1Department of Biologic and Materials Sciences, School of Dentistry, University of Michigan, Ann Arbor 48109-1078; and 2Department of Physiology, Medical School, University of Michigan, Ann Arbor, Michigan 48109-0622
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ABSTRACT |
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Koga, Tomoshige and Robert M. Bradley. Biophysical Properties and Responses to Neurotransmitters of Petrosal and Geniculate Ganglion Neurons Innervating the Tongue. J. Neurophysiol. 84: 1404-1413, 2000. The properties of afferent sensory neurons supplying taste receptors on the tongue were examined in vitro. Neurons in the geniculate (GG) and petrosal ganglia (PG) supplying the tongue were fluorescently labeled, acutely dissociated, and then analyzed using patch-clamp recording. Measurement of the dissociated neurons revealed that PG neurons were significantly larger than GG neurons. The active and passive membrane properties of these ganglion neurons were examined and compared with each other. There were significant differences between the properties of neurons in the PG and GG ganglia. The mean membrane time constant, spike threshold, action potential half-width, and action potential decay time of GG neurons was significantly less than those of PG neurons. Neurons in the PG had action potentials that had a fast rise and fall time (sharp action potentials) as well as action potentials with a deflection or hump on the falling phase (humped action potentials), whereas action potentials of GG neurons were all sharp. There were also significant differences in the response of PG and GG neurons to the application of acetylcholine (ACh), serotonin (5HT), substance P (SP), and GABA. Whereas PG neurons responded to ACh, 5HT, SP, and GABA, GG neurons only responded to SP and GABA. In addition, the properties of GG neurons were more homogeneous than those of the PG because all the GG neurons had sharp spikes and when responses to neurotransmitters occurred, either all or most of the neurons responded. These differences between neurons of the GG and PG may relate to the type of receptor innervated. PG ganglion neurons innervate a number of receptor types on the posterior tongue and have more heterogeneous properties, while GG neurons predominantly innervate taste buds and have more homogeneous properties.
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INTRODUCTION |
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Investigators of the
peripheral gustatory system have examined the properties of afferent
taste fibers by extracellular recording from dissected single fibers.
This classical approach has revealed a wealth of information on the
response characteristics of afferent taste fibers (for a review see
Smith and Frank 1993). In contrast, the current
knowledge of the biophysical and neurochemical properties of afferent
taste fibers is less complete because of the practical difficulty of
examining these fibers in vivo. Researchers of nociceptors facing a
similar problem have made profitable use of in vitro techniques to
define the biophysical and chemical sensitivity of afferent sensory
neurons supplying cutaneous receptors because the properties of the
ganglion cells of these fibers reflects the properties of the afferent
fibers and endings (Dray 1996
). The goal of the present
investigation was to apply similar techniques to afferent sensory
neurons supplying taste buds to characterize both the biophysical
properties and responses of these neurons to neurotransmitters that
have been shown using anatomical techniques to be associated with taste
receptors (Nagai et al. 1996
).
Afferent fibers transmitting gustatory information from the anterior
tongue and soft palate travel in the chorda tympani and greater
superficial petrosal nerves with cell bodies in the geniculate ganglion
(GG), while taste buds on the posterior tongue are supplied by the
glossopharyngeal nerves with cell bodies in the petrosal ganglion (PG).
Electrophysiological studies of the chorda tympani, greater superficial
petrosal, and glossopharyngeal nerves have revealed that these nerves
have heterogeneous response properties (Frank 1991;
Frank et al. 1988
; Nejad 1986
;
Ogawa et al. 1968
), but it is not known if these
differences are reflected in the biophysical properties of the ganglion
cells. In addition to having heterogeneous response properties,
afferent taste neurons also express multiple putative neurotransmitters
and neuromodulators. For example, the PG has been shown to immunostain
for substance P (SP), tyrosine hydroxylase, vasoactive intestinal
polypeptide, calcitonin gene-related peptide, galanin, glutamate, and
asparate (Czyzyk-Kreska et al. 1991
; Finley et
al. 1992
; Helke and Hill 1988
; Helke and
Niederer 1990
; Helke and Rabchevsky 1991
;
Ichikawa et al. 1991
; Okada and Miura
1992
).
