Department of Psychological and Brain Sciences, Dartmouth College, Hanover, New Hampshire 03755
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ABSTRACT |
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Stackman, Robert W., Matthew L. Tullman, and Jeffrey S. Taube. Maintenance of Rat Head Direction Cell Firing During Locomotion in the Vertical Plane. J. Neurophysiol. 83: 393-405, 2000. Previous studies have identified a subset of neurons in the rat anterodorsal thalamus (ADN) that encode head direction (HD) in absolute space and may be involved in navigation. These HD cells discharge selectively when the rat points its head in a specific direction (the preferred firing direction) in the horizontal plane. HD cells are typically recorded during free movement about a single horizontal surface. The current experiment examined how HD cell firing was influenced by 1) locomotion in the vertical plane and 2) locomotion on two different horizontal surfaces separated in height. Rats were trained in a cylindrical enclosure containing a single polarizing cue card attached to the cylinder wall, covering ~100° of arc. The enclosure contained two horizontal surfaces: the cylinder floor and an annulus around the cylinder top 76 cm above the floor. A 90° vertical mesh ladder that could be affixed at any angular position on the cylinder wall allowed the rats to locomote back and forth between the two horizontal surfaces. Rats were trained to retrieve food pellets on the cylinder floor as well as climb the mesh ladder to retrieve food pellets on the annulus. HD cell activity was monitored as the rat traversed the horizontal and vertical surfaces of the apparatus. When the angular position of the mesh corresponded to the cell's preferred firing direction, the HD cells maintained their peak discharge rate as the rat climbed up the mesh, but did not fire when the rat climbed down the mesh. In contrast, when the mesh was positioned 180° opposite the preferred firing direction, HD cells did not fire when the rat climbed up the mesh, but exhibited maximal firing when the rat climbed down the mesh. When the mesh was placed 90 or 270° from the preferred firing direction, HD cells exhibited background firing rates during climbing up or down the mesh. While preferred firing directions were maintained between the two horizontal surfaces, peak firing rate increased significantly (~30%) on the annulus as compared with the cylinder floor. These data demonstrate that HD cells continue to discharge in the vertical plane if the vertical locomotion began with the rat's orientation corresponding to the preferred firing direction. One model consistent with these data are that HD cells define the horizontal reference frame as the animal's plane of locomotion. Further, we propose that HD cell firing, as viewed within a three-dimensional coordinate system, can be characterized as the surface of a hemitorus.
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INTRODUCTION |
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Accurate spatial orientation and navigation
require the organism to estimate its current location and directional
heading in three-dimensional space. This function is likely influenced by at least two distinct neural systems. Navigation is thought to
involve the continual monitoring of internal, or idiothetic, cues
(e.g., consequences of the animal's movements: vestibular, proprioceptive, motor efference copy or motor corollary discharge, and
optic/auditory flow), together with occasional reference to familiar
external sensory cues or landmarks to correct for error accumulation
over time. For example, path integration (or movement through a novel
environment) involves an initial referral to a landmark, to define
one's initial orientation, and then navigating solely by internal cues
(Barlow 1964; Berthoz et al. 1995
;
Gallistel 1990
). Accurate path integration is thought to
require the monitoring of vestibular signals (i.e., angular and linear
acceleration cues) (Etienne et al. 1988
;
Mittelstaedt and Glasauer 1991
; Mittelstaedt and
Mittelstaedt 1980
), as lesions of the vestibular system impair spatial navigation (Matthews et al. 1989
; Miller
et al. 1983
; Ossenkopp and Hargreaves 1993
;
Potegal et al. 1977
).
Allocentric representations of spatial location and direction are
thought to be processed by an integrated multimodal neural circuit
within the limbic system (McNaughton et al. 1995). This notion is supported by evidence of two neurophysiological correlates of
allocentric space found in limbic brain regions of the freely moving
rat. Principal neurons of hippocampus discharge in accordance with the
spatial location of the rat independent of directional heading
(Muller et al. 1987
; O'Keefe and
Dostrovsky 1971
), while head direction (HD) cells discharge as
a function of the rat's head direction in the horizontal plane,
independent of current location (Taube et al. 1990a
; for
a review see Taube 1998
). The head orientation at which
a HD cell exhibits maximal firing is called the preferred firing
direction. The activity of the entire HD cell population is believed to
represent the moment-to-moment directional heading of the animal. HD
cells have been recorded in several regions of the rat brain including
postsubiculum (PoS) (Taube et al. 1990a
), anterodorsal
thalamic nuclei (ADN) (Taube 1995
), lateral dorsal
thalamic nuclei (Mizumori and Williams 1993
), retrosplenial and medial prestriate cortex (Chen et al.
1994
), striatum (Mizumori and Cooper 1995
;
Wiener 1993
), and lateral mammillary nuclei
(Stackman and Taube 1998
).
Converging evidence indicates that manipulation of environmental or
idiothetic cues influences the preferred firing directions of HD cells
(Blair and Sharp 1996; Goodridge and Taube
1995
; Knierim et al. 1995
; Taube and
Burton 1995
; Taube et al. 1990b
; for a review
see Taube et al. 1996
). These studies indicate that
familiar landmark cues exert preferential control over HD cell
activity. Taube and Burton (1995)
demonstrated that the
preferred firing directions of HD cells remain stable when the rat
self-locomotes into a novel environment, suggesting that idiothetic
cues maintain the preferred firing directions in the absence of
familiar external cues. The integration of angular
velocity information from the vestibular system may be necessary for
maintaining central representations of directional heading
(Blair and Sharp 1996
; McNaughton et al. 1995
), under circumstances in which familiar external landmark cues are not available. However, lesions of the vestibular apparatus disrupt HD cell firing in the ADN (Stackman and Taube
1997
), regardless of the presence of the familiar landmark cue.
Empirical data also suggest that motor efference copy signals influence
place cell and HD cell activity (Foster et al.
1989
; Taube 1995
; Taube et al.
1996
). Together, these data suggest that internal cues play an
essential role in the generation and maintenance of central representations of spatial orientation.
