Functional Mapping of Human Brain in Olfactory Processing: A PET Study

Ahmad Qureshy,1 Ryuta Kawashima,1,3 Muhammad Babar Imran,1 Motoaki Sugiura,1 Ryoi Goto,1 Ken Okada,1 Kentaro Inoue,1 Masatoshi Itoh,2 Thorsten Schormann,6 Karl Zilles,4,5 and Hiroshi Fukuda1,3

 1Department of Nuclear Medicine and Radiology, Institute of Development, Aging and Cancer, Tohoku University, Sendai 980-8575;  2Cyclotron and Radioisotope Center, Tohoku University, Sendai 980-8578;  3Aoba Brain Imaging Research Center, Telecommunications Advancement Organization, Sendai 980-8575, Japan;  4C. and O. Vogt-Institute of Brain Research, Heinerich-Heine University, D-40001 Dusseldorf;  5Institute of Medicine, Research Center, D-52425 Julich; and  6Institute for Neuroanatomy, Heinerich-Heine University, D-40001 Dusseldorf, Germany


    ABSTRACT
TOP
ABSTRACT
INTRODUCTION
METHODS
RESULTS
DISCUSSION
REFERENCES

Qureshy, Ahmad, Ryuta Kawashima, Muhammad Babar Imran, Motoaki Sugiura, Ryoi Goto, Ken Okada, Kentaro Inoue, Masatoshi Itoh, Thorsten Schormann, Karl Zilles, and Hiroshi Fukuda. Functional Mapping of Human Brain in Olfactory Processing: A PET Study. J. Neurophysiol. 84: 1656-1666, 2000. This study describes the functional anatomy of olfactory and visual naming and matching in humans, using positron emission tomography (PET). One baseline control task without olfactory or visual stimulation, one control task with simple olfactory and visual stimulation without cognition, one set of olfactory and visual naming tasks, and one set of olfactory and visual matching tasks were administered to eight normal volunteers. In the olfactory naming task (ON), odors from familiar items, associated with some verbal label, were to be named. Hence, it required long-term olfactory memory retrieval for stimulus recognition. The olfactory matching task (OM) involved differentiating a recently encoded unfamiliar odor from a sequentially presented group of unfamiliar odors. This required short-term olfactory memory retrieval for stimulus differentiation. The simple olfactory and visual stimulation resulted in activation of the left orbitofrontal region, the right piriform cortex, and the bilateral occipital cortex. During olfactory naming, activation was detected in the left cuneus, the right anterior cingulate gyrus, the left insula, and the cerebellum bilaterally. It appears that the effort to identify the origin of an odor involved semantic analysis and some degree of mental imagery. During olfactory matching, activation was observed in the left cuneus and the cerebellum bilaterally. This identified the brain areas activated during differentiation of one unlabeled odor from the others. In cross-task analysis, the region found to be specific for olfactory naming was the left cuneus. Our results show definite recruitment of the visual cortex in ON and OM tasks, most likely related to imagery component of these tasks. The cerebellar role in cognitive tasks has been recognized, but this is the first PET study that suggests that the human cerebellum may have a role in cognitive olfactory processing as well.


    INTRODUCTION
TOP
ABSTRACT
INTRODUCTION
METHODS
RESULTS
DISCUSSION
REFERENCES

From an evolutionary point of view, the sense of smell is among the most primitive of senses. This is an important modality by which animals interact with the environment. It helps in identification of food; enhances pleasure and warns against impending danger. Through different neural networks, the sense of smell ultimately exerts various effects on the rest of the body systems. The sense of smell is considered to be well developed in animals. Unfortunately, it is the least understood of the human senses. The lack of adequate knowledge about this sense is probably because its expression is very subjective; moreover, it is less developed in humans. Yet we know that a whiff of perfume or aroma of food elicits memories of certain objects, incidents, and emotions. To date, due to the lack of an appropriate animal model, the detailed analysis of olfactory processing was not possible.

The anatomical circuitry of the olfactory system has been well defined. The peripheral-most part of the olfactory system, coming directly in contact with the environment, consists of olfactory cells located in the superior part of each nostril. From here, the signals are transmitted to the olfactory bulb. The olfactory tract connects the olfactory bulb with the central olfactory pathways, which can be divided into three different systems (Martin 1989). The medial olfactory system is associated with the most basic olfactory reflexes. The lateral olfactory system consists of the prepiriform and piriform cortices as well as the cortical portion of the amygdaloid nuclei. From here the signals are relayed to almost all portions of the limbic system that generate emotions (Nolte 1993). A phylogenetically new pathway, through the thalamus and into the orbitofrontal cortex, is thought to be associated with the conscious analysis of odor (Guyton and Hall 1996).

Animal studies have revealed that the lateral posterior part of orbitofrontal cortex is responsive, among others, to olfactory stimuli (Rolls and Baylis 1994). It was regarded to be involved in olfactory discrimination tasks (Schoenbaum et al. 1998). Hippocampal damage results in severe memory loss as was noted in a case of temporal lobe surgery by Penfield and Milner (Rawlins 1999). Animal studies revealed that spatial and nonspatial data, olfactory memory inclusive (Eichenbaum 1998; Wood et al. 1999), are stored in the hippocampal neurons. The role of amygdala in encoding olfactory cues has been noted, and it is presumed that the orbitofrontal cortex uses this information for further selection and decision making (Schoenbaum et al. 1999). Olfaction studies in animals show only a part of the whole scenario.