Not only have these neurochemicals been shown to be present in the PG
and GG, they have been implicated in taste function at both the
peripheral and central processes of the peripheral taste system.
Serotonin (5HT) is believed to modulate chemosensory responses of taste
receptor cells (Delay and Roper 1988; Nagai et
al. 1998
) and acetylcholine (ACh),
aminobutyric acid
(GABA), and SP have been suggested to function at the synapse between taste buds and primary afferent neurons (Jain and Roper
1991
; Nagai et al. 1986
; Nagy
1982
; Paran and Mattern 1975
). Moreover, PG
neurons respond to ACh at concentrations in the physiological range
(Zhong and Nurse 1997
). At the first relay in the
central taste pathway, GABA and SP have been shown to function as
inhibitory and excitatory neurotransmitters at the synapse of the
primary afferent fibers in the nucleus of solitary tract (Davis
and Smith 1997
; Du and Bradley 1998
;
Grabauskas and Bradley 1996
, 1998a
; King et al.
1993
; Wang and Bradley 1993
).
To gain insight into the biophysical properties of afferent neurons supplying taste receptors, we have used acutely isolated GG and PG neurons innervating the tongue that were retrogradely labeled by Fluorogold injected into the area of the fungiform, foliate, and circumvallate papillae. We have also characterized the responses of PG and GG neurons to application of 5HT, SP, ACh, and GABA, which have been demonstrated to be involved in the afferent taste system.
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METHODS |
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Cell labeling and isolation
The cell labeling technique is based on methods developed in an
earlier study (Bradley et al. 1985). Female
Sprague-Dawley rats aged 20-40 days were anesthetized with an
intramuscular injection of mixture of Rompun (10 mg/kg) and ketamine
hydrochloride (90 mg/kg). The lower jaw was retracted and the tongue
depressed. Using a dissecting microscope, the single circumvallate
papilla was visualized and 10-15 µl of 5% solution of Fluorogold
(Fluorochrome, Englewood, CO) mixed in distilled water was injected
just beneath the papilla from a Hamilton syringe. To study GG neurons
innervating the anterior tongue, 5-10 µl of 5% solution of
Fluorogold was injected bilaterally just beneath both the foliate and
fungiform papillae.
After a postoperative survival time of 3-12 days, the rats were reanesthetized with sodium pentobarbital (50 mg/kg, ip) and decapitated. The skull was opened and the forebrain removed just rostral to the brainstem. Using the exit of the facial nerve from the brainstem as a landmark, the petrous portion of the temporal bone was removed to expose the GG which was then excised. The PG was exposed in the neck by following the glossopharyngeal nerve centrally and then removed. The GG or PG were placed in HEPES buffer containing (in mM) 124 NaCl, 5 KCl, 5 MgCl2, 10 sodium succinate, 15 dextrose, 15 HEPES, and 2 CaCl2, and gassed with O2. The ganglia were transferred to HEPES buffer containing 0.5 mg/ml trypsin (type III) and 0.5 mg/ml collagenase (type IVA) and incubated for 1 h at 37°C. For incubation of the GG, the enzyme concentrations were reduced by one half. The ganglia were then triturated with a series of progressively smaller diameter, fire-polished Pasteur pipettes to produce a suspension of dissociated neurons which were placed in a 35-mm-diameter plastic petri dish. The cell suspension was continuously perfused at about 2 ml/min with oxygenated HEPES buffer by gravity flow and the fluid level in the recording chamber was maintained relatively constant by suction of excess fluid. After isolation, the majority of cells were spherical, devoid of processes, and became loosely attached to the substrate after 10 min. The enzymes were obtained from Sigma and prepared before each experiment.
Electrophysiological recording
Patch electrodes, pulled in two stages from 1.5-mm OD
borosilicate filament glass, were filled with a solution containing (in
mM) 130 potassium gluconate, 10 HEPES, 10 EGTA, 1 MgCl2, 1 CaCl2, and 2 ATP.