Currently, all empirical studies of HD cells have been conducted while
the rat freely moved about the horizontal surface of enclosures and
mazes. However, in the natural setting, efficient rodent navigation
within the rat burrow requires the integration of movements in both the
horizontal and vertical planes (Calhoun 1963). Therefore
the neural mechanisms responsible for processing representations of
allocentric spatial orientation must be able to maintain accuracy
whether the organism is moving in the horizontal or vertical planes.
Interestingly, Moghaddam and colleagues (1996)
demonstrated that rats in a featureless, enclosed arena could accurately navigate to a goal location on the arena floor using the
inclination of the floor surface as a primary cue. Although qualitative
assessments have determined that moderate head pitch (head motion in
the rostral-caudal axis) or roll (head rotation along the
anterior-posterior axis) do not disrupt HD cell firing in the
horizontal plane (Stackman and Taube 1998
; Taube
et al. 1990a
), it is important to determine how the
horizontal reference frame is defined by the HD cell, and whether HD
cell firing is maintained when the rat is oriented vertically.
Therefore we examined the discharge properties of HD cells during locomotion in the vertical plane and in different horizontal planes within the same environment. Rats were trained to climb a 90° vertical mesh ladder from the floor of a cylindrical apparatus to a narrow annulus above the cylinder floor. We monitored ADN and PoS HD cell activity during locomotion in the vertical plane as the rat climbed up and down the ladder and as the rats moved freely about both horizontal surfaces. Each HD cell was recorded as the rat locomoted in the vertical plane, with the mesh ladder centered at specific positions relative to the cell's preferred firing direction. This protocol enabled an assessment of HD cell activity as the rat left one horizontal plane oriented in accordance with the preferred firing direction of the recorded cell or at the three cardinal directions orthogonal to the preferred firing direction.
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METHODS |
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Subjects and vertical plane locomotion training
Subjects were five female Long-Evans rats, weighing 250-300 g at the beginning of the experiment. Rats were maintained on a food-restricted diet (15-20 g/day) and housed separately in suspended wire mesh cages. Tap water was available ad libitum. All training, unit screening, and recording occurred during sessions in which the rats foraged for food pellets in a cylindrical apparatus (74 cm high, 76 cm diameter; see Fig. 1A). Mounted to the top of the cylinder was a 12.7-cm-wide rim, or annulus with 4 food wells equally spaced apart on the annulus surface. The annulus did not contain inside or outside walls, but was wide enough to enable rats to change their direction of movement once on it. The cylinder and annulus were constructed of wood and painted gray. Access to the annulus was provided by a floor-to-annulus vertical wire mesh "ladder" (74 cm high, 29 cm wide) placed onto the inside cylinder wall. The mesh ladder was cut from galvanized steel mesh (1.27 cm by 1.27 cm grid) and formed to fit tightly against the wall, thus allowing a 90° vertical transit up the cylinder wall. The angular position of the mesh was not fixed and could be placed anywhere on the inside surface of the cylinder wall. Finally, a white sheet of cardboard attached to the inside wall of the cylinder, and which extended from the floor to the top of the cylinder, served as the only polarizing, directional visual landmark. This cue card was positioned at 3 o'clock (see Fig. 1), and occupied ~100° of arc. Black floor-to-ceiling curtains formed a featureless circular enclosure (2 m diameter) surrounding the cylinder. Room illumination was provided by four uniformly arranged overhead DC lamps. A color video camera (Sony XC-711) was centered above the cylinder 3 m from the floor surface. The cylinder was placed on a sheet of gray photographic backdrop paper, which was replaced after each recording session. A stool was placed inside the curtain enclosure, which the experimenter used while connecting and disconnecting the rat from the recording cable. The stool was out of the rat's view while in the cylinder, but could be viewed by the rat while moving about on the annulus. The position of the stool within the curtained enclosure varied, as an attempt was made to vary the position at which the rat was released into the cylinder during each recording session.
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Initially, rats received habituation trials in the cylindrical apparatus without the mesh or annulus present. Rats were brought from the colony room into the curtained enclosure in a corrugated cardboard box (~30 × 30 cm box); no attempt was made to disorient the rats before their release into the cylinder. Rats were placed into the cylinder in pairs and received at least five habituation trials (1 trial/day) during which food pellets (20 mg, PJ Noyes, Lancaster, NH) were thrown randomly into the cylinder from outside the black curtain. By the completion of training, rats engaged in nearly continuous food pellet search behavior over the entire floor of the cylinder.
After the habituation trials, rats were trained by successive approximation to climb the mesh, from the cylinder floor to the cylinder annulus, to retrieve food pellets from the annulus cups. During training, the white cue card was always present, and its position was always fixed at the 3 o'clock position. The angular position of the mesh was changed every trial to explicitly reduce the potential association of the mesh with a spatial location and its use as an orientation cue. The mesh ladder required the rat to navigate in a vertical plane that was 90° orthogonal to the cylinder floor and annulus. During the initial training trials (1 trial/day), a circular wooden platform (64 cm high) was placed on the cylinder floor. With the platform at this height, the rats were only required to climb 10 cm to access the annulus food cups. Several food pellets were placed into the food cups of the annulus and scattered on the platform surface. The rats were placed on the platform in pairs and allowed to move about freely over the entire surface of the apparatus for 15 min. Generally, after two trials, rats habituated to the apparatus and demonstrated free movement back and forth between the platform floor and annulus. Over successive trials, the floor platform was lowered to encourage the rats to climb to greater heights to access the annulus. The platform was first lowered to 54 cm above the floor, then 25 cm above the floor, after which the platform was removed and the rats were trained to climb the full length of the mesh (76 cm) to access the annulus. During the final training trials, the annulus food cups were not rebaited until the rats returned to the cylinder floor. The criterion for training completion was that all rats readily engaged in climbing up and down the complete length of the mesh, and consumed food pellets from the annulus food cups. On completion of training, rats were implanted with driveable 10-wire microelectrode arrays directed at the ADN or PoS.
Electrode implantation
Electrode construction and implantation techniques used were
similar to that described previously (Taube 1995).
Briefly, each electrode array consisted of a bundle of 10, 25-µm-diam
nichrome wires (California Fine Wire, Grover City, CA) insulated except at the tips. The wire bundle was passed through a 26-gauge stainless steel cannula and each wire attached to a modified 11-pin Augat connector. The electrode array could be advanced in the dorsoventral plane with three screws attached to the electrode's acrylic base (Kubie 1984
). On habituation to the cylindrical
apparatus and adequate foraging behavior, each rat was anesthetized
with a ketamine-xylazine mixture (2 ml/kg im) and stereotaxically
implanted with an electrode array directed at either the PoS or ADN.
