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Department of Zoology, University of Oklahoma, Norman, Oklahoma 73019
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ABSTRACT |
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 Abstract
 Introduction
Methods
Results
Discussion
References
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Wilson, Donald A. Binaral interactions in the rat piriform cortex. J. Neurophysiol. 78: 160-169, 1997. Single-unit recordings were made from layer II/III anterior piriform cortex (aPCX) neurons in adult Wistar rats to examine odor response patterns to unilaterally and bilaterally delivered stimuli. Isoamyl acetate odor stimulation was presented either unilaterally through tubes inserted into the external nares, or bilaterally during unilateral olfactory bulb lidocaine infusions. Olfactory bulb multiunit or slow-wave activity was recorded simultaneously bilaterally to monitor selectivity of unilateral odor stimulation. The results demonstrate that 1) commissural input to aPCX neurons is sufficient to drive odor responses, and 2) aPCX neurons can be classified on the basis of spatial receptive field type. These receptive fields include cells that respond 1) selectively to ipsilateral stimulation, 2) selectively to contralateral stimulation, 3) to either ipsilateral or contralateral stimulation, and 4) selectively to bilateral stimulation. The potential functions of binaral convergence in the piriform cortex are discussed, and may include enhancement of perceived odor intensity and bilateral access to olfactory memory.
Convergence of inputs from bilateral receptive fields is a common feature of vertebrate sensory systems. This bilateral convergence allows for relative comparisons of spatially disparate inputs that can result in higher-order sensory processes such as depth perception in vision and stimulus localization in audition.
Subjects
Eleven male Wistar rats (150-350 g) obtained from Charles River Labs were used as subjects. Animals were housed in polypropylene cages lined with wood chips. Food and water were available ad libitum. Lights were maintained on a 12:12 light:dark cycle with testing occurring during the light portion of the cycle.
Electrophysiology
Animals were anesthetized with urethan (1.5 g/kg) and placed in a stereotaxic apparatus. Both olfactory bulbs were exposed through holes drilled in the dorsal surface of the skull and a third hole drilled over the aPCX, ~1 mm anterior to bregma. Recordings were made with tungsten microelectrodes (5-12 M Unilateral odor stimulation
For unilateral odor stimulation, an odor delivery device was inserted into both nares (Fig. 1). Animals were freely breathing through both nares at all times. The device consisted of a short piece of PE160 tubing connected to the narrow end of a modified Eppendorf pipetter tip (5 cm long, 0.5 cm ID at wide end). The PE160 tubing was placed into the external naris. An odor port was inserted through the wall of the pipette tip with the use of PE60 tubing. The devices were angled to create a distance of
Unilateral lidocaine application
To further examine the unilateral nature of odor response patterns, a second paradigm in which the more common method of bilateral odor stimulation with odorant delivery to the external nares was used was combined with unilateral lidocaine blockade of olfactory bulb activity. Odors were presented bilaterally through a tube ~1 cm in front of the external nares. Animals were freely breathing. A clean, humidified air stream (1 l/min) was blown constantly across the snout of the animal to exhaust odors. Stimuli were created as above, by forcing air through odorant-saturated filter paper with a syringe pump. After collection of bilateral control data, lidocaine (5-10 µl, 2% solution; Astra Pharmaceutical Products) was applied to the surface of either the ipsilateral or contralateral olfactory bulb (relative to the aPCX recording). The duration and unilateral specificity of the lidocaine effect was monitored with the olfactory bulb recordings, and odor stimulation was repeated both during the maximal lidocaine suppression and, if the stability of the aPCX recording allowed, after recovery. Spontaneous aPCX single-unit activity rates were also monitored for Response analysis
aPCX single-unit odor responses were analyzed with the use of cumulative peristimulus time histograms with either 0.1- or 0.2-s binwidths, including activity during a 4-s preodor baseline and the 4 s of odor for at least three stimulus repetitions. A single unit was determined to have responded to the stimulus if cumulative firing rate in any bin during the stimulus was above baseline variability. This measure, which has been used in the olfactory bulb (Wilson et al. 1985
A total of 70 single units was recorded from aPCX layers II/III in 10 animals. Of these, 32 cells were tested for responses to unilaterally delivered odors, and 38 cells were tested for spontaneous activity and/or odor responses during unilateral olfactory bulb lidocaine application. Simultaneously, olfactory bulb activity was recorded bilaterally in all animals to monitor and confirm the unilateral nature of odor stimulation and lidocaine actions. Olfactory bulb multiunit responses to odor were characterized by robust excitation. Olfactory bulb slow-wave responses to odors were characterized by large-amplitude waves in phase with respiration, as previously reported (Adrian 1950
Unilateral odor presentation
Figure 2 is a representative example of an aPCX single-unit response to odor stimulation of either the ipsilateral or contralateral naris. The same aPCX unit is shown for each stimulus. Bilateral odor stimulation (Fig. 2A), produced a rapid increase in this cell's activity. Similar excitatory responses were observed to unilateral odor stimulation delivered to either naris. For example, odor presented to the ipsilateral naris (Fig. 2B) selectively activated the ipsilateral olfactory bulb and excited the aPCX unit. Odor presented to the contralateral naris (Fig. 2C) selectively activated the contralateral olfactory bulb and also excited the aPCX unit. Figure 2 thus is an example of a binaral aPCX unit with bilateral receptive fields.
