Response of Anterior Parietal Cortex to Different Modes of Same-Site Skin Stimulation

M. Tommerdahl1, K. A. Delemos2, O. V. Favorov1, C. B. Metz2, C. J. Vierck Jr.3, and B. L. Whitsel1, 2

1 Department of Biomedical Engineering and 2 Department of Cell and Molecular Physiology, University of North Carolina at Chapel Hill, Chapel Hill, North Carolina 27599; and 3 Department of Neuroscience, University of Florida, Gainesville, FL 32610

    ABSTRACT
Abstract
Introduction
Methods
Results
Discussion
References

Tommerdahl, M., K. A. Delemos, O. V. Favorov, C. B. Metz, C. J. Viereck, Jr., and B. L. Whitsel. Response of anterior parietal cortex to different modes of same-site skin stimulation. J. Neurophysiol. 80: 3272-3283, 1998. Intrinsic optical signal (IOS) imaging was used to study responses of the anterior parietal cortical hindlimb region (1 subject) and forelimb region (3 subjects) to repetitive skin stimulation. Subjects were four squirrel monkeys anesthetized with a halothane/nitrous oxide/oxygen gas mixtures. Cutaneous flutter of 25 Hz evoked a reflectance decrease in the sectors of cytoarchitectonic areas 3b and/or 1 that receive input from the stimulated skin site. The intrinsic signal evoked by 25-Hz flutter attained maximal intensity <= 2.5-3.5 s after stimulus onset, remained well maintained as long as stimulation was continued, and disappeared rapidly (usually <= 2-5 s) after stimulus termination. Repetitive skin heating stimuli were delivered via a probe/thermode in stationary contact with the skin (6 temperature ramps/trial; within-trial ramp frequency 0.42 Hz; intertrial interval 180 s; initial temperature 32-36°C; maximal temperature 48-52°C; rate of temperature change 19°C/s). Skin heating led to a large-amplitude reflectance decrease within a zone of area 3a, which neighbored the region in areas 3b/1 that emitted an intrinsic signal in response to same-site 25-Hz flutter in the same subject. In three of four subjects a lower-amplitude decrease in reflectance also occurred in a region of area 4 continuous with the area 3a region that responded maximally to same-site skin heating. The reflectance decrease evoked in areas 3a/4 by skin heating consistently exceeded in both intensity and spatial extent the decrease in reflectance evoked in areas 3b/1 by same-site 25-Hz cutaneous flutter. These findings are viewed as consistent with the proposal that area 3a plays a leading role in the anterior parietal cortical processing of the afferent drive evoked by skin-heating stimuli perceived as painful. In all four subjects the reflectance decrease evoked in areas 3a/4 by skin heating was accompanied by a simultaneous but opposite change in reflectance (a reflectance increase) within a large territory located immediately posterior to the regions that responded with a decrease in reflectance---an observation that raised the possibility that skin heating evoked opposing influences on the activity of area 3a and 3b/1 regions that receive input from the stimulated skin site. This was evaluated with the method of correlation mapping. The observations obtained with correlation mapping appear consistent with demonstrations by others that skin-heating stimuli perceived as painful by conscious subjects suppress/inhibit the anterior parietal response to innocuous mechanical skin stimulation. The opposing (relative to the response of area 3a) optical response of area 1 and/or area 3b during skin heating stimulation is attributed to suppression/inhibition of area 1 and/or area 3b neuron activity.

    INTRODUCTION
Abstract
Introduction
Methods
Results
Discussion
References

Different views of the contributions of the anterior parietal (SI) cortex to nociception and pain were advanced in recent literature. One possibility receiving considerable current attention---a possibility raised by the findings reported in human positron emission tomography (PET) and functional magnetic resonance imaging (fMRI) studies---is that regions other than SI (e.g., anterior cingulate, rostral insular, and SII cortex) play the leading role in processing the sustained noxious afferent drive characteristic of clinical pain states (Jones et al. 1991; Talbot et al. 1991; Apkarian 1992, 1994 1995; Casey et al. 1994; Coghill et al. 1994; Derbyshire et al. 1994; Jones and Derbyshire 1994; Rainville et al. 1997). Although an alternative possibility was considered---that multiple, functionally interdependent cortical areas contribute to pain encoding (Rainville et al. 1997)---experimental evidence for the cortical interareal interactions required to mediate such interdependencies is lacking. It also remains unclear what it is that interdependencies between cerebral cortical areas might contribute to the processing of information about stimuli perceived as painful.