The pipette solution was adjusted to a pH of 7.2 with KOH. Electrode
resistance was between 5 and 8 M. Recordings were obtained between
30 min and 3 h after plating. The petri dish containing the
neurons was mounted on the stage of an inverted microscope equipped
with epifluorescence and Hoffman modulation optics. Fluorogold-labeled
neurons were identified by fluorescence with an exposure of sufficient
length to identify labeled neurons.
Electrodes were manipulated under visual control and membrane potential
and currents were recorded using standard whole-cell patch-clamp
recording procedures with an Axoclamp 2A amplifier (Axon Instruments).
Current- and voltage-clamp protocols, data acquisition, and analysis
were performed using pCLAMP software (Axon Instruments). Bridge balance
was carefully monitored throughout the experiments and adjusted when
necessary. The junction potential due to potassium gluconate (10 mV)
was subtracted from the recorded membrane voltages (Standen and
Stanfield 1992). Both Fluorogold and nonlabeled neurons were
investigated to determine any possible effects of the label on the
recordings. Criteria for a successful recording included a minimum of
10 min recording time with a stable resting membrane potential of >
40 mV and an action potential amplitude >70 mV. The statistics
are expressed as mean ± SD and differences between groups
measured with the Students t-test were considered
significant when P
0.05.
Drug application
Neurotransmitters or neuromodulators were prepared daily for
experiments from stock solution in HEPES buffer stored at 80°C and
diluted to the desired final concentration just before use. A three- or
seven-barrel pipette filled with a different concentration of drug was
positioned ~40 µm from the neuronal cell body. SP (0.1 µM-1 mM),
ACh (1 µM-1 mM), 5HT (0.01 µM-1 mM), GABA (1 µM-1 mM), the gamma
aminobutyric acid-A (GABAA) agonist muscimol (1 µM-1 mM), or the gamma aminobutyric acid-B
(GABAB) agonist baclofen (0.1-2 mM) was ejected
from the pipette using a Picospritzer at a low pressure (2 psi).
Concentration-response curves were fitted using the Hill equation. All
experiments were conducted at room temperature.
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RESULTS |
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Identification of neurons labeled with Fluorogold
The PG and GG were surgically removed 3-12 days after injection of Fluorogold into their receptive fields. Dissociated PG and GG neurons were round or ovoid in shape and some of them had short processes. The lengths of the long axis of the isolated PG and GG neurons were 32.2 ± 4.4 and 26.0 ± 2.6 µm, respectively (mean ± SD), while their short axes were 28.6 ± 4.2 and 23.0 ± 2.6 µm. The average diameter of the PG neurons was significantly larger than those of the GG (P < 0.001) (Fig. 3A). Figure 1 is a photomicrograph of PG neurons viewed under normal (A) and fluorescent illumination (B). Two of these neurons were strongly fluorescent. Once a labeled neuron was identified, whole-cell recordings were made under normal illumination.
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Biophysical properties of labeled dissociated cells
Recordings were made from 130 labeled PG and 103 labeled GG neurons. Recordings were also made from nonlabeled PG (n = 45) and GG (n = 26) neurons to determine if they were different from the labeled neurons. No neurons were spontaneously active. Depolarizing and hyperpolarizing currents were injected to investigate action potential and passive membrane properties, respectively.
Since the ganglion cells were obtained from animals aged 20-40 days,
it is possible that developmental changes could be still occurring to
the peripheral taste system in these animals. The full complement of
rat fungiform taste buds is present by 20 days (Mistretta
1972), but taste buds in the circumvallate papilla reach adult
numbers at 90 days (Hosley and Oakley 1987
). Thus, at
the ages studied, the receptive field of the GG neurons was probably
not changing, whereas significant changes were ongoing in the receptive
field of PG neurons. How this impacts on the biophysical properties of
the ganglion cells is not known, but no systematic differences due to
animal age were measured.
LABELED PG NEURONS.
Resting membrane potentials ranged from 40 to
78 mV, with a mean of
59 ± 8 mV. Input resistance ranged from 300 to 951 M
, with a
mean of 542 ± 147 M
. Membrane time constants, measured after a
50 pA current was injected, averaged 34.5 ± 10.2 ms. All the
neurons recorded exhibited varying degrees of membrane rectification
evident from their "sagging" voltage responses to intracellular
injections of hyperpolarizing current pulses (Fig. 2).