Electrode coordinates, with respect to bregma, were as follows: PoS:
anterior/posterior
6.6 mm, medial/lateral +2.8 mm, ventral 1.6 mm
from the cortical surface; ADN: anterior/posterior
1.4 mm,
medial/lateral +1.3 mm, ventral 3.7 mm from the cortical surface
(Paxinos and Watson 1998
). Jewelers screws, placed in
the skull plates over the cerebellar cortex, parietal cortex, and
frontal cortex, and dental cement anchored the electrode assembly in
place. All procedures were conducted according to an institutionally
approved animal care protocol. All surgical procedures were conducted
under sterile conditions, and the rats were allowed a 1-wk
postoperative recovery interval before we commenced single-unit screening.
Isolation and recording of head direction cell activity
After recovery from surgery, activity from each microelectrode
wire was assessed during daily unit screening sessions while the rat
foraged for food pellets in the cylinder. During screening sessions,
the mesh and annulus were not present. The electrode wires were
advanced over several weeks while screening for HD cell waveforms that
were of acceptable amplitude for isolation from background electrical
noise. Each rat was transported into the screening area from the animal
colony room in the corrugated cardboard enclosure; again, no attempt
was made to disorient the rats before their release into the cylinder.
The cardboard box was placed on the floor inside the curtained
enclosure next to the cylinder. A recording cable was attached to the
implanted electrode while the rat was held gently in a towel. The rat
was then released into the cylinder apparatus from a start position that varied daily in a pseudorandom manner. Unit activity was analyzed
using procedures similar to those previously described (Taube
1995; Taube et al. 1990a
). Briefly, electrical
signals were passed through a field-effect transistor (FET) (1 FET/electrode) in a source-follower configuration, through an overhead
commutator (Biela Idea Development, Anaheim, CA), amplified (Grass
Instruments P5 Series, West Warwick, RI), band-passed filtered
(300-10,000 Hz, 3 dB/octave; Peavey Electronics PME8, Meridian, MS),
and passed through a series of window discriminators (BAK Electronics
Model DDIS-1, Germantown, MD). The resultant signal was then displayed on an oscilloscope (Tektronix Model 2214, Beaverton, OR). Electrode activity was monitored while observing the rat's behavior on a video
monitor with a camera mounted 3 m above the cylinder floor. If HD
cell activity was not found, the electrodes were advanced 25-50 µm
further into the ADN or PoS, and the activity was monitored again the
next day. Screening for HD cells occurred over the course of several weeks.
When the waveform of a single cell could be sufficiently isolated from
background electrical noise, two light-emitting diodes (LEDs) were
added to the recording cable. The LED arrangement used in the present
study was a modification of that used previously (Taube
1990a). For the present study, the headstage included a red LED
positioned just posterior to the headstage, and a green LED was
positioned over the rat's back. The LEDs were spaced 5.5 cm apart
along the rostral/caudal axis of the rat. The
x,y-coordinates of the LEDs were determined at 60 Hz by a
video-tracking system (Ebtronics, Brooklyn, NY). A wide-angle video
lens (8.5 mm) was used to monitor the rat's movements in the cylinder
and on the annulus of the apparatus. During recording sessions the LED
coordinates and neuronal discharges were sampled at 60 Hz and acquired
by a data acquisition interface board (National Instruments DIO-96, Austin, TX) in a personal computer (Apple Macintosh Quadra 840AV, Cupertino, CA). Data were stored for subsequent off-line analyses using
programs written with LabView software (National Instruments, Austin,
TX). For the present data, the rat's head position was defined as the
location of the red LED. During recording sessions, white noise was
broadcast through a ceiling speaker centered above the cylinder to mask
uncontrolled auditory cues.
Vertical plane locomotion/recording protocol
The protocol used to record each HD cell during vertical plane locomotion comprised at least five 8-min recording sessions and is illustrated in Fig. 1B. Throughout all recording sessions, the cue card remained in the cylinder at the 3 o'clock position (standard position). A second charge-coupled device (CCD) color video camera was attached to a tripod positioned inside the curtained enclosure and oriented to receive the video image of the 90° vertical mesh ladder from floor-to-annulus (i.e., orthogonal to the overhead ceiling camera). An experimenter-controlled video source switching box (Electronics Shop, Dartmouth College, Hanover, NH) enabled the video signal that was fed to the video-tracking system to be manually switched between the overhead camera and the tripod-mounted camera. Generally, horizontal movements of the rat on the cylinder floor and annulus were acquired by the overhead camera, while vertical movements on the mesh (i.e., climbing up and climbing down) were acquired by the second camera. The video output from the selected camera, along with the cell's activity, was simultaneously recorded during each recording session. Each HD cell was recorded during an initial 8-min session as the rat foraged for food pellets thrown randomly into the cylinder. The mesh was not present during this initial recording session. On completion of the initial recording session, the rat was removed from the cylinder and placed into the cardboard enclosure. The HD cell's preferred firing direction was determined, and the mesh was placed into the cylinder centered at the position corresponding to the cell's preferred firing direction. This recording session will be referred to as the Mesh 0° session (i.e., the mesh ladder positioned 0° from the cell's preferred firing direction). Several food pellets were placed into each of the four food cups on the annulus. No attempt was made to disorient the rat before its return to the cylinder. The rat was returned to the floor of the cylinder, and an 8-min recording session begun. During this and following sessions, on retrieving all food pellets, the annulus food cups were not rebaited until the rat returned to the cylinder floor. This procedure facilitated the collection of HD cell activity during vertical plane locomotion, by increasing the occurrences of climbing behavior. On completion of the Mesh 0° session, the rat was removed to the cardboard enclosure. The mesh position was shifted to 180° opposite the cell's preferred firing direction, and the tripod-mounted camera was positioned accordingly. This session was designated as Mesh 180°. The rat was returned to the cylinder, and a third recording session begun. The fourth and fifth recording sessions followed, during which the mesh was positioned at 90 and 270° away from the cell's preferred firing direction. These recording sessions are referred to as Mesh 90° and Mesh 270°, respectively. In general, the recording protocol provided at least four or five episodes of climbing up and down the mesh at each of four cardinal positions relative to the HD cell's preferred firing direction.