Unilateral olfactory bulb lidocaine application
A total of 38 aPCX single units in five animals was tested for the effects of unilateral olfactory lidocaine application on spontaneous activity (ipsilateral lidocaine, n = 21; contralateral lidocaine, n = 21; 3 cells were tested in both conditions). Lidocaine application on the dorsal surface of the olfactory bulb significantly reduced olfactory bulb activity as determined by complete cessation of multiunit activity (e.g., Fig. 6) or drastic reduction in odor-evoked slow-wave activity recorded in the ventral mitral cell layer (e.g., Fig. 7). Maintenance of some slow-wave activity during lidocaine application might be expected because of the contribution of remaining subthreshold synaptic activity to this measure. The bilateral bulb recordings demonstrated that lidocaine application produced selective unilateral effects on bulb activity (e.g., Figs. 7 and 8).
The present results demonstrate that aPCX layer II/III single units can respond to unilateral odor stimulation of the contralateral naris. Furthermore, these results suggest that, similar to monocular and binocular receptive fields of mammalian visual cortical neurons, aPCX neurons have mononaral and binaral receptive fields.
Potential anatomic substrates of spatial receptive fields
The relatively similar proportion of ipsilaterally and contralaterally driven mononaral aPCX neurons was surprising given the relatively low density of commissural fibers compared with LOT fiber density in the rat aPCX (Friedman and Price 1986 Functional role of spatial receptive fields
The functional role of commissural connections and binaral receptive fields in olfactory coding is unknown. Although commissural connections are important in other sensory systems for stimulus localization, in humans simultaneous binaral comparisons do not appear sufficient to allow odorant source localization (in the absence of concomitant trigeminal activation) (Kobal et al. 1989
INTRODUCTION
Abstract
Introduction
Methods
Results
Discussion
References
).
; Luskin and Price 1983). These interhemispheric connections are carried through the anterior limb of the anterior commissure. For example, in the olfactory bulb, input from the ipsilateral naris reaches the apical dendrite of mitral/tufted cells, whereas the most direct input from the contralateral naris terminates on inhibitory interneurons called granule cells (Price and Powell 1970). In the piriform cortex, however, binaral convergence occurs within different subfields of the apical dendrites of layer II and III pyramidal neurons (Friedman and Price 1984). Fibers from the ipsilateral olfactory bulb (ipsilateral naris) compose the lateral olfactory tract (LOT) and terminate in the piriform cortex superficial layer Ia, whereas commissural fibers from the contralateral hemisphere (contralateral naris) terminate in the deeper layer Ib, along with intracortical association fibers (Haberly 1985; Haberly and Price 1978; Luskin and Price 1983). In the anterior piriform cortex (aPCX), the source of these commissural fibers is primarily the contralateral pars lateralis of the anterior olfactory nucleus, which receives direct input from the olfactory bulb (Luskin and Price 1983). Fibers in layers Ia and Ib form excitatory synaptic connections with pyramidal neurons (Haberly 1985; Haberly and Bower 1984).
that is, they may perform relative comparisons between activity in the two olfactory receptor sheetssimilar to the binocular cells of the primary visual cortex. To date, there have been relatively few single-unit examinations of mammalian piriform cortex odor response patterns and all have relied on bilaterally presented odors (Duchamp-Viret et al. 1996; Giachetti and MacLeod 1975; Haberly 1969; McCollum et al. 1991; Nemitz and Goldberg 1983; Schoenbaum and Eichenbaum 1995; Tanabe et al. 1975). These studies have found that piriform cortex units respond to odors with relatively short latencies and with a variety of temporalpatterns.
METHODS
Abstract
Introduction
Methods
Results
Discussion
References
, A-M Systems).
) was lowered from the dorsal skull surface. Placement of the recording electrode in layer II/III of the piriform was performed under physiological control by electrically stimulating (pulse duration 0.1 ms; 50-500 µA) through the ipsilateral olfactory bulb recording electrode. Recordings were performed at, or slightly dorsal to, the reversal point of the population potential evoked by ipsilateral bulb stimulation (Haberly and Shepherd 1973). Single units were isolated directly, or were extracted through template matching (sampling rate 10 kHz) with the use of Spike2 software for the Macintosh (CED). The single-unit nature of the recordings was confirmed with autocorrelograms showing a refractory period of 
3 ms.