We sought to advance current understanding of the contributions of SI to the cerebral cortical processing of afferent drive evoked by noxious skin heating. The experiments were motivated by the possibility that anterior parietal cortex may respond differentially to noxious skin heating (i.e., with area 3a and simultaneous suppression/inhibition of area 3b and/or area 1) (Tommerdahl et al. 1996). Examination of this possibility seemed timely because information can now be obtained about the SI response to skin heating at a level of spatial and temporal resolution substantially exceeding that obtainable with the methods and approaches that have been used to date to image anterior parietal cortex in humans (Apkarian 1995; Apkarian et al. 1992; Bushnell et al. 1995; Casey et al. 1994; Coghill et al. 1994; Derbyshire et al. 1994, 1995; Jones et al. 1991; Jones and Derbyshire 1994; Rainville et al. 1997) or in nonhuman primates (Tommerdahl et al. 1996).

To this end, the time course of the global SI response to skin heating and innocuous mechanical stimulation of the same skin site in squirrel monkeys was investigated with time-resolved intrinsic optical signal (IOS) imaging. The IOS imaging data were analyzed with an approach (correlation mapping) designed to detect and characterize statistically significant time-dependent interactions between the different cytoarchitectural areas (3a, 3b, 1, and 2) that receive and process the afferent drive evoked by natural mechanical and thermal skin stimuli.

    METHODS
Abstract
Introduction
Methods
Results
Discussion
References

Subjects and preparation

Adult squirrel monkeys (Saimiri sciureus; males and females; n = 4) were studied because in this species one can directly visualize most of the anterior parietal region known to receive direct input from thalamic regions that process input arising in cutaneous receptors located in the contralateral hand or foot. A second advantage derives from the fact that areas 3b and 1 in squirrel monkey were extensively investigated with both microelectrode RF mapping (Sur et al. 1982) as well as combined optical imaging and single unit recording experiments (Tommerdahl and Whitsel 1995; Tommerdahl et al. 1996). As a result, a wealth of information exists about the responses of SI in these animals to both nonnoxious mechanical and skin heating stimulation and about the sensitivity of those responses to general anesthetics.

Preparation involved 1) induction of general anesthesia, 2) insertion of a soft tube into the trachea and polyethylene cannula into the femoral vein, 3) creation of a 1 × 1 cm opening in the skull overlying the hindlimb or forelimb region of anterior parietal cortex, 4) installation with dental acrylic of a recording chamber over the skull opening, and 5) exposure of the appropriate region of SI by incision and removal of the dura. All surgical procedures were carried out under deep general anesthesia (1-4% halothane in a 50/50 mixture of oxygen and nitrous oxide), and the skin and muscle tissues within the surgical field on the head and in the vicinity of the tracheal cannula and venous catheter were infiltrated with a long-lasting local anesthetic, closed with sutures, and bandaged. Subjects were immobilized with Norcuron and ventilated with a gas mixture (a 50/50 mix of oxygen and nitrous oxide; supplemented with 0.1-1.0% halothane) with a positive pressure respirator. Respirator rate and volume were adjusted to maintain end-tidal CO2 between 2.5-4.0%; electroencephalogram (EEG) and autonomic signs (i.e., EEG slow wave content; heart rate and pupillary status) were monitored and titrated by adjustments in the halothane concentration in the anesthetic gas mixture to maintain values consistent with light general anesthesia. Rectal temperature was monitored and maintained at 37.5°C with a heating pad.

Euthanasia was accomplished by intravenous injection of pentobarbital (45 mg/kg), and intracardial perfusion with saline followed by fixative (10% formalin). Fiducial marks were placed to guide removal, blocking, and subsequent histological sectioning of the region of cortex studied. All procedures involving animal subjects were reviewed and approved in advance by an institutional committee and are in full compliance with current NIH policy on animal welfare.

