 |
 Previous Article | Next Article 
The Journal of Neuroscience, June 15, 1998, 18(12):4560-4569
Odorant Response Properties of Convergent Olfactory Receptor
Neurons
Thomas C.
Bozza and
John S.
Kauer
Department of Neuroscience, Tufts University School of Medicine,
Boston, Massachusetts 02111
 |
ABSTRACT |
Information about odorant stimuli is thought to be represented in
spatial and temporal patterns of activity across neurons in the
olfactory epithelium and the olfactory bulb (OB). Previous studies
suggest that olfactory receptor neurons (ORNs) distributed in the nasal
cavity project to localized regions in the glomerular layer of the OB.
However, the functional significance of this convergence is not yet
known, and in no studies have the odorant response properties of
individual ORNs projecting to defined OB regions been measured
directly. We have retrogradely labeled mouse ORNs connecting to
different glomeruli in the dorsal OB and tested single cells for
responses to odorants using fura-2 calcium imaging. ORNs that project
to clusters of dorsomedial (DM) glomeruli exhibit different odorant
response profiles from those that project to dorsolateral (DL)
glomeruli. DL-projecting ORNs showed responses to compounds with widely
different structures, including carvone, eugenol, cinnamaldehyde, and
acetophenone. In contrast, DM-projecting neurons exhibited responses to
a more structurally restricted set of compounds and responded
preferentially to organic acids. These data demonstrate that ORN
afferents segregate by odorant responsiveness and that the homogeneity
of ORN and glomerular input varies with different OB regions. The data
also demonstrate that a subpopulation of ORNs projecting to DM
glomeruli is functionally similar.
Key words:
olfactory receptor neurons; olfactory bulb; calcium; imaging; retrograde tracing; glomeruli; mouse; olfactory coding
| |
INTRODUCTION |
Odorant stimuli interact with a vast
array of differentially responsive olfactory receptor neurons (ORNs) in
the nasal cavity. Single-unit (Duchamp et al., 1974 ; Getchell, 1974;
Revial et al., 1978, 1982), field potential (MacKay-Sim et al., 1982;
Mackay-Sim and Kesteven, 1994; Scott et al., 1996, 1997), and
optical-imaging (Youngentob et al., 1995) studies in amphibians and
rodents have demonstrated that individual odorants differentially
activate broad regions of the olfactory epithelium (OE) and that ORNs
are "broadly tuned," responding to many, often structurally
dissimilar, molecules. In addition, ORNs that are near neighbors in the
OE respond differently to odorants (Sato et al., 1994). As a result, it
is difficult to predict the response properties of ORNs based on their
epithelial locations.
ORNs send single unbranched axons back to the olfactory bulb (OB),
where they synapse on dendrites of mitral and tufted cells in spherical
regions of neuropil called glomeruli. In mice there are ~1800
glomeruli that cover the surface of the OB (Royet et al., 1988; Pomeroy
et al., 1990). The connections between the OE and the OB lack a strict
point-to-point topography; ORNs in broad regions of the OE converge
onto localized regions of the OB (Kauer, 1980; Astic and Saucier, 1986;
Astic et al., 1987; Stewart and Pedersen, 1987; Schoenfeld et al.,
1994). Recent molecular data in rodents have shown that spatially
distributed ORNs that express the same putative odorant receptor (OR)
gene send convergent projections to pairs of glomeruli in a remarkably
precise and reproducible manner (Ressler et al., 1994; Vassar et al.,
1994; Mombaerts et al., 1996). This suggests that ORN afferents can segregate by odorant responsiveness and that local regions of the OB
may receive inputs from ORNs with similar response properties (Kauer,
1980, 1987; Shepherd, 1994; Mori, 1995). This idea is supported by
physiological data suggesting that individual odorants are represented
in part by the spatiotemporal activation of ensembles of glomeruli
(Stewart et al., 1979; Jourdan et al., 1980; Lancet et al., 1982;
Guthrie et al., 1993; Cinelli et al., 1995; Friedrich and Korsching,
1997; Joerges et al., 1997). However, there are few data relating the
expression of particular OR sequences with functional properties of
ORNs (Zhao et al., 1998), and there is presently no information
relating the differential expression of OR sequences with the spatial
representation of odorant stimuli. Furthermore, there have been no
studies in rodents that directly measure the odorant response
properties of ORNs that send convergent projections to identified
regions of the OB (Friedrich and Korsching, 1997; Joerges et al.,
1997).
In the present paper, we have studied the relationship between odorant
response properties of ORNs and their projection sites onto local OB
regions. Through a combination of retrograde tracing and calcium
imaging, we have recorded from single ORNs that project to defined,
localized regions of the dorsal OB. The data provide the first direct
characterization of the functional topography of projections between
the OE and OB in mammals assayed at the level of individual ORNs.
| |
MATERIALS AND METHODS |
Animals. All experiments were performed using
postnatal day 12-15 (P12-P15) CF-1 mice (Charles River, Wilmington,
MA). Young animals were used to facilitate the surgical manipulations
required for glomerular imaging and retrograde tracing. CF-1 mice were chosen because vital imaging of glomeruli has been performed in this
strain (LaMantia and Purves, 1989; LaMantia et al., 1992), and the
postnatal development of the OE and OB has been well studied (Pomeroy
et al., 1990).
Retrograde tracing. Mice were anesthetized with ketamine (90 mg/kg) and xylazine (18 mg/kg), and the dorsal surface of the left OB
was exposed with the dura intact by cutting a ~1.5 × 1.8 mm
window in the cranium. The windows were positioned with respect to the
midline suture between the frontal bones and the posterior end of the
OB and typically exposed 70% of the dorsal OB. Glomeruli were
visualized using methods modified from those of LaMantia et al. (1992).
The OBs were stained for 15-25 min with a 2 mg/ml solution of the
fluorescent dye RH414 (Molecular Probes, Eugene, OR). Staining was
achieved without pretreatment with hypertonic saline (LaMantia et al.,
1992) by sweeping the dura with a cotton swab before applying the dye.
Excess dye was removed, and the animals were placed in a head holder
mounted to the modified stage of an upright fluorescence microscope.
RH414-stained glomeruli and microspheres used for injections were
viewed simultaneously with a long-pass fluorescein filter set
(excitation, 490 nm; dichroic, 500 nm; and emission, >515 nm) and a
PlanApo 4× objective.
Small injections of green latex microspheres (Katz and Iarovici, 1990)
(LumaFluor, Naples, FL) were made into visualized groups of glomeruli
in the dorsomedial (DM) and dorsolateral (DL) OB using an injection
micropipette (tip size, ~5 µm) connected to a 1 ml glass syringe
via Teflon tubing. The entire assembly was filled with Fluorinert FC-77
(Sigma, St. Louis, MO) allowing small volumes of tracer to be sucked up
and expelled from the injection micropipette. Placement of the
injections was made with reference to the medial and rostral edges of
the craniotomy. Although glomerular landmarks were not used to
position injections, staining of the glomeruli greatly aided our
ability to inject the glomerular layer under fluorescence illumination
and permitted injection size to be determined in relation to glomerular
size during the injection procedure. Video images of the OBs were
acquired before and immediately after injection and before cell
dissociation (or histological analysis) to quantify the sizes of the
injections and to monitor diffusion of the tracer from the injection
site. Based on these images, the mean area of the labeled dorsal
glomeruli was 6681.8 ± 198.6 µm2 (mean ± SEM; n = 208); the injections encompassed ~21
glomeruli for the DM site (0.14 ± 0.03 mm2,
mean ± SD) and ~32 glomeruli (0.22 ± 0.08 mm2) for the DL site. Lateral tracer diffusion was
minimal during the retrograde tracing period.
In addition, the exact locations of all the injections included in the
analyses were determined from video images of the entire exposed OBs
(for example, see Fig. 2E,F). Injections
outside the DM and DL regions were not included in the analyses. The
precise location of the injection site centers with respect to the
medial and rostral edges of the OB were measured as fractions of total bulbar width and length after being scaled to a "standard" bulb of
average dimensions. The standardized locations of the injection site
centers for animals included in the physiological analyses as measured
from the medial edge and rostral tip of the OB were DM = 0.39 ± 0.09 × 1.21 ± 0.09 mm; and DL = 1.36 ± 0.11 × 1.25 ± 0.14 mm (mean ± SD). Retrograde tracing
with RH414 and microspheres was evident in the OE by 24 hr. Tracing
periods were 1-3 d for calcium imaging and 3-6 d for histology and
whole mounts.