Data analysis
Head direction of the rat was determined from the relative
positions of the two LEDs using procedures defined previously
(Taube 1995; Taube et al. 1990a
).
Head direction and unit data were sorted according to the video source
(overhead vs. tripod-mounted camera) into files of movement in the
vertical plane or horizontal plane. The vertical plane data were sorted
further according to whether the rat was climbing up, or climbing down
the mesh. The horizontal plane data were further sorted according to
whether the rat was on the cylinder floor or on the annulus. Consistent
with previous studies (Taube 1995
; Taube et al.
1990a
), each HD cell was analyzed to determine the basic firing
properties: 1) preferred firing direction,
2) peak firing rate, 3) directional
firing range, 4) directional information content, and
5) background firing rate. Information content per spike
represents a quantitative measure of the amount of information that is
conveyed by each spike generated by a cell (Skaggs et al.
1995
). In this case, the directional information
content can be thought of as a measure of the degree to which cell
firing can predict the animal's directional heading. The information
content for HD cells was calculated using the following formula
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To quantify any shift of the preferred firing direction during
locomotion on the two horizontal surfaces (annulus vs. cylinder floor),
we used a cross-correlation method described in detail previously
(Taube et al. 1990a). The firing rate versus HD tuning function of annulus horizontal plane data were shifted in 6°
increments and cross-correlated with the tuning function of the same
cell recorded during the initial standard session of the cylinder floor horizontal plane data. The degree of shift necessary to maximize the
correlation between the tuning functions of the two horizontal data
segments was considered the rotation of the preferred firing direction.
A positive shift in preferred firing direction reflects a
counterclockwise shift, whereas a negative shift reflects a clockwise shift.
All means are reported along with the standard error of the mean. At the conclusion of the analyses for a given HD cell, the electrode array was advanced further over the course of several months, and unit activity was screened for directional firing properties. Unit screening was terminated when the electrode array had been advanced ~2 mm.
Histology
At the conclusion of unit screening, rats were overdosed with pentobarbital sodium (100 mg/kg ip). Weak anodal current (15 µA for 10 s) was passed through one of the electrode wires to mark the wire location by the deposition of iron (Prussian blue reaction). The rats were perfused transcardially with 0.9% saline followed by 10% Formalin, and the brains were removed and placed into 10% Formalin for at least 48 h. The brains were then transferred to a 10% Formalin solution containing 2% potassium ferrocyanide for 24 h, returned to a 10% Formalin solution for 24 h, after which the brains were placed into a 20% sucrose solution for at least 48 h. The brains were then blocked, frozen on dry ice, sectioned coronally at 25 µm on a cryostat, and mounted onto microscope slides. The sections were stained with cresyl violet and examined under light microscopy to determine the location of recording sites. Inspection of the stained sections for each subject verified that the electrodes had passed through the PoS or the ADN.
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RESULTS |
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Thirteen HD cells were recorded from five rats (2 cells from the PoS; 11 cells from the ADN). Because the responses of PoS HD cells were markedly similar to those of ADN HD cells, the data from PoS and ADN HD cells were grouped together for analyses. Due to the short-duration of each climbing episode (1-4 s), it was necessary to combine data acquired from several episodes of climbing up and down the mesh to adequately judge the directional firing during vertical plane movement.
HD cell activity in the vertical plane
During the recording sessions, the rats engaged in several trips up and down the mesh. There were qualitative differences, however, in their behavior when climbing up the mesh as opposed to climbing down. During locomotion up the mesh, the rats followed a straight path and rarely stopped or turned around to abort an upward climb. Typically, the rats completed a climb from the cylinder floor to the annulus in ~2-3 s. In contrast, when climbing down the mesh, the rats appeared considerably more cautious, and their behavior was punctuated by several starts and stops. Behavioral observations revealed that all rats climbed down the mesh headfirst. However, the downward paths were frequently not straight, and usually cut a diagonal that often included a momentary stop in the first third of the mesh. In many cases, the rats began to climb down the mesh only to stop one-third of the way down, turn around, and then climb back up to the annulus. When rats went all the way to the floor, they usually climbed two-thirds of the way down the mesh and then jumped to the cylinder floor. Typically, the rats completed a climb from the annulus to the cylinder floor in ~3-5 s. In many cases, especially during the later sessions of the recording protocol, it was necessary to toss food pellets onto the cylinder floor to motivate the rats to climb down the mesh from the annulus.
The short latency of climbing episodes, and the fact that climbing behavior restricted the rat's head direction sampling to an arc of ~90°, compromised the ability to completely evaluate the directional discharge of HD cells over a 360° range in the vertical plane (e.g., when the rat climbed up the mesh oriented at a directional heading of 180°, the rat never oriented itself at a directional heading of 0°). Even after combining several episodes of climbing, there were still some HD bins that were not sampled. Therefore to construct firing rate plots of HD cell activity for movements in the vertical plane, it was necessary to assign head direction bins with no sampling to a firing rate of 0 spikes/s. Consequently, it was difficult to assess shifts in the preferred firing direction during climbing at each of the four angular mesh positions.
In summary, all 13 HD cells exhibited maximal firing as the rat climbed up with the mesh aligned at the Mesh 0° position, and when the rat climbed down with the mesh aligned at the Mesh 180° position. Conversely, the cells displayed minimal, if any, discharge when the rat climbed down the mesh at Mesh 0° position or climbed up the mesh at Mesh 180° position. Table 1 presents the peak firing rates of all cells exhibited during the cylinder floor session, the Mesh 0° session, the Mesh 180° session, and the annulus session. In general, there was a remarkable preservation of peak firing rates across conditions. As discussed above, due to the rat's stereotypic climbing behavior, the HD bin sampling rates were confined to within ±90° of the mesh during most climbing episodes. HD bin sampling rates during climbing episodes averaged ~30% of that observed during the standard cylinder floor session. Despite this decrease in sampling, peak firing rates during vertical locomotion (climbing up the Mesh 0°, climbing down the Mesh 180°) were not significantly different from those observed during horizontal locomotion (cylinder floor), F(2,24) = 0.17, n.s. Compared with firing rates during the cylinder floor session, the mean peak firing rates climbing up the mesh at the Mesh 0° position and down the mesh at the Mesh 180° position (expressed as a percent of the floor peak firing rate) were 104.4 ± 13.2% (mean ± SE) and 100.4 ± 10.9%, respectively. Therefore HD cell peak firing rates were not influenced by locomotion in the vertical plane when the rat's head was aligned with the cell's preferred firing direction when approaching the wall.