Stimuli and stimulus protocols

All stimuli were applied to either the contralateral volar hand or foot. Sinusoidal vertical skin displacement stimuli were applied with a servocontrolled transducer (Cantek Enterprises, Canonsburg, PA). The vibrotactile stimulator made contact with the skin via a 2-mm diameter plastic probe, which, in the absence of stimulation, indented the skin 500 µm. Skin-heating stimuli were applied with a cylindrical probe that carried a 12.5-mm diameter Peltier-type device (thermode) mounted to its end. The temperature of the thermode was regulated by a computer-interfaced controller. The controller allowed the temperature of the thermode to be changed rapidly (19°C/s), accurately, and in a highly reproducible temporal pattern under digital computer control. With the thermode held at 32-36°C, the probe was advanced with a micrometer to indent the skin by 500 µm; the probe remained in this position throughout the period during which observations on the effects of skin heating stimuli were collected. Thermode temperature was displayed and monitored throughout each experiment. Only protocols of skin-heating stimulation were employed that evoked sensations that could be tolerated by the investigators (protocols involving increases in skin temperature that did not elicit escape) and did not cause skin damage.

IOS imaging

Near-infrared (IR; 833 nm) IOS imaging was carried out with an oil-filled chamber capped with an optical window (Tommerdahl and Whitsel 1995; Tommerdahl et al. 1996). Both modes of skin stimulation were delivered to the same skin site on either the hindlimb (1 subject) or forelimb (3 subjects). A stimulus "trial" consisted of one or more applications of a preselected condition of vibrotactile or skin heating stimulation. The IOS was recorded 200 ms before each trial (prestimulus image) and every 1.2-3.0 s after trial onset (poststimulus image) until 15-30 s after the last stimulus in the trial. Thus, for each mode of stimulation, a temporal series of poststimulus images was acquired. An average poststimulus-prestimulus difference image was then generated for each time an image was acquired during a trial---providing a temporal series of average difference images ("time-resolved" IOS imaging) (see Villringer and Chance 1997).

Averaged poststimulus-prestimulus difference images typically show regions of both increased absorption (decreased reflectance) and decreased light absorption (increased reflectance), which are accompanied by increases and decreases in neuronal activation, respectively (e.g., Grinvald 1985; Grinvald et al. 1991a, 1994; Tommerdahl and Whitsel 1995; for recent comprehensive review see Ebner and Chen 1995). Thus the dark regions shown in the difference images (e.g., Fig. 1) are interpreted as regions of increased neuronal activity, and light regions are interpreted as regions of decreased neuronal activity. Difference images of the anterior parietal response to stimulation of the same skin site with nonnoxious mechanical (25-Hz cutaneous flutter) and skin-heating stimulation (temperature ramps) were obtained in the same experimental run, with trials belonging to the two different stimulus modes interleaved (alternated) in the same run to control for across-stimulus changes and/or for within-run changes in cortical "state" unrelated to stimulus conditions.


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FIG. 1. Temporal evolution of the response of anterior parietal hindlimb region to 25 Hz flutter (VT) and skin heating (48°) stimulation of the same site (filled circle on drawing of foot at bottom right) on volar surface of the contralateral foot. All images were obtained from the same hemisphere in the same subject. Anatomical orientation of each panel indicated by axes shown at bottom center. L, lateral; M, medial; A, anterior; P, posterior. Scale bar, 2 mm. Pixel values were scaled between the highest and lowest pixel values present in the entire temporal series of averaged difference images for a given stimulus mode. Top row: averaged difference images obtained during continuous cutaneous flutter stimulation (25 Hz, 400 µm, 8-s duration). Difference images were generated by subtracting the prestimulus image (collected 200 ms before trial onset) from each image collected after trial onset. Numbers above panels indicate time (number of seconds after trial onset) at which images were collected. Middle row: averaged (across trials) difference images obtained during noxious skin heating. A thermal ramp was delivered (32-48-32°C; ramp duration 1.2 s) every 2.4 s for 6 s, the first ramp started at time 0. Consequently, the second difference image (at 2.4 s) was obtained by computing the difference between the reference image (collected at -200 ms) and the image acquired at 2.4 s after initiation of the first ramp. Note that the largest increase in cortical activation is at 6.3 s. Bottom row: image at left shows cortical surface vasculature and central sulcus. Second panel from left shows locations of cytoarchitectural boundaries. Panel at bottom right shows the relative nonoverlap of the anterior parietal regions activated by the flutter (gray stippling) and skin heating (black stippling) stimuli. Panel at bottom right was obtained by thresholding the images of the response to flutter and skin heating. The threshold was set at 90%; only the pixels whose reflectance values were in the upper 10% of the values observed under each stimulus condition are shown.