3-4 cm between clean air intake sites (wide end of the pipette tip). A humidified, clean air stream (1 l/min) was directed downward over the two devices to minimize odorant diffusion out of one device to the contralateral device. Stimuli were produced with a Hamilton model 22 syringe pump controlled with a Macintosh computer and Spike2 software. The syringe pump forced air across odorant (isoamyl acetate, Sigma)-saturated filter paper to present the odor through the odor ports. Odors were presented unilaterally or bilaterally through computer-controlled solenoids that directed the odor stream (10-25 ml/min) to one or the other device. Within an animal, all stimuli were delivered at the same volume per minute, i.e., the total volume of odor delivered during bilateral stimulation matched the total volume delivered during unilateral stimulation. Stimulus duration was 4 s, and both olfactory bulb and aPCX activity during the odor were compared with a 4-s preodor baseline. Interstimulus intervals were 60 s. Each stimulus (ipsilateral, contralateral, or both) was repeated at least three times and order of presentation was varied within and between cells.
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FIG. 1.
Uninaral stimulation apparatus. Air was forced through odorant-saturated filter paper by computer-controlled activation of a syringe pump and simultaneous opening of a solenoid valve. Odor was delivered to a device inserted into the animal's naris, and inhaled by normal respiration, as described in METHODS. Humidified clean air was directed over the open end of the delivery devices.
100 s before and 5 min after lidocaine application.
), was chosen because, as previously reported (McCollum et al. 1991) and as described below, aPCX single-unit odor responses were frequently very brief and habituated rapidly. Furthermore, preodor spontaneous activity was often very slow or silent (especially in the lidocaine experiments), thus precluding statistical comparison of preodor versus postodor firing rates. Because our objective was to describe the frequency of occurrence of ipsilaterally and contralaterally driven responses, the most important requirement was to apply a constant, sensitive criterion in all circumstances. This measure satisfies that requirement. Because of variability of responses, no attempt was made to quantify response magnitude for comparisons of relative effectiveness of ipsilateral versus contralateral inputs.
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FIG. 2.
Anterior piriform cortex (aPCX) single-unit response to isoamyl acetate delivered bilaterally and unilaterally through tubes inserted into the nares. A: bilateral odor presentation produced a robust aPCX excitatory response. B: unilateral odor presentation to ipsilateral naris selectively activated ipsilateral main olfactory bulb (MOB) as shown in histograms of multiunit activity recorded simultaneously in ipsilateral and contralateral MOB. aPCX single unit responded to this ipsilateral stimulation. C: unilateral odor presentation to contralateral naris selectively activated contralateral MOB. aPCX unit also responded to this contralaterally delivered odor stimulus. D: single-unit nature of this and all recordings was verified with autocorrelation analysis.
RESULTS
Abstract
Introduction
Methods
Results
Discussion
References
). Both of these measures were reliable and sensitive indicators of odor stimulation. aPCX single units responded to odor stimulation most frequently with excitation, and very rarely with suppression in our sample. This excitation generally did not last through the entire 4-s odor stimulus (e.g., Figs. 3 and 7).
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FIG. 3.
Unilateral odor presentation could result in "cross talk" between nasal passages. Stimulus rates were adjusted within animals to eliminate cross talk. This figure shows simultaneous bilateral multiunit recording of MOB activity and single-unit recording of aPCX activity in response to odor presented to the naris ipsilateral to the aPCX recording (stimulus presentation marked by horizontal bar). Top: when the odor was presented at a high rate (25 ml/min), leakage occurred between nasal passages, possibly through the septal window, such that both MOBs responded. Bottom: at lower stimulus rates (10 ml/min) no leakage occurred between hemispheres. Note aPCX unit responded to odor in both conditions.
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FIG. 7.
aPCX single-unit and simultaneous MOB slow-wave response to bilateral odor presentation before and after lidocaine application on ipsilateral MOB. Ipsilateral lidocaine dramatically reduced aPCX spontaneous activity, but did not block the aPCX odor response despite dramatically reducing ipsilateral MOB slow-wave activity. Fast Fourier transform (FFT) analysis showed that contralateral MOB slow-wave activity was not reduced by lidocaine infusion.
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FIG. 4.
aPCX single-unit odor response patterns to bilateral and unilateral stimulation through naris tubes. Selective response patterns including cells that responded to ipsilateral stimulation but not contralateral stimulation, that responded to contralateral stimulation but not ipsilateral stimulation, that responded to either ipsilateral or contralateral stimulation, and that responded only to bilateral stimulation. Values are means ± SE.
; Nemitz and Goldberg 1983).