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Figure 2 provides two examples of typical HD cell responses from two different rats during an initial standard session in the cylinder, and during the four subsequent recording sessions in which the rat climbed up and down the mesh, with the mesh at each respective angular position. Figure 2, A and F, presents firing rate by HD tuning functions during 8-min sessions as the rats foraged for food pellets on the cylinder floor.
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MESH 0°.
Across the 13 HD cells recorded, rats made an average of 5 transits up
and 4 transits down the mesh, when it was positioned at Mesh 0°.
Figure 2, B and G, depicts typical responses of
HD cells during vertical plane locomotion when the center of the mesh
was positioned on the cylinder wall in alignment with the HD cell's
preferred firing direction (Mesh 0°). The preferred firing directions
of these two cells were 270 and 200°, respectively. In both cases,
directional discharge was maintained as the rat climbed up () the
mesh, but not when it climbed down (- - -). Moreover, cell
discharge was consistently present throughout the period when the rat
climbed up the mesh. In the cases where the rat climbed down the mesh,
the rat's approach path from the annulus to the mesh was one in which
the animal's directional heading was 180° opposite the cell's
preferred firing direction. Therefore the cells did not fire at this
directional heading and remained silent during the entire course of
travel down the mesh.
MESH 180°.
Across the 13 cells, there were an average of 3 transits up and 3 transits down the mesh when it was positioned at Mesh 180°. Figure 2,
C and H, depicts the typical responses of HD
cells during vertical plane locomotion when the mesh was positioned
180° opposite the cells' preferred firing directions (Mesh 180°).
The tuning curves indicate that when the rats climbed down the mesh
(- - -) the HD cells continued to fire, but did not fire when the
rats climbed up the mesh (). Despite a decrease in the peak firing rate when the rat climbed down the mesh (compare Fig. 2C
with Fig. 2, A and B), the two plots illustrate
that the cell's directional coding was still maintained during the
downward trip. Table 1 shows that most cells did not exhibit a
significant change in peak firing rate during vertical locomotion when
the rat climbed down the mesh at this position (e.g., compare Fig.
2H with Fig. 2, F and G). In general,
during the Mesh 180° session, as the rat approached the mesh from the
annulus, the rat's head orientation was in alignment with the cell's
preferred firing direction. Thus the cell discharged maximally and
continued to fire at, or near, its peak firing rate as the rat climbed
down the mesh. When the rat was on the cylinder floor and approached
the mesh, the rat's head orientation was 180° opposite the cell's
preferred firing direction. Therefore the cell did not fire at this
directional heading and continued to exhibit no discharge as the rat
climbed up the mesh.
MESH 90° AND 270°. Across the 13 cells, there were an average of 3 transits up and 3 transits down the mesh, when it was positioned at Mesh 90° or 270°. With the mesh positioned 90 or 270° from the cell's preferred firing direction, all HD cells exhibited negligible firing when the rats climbed up or down the mesh. Figure 2, D and I, depicts the typical responses of HD cells during vertical plane locomotion in the Mesh 90° sessions; and Fig. 2, E and J, depicts HD cell activity during the Mesh 270° sessions. These tuning functions show that minimal, if any, firing was detected as the rats moved up or down the mesh at these positions. During these sessions the approach paths to the mesh, whether from the cylinder floor or the annulus, would lead the rat's head orientation to be ±90° away from the preferred firing direction of the recorded cells.
In climbing down the mesh the rats would often stop one-third of the way down and turn to look around before continuing down. Such behavior would occasionally result in the rat's head position becoming aligned with the preferred firing direction of the HD cell. In these instances, HD cells would exhibit an increase in firing rate. For example, Fig. 3A depicts the directional tuning properties of a HD cell as the rat foraged for food pellets on the cylinder floor. During the subsequent Mesh 90° recording session when the rat climbed down the mesh, the rat oriented its head toward the preferred firing direction. In this event, as shown in Fig. 3B (- - -), the cell discharged at its peak firing rate. In contrast, the cell exhibited negligible firing during the more stereotypic straight running path when the rat climbed up (
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HD cell activity in the elevated horizontal plane
In general, HD cells continued to exhibit direction-specific discharge when the rat moved about on the annulus and pointed its head in the cell's preferred firing direction, as defined by the reference frame of the cylinder floor. In some cases, once the rat climbed onto the annulus it was necessary to remove the mesh, to increase the annulus sampling time. In such cases, we found no differences in HD cell discharge properties, and data were combined across all annulus sessions. To compare HD cell firing rates on the floor versus the annulus, we combined all the episodes on the annulus into one session for each cell. Average sampling time per episode on the annulus was 15.8 s (range: 5.0-29.9); the average total time spent on the annulus for each rat over the entire recording session was 10.13 min (range: 4.82-19.62).
Eleven of the 13 HD cells exhibited an increase in peak firing rate
during locomotion on the annulus compared with the activity displayed
on the cylinder floor (see Fig. 4,
A and B). Table 2 presents the mean peak firing rate, directional firing range, directional information content, background firing rate, and
signal-to-noise ratio of the HD cells during movement on the two
horizontal surfaces. A paired two-tailed t-test indicated
that there was a significant difference in HD cell peak firing rates
between the cylinder floor and annulus, t (12) = 3.36, P < 0.001. Despite a 20% increase in the mean
background firing rate of HD cells on the annulus, a
t-test found that this difference was not significant
(P > 0.05). Similar analyses of directional firing
range, directional information content, and signal-to-noise ratio
indicated no significant differences in these parameters (all
Ps >0.05). These results indicate that HD cells exhibit a
significant increase (~30%) in peak discharge while the rats
locomote about the annulus surface as compared with the peak discharge
rates exhibited on the cylinder floor surface.