Our knowledge of the olfactory system in humans was mainly restricted to that gleaned from clinical and pathological studies (Jones-Gotman and Zatorre 1993; Wu et al. 1993). In the previous decade, attempts were carried out to investigate olfactory disorders using X-ray computed tomography (x-CT) and magnetic resonance imaging (MRI). These techniques have proved useful in the assessment of the peripheral causes of olfactory deficits only (Li et al. 1994). Brain perfusion single photon emission tomographic (SPECT) imaging has also been used to identify the areas involved in odor identification (Malaspina et al. 1998). However, this technique lacks the dynamicity required for detailed assessment of olfaction. Recent advanced functional neuroimaging techniques, i.e., positron emission tomography (PET) and functional magnetic resonance imaging (fMRI) have added to our understanding of olfactory processing in the human brain (Levy et al. 1997; Zatorre et al. 1992). PET has been found to be superior for evaluation of central neural circuits involved in the olfactory processing (Jolles et al. 1989). The main thrust of olfactory fMRI studies until now has been on smell perception only (Levy et al. 1998a,b; Sobel et al. 1997; Yousem et al. 1997). Recently, there is a trend of studying olfactory cognition by fMRI (Sobel et al. 1998a,b, 1999). However, the fMRI techniques suffer from certain setbacks such as the presence of air sinuses near the base of the brain, where the relevant anatomical structures are located. The studies so far reported have mainly concentrated only on the brain regions classically attributed to the olfactory system (Levy et al. 1998a; Sobel et al. 1998a).

There are very few human PET studies conducted that can elucidate olfactory processing. Simple odor presentation to subjects resulted in significant activation of the piriform cortices bilaterally, the right orbitofrontal cortex, the left inferomedial frontal cortex, and the insula/claustrum (Zatorre et al. 1992). In their study, the cognitive aspects involved in olfactory processing were not investigated, but it was concluded that the piriform cortex is the functional primary olfactory cortex and the right orbitofrontal cortex is the secondary olfactory cortex because it showed lateralization. In another human olfactory study, the amygdala and left orbitofrontal cortex showed increased regional cerebral blood flow (rCBF) in response to aversive odorants (Zald and Pardo 1997). Recently, attempts have been made to map the brain regions involved in the semantic analysis of olfactory stimuli. It was seen that the piriform and orbitofrontal areas were involved in olfactory recognition tasks, but a paucity of temporal lobe activation was noted (Dade et al. 1998). Additionally, the prefrontal cortex was involved in the short-delay olfactory recognition task. In another study on the comestibility judgment based on odors, the visual areas and frontal cortex were found to be selectively activated, while the olfactory familiarity task involved, among others, the orbitofrontal cortex and frontal gyri (Royet et al. 1999).

Therefore this study was designed to map out the brain regions that are involved in short- and long-term olfactory memories. In olfactory naming task (ON), using familiar odors of common edible items, the source of the odor was to be identified. According to our hypothesis, this would involve long-term recognition memory of odors, which a person has encountered at some time during his life. Most probably, such odors are verbalized or labeled, i.e., associated with an item or a name. In olfactory matching task (OM), an unfamiliar odor introduced only recently was to be identified among certain number of unfamiliar odors. As such odors were not associated with any certain item, this task involves brain regions that control our ability to differentiate one odor from another. In our opinion, these naming and matching tasks will activate brain regions where higher semantic processing of odor information occurs. Two baseline tasks, one with no olfactory stimulation and one with olfactory stimulation without cognition, were used to provide appropriate comparison with the main olfactory tasks. Additionally corresponding visual tasks were also included to see the effect of pure olfactory-related cognition. Thus our study is an attempt to clarify the mechanisms underlying human olfaction left unexplored by previous studies.


    METHODS
TOP
ABSTRACT
INTRODUCTION
METHODS
RESULTS
DISCUSSION
REFERENCES

Subjects

Eight subjects participated in the study. All subjects were right-handed, male native Japanese volunteers selected among local university students who responded to our advertisement. Their ages ranged from 18 to 26 yr (20.7 ± 2.4 yr). Written informed consent was obtained from each subject after full technical explanation regarding the experiment and the potential risks involved, in accordance with the principles of the Declaration of Helsinki. The study was approved by the Medical Ethics Committee of the Institute of Development, Aging and Cancer (IDAC), Tohoku University.

The subjects were selected after checking for strong right-handedness and normal color vision. Right-handedness was assessed by the Edinburgh Handedness Inventory (Oldfield 1971). None of the subjects had a history of rhinal surgery, head injury, hypertension, diabetes mellitus, any recent upper respiratory tract infection or related medication, alcoholism, or excessive smoking. On the day of the PET study, each subject was screened for nasal congestion. The subjects were instructed to abstain from any oral intake for about 4 h prior to the study and not to use perfume, mouthwash, etc. before the study. All the studies were performed in the morning hours to avoid the high variability in the odor threshold (Lötsch et al. 1997).