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LABELED GG NEURONS.
Resting membrane potentials ranged from 43 to
84 mV, with a mean of
64 ± 9 mV. Input resistance ranged from 322 to 975 M
, with a
mean of 574 ± 137 M
. Membrane time constants averaged 25.0 ± 6.0 ms. Like PG neurons, all of the GG neurons had a
sagging phase in response to hyperpolarizing currents (Fig.
2B).
Comparison of GG and PG neuron biophysical properties
Membrane time constant (Fig.
3B) and input resistance (Fig.
3C) of labeled PG and GG neurons were measured from the
response to a 50 pA current injection. The characteristics of the
action potentials were measured in the first spike at the threshold
depolarizing current injection. Values measured were action potential
threshold (Fig. 3D), action potential decay time (Fig.
3E), action potential half-width (Fig. 3F), and
action potential area (Fig. 3G). The mean time constant of
GG neurons was significantly less than those of PG neurons. However,
there was no difference in input resistance measured at the steady
phase of membrane potential between the two groups. The mean spike
threshold of GG neurons was significantly (P < 0.05)
lower than that of PG neurons.
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The most remarkable difference between the biophysical properties of PG and GG neurons was the shape of the action potential. As described above, no hump neurons were detected in GG labeled neurons. All GG neurons had narrow spikes and the decay time was much shorter (Fig. 3E). The action potential amplitude of GG neurons was significantly (P < 0.05) smaller than that of PG neurons (Fig. 3G).
Comparison between biophysical properties of labeled and nonlabeled neurons
Differences in the biophysical properties of labeled and nonlabeled neurons were examined by recording from an additional 45 PG and 26 GG nonlabeled neurons. Forty-two percent of the nonlabeled PG neurons were classified as hump neurons, and this ratio was similar to that of the PG labeled neurons. In addition, no significant differences were found in any biophysical properties between PG labeled and nonlabeled neurons, indicating that the label did not alter the biophysical properties of the neurons. In nonlabeled GG neurons, no differences were detected in passive membrane properties. However, 3 of 26 GG nonlabeled neurons had a hump in their action potential. As a result, the decay time and action potential area of nonlabeled neurons were significantly larger than labeled GG neurons. Similarly, the mean half-width of nonlabeled neurons was larger than the labeled neurons, but the difference was not significant.
Effects of putative neurotransmitters
ACETYLCHOLINE (ACH).
Sensitivity to ACh was investigated in 48 PG and 32 GG neurons. The
sensitivity to ACh was examined during 100 pA, 100 ms, hyperpolarizing current injection at 1 Hz to determine changes in
membrane potential and input resistance. Neurons were considered responsive if they depolarized with a decrease in input resistance (current clamp) or produced an inward current (voltage clamp at
60 mV
holding potential) during application of ACh concentrations up to 0.5 mM. Thus, if neurons did not respond to 0.5 mM ACh, they were
considered to be insensitive. Concentrations of ACh were selected
between 1 µM and 1 mM, because 0.3 mM ACh is reported to evoke almost
maximal current measured under voltage clamp in PG neurons
(Zhong and Nurse 1997
). The duration of the ACh
application was 5 s.
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SEROTONIN (5HT). Sensitivity to 5HT was examined in 30 PG and 21 GG neurons. 5HT rapidly depolarized over 60% of the PG neurons and initiated action potentials (Fig. 5A). The amplitude of the response to a second application of 5HT decreased when compared with the first application, especially with high concentrations of 5HT. Thus, a 5-min interval between the first and second applications was required to obtain consistent responses. The membrane potential and input resistance changes in response to 5HT were different in each cell. However, 11 of 30 PG neurons were insensitive to application of relatively high concentrations of 5HT (0.5-1 mM). Figure 5D shows the 5HT dose-response relation recorded under voltage clamp. The mean peak current at each 5HT concentration was normalized to that elicited by 100 µM 5HT. This concentration of 5HT elicited a saturated response (Fig. 5, C and D). The complete dose-relation curve was also described by the Hill equation. The EC50 for receptor activation was approximately 4.6 µM and the Hill coefficient was 0.93, suggesting one 5HT agonist binding site per receptor. The 5HT-induced currents were activated more rapidly at higher concentrations (Fig. 5C). A typical example of the activation and short-term desensitization phases of an inward current is indicated in 100 and 1000 µM 5HT.