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To evaluate the increased firing rates on the annulus, an annulus:cylinder floor peak firing rate ratio was calculated for each HD cell. A ratio value of 1.0 would indicate equivalent peak firing rates across both surfaces, whereas a ratio <1.0 would indicate higher firing rates on the cylinder floor, and a ratio >1.0 would indicate higher firing rates on the annulus. The mean firing rate ratio for the 13 HD cells was 1.31 ± 0.08 (range: 0.69-1.76), and Fig. 4C illustrates the distribution of the firing rate ratios. A paired, two-tailed t-test showed that this value is significantly different from the expected ratio of 1.0 if there were no difference in peak firing rate between the two surfaces [t (12) = 4.04, P = 0.002]. As mentioned above and illustrated in Fig. 4C, only one HD cell exhibited a decrease in peak firing rate on the annulus as compared with the cylinder floor (i.e., annulus: floor ratio <1.0). In two cases, two HD cells were recorded simultaneously over the course of the entire recording protocol. In these cases, the annulus:floor ratios of the two simultaneously recorded cells were Case 1: cell 1 = 1.06, cell 2 = 1.59; Case 2: cell 1 = 1.46, cell 2 = 1.65. Thus in one of these cases, the two cells did not exhibit equivalent increases in peak firing rate on the annulus and suggests that the magnitude to which the peak firing rate increases while the rat is on the annulus may be cell specific.
To determine whether the difference in peak firing rate between the two horizontal surfaces was consistent across episodes, we calculated the annulus:cylinder floor peak firing rate ratio for each episode of transit between the two surfaces. This analysis was only conducted on 11 of the 13 HD cells because two cells were recorded before the use of the video switchbox. The analysis was further hampered by the fact that in some cases, the rat would climb up to the annulus surface, turn around, and immediately return to the cylinder floor. Such transit episodes were not included in the analyses because they were of too short-duration to accurately evaluate the peak firing rate. Depending on the rat's activity, episodes as short as 1-2 min were often sufficient to determine the HD cell's firing properties. For each of the 11 cells, we recorded at least 12 episodes of transit between the two surfaces that contained sufficient sampling times. Figure 4D depicts the mean change in peak firing rate ratio across the 12 episodes of transit. The mean annulus:cylinder floor peak firing rate ratio across all episodes was 1.30 ± 0.07. A repeated measures ANOVA, conducted to examine the per episode variability in peak firing rate ratio, found a nonsignificant repeated measures effect of episode on the ratio value, F(11, 110) = 1.12, n.s. These data confirm that the peak firing rate ratio did not significantly change across 12 transits between the 2 horizontal surfaces and indicates that the increased peak firing rate on the annulus surface was consistent across multiple episodes.
To further determine whether an increase in background firing rate contributed to the observed increase in peak firing rate on the annulus, we computed an annulus:floor background firing rate ratio for each cell. The mean background firing rate ratio for the 13 HD cells was 1.59 ± 0.09 (range: 0.58-4.99), and Fig. 4E illustrates the distribution of the ratios. Although this value is greater than the ratio observed for peak firing rate (1.31), a paired, two-tailed t-test showed that this value was not significantly different from the expected ratio of 1.0 if there were no difference in background firing rate between the two surfaces [t (12) = 1.85, n.s.]. The lack of significance can be attributed to two large individual ratio values (2.65 and 4.99); without these two values, the overall mean background firing rate ratio is 1.18 ± 0.11. Figure 4F depicts the mean change in background firing rate ratio across the 12 episodes of transit. The mean annulus:cylinder floor peak firing rate ratio across all episodes was 1.06 ± 0.06. A repeated measures ANOVA found a nonsignificant repeated measures effect of episode on the ratio value, F(11, 110) = 0.75, n.s. These data confirm that the background firing rate ratio did not significantly change across 12 transits between the 2 horizontal surfaces and suggest that the observed increase in peak firing rate on the annulus was not simply due to an overall increase in the cell's activity.
We also examined the variability in peak firing rates of the 11 HD
cells between at least 2 standard cylinder sessions in the absence of
the mesh, to determine whether peak firing rates were stable on the
cylinder floor. This analysis also enabled us to determine whether the
increase in peak firing rate on the annulus was attributed to random
variability in peak firing rate across sessions. The mean (±SE)
cylinder floor standard session:standard session peak firing rate ratio
was 1.01 ± 0.02. A paired, two-tailed t-test assessed
whether the ratio of peak firing rate between the floor and annulus was
greater than the ratio between successive cylinder floor sessions. This
analysis revealed a significant difference between the two mean peak
firing rate ratios, t (10) = 3.62, P = 0.005. These data indicate that HD cell peak firing rates on the
cylinder floor and on the annulus are consistent, but that there is a
significant increase in annulus peak firing rate over cylinder floor
peak firing rate.
Minor shifts of the preferred firing direction between the cylinder floor and annulus
The preferred firing directions of the HD cells were
stable across standard recording sessions when the rat was on the
cylinder floor and were not influenced by the session-to-session change in the angular position of the mesh on the cylinder wall. The preoperative training regimen involved the explicit attempt to decrease
apparent stability of mesh orientation by varying its angular position
across trials. Comparison of preferred firing directions between the
annulus and cylinder floor revealed a mean absolute shift of 12.9 ± 3.2° (range: 0-36°). This shift is significantly different from
that reported by Taube (1995) between two standard cylinder sessions (4.7 ± 1.8°) [t (25) = 2.25, P = 0.03]. The increased shift on the annulus may
reflect the influence of additional cues and environmental features
that are available to the rat while foraging on the annulus. Although
12.9° is > 4.7°, this shift magnitude is substantially smaller
than that observed following a change of apparatus. For example,
Dudchenko and Taube (1997)
observed a mean absolute
shift in preferred firing direction of 40.7 ± 17.4° between a
standard cylinder session and a session on an elevated radial-arm maze,
and Taube et al. (1990b)
reported a mean shift of
83.3 ± 15.6° between a cylinder and rectangular apparatus. The
second apparatus in each case was also centered in the same recording
room as the cylinder sessions. Thus the shift between the cylinder
floor and the annulus, although significantly different, should be
considered relatively minor.