All the subjects underwent MRI scanning using a 0.5 Tesla Signa Contour GE/YMS Co. system within a week of the PET study. A T1-weighted spoiled gradient echo (SPGR) sequence was used with repetition time (TR) = 40 ms, echo time (TE) = 7 ms, a flip angle of 30°, voxel size of 1 × 1 × 1.5 mm, and matrix size of 256 × 256 × 128. All had normal brain MRI scans.

PET scanner description and study procedure

The rCBF was determined by PET during the performance of six tasks (2 control and 4 activation tasks) by all of the eight subjects. PET was performed using a Shimadzu SET2400SW Scanner in three-dimensional (3-D) mode with a scatter shield in place and septa retracted (Fujiwara et al. 1997). This PET imaging system has axial and transverse fields of view (FOV) of 200 and 590 mm, respectively. Prior to the emission scans, a transmission scan with an external source of 68Ge 68Ga was obtained.

For each task, 200 MBq H215O was administered via a cannula introduced into the right antecubital vein. Data were obtained from the time the peak count rate was observed in the head region, and data were collected for the next 60 s. For radioactive decay, an interval of 10 min was allowed between two consecutive studies. The subjects had no earplugs and were positioned supine on the PET scanner. Light and auditory disturbances in the room were kept to a minimum.

Task procedures

A pre-PET psychophysical test was carried out on a separate group of 15 subjects to rate the ability to identify, the familiarity, the hedonicity, and the intensity of 50 odors available. Irritating odors were excluded from the PET study (Yousem et al. 1997). Among the remaining odors, one group was found familiar to the general population as these odors were relatively easy to name. This group included odors of fruits, flowers, and edible items. The other group consisted of odors that were unfamiliar, difficult to identify and did not elicit any memories (Table 1). The visual stimuli were chosen among the commonly available edible items, and their identification was not difficult for the subjects (Fig. 1A, Table 2).


                              
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Table 1. Odors presented during the PET tasks



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Fig. 1. Visual stimuli used in the positron emission tomography (PET) tasks. A: example of a picture used in visual naming (VN) task. B: example of the scrambled pictures used.


                              
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Table 2. Visual stimuli presented during the visual naming task (VN)

In each PET task, eight pairs of olfactory and visual stimuli were presented. Each pair consisted of an odor and an image presented for 6 s. The presentation and withdrawal of these stimuli were simultaneous. This was followed by a 4 s interstimulus interval to allow response from the subject. Based on our experience with pre-PET psychophysical study, the duration of odor presentation and withdrawal was tailored to avoid desensitization of olfactory fibers and to allow time for recovery (Hummel et al. 1996). The adequacy of the duration of odor exposure conformed with other studies (Hummel et al. 1997; Royet et al. 1999). Only nose breathing at the normal respiratory rate was allowed; sniffing was not allowed since it is known to induce different activation pattern in the brain (Sobel et al. 1998a). The subject was instructed to fix his gaze on a red cross presented against a white background during the interstimulus interval. Each task was started 20 s prior to the PET scan.

Odor was presented birhinally using a "sniffing stick" (Hummel et al. 1997), which was held 2-3 cm in front of the anterior nares of the subject. Colored two-dimensional visual stimuli were presented on a head-mounted display, (HMD; Eyetrek FMD011F, Olympus, Tokyo), with the signals relayed from a Macintosh PowerBook G3 computer. The visual stimulus was presented binocularly. The size of the picture was set within a 5° visual angle, so that the subject did not have to move his eyes excessively to see the picture in its entire detail. The subjects were also asked to fix their gaze on the red cross presented at the center of the field of view to keep the eye movements to a minimum.

The subjects were subjected to dummy runs with HMD and mock odor presentations prior to the PET study. This was carried out to familiarize them thoroughly with the task design and requirements. Randomization of the tasks across the eight subjects was carried out.

Certain instructions were common to all the tasks including the following: "Breathe through your nose normally. Please do not breathe through your mouth or sniff. Keep your eyes fixed on the red cross at the center of the screen."

Control task 1 (C1) consisted of fixing the eyes on the red cross, with no cognition involved. No olfactory or other visual stimuli were presented. The subjects were instructed as follows: "Do not concentrate on anything in particular."

Control task 2 (C2) consisted of gazing at a scrambled picture (without any specific shape or design) and smelling of an unfamiliar odor. No cognition was involved in this task either. For this task instructions were as follows: "Do not think of anything particular and do not concentrate on the stimuli."

In the ON task, a familiar olfactory stimulus was presented simultaneously with a scrambled picture. These odors were those of common easily recognizable items (Table 1). The subject was instructed to concentrate on the olfactory stimulus only and to name or categorize it within the next 4 s or say "Pass" if he could not. In this task additional instructions were as follows: "Each odor will be presented for the same duration as that of a simultaneously presented scrambled picture. Concentrate on the odor only and try to visualize and name it. Identify the odor after the picture is withdrawn. If unable to answer, please say `Pass'."