Histology. Labeling was viewed in unfixed tissue, because
RH414 fluorescence is abolished by short periods of paraformaldehyde fixation. For cryosections, the intact septum and turbinates were removed and washed in PBS and cryoprotected in PBS and 30% sucrose overnight. The tissue was then embedded and frozen in O.C.T. and cut
into 20 µm sections in a freezing microtome. For whole mounts, the
septum and turbinates were removed and placed on depression slides in
Ringer's solution (below). Whole mounts and sections were viewed and
photographed using standard rhodamine and fluorescein filter sets. All
images were digitized and assembled for display using Adobe Photoshop
4.0 (Adobe Systems) using the minimum digital image processing
necessary to accurately reproduce the data.
Cell dissociation and calcium imaging. Procedures were
modified from those of Restrepo et al. (1993). Animals were killed, and
the OE was dissected and minced at 4°C in low-Ca2+
Ringer's solution (in mM: 140 NaCl, 5 KCl, 10 HEPES, 1 EDTA, 10 glucose, and 1 sodium pyruvate, pH 7.2) supplemented with 1 mM cysteine. The tissue was incubated in 1 U/ml papain
(Sigma) for 4 min at room temperature, and the reaction was terminated with 1 ml of Ringer's solution (in mM: 140 NaCl, 5 KCl, 1 CaCl2, 1 MgCl2, 10 HEPES, 10 glucose, and 1 sodium pyruvate, pH 7.2) supplemented with 0.1 mg/ml
BSA, 200 µg/ml leupeptin (Sigma), and 0.025 mg/ml DNase I
(Sigma). Cells were washed, resuspended in Ringer's solution with
8 µM fura-2 AM and 80 µg/ml Pluronic F-127 (Molecular
Probes), gently triturated through a flame-polished Pasteur pipette,
plated on glass coverslips (coated with Con A type V, 2 mg/ml; and
poly-L-lysine, 1 mg/ml), and loaded for 45 min-1 hr at
room temperature. The coverslips formed the bottom of a Teflon
recording chamber (Biophysica, Sparks, MD). Dissociated preparations
were constantly perfused with normal Ringer's solution (~3 ml/min)
except during stimulus presentations.
Stimuli were bath-applied for 4-10 sec. Complete washout of the
chamber required 1-2 min. The depolarizing stimulus was a high-K+ Ringer's solution (in mM: 40 NaCl, 100 KCl, 1 CaCl2, 1 MgCl2, 10 HEPES, 10 glucose, and 1 sodium pyruvate, pH 7.2). The odorant stimuli were a set of six mixtures (mixes A-F) of 32 different molecules (Fig. 1). Mixes A-D were
designed so that each mixture included molecules with various chemical
structures and odor qualities. Mix B was also designed as a mixture of
highly water-soluble odorants. Mixes E and F were designed to contain
odorants that have been reported to stimulate cAMP and IP3
pathways, respectively (Sklar et al., 1986; Breer and Boekhoff, 1991;
Restrepo et al., 1993). Stimulus solutions were prepared as 1 mM stocks in Ringer's solution and diluted to a final
concentration of 50 µM for each odorant in the solution.
This stimulus concentration is well within the range (1 nM-1 mM) of those used in previous studies on
dissociated mammalian ORNs (Restrepo et al., 1993; Sato et al., 1994;
Tareilus et al., 1995). Stocks were vortexed and sonicated before and
after dilution to ensure that all components were in solution.
View larger version (21K):
[in this window]
[in a new window]
|
Figure 1.
Molecular composition of the six odorant mixtures
(A-F). Each mixture was designed to include many
different classes of molecules (see Materials and Methods). Mix
E and Mix F are mixtures of molecules that have
been reported to increase cAMP and IP3, respectively
(Sklar et al., 1986; Breer and Boekhoff, 1991). Four of the molecules
in Mix E are also found in other mixtures: geraniol and
acetophenone are in A and E, eugenol is
in C and E, and citralva is in
D and E.
|
|
Optical recordings were obtained using an inverted microscope with a
CCD video camera coupled to an image intensifier; the light source was
a 75 W xenon lamp attenuated with neutral density filters. ORNs labeled
with RH414 and/or green microspheres could be viewed simultaneously
using a long-pass fluorescein filter set. Dual-excitation calcium
imaging was performed using a standard fura-2 filter set (Omega
Optical, Brattleboro, VT). Data were acquired every 4 sec; the average
pixel values over the cell bodies of selected cells were calculated at
340 and 380 nm excitation, and ratios were obtained. Filter wheel
control and image acquisition were performed using software designed in
our laboratory.
Ratiometric imaging controlled for occasional movement artifacts and
allowed us to estimate the magnitude of the stimulus-induced [Ca2+]i changes in a sample of
responsive ORNs. [Ca2+]i was estimated
from the ratios using standard methods (Grynkiewicz et al., 1985). The
Kd of fura-2 for Ca2+ was
taken to be 220 nM; Rmax and
Rmin were determined in intact cells by adding 5 µM ionomycin in Ringer's solution (1 mM
Ca2+) to saturate the fura-2 and by subsequently
bathing the cell in low-Ca2+ Ringer's solution
supplemented with 5 mM EGTA. The mean resting [Ca2+]i was 68 ± 6 nM (mean ± SEM; n = 4), and the peak responses
to odorants were within the 400-600 nM range. These
numbers should be considered approximate estimates of absolute
[Ca2+]i, because the
calibration is prone to several errors, including variation in the
apparent Kd of fura-2 attributable to pH and ionic strength as well as incomplete equilibration of internal and
external Ca2+ using ionomycin.
Dissociated ORNs were identified if they were retrogradely labeled and
exhibited a fast onset, rapidly recovering Ca2+
response to KCl depolarization. Many traced ORNs retained
characteristic morphology, including a dendrite and cilia; however, the
tendency was for ORNs to retract their dendrites, forming a bowling
pin-like shape. Unlabeled cells were considered to be ORNs based on
morphological criteria and because they showed fast KCl and
odorant-induced Ca2+ transients.
Analysis. All responses were repeated at least once; we took
the largest response to a given stimulus to include in the analysis. Responses were normalized to the KCl response within cells to control
for imaging artifacts present in some recordings. Stepwise logistic
regression was used for pair-wise comparisons among the four groups
[DM, DL, dorsal (D), and unlabeled]. This procedure tests for
differences among populations in which the individual members are
defined by a set of variables, in this case the normalized response
amplitudes to each of the six mixtures. The analysis finds the
combination of independent variables that best discriminate between two
groups and estimates the probability that the groups differ.
Significant differences are reported along with the variables (odorant
mixtures) included in the model. Data were analyzed using SPSS 8.0 software and plotted using Mathematica 3.0 software (Wolfram Research).
| |
RESULTS |
Retrograde tracing from visualized groups of glomeruli
Focal injections of fluorescein-labeled microspheres were made
into clusters of visualized glomeruli in the DM or DL OB (Fig. 2A,B, insets). DM
injections typically involved ~20 glomeruli, and DL injections
involved ~30 glomeruli (see Materials and Methods); this represents
2-3% of the total number of glomeruli (~1000) in 2-week-old CF-1
mice (Pomeroy et al., 1990). Smaller injections (one to four glomerular
widths) were made in preliminary experiments but resulted in too few
labeled ORNs to make physiological recordings feasible. Injections were
localized using the anterior and medial edges of the craniotomy as
landmarks, and care was taken to inject the same region of the DM
or DL OB across animals (see Materials and Methods). The width of the
OB is ~1.8 mm at this age, and DL and DM sites were separated by ~1
mm (Fig. 2A,B, insets, E,F).
View larger version (76K):
[in this window]
[in a new window]
|
Figure 2.
Distribution of retrogradely labeled ORNs in the
OE. A, B, insets, Fluorescence images of the dorsal OBs
of living mice taken immediately after injection of fluorescein-labeled
latex microspheres (yellow) into the DL
(A) and DM (B) sites.
Glomeruli are seen (orange) in the regions stained with
RH414. A-D, Whole mounts of the septum (A,
B) and the medial face of the turbinates (C, D)
show the distribution of microsphere label 4 d after injection.