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DISCUSSION |
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Locomotion in the vertical plane
This experiment was designed to determine how the horizontal reference frame is reflected in the activity of HD cells. HD cell firing properties were monitored as the rat moved from a horizontal surface onto a 90° vertical surface that was positioned at discrete angular orientations with respect to the cell's preferred firing direction. The study also enabled a determination of the differences in HD cell activity between the two distinct horizontal surfaces that were separated in height by ~1 m.
In general, HD cell discharge properties were maintained during
locomotion in both vertical (up and down) and horizontal (cylinder floor and annulus) planes. The present results indicate that when the
rat approached the mesh either from the cylinder floor or annulus with
its head direction corresponding to the cell's preferred firing
direction, and maintained this head orientation during vertical
movement (either up or down the mesh), cell discharge was sustained
throughout vertical movement. Thus regardless of body position, as long
as the head orientation corresponded to the cell's preferred firing
direction (within ±90° roll of the head), whether +90° vertical
(as during climbing up the mesh) or 90° vertical (as during
climbing down the mesh), HD cells will discharge at maximal rates.
These results indicate that HD cells are capable of maintaining their
discharge in planes outside earth horizontal. Furthermore, when
locomoting in the vertical plane, HD cell firing is dependent on the
directional orientation of the rat as it moves out of the horizontal
plane. These data are consistent with the notion that the cell defines
the horizontal reference frame as the rat's plane of locomotion, but
leaves open the question of which cue source(s) determine the
horizontal reference frame: vestibular, somatosensory, visual, motor
cues, or some combination of these sources.
Directional cells of the ADN and PoS are thought to be components of a
neural circuit that supports landmark-based navigation and inertial
navigation or path integration. In the absence of landmark cues, the
firing properties of HD cells may be maintained by internal cue sources
(Goodridge and Taube 1995; Taube and Burton 1995
). Moreover, as mentioned earlier, vestibular input to the HD cell circuit is necessary for the directional discharge in the ADN,
even in the presence of familiar landmark cues (Stackman and
Taube 1997
). This result indicates that idiothetic cue sources may be essential for the generation of the HD cell signal. In light of
the fact that HD cell activity is dependent on vestibular information,
it is interesting to note that during movement in the vertical plane
the HD cell circuitry is receiving different vestibular/kinesthetic
input as compared with that during movement in the horizontal plane.
Thus despite the different input from the semicircular canals and
otolith organs during vertical plane transits, HD cell activity was
maintained. There may be additional inputs to HD cells that provide
information about head position in the vertical plane. For example, we
have recently demonstrated the presence of head pitch cells in the
lateral mammillary nuclei that discharged when the rat's head was
orthogonal to the horizontal plane (Stackman and Taube
1998
). Via the mammillothalamic tract, information regarding
head pitch may be conveyed to the ADN and integrated to enable a
representation of head direction in three dimensions. Therefore it is
conceivable that the maintained discharge of HD cells in the vertical
plane was a consequence of monitoring angular and linear head movements
through vestibular signals and motor efference copy. However, it is
important to consider that the cue card was present throughout all
recording sessions, and therefore the maintained HD cell discharge may
have simply been a consequence of referencing this familiar landmark.
Our experiment does not distinguish between these two possibilities.
Three-dimensional model of HD cell firing
How the horizontal reference frame is defined by HD cells is an
important theoretical issue. There is considerable evidence in humans
to indicate that the perception of horizontality and verticality are
dependent on visual, vestibular, and proprioceptive systems
(Howard and Templeton 1966). In the dark, vestibular
deficient humans are impaired in aligning a visual target to the
Earth-horizontal as compared with intact humans (Miller and
Graybiel 1966
; Friedmann 1970
). In intact
humans, the perception of horizontality and verticality can be altered
by simply lying in a supine position (Miller and Graybiel
1966
) or by tilting the body to one side (Werner et al. 1951
). These findings illustrate the importance that vestibular signals from the otolith organs play in judging spatial orientation.
The present findings indicate that HD cells appear to maintain a
representation of directional heading despite the vertical body
positioning. However, it remains to be determined whether the shift in
body position alters the animal's sense of spatial orientation.
Nonetheless, because HD cell discharge was maintained in planes outside
the earth horizontal, our results indicate that the HD cell signal can
be represented by a three-dimensional model (see Fig.
5). Figure 5A illustrates in
polar coordinates a three-dimensional representation of a hypothetical
HD cell with a preferred firing direction of 90° in the horizontal
plane (defined as the x-y plane). Figure 5B
illustrates the typical firing rate by HD tuning function of this cell
during movement about the horizontal cylinder floor. In Fig.
5A, the activity of this HD cell on the horizontal surface is represented as the thick elliptical plot centered at an HD of 90°
horizontal, 0° vertical. The firing rates of the hypothetical cell
when the rat's head is aligned with the preferred firing direction but
at varying vertical orientations are depicted by the elliptical plots
(a, b, c, d, and e). If one were to superimpose the polar plots for all possible vertical orientations, from +90 to
90, the cell's firing could be characterized as a
hemitorus1 surface
as depicted in the drawing of Fig. 5C. Such a model is consistent with the present results and the notion that the HD cell
will exhibit peak discharge rates whenever the head orientation corresponds to the cell's preferred firing direction, irrespective of
body orientation. It is interesting to note that this model makes the
prediction that HD cell firing will cease during movement in a vertical
plane that is greater than +90°. In the extreme case the question
becomes how an HD cell would respond if the rat approached a +90
vertical surface aligned with the preferred firing direction, climbed
this surface to the ceiling, and then continued onto the ceiling. In
this case, the rat's path would culminate in climbing upside down on
the ceiling heading in the opposite direction from which it started. We
are currently examining whether an HD cell continues to fire, or
whether the activity of the cell is attenuated, in such a situation.
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It is tempting to suggest that the maintenance of directional firing in
the vertical plane indicates the preservation of the directional sense
while locomoting in the vertical plane. To verify this notion, however,
it would be necessary to test the animal on a navigational task that
requires vertical plane movements. Interestingly, Grobety and
Schenk (1992a) showed that rats trained on a radial-arm maze
containing arms of varying vertical tilt (from 0 to 25°) acquired the
task faster than rats trained on a standard version with all arms
horizontal. Moreover, in a three-dimensional cubic maze, where
accessing the goal location required movements in the horizontal and
vertical planes, rats appeared to acquire the vertical coordinate of
the goal location before successfully learning the horizontal
coordinate (Grobety and Schenk 1992b
). These data
demonstrate the ability of rats to process spatial information
concerning their three-dimensional orientation. The added kinesthetic
information that results from movements in the vertical dimension may
facilitate the acquisition of spatial learning and navigation tasks by
providing an additional cue source.