In the visual naming task (VN), a picture of an object was presented simultaneously with an unfamiliar olfactory stimulus. The pictures of common edible objects were shown in this task (Fig. 1A, Table 2). The subject was required to concentrate on the picture only and name it within the next 4 s or say "Pass" if he could not. For this task, additional instructions were as follows: "Each picture will be presented simultaneously with an odor. Please concentrate on the picture only and try to name it. Name the picture after it is withdrawn. If unable to answer, please say `Pass'." The answer in the naming tasks was required to be a single word in the Japanese language.

In the OM task, an unfamiliar target odor was presented around 2 min before the actual task. The subject was required to "memorize" it. During the task, the target odor was presented thrice among eight cycles of olfactory-visual pairs. Attention to the simultaneously presented scrambled visual stimulus was not required. For this task additional instructions were as follows: "Each odor will be presented for the same duration as that of a simultaneously presented scrambled picture. Please concentrate on the odor only and try to match it with the target odor sampled a while ago. If the presented and target odors appear the same, say `Yes,' if different say `No,' and if undecided, say `Pass.' Answer when the picture is withdrawn."

In the visual matching task (VM), two scrambled target pictures were presented around 2 min before the actual task (Fig. 1B). The subject was required to memorize their characteristics (colors and design). During the task, the target visual stimuli were presented thrice among eight cycles of olfactory-visual pairs. Attention to the simultaneously presented olfactory stimulus was not required. For this task additional instructions were as follows: "Each picture will be simultaneously presented with an odor. Please concentrate on the picture only and try to match it with target pictures shown a while ago. If the presented and target pictures appear the same, say `Yes,' if different say `No,' and if undecided say `Pass.' Answer when the picture is withdrawn."

In the OM and VM tasks, the expected answer was "yes," "no," or "pass" when the presented stimulus matched the target stimulus, did not match the target stimulus, or when the subject could not decide, respectively. These tasks were the same as previously described delayed matching to sample tasks, requiring short-term memory retrieval (Hummel et al. 1997), and were different from n-back memory tasks (Carlson et al. 1998; Smith and Jonides 1999). In our tasks, the subject was thoroughly familiarized with the target stimulus prior to the actual PET study. In the OM task, only one unfamiliar odor was introduced for short-term memory task. In the pre-PET psychophysical tests it was observed that introducing more than one odor for short-term memory task greatly reduced the percentage correct responses from the subjects. It is known that unlabeled odors are difficult to recognize as compared with those associated with some verbal labels (Jones-Gotman and Zatorre 1993). As the group 2 odors (Table 1) were presumed not to be associated with any specific names, encoding was based purely on the characteristics of the odor itself.

The group 1 odors (Table 1) were exclusively used during ON task and not in other tasks. Similarly, the target odor used in OM task was not included in any other tasks. Due to the limited number of odors available, repetition of certain odors was unavoidable; however, care had been taken to do so only in the VN and VM tasks with different order of presentation.

Image processing and statistical analysis

The raw PET images were corrected for attenuation by using the 68Ge 68Ga transmission data. Each subject's MRI was normalized to the standard brain MRI of the Human Brain Atlas (HBA) (Roland et al. 1994) using affine transformation of Automated Image Registration (AIR) (Woods et al. 1998), followed by application of the 3-D deformation field of the Elastic transformation (Schormann et al. 1996). These parameters were subsequently used to transform each subject's PET image and MRI into the standard brain anatomy. Statistical Parametric Mapping (SPM96, Wellcome Department of Cognitive Neurology, London) (Friston et al. 1995) software was used for smoothing and statistical analyses. A 3-D Gaussian filter of 16 mm was used for image smoothing.

For the same-task analysis, the cognitive contrasts of respective tasks with control conditions were computed by SPM96 (Tables 5 and 6). The masking (P < 0.01) was applied with appropriate contrasts to elimate the voxels that did not survive corrections for multiple comparison (P < 0.05). For the cross-task analysis, the cognitive contrasts of respective olfactory and visual tasks were computed without masking, with the uncorrected extent threshold value set at P < 0.05 (Tables 5 and 6).

For each comparison, voxels with Z values > 3.09 (P < 0.001) were considered to denote regions of significantly increased rCBF. Anatomical localization of the activated areas was performed in relation to the mean reformatted MRI of all the eight subjects, and expressed in the stereotactic space defined by Talairach and Tournoux (1988).


    RESULTS
TOP
ABSTRACT
INTRODUCTION
METHODS
RESULTS
DISCUSSION
REFERENCES

The performance of the subjects during each task is shown in Table 3. The one-way ANOVA test was performed to check the individual performance of the subjects across the tasks, and it was found to be significantly different (F = 7.2, P < 0.001). The ON task was the most difficult, in contrast to the VN, which was the easiest task to perform. In post hoc analysis, the mean subject performance, in terms of the number of correct responses, between these two tasks was significantly different (P = 0.002: paired t-test). The mean subject performance, however, was not significantly different between the OM and VM tasks (P = 0.66: paired t-test).


                              
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Table 3. The task performance of the subjects

The significantly activated areas based on the computed cognitive contrasts are tabulated in Tables 4-6.