The anterior lateral OE is reflected dorsally in A and
B. Turbinates are numbered
II-IV in C and
D. DL labeling was detectable in the anterior septum
(A) and in turbinates II and
II' (C). DM labeling formed an
anterior to posterior band on the septal OE (B)
and in the dorsal turbinates (D). Labeling in
turbinate IV in D is out of the place of
focus. E, F, Combined fluorescence and
bright-field video images of both OBs exposed 2 d after injection
with microspheres. In these images anterior is left, and
the DL (E) and DM
(F) injection sites are shown as
bright spots in the left OBs.
G, Coronal 20-µm-thick section through the nasal
cavity showing microspheres (yellow) and RH414
(orange) label in the OE 3 d after a large
injection into the dorsocentral OB. A, Anterior;
M, medial; S, septum. Scale bar:
A-D, insets, 200 µm; E, F, 230 µm;
G, 100 µm.
|
|
The microspheres were sufficiently bright to allow the tracing patterns
to be observed in whole mounts of the septum and turbinates (DM,
n = 12; DL, n = 8). Microsphere-labeled
ORNs that were retrogradely traced from the two sites (DM-projecting
and DL-projecting ORNs) were found in distinct but overlapping regions
of the septum and turbinates of the OE (Fig. 2A-D).
DL-projecting ORNs were found exclusively in the anterior dorsal recess
and anterior turbinates II and II' (Fig. 2A,C). These
data are consistent with patterns observed with injections into the
DL quadrant of the OB in rat and hamster (Astic and Saucier, 1986;
Astic et al., 1987; Schoenfeld et al., 1994). In contrast,
DM-projecting ORNs were found in a stripe occupying the entire anterior
to posterior extent of the dorsal septum and turbinates (Fig.
2B,D). Interestingly, an anteroposterior band of
labeling was only observed if the retrograde tracer had access to the
extreme medial edge of the OB. Injections placed ~400 µm lateral to
the DM site resulted in a DL pattern of labeling or in a DL pattern in
which a small fraction of the total label could be found in the caudal
OE (n = 3; data not shown). Thus the DM site seems to
be close to a transition point in the anatomical projection patterns
between the OB and OE (Schoenfeld et al., 1994).
Surprisingly, RH414, which was used to visualize glomeruli, also acted
as a retrograde tracer (Tsau et al., 1996) and could be observed in
whole mounts (data not shown) and sections of the OE (Fig.
2G). Sections of OE and OB clearly demonstrated that RH414
stained only the dorsal half (and preferentially the DL surface) of the
OB (data not shown). This was likely attributable to differential
access of the dye to regions of the bulb through the dura, because
breaching the dura with high-osmolarity treatment resulted in more
uniform staining of the dorsal OB (LaMantia et al., 1992). The tracing
pattern observed with RH414 was almost identical to the DL projection
pattern seen with the smaller microsphere injections. However, within
the labeled regions, many more ORNs were labeled with RH414 than with
microspheres. This fortuitous result allowed us to use RH414 as a
marker for ORNs that project to a large region of the D half of the
OB.
Responses of traced ORNs to odorant mixtures
Dissociated ORNs were loaded with fura-2 for measuring
odorant-induced calcium responses (Restrepo et al., 1990). The stimuli were a set of six odorant mixtures (mixes A-F), each of which was
designed to include molecules with disparate chemical structures (see
Materials and Methods, Fig. 1). We wished to test for response biases
of ORNs based on where they project to the OB without making a
priori assumptions as to which molecules might stimulate the different anatomical groups. The use of mixtures permitted initial screening of ORNs with many structurally diverse odorants using relatively few stimulus applications. After initial characterization, the mixtures were then broken down to identify individually effective stimuli.
DM- or DL-projecting microsphere-labeled ORNs were easily identified in
the dissociated preparations by bright green, punctate fluorescence
(Fig. 3A,B), whereas the
D-projecting, RH414-labeled ORNs were identified by red, punctate
fluorescence (data not shown). We recorded from 200 retrogradely
labeled ORNs, each of which was stimulated with KCl (100 mM) to assess viability and subsequently tested with all
six mixtures. Of these, 12 of 33 (36.4%) D-projecting, 21 of 99 (21.2%) DL-projecting, and 20 of 68 (29.4%) DM-projecting ORNs
responded to at least one of the mixtures. Stimulation with KCl or
odorants elicited short-latency increases in
[Ca2+]i that typically lasted 1-2 min
(Fig. 3C). The durations of the odorant responses were
similar and in some cases shorter than the somatic
Ca2+ transients seen in previous studies of
dissociated ORNs (Restrepo et al., 1993; Sato et al., 1994; Tareilus et
al., 1995; Leinders-Zufall et al., 1997) (see Discussion).
View larger version (55K):
[in this window]
[in a new window]
|
Figure 3.
Odorant responses of a retrogradely labeled ORN
tested with odorant mixtures. A, Dissociated
odorant-responsive ORN viewed under Nomarsky optics showing a short
dendrite and cilia. B, Fluorescence image of the same
field showing the microsphere labeling (white dots).
Microsphere-labeled ORNs were often but not always labeled with RH414.
Scale bar: A, B, 10 µm. C, Fura-2
measurements in a single retrogradely labeled ORN (dorsolateral
#8a in Fig. 4A) stimulated with 10 sec pulses (black bars) of six odorant mixtures
(A-F) and an OA mix, as well as with a 4 sec
pulse of KCl. Inset, "Dot" plot of the odorant
response profile derived from the analog data for this cell. The
dot area represents the response amplitude as a
percentage of the KCl response; the largest responses observed to each
mixture were used for these plots. Dashes indicate no
response. This cell responded to mixes A and E.
|
|
We first asked whether selecting ORNs that project to different
discrete regions of the dorsal OB biases the responses to our stimulus
set. DM- and DL-projecting ORNs exhibited markedly different response
profiles, preferentially responding to different odorant mixtures (Fig.
4, compare A, B). Overall, the
DL group responded most often to mixes C and E, with the most common
response profile being "A, C, E." In fact, there was a significant
correlation between C and E responses (Pearson correlation coefficient,
r = 0.613; p < 0.01). We reasoned that
this might be caused by the fact that mixes C and E both contain
eugenol (see below). In contrast, the DM group responded best to mix B,
with the most common profile being "B" only. The most striking
differences between the two groups were that responses to mixes C and E
observed in the DL population were completely absent from the DM
population and that the number of responses to mix B observed in the DM
population was significantly higher than in the DL population
(Fisher's exact test, p < 0.001). The two populations
could be separated with respect to C and E responsiveness (logistic
regression analysis, p < 0.001). These data
demonstrate directly that individual ORNs that are partially
intermingled in the OE and that project to two different loci in
the OB exhibit different response profiles to the same stimulus
set.
View larger version (21K):
[in this window]
[in a new window]
|
Figure 4.
Odorant response profiles for ORNs responsive to
the odorant mixtures. A, DL-projecting
microsphere-labeled ORNs (n = 21).
B, DM-projecting microsphere-labeled ORNs
(n = 20). C, RH414-labeled,
D-projecting ORNs (n = 12). D,
Unlabeled ORNs (n = 17). Each row
shows the responses of a single ORN to the six mixtures
(A-F) as well as to the OA mix, as in
Figure 3C. Blank spaces indicate that the
stimulus was not tested, and dashes indicate no
response. The percentage of responsive cells responding to each mixture
is indicated below each plot. Cell identification
numbers are to the left; within each
panel, cells with the same number are from the same animal. Cells are
ordered by increasing breadth of response.
|
|
We next asked whether the response properties of DM- or DL-projecting
ORNs were distinguishable from those of ORNs projecting to more broad
regions of the OB. These included RH414-labeled, D-projecting ORNs and
17 unlabeled ORNs, which likely project to regions of the OB (ventral
half) not exposed to either RH414 or microspheres (Fig.
4C,D). Responses seen in the DL-projecting ORNs were not
statistically different from responses in 12 D-projecting ORNs but were
significantly different from 17 unlabeled ORNs with respect to D and E
responses (logistic regression, p < 0.001). Thus, with
respect to this stimulus set, responses of ORNs that project to a small
DL region of the OB were indistinguishable from those projecting to a
more broad dorsal OB region.
