The Effect of VR1 Blockers on Peripheral Trigeminal Nerve Responses to Irritants Sallie Allgood and Wayne L. Silver Department of Biology, Wake Forest University, Winston-Salem, NC 27109

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The Effect of VR1 Blockers on Peripheral
Trigeminal Nerve Responses to IrritantsSallie
Allgood and Wayne L. SilverDepartment of
Biology, Wake Forest University, Winston-Salem,
NC 27109
Introduction The trigeminal nerve innervates the
head and face and terminates in free nerve
endings in the nasal mucosa, oral cavity and the
eyes. These nerve endings detect noxious
chemical stimuli and act as a warning system to
help prevent damage to these areas. On the
trigeminal nerve fibers, there are many receptors
that respond to different types of stimuli. One
of these receptors is the Vanilloid Receptor 1
(VR1). VR1 is known to respond to heat, acids as
well as capsaicin (the active ingredient in chili
peppers), but exactly how it contributes to
trigeminal stimulation by compounds other than
capsaicin is not well known. These preliminary
experiments were preformed to determine how VR1
may contribute to the trigeminal response to
irritating compounds. Whole-nerve recordings
were obtained from Sprague-Dawley rats and
responses to irritating compounds were recorded
both in the presence and absence of the VR1
blockers capsazepine (a selective VR1 antagonist)
and ruthenium red (a nonselective inhibitor that
blocks Ca2 transport through membrane channels).
Discussion These preliminary experiments show
how the ethmoid nerve responds to various
compounds in solution, and how VR1 blockers
affect this response. The response to capsaicin
decreases in the presence of the VR1 blockers.
Thus, capsaicin exerts its effect on the
trigeminal nerve through VR1s. Additionally,
because capsaicins effect was almost eliminated
with high concentrations of capsazepine,
capsaicin appears to exert its effect solely
through VR1s. The response to high
concentrations of propionic acid is also
decreased in the presence of the VR1 blockers
suggesting that this compound stimulates
trigeminal nerve fibers partly through VR1s.
Other acid receptors (ASIC and P2X) may also play
a role. The VR1 blockers have no affect on the
ethmoid nerves response to nicotine and
cyclohexanone. This suggests that these compounds
do not stimulate trigeminal nerve through VR1s.
Future experiments will use this experimental
protocol to examine the role of VR1s in response
to additional irritating trigeminal stimuli.
Blockers for other known receptors on the
trigeminal nerve will also be tested.
Methods Adult Sprague-Dawley rats were
anesthetized with Urethane (ethyl carbamate 1.0
g/kg injected i.p.). When they were fully
anesthetized, two cannulae were placed in the
trachea. One cannula was inserted toward the
lungs, to allow for breathing, and the second was
inserted up into the nasopharynx. Then the
rats head was restrained and the ethmoid nerve
was isolated and placed on two platinum-iridium
electrodes. Rat Ringers (5.4mM KCl, 5mM HEPES,
135mM NaCl, 1.8mM CaCl2 ) was flowed through the
nasal cavity continuously (10 mL/min) using the
nasophayngeal cannula. Irritants (capsaicin,
nicotine, cyclohexanone and propionic acid) were
injected into the Ringers in 0.5 cc aliquots.
These stimuli were delivered alone or along with
either capsazepine or ruthenium red in order to
block the response from VR1. The whole nerve
response was integrated and stored and analyzed
using a data acquisition system. The height of
the integrated response was used to determine the
strength of the response.
Figure 2 Concentration-response curves for the
four compounds tested. Concentrations which
elicited similar responses were used in the
subsequent experiments. Figure 3 An example of a
neural recording. The top trace is the raw
neural response and the bottom trace is the
integrated response. This example shows the
response of the ethmoid nerve to 10µM capsaicin
decreasing in the presence of capsazepine. Figures
4-11 These figures show effect of ruthenium red
and capsazepine on ethmoid nerve responses to
capsaicin, nicotine, cyclohexanone and propionic
acid. Each stimulus was presented alone or with
one of three different concentrations of either
capsazepine or ruthenium red. The response is
calculated as the percent height of the
integrated response as compared to the response
of either 10 uM capsaicin, 0.308 mM nicotine,
114.1 mM cyclohexanone or 36.4 mM propionic acid
when presented alone. Error bars (when present)
represent one standard error of the mean (n1 or
2).
Figure 1 Experimental Set-Up. Stimuli were
injected into the Ringers flow with a syringe in
0.5cc aliquots. The solutions flowed through the
nasal cavity and dripped out of the nose. Neural
responses were recorded from the ethmoid nerve
and the integrated response was used for data
analysis.