                              
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Table 4. Talairach coordinates and Z scores of peak activation for the subject-averaged results for olfactory and visual perception without cognition


                              
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Table 5. Talairach coordinates and Z scores of the peak activation for the subject-averaged results in contrast analysis for olfactory naming and matching


                              
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Table 6. Talairach coordinates and Z scores of the peak activation for the subject-averaged results in contrast analysis for visual naming and matching

The contrast C2-C1 shows the brain regions involved in olfactory and visual perception, without any cognitive effort required. The areas found to be activated were the left middle frontal lobe, the left orbitofrontal lobe, the right piriform cortex, the right lateral occipital lobe, and the left cuneus (Table 4).

The contrast ON-C1 masked by contrast ON-C2 showed significant activation of the left cuneus, the right anterior cingulate gyrus, the left insula, the left anterolateral cerebellum, and the right posteromedial cerebellum (Table 5, Fig. 2). This analysis reveals the areas involved in olfactory naming.



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Fig. 2. Human neural substrates of olfactory naming. A: left cuneus. B: right anterior cingulate gyrus. C: left insula. D: left anterolateral cerebellum. E: right posteromedial cerebellum. The cerebellar activation foci are bilateral and prominent. The slices are displayed according to radiological convention.

In the contrast OM-C1 masked with OM-C2, significant activation was determined in the left cuneus, the left anterolateral cerebellum, and the right posteromedial cerebellum (Table 5, Fig. 3). This analysis reveals the areas activated during olfactory matching.



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Fig. 3. Human neural substrates of olfactory matching. A: left cuneus. B: left anterolateral cerebellum. C: right posteromedial cerebellum. The cerebellar activation foci are bilateral. The slices are displayed according to radiological convention.

In the cross-task contrast analysis of ON-VN, significant activation was determined in the left cuneus (Table 5, Fig. 4). This analysis shows the specific area for olfactory naming. The contrast analysis of OM-VM did not reveal significant activation of any brain area.



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Fig. 4. Human neural substrate specific for olfactory naming as revealed by the cross-task contrast analysis. The left cuneus is shown. The slice is displayed according to radiological convention.

The contrast VN-C1 masked with VN-C2 revealed significant activation in the left orbitofrontal cortex, the left fusiform gyrus, and the right anterior cingulate gyrus (Table 6). The VN task is primarily an object naming task combined with passive odor perception. The above analysis reveals the areas involved in visual naming.

In the contrast VM-C1 masked with VM-C2, significant activation was determined in the right inferior temporal gyrus, the left cuneus, the left anterolateral cerebellum, and the right anterior cingulate gyrus (Table 6). This analysis shows the areas involved in the visual matching.

In the cross-task contrast analysis of VN-ON, significant activation was determined in the right inferior parietal lobule, the left precuneus, the right superior temporal gyrus, the bilateral middle occipital gyri, the left fusiform gyrus, and the left anterior cingulate gyrus (Table 6). This analysis shows the areas activated specifically during visual naming. The contrast analysis of VM-OM did not reveal significant activation in any brain area.

In comparison of results of olfactory and visual matching, it was noted that the occipital activation foci were greater in number during the visual task and also included a part of the lingual gyrus. The cerebellum was bilaterally activated during the olfactory task, while only the right cerebellum was activated during the visual matching. There was a common site of activation in the cerebellum during the olfactory and visual tasks, i.e., the anterolateral part of the cerebellum (Tables 5 and 6).


    DISCUSSION
TOP
ABSTRACT
INTRODUCTION
METHODS
RESULTS
DISCUSSION
REFERENCES

This study highlighted the neural substrates for human olfactory perception, long- and short-term memories. The tasks were designed to analyze simple detection as well as a more comprehensive semantic analysis of the olfactory stimuli. It was noted that semantic analysis of the odors involved brain regions that were not part of the classically defined olfactory regions. The most significant finding of this study was that performing semantic analysis resulted in, among others, the recruitment of the occipital cortex and the cerebellum. Although the activation of classically described olfactory areas during simple olfactory stimulation such as the piriform and orbitofrontal cortices was observed, their role in semantic analysis may not be as important as previously reported.

Task design

In this study, the odors were categorized and selected by a pre-PET psychophysical test. Those odors found to be familiar and relatively easy to recognize by the general population were used in the ON task. The ON task involved recognition of olfactory stimuli, using long-term olfactory memory, and naming. This places a greater semantic load on the brain areas involved. The option of saying "pass" was provided to prevent the factor of guessing. The odors were dispensed with sniffing sticks instead of squeeze bottles. It is known that headspace-air in the bottle produces tactile sensations in the nasal epithelium (Hummel et al. 1997), which was expected to change the true activation patterns.

As shown in Table 3, the mean percentage of correct answers in the ON task is 50.1%. This is in contrast to the 89.3% correct response in the VN task. The difference between the two reflects the degree of task load inherent in the ON and VN tasks. The VN task is easier to perform and involves the naming of common edible items, while the ON task involves some degree of mental imagination as well. In the OM and VM tasks, the percentages of correct responses were 73.9 and 79%, respectively, which show that the semantic load for these tasks is nearly the same.