In contrast, DM-projecting ORNs were separable from both D-projecting
(p < 0.001, with respect to C and B responses)
and unlabeled ORNs (p < 0.001, with respect to
C and D responses). Interestingly, when compared with the other three
groups, DM neurons exhibited the least variability in the number of
different profiles per sample size, implying that this group is the
most homogeneous of all the samples. This holds true when comparing
populations of ORNs that project to similar numbers of glomeruli; for
DL-projecting ORNs (n = 21) there were 12 different
response profiles with respect to the six mixtures, although for
DM-projecting ORNs (n = 20) there were only five
profiles (Fig. 4A,B). These data suggest that the
homogeneity of convergent ORNs may vary for different regions of the
OB.
The response profiles of DM cells consisted exclusively of responses to
mixtures containing organic acids (mixes A, B, and F). This led us to
ask whether DM-projecting ORNs respond preferentially to the acids in
our stimulus set. As a first step in testing this, 18 ORNs stimulated
with mixes A-F were also tested with an organic acids (OA) mixture
containing only the acids from the original six mixtures. Seven of nine
DM-projecting and two of nine DL-projecting ORNs responded to the OA
mix (Fig. 4A,B). These data suggest that a higher
proportion of DM-projecting ORNs responds to the organic acids in the
mixtures compared with DL-projecting ORNs.
Responses of traced ORNs to mixture components
To identify individual odorants that activate ORNs, we tested
cells with components of the individual mixtures. Because a majority of
responsive DL-projecting ORNs (86%) responded to mix C, we sought to
identify the molecular species in mix C that stimulated this group. To
do this, we performed an experiment in which another 92 DL-projecting
ORNs were stimulated with mix C and its components (carvone, eugenol,
cinnamaldehyde, limonene, citral, and ethyl-fenchol) as well as
geraniol and acetophenone (Fig.
5A). The latter two stimuli
are components of mixes A and E that we reasoned DL-projecting ORNs
might respond to given the prevalence of A, C, E profiles in the
mixture data. In this experiment, 19 cells responded to at least one
stimulus, and 17 responded to mix C. Of the 17 mix C-responsive
ORNs, 9 responded to carvone, eugenol, or cinnamaldehyde only, 5 responded to more than one of the individual mix C components, and 3 did not respond to any of the components (Fig. 5C). There were no responses observed with limonene, geraniol, or ethyl-fenchol, and mix C-responsive ORNs typically did not respond to the mix A and E
components geraniol and acetophenone. Interestingly, responses to a
given molecule (carvone, for instance) were observed in the context of
a number of different response profiles, providing a clear
demonstration that, within this anatomically restricted set of ORNs, a
given odorant induces responses in cells with different but overlapping
response properties.
View larger version (33K):
[in this window]
[in a new window]
|
Figure 5.
Odorant responses of retrogradely labeled ORNs to
individual mixture components. A, Calcium responses from
a single DL-projecting ORN tested with mix C and its components,
(+)-carvone (Car), eugenol (Eug),
cinnamaldehyde (Cin), ( )-limonene
(Lim), citral (Cit), and 2-ethyl fenchol
(Fen) and with geraniol (Ger) and
acetophenone (Ace), two components of mixes A and E. Rin and KCl mark stimulations with normal
and high-K+ Ringer's solution, respectively.
B, Calcium responses from a single DM-projecting ORN
tested with mix B and its components, n-valeric acid
(5A), n-heptanoic acid
(7A), n-pelargonic acid
(9A), n-heptanol
(7OH), n-hexanol
(6OH), n-butanol
(4OH), and benzyl alcohol
(BOH) and with mix C. Dot response profiles for
the two cells are shown in the insets. The small dip in
the ratio after 4OH in B is an optical
artifact. C, D, Summary of response
profiles (as in Fig. 4) for 19 DL-projecting and 18 DM-projecting ORNs.
Blank spaces indicate that the stimulus was not tested,
and dashes indicate no response.
|
|
For the DM group, we had shown that mix B was the most effective
stimulus, eliciting responses in 85% of the responsive ORNs. As a
result, a new group of 98 DM-projecting ORNs was stimulated with mix B
and its component odorants (valeric acid, heptanoic acid, pelargonic
acid, heptanol, hexanol, butanol, and benzyl-alcohol) and mix C (Fig.
5B). In this experiment, 18 cells responded to at least one
of the stimuli. Of 17 mix B-responsive cells, 16 responded to at least
one of the individual organic acids, and 14 responded to the acids but
not the alcohols (Fig. 5D).
It has been shown previously that ORNs isolated from mouse septal OE
respond to carboxylic acids and alcohols of similar carbon chain
length, but that a majority do not discriminate acids from alcohols
when tested over a range of concentrations (Sato et al., 1994).
Interestingly, 88% of the acid-responsive DM-projecting ORNs, which
lie in a similar region of the septal OE, discriminated the acids from
alcohols of similar carbon chain length at the single concentration
used here. Most strikingly, of the nine heptanoic acid-responsive
neurons, only one responded to heptanol. Although it is possible
that higher stimulus concentrations might have induced alcohol
responses in these ORNs, our stimulus concentrations were already at
the high end of those that might be encountered under physiological
conditions.
Taken with the results using mixtures, these data show that a majority
of responsive DM-projecting ORNs respond to similar organic acids
in our stimulus set. Thus, the DM OB likely receives a large proportion
of its inputs from organic acid-responsive ORNs.
| |
DISCUSSION |
The goal of the present study was to characterize the spatial
representation of odorant information in the projections between the OE
and the OB in a species in which there are anatomical and OR gene
expression data. Our results provide direct evidence that ORNs
projecting to two regions of the dorsal OB respond best to individual
molecules that are structurally different, and that these regions
appear to differ in the homogeneity of their response profiles. The
data also indicate that a subset of distributed ORNs that converge onto
a small region of the DM OB can be shown to respond to odorants with
similar chemical structures. These findings are consistent with
molecular data showing that ORNs expressing the same OR gene converge
onto glomeruli in the OB. Moreover, given the lack of information
on the relationship between the OR genes and the ligand-binding
properties of their respective proteins (Zhao et al., 1998), these
studies provide a view of the functional topography of the olfactory
system not provided by previous molecular investigations.
Retrograde tracing patterns
Several lines of evidence suggest that there may be a mediolateral
symmetry to the organization of the mammalian OB. ORNs that express the
same OR gene converge onto medial and lateral glomeruli (Ressler et
al., 1994; Vassar et al., 1994; Mombaerts et al., 1996), and the medial
and lateral "hemispheres" of the OB are connected by an intrabulbar
association pathway connecting homotopic regions of the OB (Schoenfeld
et al., 1985). However, the true mediolateral and dorsoventral axes of
the OB are thought to be rotated medially about the anteroposterior
axis (Schoenfeld et al., 1985, 1994). With respect to these
"corrected" axes, our DL site is within the lateral half of the OB,
while our DM site is located at or near the true dorsal meridian. Thus,
DM injections likely labeled ORNs projecting to both the medial and
lateral hemispheres, whereas DL injections likely labeled only ORNs
projecting to the lateral hemisphere. The DM and DL tracing patterns
were strikingly different but did overlap considerably in the
anterior OE. The DM pattern resembled the anteroposterior stripe of OR gene expression in the dorsal-most OE region ("zone 1") observed in
in situ hybridization studies (Ressler et al., 1993; Vassar et al., 1993). In contrast, the DL pattern formed only the rostral half
of a more ventrally located stripe. These data support the idea that a
complement of both medially and laterally projecting ORNs is required
to produce a complete anteroposterior stripe of ORNs on either the
septum or turbinates (Schoenfeld et al., 1994; Mombaerts et al.,
1996).
Response properties of retrogradely traced ORNs
Our experiments required the use of enzymes to dissociate ORNs.
The cells included in our analyses maintained normal physiological [Ca2+]i, exhibited robust
calcium transients to KCl and odorant stimulation, and responded
differentially to odorants. These observations attest to the health of
the neurons. Our findings are consistent with studies showing that
somatic Ca2+ responses typically last 1-2 min after
brief odorant or IBMX stimulation (Restrepo et al., 1993; Sato et al.,
1994; Tareilus et al., 1995; Leinders-Zufall et al., 1997). Thus, the
time course of our Ca2+ responses is unlikely to be
attributable solely to stimulus washout time but may also reflect
intrinsic Ca2+ dynamics in ORNs (e.g.,
Ca2+ release from intracellular compartments;
Leinders-Zufall et al., 1997).