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FIG. 6.
aPCX single-unit and ipsilateral MOB multiunit response to bilateral odor presentation before (top) and after (bottom) lidocaine application on ipsilateral MOB. Lidocaine resulted in total blockade of ipsilateral MOB multiunit activity (bottom left), yet the aPCX single unit continued to respond to odor stimulus (bottom right).
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FIG. 8.
aPCX single-unit and simultaneous MOB slow-wave response to bilateral odor presentation before and after lidocaine application on ipsilateral MOB. In this example, the aPCX single-unit spontaneous activity and odor response were blocked by ipsilateral MOB lidocaine infusion. Responses returned as lidocaine effect diminished.
92.9, P < 0.001; contralateral lidocaine, t(20) = 54.5, P < 0.001; Fig. 5B).
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FIG. 5.
aPCX single-unit spontaneous activity following unilateral lidocaine surface application onto either ipsilateral or contralateral MOB. Ipsilateral application resulted in a nearly complete cessation of aPCX activity, whereas contralateral applications resulted in a 50% decrease in spontaneous activity.
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FIG. 9.
Percent of aPCX single units responding to bilateral odor stimuli before and after lidocaine application to ipsilateral OB. Despite elimination of detectable ipsilateral olfactory bulb activity, 21% of cells maintained odor responses.
DISCUSSION
Abstract
Introduction
Methods
Results
Discussion
References
; von Baumgarten et al. 1962). In the present study, evidence of this interbulbar inhibition was seen in the lidocaine experiments. In several cases, unilateral olfactory bulb blockade with lidocaine enhanced the magnitude of contralateral bulb odor responses, as seen quantitatively in the FFT analysis in Fig. 7 and qualitatively in the slow-wave amplitude of olfactory bulbs contralateral to lidocaine-suppressed bulbs (Figs. 7 and 8). Thus, if the interbulbar interaction influenced aPCX activity, it might be expected to reduce the probability of observing aPCX unit odor responses. Instead, we observed contralaterally driven excitation of aPCX units. Furthermore, as shown above, aPCX units continued to respond to odors despite lidocaine suppression of the ipsilateral bulb.
).
). However, as stated above, commissural fibers terminate in the proximal half of the apical dendritic tree, and thus are electrotonically closer to the axon hillock than LOT fiber terminals. In addition, LOT fibers, but not commissural fibers, are believed to activate feedforward inhibitory interneurons, which could produce a relative weakening of ipsilateral input (Kanter et al. 1996).
; Heimer and Kalil 1978). Thus semilunar cells might be expected to receive strong input from the ipsilateral LOT and limited input from associational/commissural fibers. In fact, bulbectomy results in a selective, rapid die-off of semilunar cells in the ipsilateral aPCX (Heimer and Kalil 1978). As another example, although most layer II/III pyramidal neurons have apical dendrites extending through both layers Ib and Ia, a subclass of layer III pyramidal cells has apical dendrites that end in profuse branching at the layer Ib/Ia border (Haberly 1983). Thus these cells might be expected to receive primarily association/commissural input and very limited ipsilateral LOT input.
). Thus the relative proportion of the four described spatial receptive fields as shown in Fig. 4 might be expected to vary with location within the piriform cortex.
). However, lesion studies of olfactory guided behavior have suggested that the anterior commissure may be involved in at least two specific aspects of olfaction. First, lesions of the anterior commissure increase odor detection thresholds in rats (Bennett 1968). These results were interpreted as suggesting that intact commissural connections increase perceived odor intensity, perhaps through an additive effect of bilateral stimulation on higher structures. The present finding that 10% of aPCX neurons required bilateral stimulation to produce a detectable response (Fig. 4) supports this hypothesis. Second, the anterior commissure appears to be critically involved in transfer of, or access to, olfactory memories acquired unilaterally in rats (Kucharski and Hall 1987; cf. Olsson and Cain 1996 for humans). The observation here that aPCX neurons have binaral receptive fields may help account for this apparent information transfer. Unilateral olfactory training may modify both ipsilateral and contralateral aPCXs.
)? 2) Do binaral aPCX units have similar odor receptive fields on both sides (i.e., do they respond to the same set of odors ipsilaterally and contralaterally)? 3) Does habituation of aPCX unit responses to ipsilateral odors produce habituation to contralateral odors and vice versa?
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ACKNOWLEDGEMENTS |
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The author thanks Drs. Joseph Bastian and Regina Sullivan for comments on an earlier draft of this manuscript.
This work was supported by National Institute of Deafness and Other Communications Disorders Grant DC-01674 to D. A. Wilson.
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FOOTNOTES |
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Received 7 November 1996; accepted in final form 14 March 1997.
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REFERENCES |
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Abstract
Introduction
Methods
Results
Discussion
References
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