Our study involved exposure of each subject to at least six olfactory stimuli for each task. The duration of each task was optimal for a standard PET study, i.e., 60 s. The number of olfactory stimuli in each task could have been more, but it was not adopted due to the logistical problems of increasing the scan time and the ever increasing olfactory fatigue. The olfactory stimuli were presented to the subjects manually. Recently the automated synchronized delivery of olfactory stimuli has been used for fMRI imaging (Sobel et al. 1997). Probably the use of an olfactometer for delivery of the olfactory stimuli would have yielded better results.

Brain activations

The C2-C1 contrast analysis revealed these brain regions that are involved in the olfactory and visual perception only. It is important to note that no cognitive effort was required in this task design. The significantly activated regions included the left middle frontal cortex, the left orbitofrontal region, and the right piriform cortex. The piriform cortex is among the primary receiving sites of information from the olfactory bulb. Hence, it is considered to be part of the primary olfactory cortex (Nolte 1993). In a pioneer PET study on olfaction, Zatorre et al. (1992) have shown that simple presentation of odors resulted in bilateral activation of the piriform cortex. There was activation of the right orbitofrontal cortex as well, which was designated as the secondary olfactory cortex by the authors. Their theory was that the unilateral activation of orbitofrontal cortex indicated its functional specialization and that probably it was involved in higher order processing of odor. However, their study design apparently did not involve any cognitive analysis of odors. In our study, the left orbitofrontal cortex was also activated, but again, there was no cognitive effort required by our subjects. The bilateral activation of the occipital cortex in C2-C1 contrast is most likely due to visual perception.

Olfactory tasks

As stated earlier, the main purpose of our study was to analyze the cognitive component of the olfactory processing in the human brain. To our knowledge, there are few PET studies reported that elaborate the neural circuits involved in olfactory semantic processing and cognition by the human brain. Moreover, the main thrust of fMRI studies has been on olfactory perception.

The brain areas activated during olfactory naming were located in the left cuneus, the right anterior cingulate gyrus, the left insula, the left anterolateral cerebellum, and the right posteromedial cerebellum (Table 5, Fig. 2). During olfactory naming, the piriform and orbitofrontal cortices were not activated. The orbitofrontal cortex was considered to be the area where olfactory discrimination and identification occurs (Zatorre and Jones-Gotman 1991). However, our data do not support this concept. Our findings are partially supported by another olfactory study in which comestibility judgment about odors did not evoke activation of the orbitofrontal cortex (Royet et al. 1999).

In our study, another interesting finding was the left insular activation. Bilateral insular activation during olfactory stimulation was also reported by Zatorre et al. (1992). Sobel et al. (1997) also reported the activation of the peri-insular areas during olfactory tasks. This activation was observed even when there was no conscious perception of odor by the subject (Sobel et al. 1999). In our study, it is noteworthy that insular activity has been localized in the left side only during olfactory naming. This activation was not noted during olfactory matching, indicating that the insula has some role in olfactory naming. The insula is also involved in gustatory tasks (Cerf et al. 1998). The authors showed that the lower part of the insula showed strong lateralization related to handedness. In our institute, a previous PET study revealed significant left insular activation during a taste discrimination task. In that study, Kinomura et al. (1994) suggested that taste and smell modalities converge in a specific area in the insular cortex. In our study, breathing through the mouth was strictly prohibited so that the taste buds were not activated during any of the various tasks. The insular activation observed in our olfactory study seems to lend support to their hypothesis. The lateralization of the insula to the left side of the brain in our study is probably because all our subjects were strongly right-handed with left hemisphere dominance.

The area found specific for olfactory naming was the left cuneus (Table 5, Fig. 4). This provides further credence to the role of the occipital lobe in olfactory memory. The activation of the left cuneus during olfactory naming indicates that this task may involve some imagination to ascertain the origin of the odor. It has been observed in many cognitive neuroimaging studies that the visual cortex is involved in mental imagery tasks (Kosslyn 1988; Royet et al. 1999).

The brain regions involved in olfactory matching were the left cuneus and bilateral cerebellar regions (Table 5, Fig. 3). In this task, the odors presented were unfamiliar to the general population. It was assumed that because these odors do not carry any labels, i.e., these are not associated with any particular object, so these can be used for study of olfactory short-term memory. No area was specifically activated during olfactory matching as determined by cross-task analysis.

It is interesting to note that the left cuneus and bilateral cerebellar activation was noted during the OM and ON tasks. One factor common to both tasks was the requirement of cognitive effort for olfactory assessment. Although the C2-C1 contrast also involved olfactory perception, no cognitive effort was required. As mentioned above, no cerebellar activation was seen in C2-C1 contrast analysis.

In our study, the most significant finding was the presence of cerebellar activation during the olfactory tasks. As reported by Yousem et al. (1997), cerebellar activation during the olfactory tasks has been noted in PET studies, but we were not able to find any detailed published reports. There was one preliminary study report by Savic et al. (1999) in which right cerebellar activation was noted during the olfactory tasks, but its importance was not discussed. In the most recent PET studies, Dade et al. (1998) and Royet et al. (1999) have not found nor discussed the cerebellar role in olfaction.