There are few studies that have characterized the response breadth of
individual mammalian ORNs using broad classes of odorants (Sicard,
1986). Data from amphibians show that the response selectivity of ORNs
varies considerably (Duchamp et al., 1974; Getchell, 1974; Revial et
al., 1978, 1982). Similarly, in the present experiments, some mouse
ORNs responded to stimuli with quite different structures, whereas
others displayed a remarkable ability to discriminate among molecules
that differed by small changes in carbon chain length or functional
group (Sato et al., 1994). Thus, as in other species, odorant
information in the mouse is likely encoded by populations of variably
selective ORNs.
Our view of the tuning characteristics among groups of traced ORNs may
be dependent on the odorant set used. This is illustrated by the fact
that ~75% of the KCl-responsive ORNs we tested failed to respond to
odorants, either because they were damaged by the dissociation
procedure or were not presented with an adequate stimulus. Thus, some
undefined proportion of potentially responsive cells were not
characterized, because they did not respond to any of the odorants
presented. Therefore, the data represent one view of a subpopulation of
anatomically defined ORNs selected by responsiveness to this particular
odorant set. We consider it significant, however, when starting with a
diverse odorant set chosen more or less randomly, that prominent
response differences among anatomically selected ORNs emerged. This
suggests that the overall differences in responsiveness between the DM
and DL sites is robust and can likely be detected using a number of
stimulus sets.
The assessment of variability between groups of traced ORNs is likely
to be influenced strongly by the choice of stimuli and by variability
in injection sizes and locations. It is worth noting that DL injections
involved a few more glomeruli and varied more in location across
animals than did DM injections. Consequently, the differences in
heterogeneity between the groups may be related to differences in
variability in placing the injection sites. Both sets of injections,
however, were similar with respect to percentage of the entire
glomerular sheet labeled.
Another possible difference might be in the developmental stages of
ORNs projecting to the DM and DL regions in young animals. This would
appear less likely for several reasons. The available data suggest that
ORNs are distributed in adult-like patterns and express molecules
involved in odorant transduction before birth (Margalit and Lancet,
1993; Menco et al., 1994; Sullivan et al., 1995), and that potential
effects of immaturity on odorant responsiveness are gone by embryonic
day 19 in rats (Gesteland et al., 1982). In addition, medially and
laterally projecting neurons expressing the P2 OR gene are
qualitatively similar in P0 and adult animals at the gross
morphological level (Mombaerts et al., 1996), and ORN terminals within
glomeruli at this time are also morphologically indistinguishable from
adults (Klenoff and Greer, 1998). If the DM population is in fact more
homogeneous, an interesting conjecture is that the homogeneity of ORNs
projecting to groups of glomeruli may vary with location relative to
natural anatomical boundaries in the OB (see above).
Functional topography of the mammalian OB
Much of what we know about the distribution of odorant-induced
activity across the glomerular layer in mammals has come from summed
activity measures or from recordings of OB output neurons. Our data are
the first to address this issue at the level of individual ORNs.
Studies of 2-deoxyglucose uptake in rats have demonstrated a DM focal
region of bulbar activation after stimulation with propionic acid (Bell
et al., 1987), a component of mix A. However, DM-projecting ORNs were
not preferentially responsive to mix A as might be expected. Although
it is possible that a high proportion of propionic acid-responsive
cells project to regions near the DM site, this will have to be tested
explicitly. Our data fit well with single-unit recordings from rabbit
mitral and tufted cells showing that neurons in the DM OB are activated
in response to longer-chain carboxylic acids (and aldehydes) but not
alcohols of similar carbon chain length (Imamura et al., 1992; Mori et al., 1992). Our data also provide support for the hypothesis that the
response bias of DM-situated mitral cells may be attributed to direct
input from organic acid-responsive ORNs. Unlike for the DM OB, the
odorant response properties of individual mitral cells in the DL OB are
not known. Our studies have identified a set of molecules (carvone,
eugenol, and cinnamaldehyde) to which mitral cells in the DL mouse
OB may be preferentially responsive. The correspondence between our
data from young mice and these data from adult rabbits suggests that
the response biases observed in both studies may reflect a general
feature of the organization of mammalian OBs.
Functional convergence of ORN afferents
DM-projecting ORNs were clearly biased toward responding to the
organic acids in our stimulus set. It is important to realize that the
data do not demonstrate that DM-projecting ORNs respond only to organic
acids for several reasons. First, as noted above, some proportion of
the unresponsive ORNs may have responded to odorants that were not
included in our stimulus set. Second, the data show that two
DM-projecting ORNs did not respond to acids (Fig. 4B, #8,
#7a) and four DM-projecting ORNs also responded to nonacid stimuli
(Fig. 5D, #2a, #9b, #6, #10b). Taken together, the data
suggest that acid-responsive ORNs can synapse in close proximity to
ORNs that respond to other types of molecules. Nonetheless, the
prevalence of the acid-responsive profiles demonstrates that the DM OB
is a site of convergence of a large proportion of acid-responsive ORNs.
Because our injection sites encompass ~20 glomeruli, it is likely
that several glomeruli in this region receive inputs from
acid-responsive ORNs. These observations indicate that some groups of
glomeruli receive afferents from ORNs that carry similar information
(Friedrich and Korsching, 1997).
One of the operations that OB circuitry may perform on incoming
afferent activity is a modification of inputs to neighboring glomeruli
via lateral inhibition (Yokoi et al., 1995). It is generally thought
that glomeruli may be functional units (Jourdan et al., 1980; Kauer,
1980, 1987; Lancet et al., 1982; Guthrie et al., 1993; Kauer and
Cinelli, 1993; Shepherd, 1994; Mori, 1995), receiving inputs from ORNs
with similar response properties. If this is true, a key issue in
olfactory coding will be to understand how similar the response
properties are of ORNs projecting to a single glomerulus or groups of
glomeruli among which lateral interactions occur. This question bears
directly on the kind of processing that takes place in local OB
circuits.
| |
FOOTNOTES |
Received Jan. 12, 1998; revised March 16, 1998; accepted March 25, 1998.
This work was supported by a grant from National Institutes of Health,
National Institute on Deafness and Other Communication Disorders. We
thank Diego Restrepo for advice and assistance with calcium imaging, A. LaMantia for advice concerning glomerular imaging, and William Rand for
help with statistical analyses. We thank Dale Hunter, Joel White,
Kathleen Dorries, and Tarik Alkasab for valuable discussion and
assistance in preparing this paper.
Correspondence should be addressed to Dr. John S. Kauer, Department of
Neuroscience, Tufts University School of Medicine, 136 Harrison Avenue,
Boston, MA 02111.
Dr. Bozza's present address: The Rockefeller University, 1230 York
Avenue, New York, NY 10021.
| |
REFERENCES |
-
Astic L,
Saucier D
(1986)
Anatomical mapping of the neuroepithelial projection to the olfactory bulb in the rat.
Brain Res Bull
16:445-454[Web of Science][Medline].
-
Astic L,
Saucier D,
Holley A
(1987)
Topographical relationships between olfactory receptor cells and glomerular foci in the rat olfactory bulb.
Brain Res
424:144-152[Web of Science][Medline].
-
Bell GA,
Laing DG,
Panhuber H
(1987)
Odor mixture suppression: evidence for a peripheral mechanism in human and rat.
Brain Res
426:8-18[Web of Science][Medline].
-
Breer H,
Boekhoff I
(1991)
Odorants of the same odor class activate different second messenger pathways.
Chem Senses
16:19-29[Abstract/Free Full Text].
-
Cinelli AR,
Hamilton KA,
Kauer JS
(1995)
Salamander olfactory bulb neuronal activity observed by video rate, voltage-sensitive dye imaging: III. Spatial and temporal properties of responses evoked by odorant stimulation.
J Neurophysiol
73:2053-2071[Abstract/Free Full Text].
-
Duchamp A,
Revial MF,
Holley A,
MacLeod P
(1974)
Odour discrimination by frog olfactory neurons.
Chem Senses
1:213-233[Abstract/Free Full Text].
-
Friedrich RW,
Korsching SI
(1997)
Combinatorial and chemotopic odorant coding in the zebrafish olfactory bulb visualized by optical imaging.
Neuron
18:737-752[Web of Science][Medline].
-
Gesteland RC,
Yancey RA,
Farbman AI
(1982)
Development of olfactory receptor neuron selectivity in the rat fetus.