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SUBSTANCE P (SP). The effect of SP was first examined at concentration of 100 nM-1 µM in PG neurons. A 10-s application of 1 µM SP did not produce an observable response. However, a longer application of the same concentration of SP (30-40 s) was an effective stimulus (Fig. 6A), producing either a membrane depolarization with an input resistance decrease (Fig. 6, Aa and c), or a membrane hyperpolarization with an increase in input resistance (Fig. 6Ab). More than 80% of both PG and GG neurons were sensitive to SP.
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GAMMA AMINOBUTYRIC ACID (GABA). The sensitivity of PG and GG neurons to GABA as well as the GABAA and GABAB receptor agonists muscimol and baclofen was investigated to determine differences in distribution of each GABA receptor subtype.
Sensitivity to GABA was examined in 17 PG and 11 GG neurons. Six of the 17 PG neurons were hyperpolarized with a decrease in input resistance (Fig. 7Ab). In eight PG neurons, however, GABA produced a small depolarization associated with a decrease in input resistance (Fig. 7Aa). In contrast, most GG neurons were hyperpolarized with a marked decrease in input resistance (Fig. 7B). In summary, 82% of PG neurons and all GG neurons were sensitive to GABA accompanied with a decrease in input resistance. The ratio of response patterns is summarized in Fig. 7C.
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MUSCIMOL. The GABAA agonist muscimol was applied to 13 PG and 18 GG neurons. Most of PG neurons (9/13) and all of GG neurons were hyperpolarized by muscimol (Fig. 8B). Some PG neurons were depolarized by muscimol; however, the membrane resistance of all neurons was decreased in a dose-dependent manner (Fig. 8, A, C, and D). The concentration-response curves were fitted using the Hill equation (Fig. 8D). The EC50 of PG and GG neurons were approximately 36.8 and 27.1 µM, respectively. The Hill coefficient of PG and GG neurons were 1.02 and 0.98, respectively, suggesting that one molecule of muscimol binds to one ion channel. In summary, the membrane resistance of all PG and GG neurons was decreased by muscimol in a dose-dependent manner.
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BACLOFEN. The GABAB receptor agonist baclofen was applied to seven PG and 10 GG neurons. A slight hyperpolarization with membrane conductance increase was observed only at high concentration (1 mM) in two PG and six GG neurons. An example of response is indicated in Fig. 8E. However, no obvious effect was observed in all neurons at concentrations of <100 µM for 10-s application. Therefore, GABAB receptors do not play a major role in the GABA response of these neurons.
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DISCUSSION |
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Compared with the dorsal root, nodose, and trigeminal ganglia, the
biophysical and neuropharmacological properties of the PG and GG
ganglia have received little attention. Moreover, when these ganglia
have been studied, investigators have not always recorded from a
selected population of ganglion cells even though there is evidence
from immunochemical and anatomical studies that these ganglia are
heterogeneous and innervate very different populations of receptors
(Hall et al. 1997). The present study is thus the first
in which an attempt has been made to record from a defined population
of ganglion cells similar to recent experiments on tooth pulp neurons
in the trigeminal ganglion (Chiego et al. 1997
; Cook et al. 1998
).
The results indicate that there are significant differences between the biophysical and neuropharmacological properties of neurons in the PG and GG ganglia. For example, neurons in the PG have both sharp and humped action potentials and PG neurons respond to ACh, 5HT, SP, and GABA, whereas GG neurons have sharp action potentials and only respond to SP and GABA. In addition, the properties of GG neurons are more homogeneous than those of the PG because all the neurons have sharp spikes and when responses to neurotransmitters occurred, either all or most of the neurons responded.
In other sensory ganglia, a relationship has been established between
axon diameter, action potential duration, and the type of receptor
innervated (Djouhri et al. 1998; Harper and
Lawson 1985
; Koerber et al. 1988
; Rose et
al. 1986
; Villière and McLachlan 1996
).