Our data reveal bilateral cerebellar activation during the olfactory tasks and unilateral activation during the visual matching (Tables 5 and 6). The number of activated sites is greater, as shown both in terms of voxel numbers and the cluster size, during the olfactory tasks as compared with visual task. Over the last two centuries, the role of the cerebellum had been mainly thought to be as a controller of posture and movement. In fact, previous historical studies by Babinski and Holmes et al. in the early part of the century suggested that the cerebellum is not involved in any sort of cognitive function (Thach 1996). Isolated cerebellar lesions lead to motor deficits, but the cognitive and sensory domains remain unaffected. On the other hand, many diseases that affect the CNS including the cerebellum, e.g., Parkinsonism, Alzheimer's disease, and Multiple sclerosis, are characterized by olfactory dysfunction as well (Doty 1997). Recent research has indicated that the cerebellum also has a role in nonmotor and learning behavior (Allen et al. 1997). Cerebellar activations have been noted in sensory acquisition and discrimination tasks (Gao et al. 1996; Lejeune et al. 1997). The neocerebellum, which includes the entire posterior cerebellar lobe, except for the uvula and pyramid, was thought to control fine motor voluntary movements. A review of literature on the cerebellar involvement in cognitive tasks has revealed that the neocerebellum is also activated during movement imagination (Jueptner et al. 1997). Kim et al. (1999) reported that the cerebellum may play a role in recognition processes as well.

Recent fMRI studies have also shown the cerebellar activation during olfactory tasks (Sobel et al. 1998b; Yousem et al. 1997). Sobel et al. (1998b) concluded that cerebellar activation of the anterior medial region resulted from active sniffing, regardless of presence of odor, while posterolateral activation occurred in the mere presence of odor.

In our study, sniffing was not allowed, and the subjects did not perform any motor movements. In the C2-C1 analysis where no cognition was involved, no cerebellar involvement was observed (Table 4). However, widespread activation of different parts of the cerebellum was noted during the olfactory naming and matching analyses, i.e., where cognitive or semantic analysis was involved. The cognitive analysis for olfactory naming and matching revealed activation foci in the right posteromedial and the left anterolateral portions of the cerebellum (Table 5, Figs. 2 and 3), while the visual matching activated the left anterolateral part of the cerebellum (Table 6). Among the ON, VN, and VM tasks, the common cognitive features are semantic analysis and speech. It has been reported that the phonological retrieval of names of visually presented stimuli activates, among others, the midline cerebellum (Price and Friston 1997). This cerebellar area is different from the one activated in our study. In our data, the right posteromedial cerebellum was also activated during visual naming, but this activation did not survive the correction for multiple comparisons. Whereas Sobel et al. (1998b) concluded that the cerebellum is involved in olfaction in humans, our results indicate that this involvement is at the level of cognition as well. It is known that the cerebellum is connected to the frontal, parietal, temporal, and occipital lobes via the efferent fibers (Snell 1992). Our data demonstrates that the cortico-cerebellar connections are also part of the extensive neural network used for odor memory.

There is evidence that the cerebellum has a certain role in memory retrieval without requiring any sensory or motor input or output (Andreasen et al. 1999). The authors were able to find robust bilateral cerebellar activations, more on the right side, during silent recall of a consciously retrieved episodic memory. Contents of the retrieved memory have not been mentioned in the paper. It was interesting to note that the sites of the cerebellar activations during our olfactory tasks were almost the same as those noted by Andreasen et al. (1999). In our tasks, the retrieval of short- and long-term memories was required, and it may be possible that the neural circuits involved in the conscious episodic memory retrieval, as described by the authors, were also involved in our tasks. The next point to be explored is whether these circuits are specific for olfactory tasks only or a part of the general conscious memory recall. This question, probably, cannot be answered based on our data and needs to be further studied.

In our olfactory tasks, we could not find any activation of the medial temporal region. This included the long- as well as short-term memory storage. The role of the medial temporal lobe in a variety of memory encoding and retrieval tasks had been discussed in detail (Schacter and Wagner 1999; Schacter et al. 1999; Zola et al. 2000). A closer investigation showed no olfactory memory study among them. The medial temporal lobe contains structures that receive fibers directly from the olfactory bulb (Squire and Zola 1996). The odor and taste memory has been studied by fMRI (Levy et al. 1999a,b). In these studies, the actual stimulus or its imagination resulted in activation of the temporal regions. These included the inferior temporal region as well as parahippocampal areas. However, in these studies scanning was limited to the temporal areas only, not covering the entire brain. Attention was specifically focused on the temporal lobe based on a priori hypothesis of its importance in memory retrieval (Levy et al. 1997). In the limited number of PET studies available in literature, concerning the olfactory memory and semantic analysis, a lack of temporal lobe activation was noted (Dade et al. 1998; Royet et al. 1999). Royet et al. (1999) have attributed this absence of temporal lobe activation to the lack of a true nonstimulated baseline in their task design. In our task design a true nonstimulated baseline condition (the C1 task) was included for better comparison among the activation tasks. Still, we were not able to find any temporal lobe activation during the odor naming and matching tasks. On the other hand, in visual naming and matching, medial temporal/parahippocampal activations with high Z values were noted, although these did not survive the correction for multiple comparisons (data not shown).