Neuroscience
7:3127-3136[Web of Science][Medline].
-
Getchell TV
(1974)
Unitary responses in frog olfactory epithelium to sterically related molecules at low concentrations.
J Gen Physiol
64:241-261[Abstract/Free Full Text].
-
Grynkiewicz G,
Poenie M,
Tsien RY
(1985)
A new generation of Ca2+ indicators with greatly improved fluorescence properties.
J Biol Chem
260:3440-3450[Abstract/Free Full Text].
-
Guthrie KM,
Anderson AJ,
Leon M,
Gall C
(1993)
Odor-induced increases in c-fos mRNA expression reveal an anatomical "unit" for odor processing in olfactory bulb.
Proc Natl Acad Sci USA
90:3329-3333[Abstract/Free Full Text].
-
Imamura K,
Mataga N,
Mori K
(1992)
Coding of odor molecules by mitral-tufted cells in rabbit olfactory bulb. I. Aliphatic compounds.
J Neurophysiol
68:1986-2002[Abstract/Free Full Text].
-
Joerges J,
Küttner A,
Galizia G,
Menzel R
(1997)
Representation of odours and odour mixtures visualized in the honeybee brain.
Nature
387:285-288.
-
Jourdan F,
Duveau A,
Astic L,
Holley A
(1980)
Spatial distribution of [14C]2-deoxyglucose uptake in the olfactory bulbs of rats stimulated with two different odours.
Brain Res
188:139-154[Web of Science][Medline].
-
Katz LC,
Iarovici AM
(1990)
Green fluorescent latex microspheres: a new retrograde tracer.
Neuroscience
34:511-520[Web of Science][Medline].
-
Kauer JS
(1980)
Some spatial characteristics of central information processing in the vertebrate olfactory system.
In: Olfaction and taste VII (van der Starre H,
ed), pp 227-236. London: IRL.
-
Kauer JS
(1987)
Coding in the olfactory system.
In: Neurobiology of taste and smell (Finger TE,
Silver WL,
eds), pp 205-231. New York: Wiley.
-
Kauer JS,
Cinelli AR
(1993)
Are there structural and functional modules in the vertebrate olfactory bulb?
Microsc Res Tech
24:157-167[Web of Science][Medline].
-
Klenoff JR,
Greer CA
(1998)
Postnatal development of olfactory receptor cell axonal arbors.
J Comp Neurol
390:256-267[Web of Science][Medline].
-
LaMantia AS,
Purves D
(1989)
Development of glomerular pattern visualized in the olfactory bulbs of living mice.
Nature
341:646-649[Medline].
-
LaMantia AS,
Pomeroy SL,
Purves D
(1992)
Vital imaging of glomeruli in the mouse olfactory bulb.
J Neurosci
12:976-988[Abstract].
-
Lancet D,
Greer CA,
Kauer JS,
Shepherd GM
(1982)
Mapping of odor-related neuronal activity in the olfactory bulb by high-resolution 2-deoxyglucose autoradiography.
Proc Natl Acad Sci USA
79:670-674[Abstract/Free Full Text].
-
Leinders-Zufall T,
Rand MN,
Shepherd GM,
Greer CA,
Zufall F
(1997)
Calcium entry through cyclic nucleotide-gated channels in individual cilia of olfactory receptor cells: spatiotemporal dynamics.
J Neurosci
17:4136-4148[Abstract/Free Full Text].
-
MacKay-Sim A,
Kesteven S
(1994)
Topographic patterns of responsiveness to odorants in the rat olfactory epithelium.
J Neurophysiol
71:397-398.
-
MacKay-Sim A,
Shaman P,
Moulton DG
(1982)
Topographic coding of olfactory quality: odorant specific patterns of epithelial responsivity in the salamander.
J Neurophysiol
48:584-596[Abstract/Free Full Text].
-
Margalit T,
Lancet D
(1993)
Expression of olfactory receptor and transduction genes during rat development.
Dev Brain Res
73:7-16[Medline].
-
Menco BPM,
Tekula FD,
Farbman AI,
Danho W
(1994)
Developmental expression of G-proteins and adenylyl cyclase in peripheral olfactory systems. Light microscopic and freeze-substitution electron microscopic immunocytochemistry.
J Neurocytol
23:708-727[Web of Science][Medline].
-
Mombaerts P,
Wang F,
Dulac C,
Chao SK,
Nemes A,
Mendelsohn M,
Edmondson J,
Axel R
(1996)
Visualizing an olfactory sensory map.
Cell
87:675-686[Web of Science][Medline].
-
Mori K
(1995)
Relation of chemical structure to specificity of response in olfactory glomeruli.
Curr Opin Neurobiol
5:467-474[Web of Science][Medline].
-
Mori K,
Mataga N,
Imamura K
(1992)
Differential specificities of single mitral cells in rabbit olfactory bulb for a homologous series of fatty acid odor molecules.
J Neurophysiol
67:786-789[Abstract/Free Full Text].
-
Pomeroy SL,
LaMantia AS,
Purves D
(1990)
Postnatal construction of neural circuitry in the mouse olfactory bulb.
J Neurosci
10:1952-1966[Abstract].
-
Ressler KJ,
Sullivan SL,
Buck LB
(1993)
A zonal organization of odorant receptor gene expression in the olfactory epithelium.
Cell
73:597-609[Web of Science][Medline].
-
Ressler KJ,
Sullivan SL,
Buck LB
(1994)
Information coding in the olfactory system: evidence for a stereotyped and highly organized epitope map in the olfactory bulb.
Cell
79:1245-1255[Web of Science][Medline].
-
Restrepo D,
Miyamoto T,
Bryant BP,
Teeter JH
(1990)
Odor stimuli trigger influx of calcium into olfactory neurons of the channel catfish.
Science
249:1166-1168[Abstract/Free Full Text].
-
Restrepo D,
Okada Y,
Teeter JH
(1993)
Odorant-regulated Ca2+ gradients in rat olfactory neurons.
J Gen Physiol
102:907-924[Abstract/Free Full Text].
-
Revial MF,
Duchamp A,
Holley A
(1978)
Odour discrimination by frog olfactory receptors: a second study.
Chem Senses
3:7-21[Abstract/Free Full Text].
-
Revial MF,
Sicard G,
Duchamp A,
Holley A
(1982)
New studies on odour discrimination in the frog's olfactory receptor cells. I. Experimental results.
Chem Senses
7:175-190[Abstract/Free Full Text].
-
Royet JP,
Souchier C,
Jourdan F,
Ploye H
(1988)
Morphometric study of the glomerular population in the mouse olfactory bulb: numerical density and size distribution along the rostrocaudal axis.
J Comp Neurol
270:559-568[Web of Science][Medline].
-
Sato T,
Hirono J,
Tonoike M,
Takebayashi M
(1994)
Tuning specificities to aliphatic odorants in mouse olfactory receptor neurons and their local distribution.
J Neurophysiol
72:2980-2989[Abstract/Free Full Text].
-
Schoenfeld TA,
Marchand JE,
Macrides F
(1985)
Topographic organization of tufted cell axonal projections in the hamster main olfactory bulb: an intrabulbar association system.
J Comp Neurol
235:503-518[Web of Science][Medline].
-
Schoenfeld TA,
Clancy AN,
Forbes WB,
Macrides F
(1994)
The spatial organization of the peripheral olfactory system of the hamster: part I, Receptor neuron projections of the main olfactory bulb.
Brain Res Bull
34:183-210[Web of Science][Medline].
-
Scott JW,
Davis LM,
Shannon D,
Kaplan C
(1996)
Relation of chemical structure to spatial distribution of sensory responses in rat olfactory epithelium.
J Neurophysiol
75:2036-2049[Abstract/Free Full Text].
-
Scott JW,
Shannon DE,
Charpentier J,
Davis LM,
Kaplan C
(1997)
Spatially organized response zones in rat olfactory epithelium.
J Neurophysiol
77:1950-1962[Abstract/Free Full Text].
-
Shepherd GM
(1994)
Discrimination of molecular signals by the olfactory receptor neuron.
Neuron
13:771-790[Web of Science][Medline].
-
Sicard G
(1986)
Electrophysiological recordings from olfactory receptor cells in adult mice.
Brain Res
397:405-408[Web of Science][Medline].