Locomotion in a horizontal plane at a different height
Although the preferred firing direction of HD cells were similar between the two horizontal surfaces (±12°), peak firing rates were increased by an average of 33% during locomotion on the annulus as compared with the cylinder floor. During movement on the annulus, there were no significant differences in other HD cell firing parameters. These results suggest that there are additional influences or environmental features available to the rat that affect HD cell directional coding while on the annulus that are not available to it while on the cylinder floor. Alternatively, the changes in firing rates may be a consequence of alterations in the motoric demand between the two surfaces.
It is tempting to speculate that the increase in peak firing rate on the annulus reflects an additional encoding of height from a horizontal surface by HD cells. During acquisition training rats received considerable exposure to the apparatus and cues within the recording room that could be seen from the annulus (i.e., the laboratory floor, stool, etc.), therefore it is unlikely that such cues were responsible for increased peak firing rate when the rat was on the annulus. It is also unlikely that the presence of the second camera influenced HD cell activity, because the preferred firing direction of each HD cell was stable despite moving the second camera from session to session. Such stability suggests that the rats did not use the second camera as a reference landmark. In addition, it is unlikely that the increased firing rate on the annulus reflects food consumption from the annulus food wells because the consumption of food pellets in the cylinder did not increase peak firing rate. Therefore the rats consumed food pellets on both horizontal surfaces, and we have not previously observed a consumption-dependent increase in HD cell firing rates.
Other factors also need to be considered when accounting for the
increased firing rate on the annulus. For instance, the annulus was
narrower than the cylinder floor and contained no inside or outside
walls. Rats routinely moved about the annulus surface in a ballistic
manner (intermittent episodes of forward locomotion followed by eating
at the food cups, or peering over the inside or outside edges of the
annulus). Thus locomotion on the annulus restricts the rat's movement
somewhat and may require a heightened demand for coordination and
balance. Given the importance of vestibular signals for HD cell
activity, it is possible that movement on the annulus influences the
vestibular system in a way that is then reflected in the increased peak
firing rates of HD cells. However, Dudchenko and Taube
(1997) failed to observe any significant change in HD cell
firing rates as rats performed a radial arm maze task on a platform
that was elevated 34 cm above the floor. In sum, the differences in
peak firing rate between the two horizontal surfaces can be attributed
to either, an increased attention to balance while on the annulus or
the restriction of motor behavior by the physical dimensions of the annulus.
It is also possible that the increased firing rate on the annulus
represents a "gain field" in the HD cell, analogous to the spatial
gain fields observed in visual receptive fields for posterior parietal
cortex neurons (Andersen et al. 1985). Because the peak firing rate was increased when the rat was on the annulus, this finding
suggests that the HD cell's gain field would vary along only one
dimension: the height of the animal above the floor. In this case, the
cell's peak firing rate when the animal is on the floor would set the
minimal point of the gain field. Whether HD cells contain a gain field
comparable to posterior parietal neurons could be easily tested by
varying the height of the annulus from the floor and determining
whether the cell's peak firing rate varies proportionately.
Converging evidence indicates that HD cell activity is influenced by
the ongoing motor behavior of the animal. For example, HD cell firing
rates are decreased when the rat is securely restrained and passively
rotated through the cell's preferred firing direction (Knierim
et al. 1995; Taube 1995
; Taube et al.
1990b
). Furthermore, there is a small but significant positive
correlation between HD cell firing rates and the rat's linear velocity
over the cylinder floor (Stackman and Taube 1998
;
Taube 1995
; Taube et al. 1990a
). Together, these results suggest a continuum of the influence of motor
behavior on HD cell firing rates. Denial of volitional motor behavior
during restraint causes a decrease in HD cell firing rate, whereas the
heightened awareness of, or necessity for cautious coordinated
movements on the narrow annulus, causes an increase in HD firing rates.
Thus it would be interesting to parametrically manipulate not only the
height of the annulus, but also its width, to alter motor behavior and
examine the resulting influences on HD cell firing rates to further
define this relationship.
In summary, HD cells maintained their directional firing during traverses in the vertical plane when the animal approached and climbed into the plane facing the cells' preferred firing direction. Cell firing was negligible for all other orientations. These results suggest that the HD cell defines the horizontal reference frame as the rat's plane of current locomotion.
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ACKNOWLEDGMENTS |
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The authors thank R. Dauphinais for constructing the cylinder and annulus apparatus, D. Whitmer for expert technical assistance in training rats to climb the vertical mesh, D. Collins and J. Noll of the Electronics Shop at Dartmouth College for design and construction of the video source switchbox, Dr. Charles Oman of Massachusetts Institute of Technology, Cambridge, MA, for numerous discussions of the hemitorus model as a 3D descriptor of HD cell firing, and The Bagel Basement of Hanover, NH, for providing a fresh supply of "torus models" to facilitate descriptions of HD cell firing in 3D. A preliminary report of this research was presented at the 27th Annual Society for Neuroscience meeting in New Orleans, LA, in 1997.
This research was supported by National Institutes of Health Grant DC-00236 to R. W. Stackman and MH-48924 and MH-01286 to J. S. Taube.
Present address of R. W. Stackman: Dept. of Behavioral Neuroscience, L470, Oregon Health Sciences University, 3181 SW Sam Jackson Park Rd., Portland, OR 97201-3098.
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FOOTNOTES |
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Address for reprint requests: J. S. Taube, Dept. of Psychological and Brain Sciences, Dartmouth College, 6207 Moore Hall, Hanover, NH 03755-3549.
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.
1 A torus, defined in geometry, is a surface generated by rotating a circle around an axis that is in the plane of the circle, but does not intersect the circle (i.e., a doughnut). A hemitorus is the surface generated by rotating the circle 180° about the axis.
Received 5 April 1999; accepted in final form 22 September 1999.
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REFERENCES |
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