL SYSTEM; BEHAVIOR
AND PSYCHOPHYSICS; ODORANT
RECEPTORS & OLFACTION PERIPHERY
Genetic Variation in Bitter Receptor Genes is Associated with
Cough Threshold in Human Subjects
Paul M Wise, Danielle R Reed, Julie A Mennella,
Liang-Dar Hwang, Susana Finkbeiner
Monell Chemical Senses Center Philadelphia, PA, USA
Cough is a protective response that clears mucus, irritant chemicals,
and other xenobiotics from the airways. Individuals differ in cough
sensitivity, which impacts the ability to respond appropriately to
airborne threats. We report a preliminary investigation of the
relationship between individual differences in cough reflex
sensitivity and polymorphisms in the genes that encode
chemosensory receptor proteins expressed in the airways. Genes of
interest included those that encode TRP channels known to play a
role in generating chemically evoked cough (TRPA1, TRPV1) and
bitter receptors (TAS2R) known to modulate other respiratory
responses. Cough thresholds, i.e., the minimum concentration of
capsaicin needed to trigger at least 2 coughs, were measured in 65
healthy subjects (adolescents and their mothers). Saliva samples
were collected from each subject; DNA was extracted and
genotyped at 20 markers within or near the
TRPV1
,
TRPA1
, and
TRPM8
genes, as well as members of the
TAS2R
gene family.
Preliminary analyses revealed that two variant sites within a bitter
receptor cluster on chromosome 12 (centering on
TAS2R20
) were
related to cough thresholds (
rs10845279
, χ
2 (1)
= 7.18, p = 0.03;
rs12226920
; χ
2 (1)
= 6.47, p = 0.04). These results suggest that
people with particular genotypes differ in response to inhaled
capsaicin. It remains to be seen whether these effects generalize to
other irritant compounds and to other airway threats such as
bacteria, viruses, or toxins. Acknowledgements: The project
described was funded, in part, by a grant from the Pennsylvania
Department of Health, Award Number R01HD37119 from the
Eunice Kennedy Shriver National Institute of Child Health and
Human Development, and 1P30DC011735-01 from NIDCD. The
content is solely the responsibility of the authors and does not
necessarily represent the official views of the Eunice Kennedy
Shriver National Institute of Child Health and Human Development
or the National Institutes of Health. The Pennsylvania Department
of Health specifically disclaims responsibility for any analyses,
interpretations, or conclusions.
Abstracts | 87
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