-
Sklar PB,
Anholt RRH,
Snyder SH
(1986)
The odorant-sensitive adenylate cyclase of olfactory receptor cells.
J Biol Chem
261:15538-15543[Abstract/Free Full Text].
-
Stewart WB,
Pedersen PE
(1987)
The spatial organization of olfactory nerve projections.
Brain Res
411:248-258[Web of Science][Medline].
-
Stewart WB,
Kauer JS,
Shepherd GM
(1979)
Functional organization of rat olfactory bulb analyzed by the 2-deoxyglucose method.
J Comp Neurol
185:715-734[Web of Science][Medline].
-
Sullivan SL,
Bohm S,
Ressler KJ,
Horowitz LF,
Buck LB
(1995)
Target-independent pattern specification in the olfactory epithelium.
Neuron
15:779-789[Web of Science][Medline].
-
Tareilus E,
Noe J,
Breer H
(1995)
Calcium signals in olfactory neurons.
Biochim Biophys Acta
1269:129-138[Medline].
-
Tsau Y,
Wenner P,
O'Donovan MJ,
Cohen LB,
Loew LM,
Wuskell JP
(1996)
Dye screening and signal-to-noise ratio for retrogradely transported voltage-sensitive dyes.
J Neurosci Methods
70:121-129[Web of Science][Medline].
-
Vassar R,
Ngai J,
Axel R
(1993)
Spatial segregation of odorant receptor expression in the mammalian olfactory epithelium.
Cell
74:309-318[Web of Science][Medline].
-
Vassar R,
Chao SK,
Sitcheran R,
Nuñez JM,
Vosshall LB,
Axel R
(1994)
Topographic organization of sensory projections to the olfactory bulb.
Cell
79:981-991[Web of Science][Medline].
-
Yokoi M,
Mori K,
Nakanishi S
(1995)
Refinement of odor molecule tuning by dendrodendritic synaptic inhibition in the olfactory bulb.
Proc Natl Acad Sci USA
92:3371-3375[Abstract/Free Full Text].
-
Youngentob SL,
Kent PF,
Sheehe PR,
Schwob JE,
Tzoumaka E
(1995)
Mucosal inherent activity patterns in the rat: evidence from voltage-sensitive dyes.
J Neurophysiol
73:150-160.
-
Zhao H,
Ivic L,
Otaki JM,
Hashimoto M,
Mikoshiba K,
Firestein S
(1998)
Functional expression of a mammalian odorant receptor.
Science
279:237-242[Abstract/Free Full Text].
Copyright © 1998 Society for Neuroscience 0270-6474/98/18124560-10$05.00/0
This article has been cited by other articles:
|
|
|
|
 
S. P. Saidu, S.D. Weeraratne, M. Valentine, R. Delay, and J. L. Van Houten
Role of Plasma Membrane Calcium ATPases in Calcium Clearance from Olfactory Sensory Neurons
Chem Senses,
May 1, 2009;
34(4):
349 - 358.
[Abstract]
[Full Text]
[PDF]
|
|
|
|
|
|
|
M. Maritan, G. Monaco, I. Zamparo, M. Zaccolo, T. Pozzan, and C. Lodovichi
Odorant receptors at the growth cone are coupled to localized cAMP and Ca2+ increases
PNAS,
March 3, 2009;
106(9):
3537 - 3542.
[Abstract]
[Full Text]
[PDF]
|
|
|
|
|
|
|
S. D. Weeraratne, M. Valentine, M. Cusick, R. Delay, and J. L. Van Houten
Plasma Membrane Calcium Pumps in Mouse Olfactory Sensory Neurons
Chem Senses,
October 1, 2006;
31(8):
725 - 730.
[Abstract]
[Full Text]
[PDF]
|
|
|
|
|
|
|
T. A. Schoenfeld and T. A. Cleland
Anatomical Contributions to Odorant Sampling and Representation in Rodents: Zoning in on Sniffing Behavior
Chem Senses,
February 1, 2006;
31(2):
131 - 144.
[Abstract]
[Full Text]
[PDF]
|
|
|
|
|
|
|
J. Li, J. A. Mack, M. Souren, E. Yaksi, S.-i. Higashijima, M. Mione, J. R. Fetcho, and R. W. Friedrich
Early Development of Functional Spatial Maps in the Zebrafish Olfactory Bulb
J. Neurosci.,
June 15, 2005;
25(24):
5784 - 5795.
[Abstract]
[Full Text]
[PDF]
|
|
|
|
|
|
|
G. Gomez, F. W. Lischka, M. E. Haskins, and N. E. Rawson
Evidence for Multiple Calcium Response Mechanisms in Mammalian Olfactory Receptor Neurons
Chem Senses,
May 1, 2005;
30(4):
317 - 326.
[Abstract]
[Full Text]
[PDF]
|
|
|
|
|
|
|
P.-M. Lledo, G. Gheusi, and J.-D. Vincent
Information Processing in the Mammalian Olfactory System
Physiol Rev,
January 1, 2005;
85(1):
281 - 317.
[Abstract]
[Full Text]
[PDF]
|
|
|
|
|
|
|
T. A. Christensen
Making Scents Out of Spatial and Temporal Codes in Specialist and Generalist Olfactory Networks
Chem Senses,
January 1, 2005;
30(suppl_1):
i283 - i284.
[Full Text]
[PDF]
|
|
|
|
|
|
|
S. E. Hall, W. B. Floriano, N. Vaidehi, and W. A. Goddard III
Predicted 3-D Structures for Mouse I7 and Rat I7 Olfactory Receptors and Comparison of Predicted Odor Recognition Profiles with Experiment
Chem Senses,
September 1, 2004;
29(7):
595 - 616.
[Abstract]
[Full Text]
[PDF]
|
|
|
|
|
|
|
W. B. Floriano, N. Vaidehi, and W. A. Goddard III
Making Sense of Olfaction through Predictions of the 3-D Structure and Function of Olfactory Receptors
Chem Senses,
May 1, 2004;
29(4):
269 - 290.
[Abstract]
[Full Text]
[PDF]
|
|
|
|
|
|
|
R. C. Araneda, Z. Peterlin, X. Zhang, A. Chesler, and S. Firestein
A pharmacological profile of the aldehyde receptor repertoire in rat olfactory epithelium
J. Physiol.,
March 15, 2004;
555(3):
743 - 756.
[Abstract]
[Full Text]
[PDF]
|
|
|
|
|
|
|
P. A. Godfrey, B. Malnic, and L. B. Buck
The mouse olfactory receptor gene family
PNAS,
February 17, 2004;
101(7):
2156 - 2161.
[Abstract]
[Full Text]
[PDF]
|
|
|
|
|
|
|
L. Lagostena and A. Menini
Whole-cell Recordings and Photolysis of Caged Compounds in Olfactory Sensory Neurons Isolated from the Mouse
Chem Senses,
October 1, 2003;
28(8):
705 - 716.
[Abstract]
[Full Text]
[PDF]
|
|
|
|
|
|
|
F. Xu, N. Liu, I. Kida, D. L. Rothman, F. Hyder, and G. M. Shepherd
Odor maps of aldehydes and esters revealed by functional MRI in the glomerular layer of the mouse olfactory bulb
PNAS,
September 16, 2003;
100(19):
11029 - 11034.
[Abstract]
[Full Text]
[PDF]
|
|
|
|
|
|
|
M. L. Fletcher and D. A. Wilson
Olfactory Bulb Mitral-Tufted Cell Plasticity: Odorant-Specific Tuning Reflects Previous Odorant Exposure
J. Neurosci.,
July 30, 2003;
23(17):
6946 - 6955.
[Abstract]
[Full Text]
[PDF]
|
|
|
|
|
|
|
H. Hamana, J. Hirono, M. Kizumi, and T. Sato
Sensitivity-dependent Hierarchical Receptor Codes for Odors
Chem Senses,
February 1, 2003;
28(2):
87 - 104.
[Abstract]
[Full Text]
[PDF]
|
|
|
|
|
|
|
N. H. Kulkarni, A. H. Yamamoto, K. O. Robinson, T. F. C. Mackay, and R. R. H. Anholt
The DSC1 Channel, Encoded by the smi60E Locus, Contributes to Odor-Guided Behavior in Drosophila melanogaster
Genetics,
August 1, 2002;
161(4):
1507 - 1516.