No prefrontal cortex activation was detected in any of our olfactory or visual analyses. The review of literature shows that prefrontal activity is associated with memory encoding, recognition, memory retrieval effort and memory retrieval (Henson et al. 1999). Klinberg and Roland (1998) have shown a lack of right prefrontal activity during nonverbal memory retrieval task. They attributed it to the near-perfect performance (<5% error rate, low reaction time ~0.9 s) of their subjects during the retrieval task. In their opinion, the fast and accurate long-term memory retrieval may result in prefrontal cortex nonactivation. In our study, the cognitive effort required during the naming and matching tasks was expected. We did not take note of the reaction time of our subjects precisely, and the correct responses were quite low, specially during the olfactory naming task. Therefore in our case, the absence of prefrontal activity cannot be attributed to such automatic responses. One plausible explanation may be the low frequency of stimuli presentation in our task. Our study consisted of six stimuli presentations for each task. A greater number of stimuli presentations in each task would probably reveal the rCBF changes in the prefrontal area.

Activation of the anterior cingulate gyrus has been noted in many olfaction-related studies (Kiyosawa et al. 1996; Levy et al. 1998a; Royet et al. 1999; Sobel et al. 1997, 1999). In a PET study by Posner et al. (1988), the rCBF of the anterior cingulate gyrus was found to increase when the number of targets were increased. Various tasks, evoking anterior cingulate activation have a few common characteristics: either those were willful, nonautomatic, attention-seeking, or unique (Lenz et al. 1998; Pardo et al. 1990; Posner 1990). Murtha et al. (1996) have shown that simple anticipation during a PET study causes an increase in the rCBF to the anterior cingulate cortex. In our study, an increase in the rCBF to the right anterior cingulate gyrus is observed in olfactory- as well as visual-stimulus analyses. This may be attributable to simple anticipation. The presence of the right anterior cingulate gyrus activation in all these analyses was expected as the tasks required a high degree of attention.

Visual tasks

The main emphasis of our study was to map the olfactory processing regions. The visual processing tasks were included in the study design to provide a comparison with the olfactory studies. The results of the visual processing are discussed here briefly.

The brain areas activated during visual naming were in the left orbitofrontal cortex, the left fusiform gyrus, and the right anterior cingulate gyrus. The cross-task analysis of contrast VN-ON reveal the areas specific for visual naming (Table 6). These significantly activated areas include, among others, the left fusiform gyrus and the bilateral middle occipital gyri. A number of functional imaging studies have investigated the brain regions associated with visual naming tasks (Gorno Tempini et al. 1998; Kim et al. 1999; Kiyosawa et al. 1996; Petersen et al. 1990; Vandenberghe et al. 1996). Our results of visual naming are congruent with those of previously reported studies.

In human studies the role of the fusiform gyrus in visual recognition has been well established. Within the fusiform cortex, the area specific for face perception has been outlined (Gorno Tempini et al. 1998; Haxby et al. 1994; Malaspina et al. 1998). The object recognition area lies immediately medial to this face recognition area (Kanwisher et al. 1997).

The orbitofrontal cortex has been designated as the secondary olfactory cortex (Zatorre et al. 1992), but it appears to be a multimodal area as its activation was seen in our visual naming analysis. Animal studies have shown that the orbitofrontal cortex receives input from the visual, gustatory, and olfactory modalities. It contains uni- as well as bimodal neurons that respond to one or more than one modality (Rolls and Baylis 1994).

In our study, visual matching involved activation of the right inferior temporal gyrus, the left cuneus, the left anterolateral cerebellum, and the right anterior cingulate gyrus. No area specific for visual matching was found in cross-task analysis.

In our study, the visual matching stimuli were software generated abstract pictures (Fig. 1B). The bilateral primary visual cortices and the surrounding areas were found to be active during long-term memory retrieval of colored nonfigurative pictures (Klinberg and Roland 1998). The left inferotemporal cortex has been found to be involved in visual pattern discrimination tasks as well (Kawashima et al. 1998; Kikuchi and Iwai 1980; Roland and Guylás 1995).


    ACKNOWLEDGMENTS

This research was supported by Grants-in-Aid for Scientific Research on priority from the Japanese Ministry of Education, Science, Sports and Culture (1145204, 09207102), from Research for the Future from the Japan Society for the Promotion of Science (JSPS-RFTF 97L00202), and from Comprehensive Research on Aging and Health from the Ministry of Health and Welfare, Japan. The supporting grant for the ELAST Transformation Software development (SFB 194/A6) was provided by BIOTECH Program, EC.


    FOOTNOTES

Address for reprint requests: A. Qureshy, Dept. of Nuclear Medicine and Radiology, Institute of Development, Aging and Cancer, Tohoku University, 4-1 Seiryomachi, Aoba Ku, Sendai 980-8575, Japan (E-mail: ahmad{at}idac.tohoku.ac.jp).

The costs of publication of this article were defrayed in part by the payment of page charges. The article must therefore be hereby marked "advertisement" in accordance with 18 U.S.C. Section 1734 solely to indicate this fact.

Received 4 October 1999; accepted in final form 17 April 2000.


    REFERENCES
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ABSTRACT
INTRODUCTION
METHODS
RESULTS
DISCUSSION
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