Even when a single receptor type is innervated, the ganglion cells can
have different membrane properties (Belmonte and Gallego 1983
). Afferent fibers in the rat and hamster glossopharyngeal and chorda tympani nerves range in diameter from 0.2 to 4.1 µm and
innervate a number of different receptors (Farbman and Hellekant 1978
; Jang and Davis 1987
; Miller et al.
1978
). By labeling a selective receptive field, the types of
receptor innervated are reduced. Thus, a label injected into the
posterior tongue will label PG cells that innervate taste and
somatosensory receptors and a label injected into the anterior tongue
will label GG cells that innervate taste buds (Krimm and Hill
1998
). The more heterogeneous properties of the PG neurons
probably reflects its role in innervating taste and somatosensory
receptors, while the homogeneous response characteristics of the GG
neurons reflects its main role in innervating taste buds on the
anterior tongue.
However, even though GG neurons innervating the anterior tongue have
homogeneous biophysical and neuropharmacological properties, they
differ in other characteristics. For example, fibers that innervate
taste buds on the anterior tongue are of different diameters, the
receptive field properties of chorda tympani fibers differ (Nagai et al. 1988), and the response properties of
chorda tympani fibers to taste stimuli applied to the tongue are not
homogeneous (Frank et al. 1988
). Chorda tympani fibers
with small receptive fields respond best to NaCl, whereas fibers with
large receptive fields respond to a larger number of salts
(Nagai et al. 1988
). Additional experiments in which
intracellular recordings are made from the whole ganglion while
stimulating the afferent input are needed to determine if GG neurons
with fibers of different diameters have different membrane properties.
While little is known about the biophysical and neuropharmacological
properties of the GG, there is information on the PG. In early
pioneering work, Gallego and his co-workers performed the first
intracellular analysis of PG ganglion neurons (Belmonte and
Gallego 1983; Gallego 1983
). These investigators
were particularly interested in neurons innervating chemoreceptors and
baroreceptors involved in the cardiovascular system. Intracellular
recordings with sharp electrodes were made using an in vitro
preparation of the whole ganglion allowing electrical stimulation of
the carotid sinus nerve to isolate neurons innervating baroreceptors or
carotid body chemoreceptors. As in the present study, neurons with both sharp and humped action potentials were described. All the neurons innervating the carotid body had humped action potentials and even with
prolonged depolarization only produced a single spike. In contrast, the
neurons innervating baroreceptors were of two types based on the
conduction velocity of their axons and these neurons were capable of
firing multiple action potentials when depolarized. It is possible,
therefore, that the ganglion cells innervating the posterior tongue
with humped and sharp action potentials innervate different populations
of receptors on the posterior tongue. In other ganglia, the humped
neurons have small diameter, slow conducting axons, and innervate
nociceptors (Koerber et al. 1988
), so it is possible
that the PG neurons with humped spikes also innervate posterior tongue
nociceptors, and the neurons with sharp action potentials innervate
taste receptors. This possibility is supported by the results from the
labeled anterior tongue neurons in the GG which supply taste receptors
and all have sharp spikes.
More recently, Nurse and co-workers have examined the sensitivity of
cultured PG cells to application of ACh and 5HT (Zhong et al.
1999; Zong and Nurse 1997
). Application of ACh
resulted in a rapid depolarization accompanied by an increase in
membrane conductance in 68% of the neurons which is similar to the
results obtained in the present study. In addition, the
EC50 for receptor activation in the present study
was 40.5 µM and the Hill coefficient was 1.54 µM, which is similar
to those reported by Zhong and Nurse (1997)
, who
suggested that more than two ACh agonist binding sites exist per
receptor from the value of the Hill coefficient. 5HT depolarized 43%
of the PG cells with an increase in membrane conductance, but in a
small percentage of neurons (6%) the depolarization was accompanied
with a decrease in membrane conductance. The EC50 for 5HT receptor activation in the present study was 4.6 µM and the
Hill coefficient 0.93 µm. Zhong et al. (1999)
report
similar results and suggested that more than two binding sites are
required for maximal activation of the receptor.