[Abstract]
[Full Text]
[PDF]
|
|
|
|
|
|
|
T. Bozza, P. Feinstein, C. Zheng, and P. Mombaerts
Odorant Receptor Expression Defines Functional Units in the Mouse Olfactory System
J. Neurosci.,
April 15, 2002;
22(8):
3033 - 3043.
[Abstract]
[Full Text]
[PDF]
|
|
|
|
|
|
|
A. R. Goriely, T. W. Secomb, and L. P. Tolbert
Effect of the Glial Envelope on Extracellular K+ Diffusion in Olfactory Glomeruli
J Neurophysiol,
April 1, 2002;
87(4):
1712 - 1722.
[Abstract]
[Full Text]
[PDF]
|
|
|
|
|
|
|
T. K. Alkasab, J. White, and J. S. Kauer
A Computational System for Simulating and Analyzing Arrays of Biological and Artificial Chemical Sensors
Chem Senses,
March 1, 2002;
27(3):
261 - 275.
[Abstract]
[Full Text]
[PDF]
|
|
|
|
|
|
|
H. U. Fried, S. H. Fuss, and S. I. Korsching
Selective imaging of presynaptic activity in the mouse olfactory bulb shows concentration and structure dependence of odor responses in identified glomeruli
PNAS,
February 14, 2002;
(2002)
52658399.
[Abstract]
[Full Text]
[PDF]
|
|
|
|
|
|
|
A. A. Nikonov and J. Caprio
Electrophysiological Evidence for a Chemotopy of Biologically Relevant Odors in the Olfactory Bulb of the Channel Catfish
J Neurophysiol,
October 1, 2001;
86(4):
1869 - 1876.
[Abstract]
[Full Text]
[PDF]
|
|
|
|
|
|
|
J. Reisert and H. R Matthews
Simultaneous recording of receptor current and intraciliary Ca2+ concentration in salamander olfactory receptor cells
J. Physiol.,
September 15, 2001;
535(3):
637 - 645.
[Abstract]
[Full Text]
[PDF]
|
|
|
|
|
|
|
D. A. Wilson
Receptive Fields in the Rat Piriform Cortex
Chem Senses,
June 1, 2001;
26(5):
577 - 584.
[Abstract]
[Full Text]
[PDF]
|
|
|
|
|
|
|
L. Belluscio and L. C. Katz
Symmetry, Stereotypy, and Topography of Odorant Representations in Mouse Olfactory Bulbs
J. Neurosci.,
March 15, 2001;
21(6):
2113 - 2122.
[Abstract]
[Full Text]
[PDF]
|
|
|
|
|
|
|
M. Mezler, J. Fleischer, and H. Breer
Characteristic features and ligand specificity of the two olfactory receptor classes from Xenopus laevis
J. Exp. Biol.,
January 9, 2001;
204(17):
2987 - 2997.
[Abstract]
[Full Text]
[PDF]
|
|
|
|
|
|
|
D. A. Wilson
Comparison of Odor Receptive Field Plasticity in the Rat Olfactory Bulb and Anterior Piriform Cortex
J Neurophysiol,
December 1, 2000;
84(6):
3036 - 3042.
[Abstract]
[Full Text]
[PDF]
|
|
|
|
|
|
|
Q. Yuan, C. W. Harley, J. C. Bruce, A. Darby-King, and J. H. McLean
Isoproterenol Increases CREB Phosphorylation and Olfactory Nerve-Evoked Potentials in Normal and 5-HT-Depleted Olfactory Bulbs in Rat Pups Only at Doses That Produce Odor Preference Learning
Learn. Mem.,
November 1, 2000;
7(6):
413 - 421.
[Abstract]
[Full Text]
|
|
|
|
|
|
|
M. Ma and G. M. Shepherd
Functional mosaic organization of mouse olfactory receptor neurons
PNAS,
October 23, 2000;
(2000)
220301797.
[Abstract]
[Full Text]
|
|
|
|
|
|
|
D. A. Fadool, K. Tucker, J. J. Phillips, and J. A. Simmen
Brain Insulin Receptor Causes Activity-Dependent Current Suppression in the Olfactory Bulb Through Multiple Phosphorylation of Kv1.3
J Neurophysiol,
April 1, 2000;
83(4):
2332 - 2348.
[Abstract]
[Full Text]
[PDF]
|
|
|
|
|
|
|
R. M. Costanzo
Rewiring the Olfactory Bulb: Changes in Odor Maps following Recovery from Nerve Transection
Chem Senses,
April 1, 2000;
25(2):
199 - 205.
[Abstract]
[Full Text]
[PDF]
|
|
|
|
|
|
|
F. Zufall, T. Leinders-Zufall, and C. A. Greer
Amplification of Odor-Induced Ca2+ Transients by Store-Operated Ca2+ Release and Its Role in Olfactory Signal Transduction
J Neurophysiol,
January 1, 2000;
83(1):
501 - 512.
[Abstract]
[Full Text]
[PDF]
|
|
|
|
|
|
|
J. W. Scott and T. Brierley
A Functional Map in Rat Olfactory Epithelium
Chem Senses,
December 1, 1999;
24(6):
679 - 690.
[Abstract]
[Full Text]
[PDF]
|
|
|
|
|
|
|
J. H. McLean, C. W. Harley, A. Darby-King, and Q. Yuan
pCREB in the Neonate Rat Olfactory Bulb Is Selectively and Transiently Increased by Odor Preference-Conditioned Training
Learn. Mem.,
November 1, 1999;
6(6):
608 - 618.
[Abstract]
[Full Text]
|
|
|
|
|
|
|
K. Mori, H. Nagao, and Y. Yoshihara
The Olfactory Bulb: Coding and Processing of Odor Molecule Information
Science,
October 22, 1999;
286(5440):
711 - 715.
[Abstract]
[Full Text]
|
|
|
|
|
|
|
G. Laurent
A Systems Perspective on Early Olfactory Coding
Science,
October 22, 1999;
286(5440):
723 - 728.
[Abstract]
[Full Text]
|
|
|
|
|
|
|
J. R. Murrell and D. D. Hunter
An Olfactory Sensory Neuron Line, Odora, Properly Targets Olfactory Proteins and Responds to Odorants
J. Neurosci.,
October 1, 1999;
19(19):
8260 - 8270.
[Abstract]
[Full Text]
[PDF]
|
|
|
|
|
|
|
P. Duchamp-Viret, M. A. Chaput, and A. Duchamp
Odor Response Properties of Rat Olfactory Receptor Neurons
Science,
June 25, 1999;
284(5423):
2171 - 2174.
[Abstract]
[Full Text]
|
|
|
|
|
|
|
A Gelperin
Oscillatory dynamics and information processing in olfactory systems
J. Exp. Biol.,
January 7, 1999;
202(14):
1855 - 1864.
[Abstract]
[PDF]
|
|
|
|
|
|
|
F. Zufall and T. Leinders-Zufall
Calcium Imaging in the Olfactory System: New Tools for Visualizing Odor Recognition
Neuroscientist,
January 1, 1999;
5(1):
4 - 7.
[Abstract]
[PDF]
|
|
|
|
|
|
|
R. W. Friedrich and S. I. Korsching
Chemotopic, Combinatorial, and Noncombinatorial Odorant Representations in the Olfactory Bulb Revealed Using a Voltage-Sensitive Axon Tracer
J. Neurosci.,
December 1, 1998;
18(23):
9977 - 9988.
[Abstract]
[Full Text]
[PDF]
|
|
|
|
|
|
|
Y. Cao, B. C. Oh, and L. Stryer
Cloning and localization of two multigene receptor families in goldfish olfactory epithelium
PNAS,
September 29, 1998;
95(20):
11987 - 11992.
[Abstract]
[Full Text]
[PDF]
|
|
|
|
|
|
|
H. U. Fried, S. H. Fuss, and S. I. Korsching
Selective imaging of presynaptic activity in the mouse olfactory bulb shows concentration and structure dependence of odor responses in identified glomeruli
PNAS,
March 5, 2002;
99(5):
3222 - 3227.
[Abstract]
[Full Text]
[PDF]
|
|
|
|
|
|
|
M. Ma and G. M. Shepherd
Functional mosaic organization of mouse olfactory receptor neurons
PNAS,
November 7, 2000;
97(23):
12869 - 12874.
[Abstract]
[Full Text]
[PDF]
|
|
|
|
Top
Abstract
Introduction