A number of investigators using immunohistochemical methods have
demonstrated SP-like immunoreactivity in PG neurons and there is one
report of intense SP immunoreactive neurons in the GG (Ichiyama et al. 1997). However, there are no reports of
electrophysiological studies with SP. Guinea-pig trigeminal ganglion
neurons and bullfrog dorsal root ganglion neurons are depolarized by
exogeneous application of SP (Dray and Pinnock 1982
;
Ishimatsu 1994
; Spiegelman and Puil 1990
). However, in a recent study in ferrets, SP in nanomolar concentrations were found to hyperpolarize the membrane potential of
nodose ganglion neurons by activation of an outward calcium-dependent potassium current (Jafri and Weinreich 1998
).
To our knowledge, this is the first study of the sensitivity of PG and
GG to GABA. Electrophysiological studies have been performed to study
the sensitivity of chick dorsal root ganglion cells (Choi and
Fishbach 1981; Choi et al. 1981
), rat
sympathetic ganglion cells (Adams and Brown
1998
; Bowery and Brown 1974
), and rabbit
nodose ganglion cells (Higashi et al. 1982
) to GABA. In
all these ganglia, GABA depolarizes the cell membrane accompanied by an
increase in membrane conductance, whereas in the present study GABA
hyperpolarizes the majority of the ganglion cells in both the PG and GG
with an increase in membrane conductance. However, the earlier studies
were on different ganglia and used both extracellular and intracellular
recording techniques and were accomplished before the use of whole-cell
recording. Not all the neurons were hyperpolarized by GABA and neurons
with lower resting membrane potentials were in fact hyperpolarized by
GABA (Adams and Brown 1998
).
Functional significance
SP has been shown to be present in centrally projecting fibers of
the glossopharyngeal nerve (Helke and Hill 1988) and in the terminal fields of the glossopharyngeal nerve in the nucleus of the
solitary tract (Cuello and Kanazawa 1978
; Hokfelt
et al. 1975
). Moreover, King et al. (1993)
have
shown that SP excites neurons of the rostral gustatory portion of the
nucleus of the solitary tract (rNST), suggesting that SP may be
involved with afferent transmission of taste information at the first
central synapse in the taste pathway. Similarly, GABA has been shown to have a major influence on neurons processing taste information in the
rNTS (Grabauskas and Bradley 1996
, 1998b
; Wang
and Bradley 1993
, 1995
) and this influence may be partially
acounted for via presynaptic inhibition at the primary afferent
excitatory synapse.
At present, the neurochemical characteristics of afferent taste fibers
is not known and mechanisms of synaptic transmission between the taste
bud and the primary afferent fiber are also unknown. Investigators of
peripheral cutaneous afferent fibers have determined their
chemosensitive properties using isolated dorsal root ganglion neurons
(Dray 1996). It is possible, therefore, that the
receptors on the distal afferent fibers innervating taste buds are
similar to those characterized in the present study on isolated PG and
GG ganglion neurons. Thus, neural transmission between glossopharyngeal
nerves and taste buds could be mediated by ACh, SP, 5HT, and GABA,
whereas transmission in taste buds innervated by the facial nerve could
be mediated by 5HT and GABA but not by ACh and SP. Of course, other
untested neurotransmitters could also be involved. The approach of
using isolated ganglion cells to examine neurotransmission in taste
buds provides a practical approach to what so far has proved to be a
difficult technical problem.
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ACKNOWLEDGMENTS |
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This work was supported by National Institute on Deafness and Other Communication Disorders Grant DC-00288 to R. M. Bradley. T. Koga was the recipient of a subsidy from the Nakayama Foundation for Human Science.
Present address of T. Koga: Dept. of Restorative Science, Kawasaki University of Medical Welfare, Kurashiki 701-0193, Japan.
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FOOTNOTES |
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Address for reprint requests: R. M. Bradley, Dept. of Biologic and Materials Sciences, Room 6228, School of Dentistry, University of Michigan, Ann Arbor, MI 48109-1078 (E-mail: rmbrad{at}umich.edu).
The costs of publication of this article were defrayed in part by the payment of page charges. The article must therefore be hereby marked "advertisement" in accordance with 18 U.S.C. Section 1734 solely to indicate this fact.
Received 3 February 2000; accepted in final form 30 May 2000.
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REFERENCES |
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