The decision to employ IOS imaging in this study was motivated in part by the fact that, of the variety of currently available recording methods, only optical imaging methods allow one to detect and quantitatively characterize at relatively high temporal and spatial resolution the spatially distributed cortical response to natural sensory stimulation. The decision to employ IOS imaging rather than imaging with a voltage-sensitive dye was based on the published demonstrations that the former can be utilized repeatedly and over prolonged time periods (Grinvald 1985; Grinvald et al. 1991a,b, 1994; Lieke et al. 1989; Narayan et al. 1994) without substantial negative impact on cerebral cortical function. An important property of the intrinsic signal detected in the IR range is that it is relatively independent of changes in blood flow (Haglund et al. 1993). In addition, the IOS 1) reflects a variety of factors, but most significantly changes in the volume of the extracellular fluid compartment attributable to stimulus-evoked changes in extracellular [K+] and/or neurotransmitter release (Cohen 1973; Lieke et al. 1989; MacVicar and Hochman 1991); 2) is attenuated in a dose-dependent manner by cortical alpha -amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid (AMPA) and N-methyl-D-aspartate (NMDA) receptor block (Haglund et al. 1992, 1993; MacVicar and Hockman 1991); 3) arises chiefly from activity in the dendrites of cortical pyramidal neurons (Grinvald et al. 1994); and 4) is not a direct reflection of either neuronal spike discharge activity or more generally the local cortical voltage changes evoked by a sensory stimulus---the IOS has a much slower onset and decay than the neuroelectrical responses of single neurons or neuronal populations, and it can be severely attenuated by drugs, e.g., by NMDA receptor blocker, that do not substantially modify the sensory cortical neuron spike discharge activity evoked by direct, short-latency thalamocortical drive (Tommerdahl and Whitsel 1995).

A correlation map was generated for each stimulus condition for each subject; it was made by choosing a reference region within the imaged field and computing the intensity correlation rij between the reflectance value of each pixel (i, j) and the average reflectance value within the reference region over time after stimulus onset. The region selected as the reference was the region defined by the boxel (1 × 1 mm2 area) for which the average reflectance underwent the largest decrease in response to stimulation. Each pixel (i, j) on the correlation map is represented by a correlation value rij (-1 <r <1; -1 indicates negative correlation; +1 indicates positive correlation). The statistical significance of each of the correlations was tested with the standard t-test.

Histological procedures and method for relating cytoarchitectural boundaries to functional observations

Blocks of the regions of cerebral cortex studied with the IOS imaging method were prepared subsequent to euthanasia and intracardial perfusion. The blocks then were postfixed, cryoprotected, frozen, and sectioned serially in the sagittal plane at 30 µm. Sections were stained with cresyl fast violet and inspected microscopically to distinguish anterior parietal regions on the basis of established cytoarchitectonic criteria (Powell and Mountcastle 1959a; see also Jones and Porter 1980).

The boundaries between adjacent anterior parietal cytoarchitectonic areas were identified by microscopic inspection of sections obtained at regular intervals (at least once per 500 µm) within the region of cortex studied, and a drawing of the boundaries identified in each section was prepared with a microscope and drawing tube. For each experiment the drawings were then used to generate a reconstruction of the boundaries of the cytoarchitectonic areas within the region from which IOS images were obtained. As the final step, the reconstruction of cytoarchitectonic boundaries was transferred to the images of the stimulus-evoked intrinsic signal obtained from the same subject, with fiducial marks as well as morphological landmarks (e.g., blood vessels and sulci evident both in optical images and in histological sections). To ensure that determinations of the locations of cytoarchitectonic boundaries within the imaged cortical region were uninfluenced by information about the location of intrinsic signal, the identity of the experiment was concealed from the two investigators (BW and OF), who determined the locus of the boundaries, and the relationship between cortical cytoarchitecture and the stimulus-evoked IR reflectance patterns for a subject was not evaluated until those investigators completed the map of cytoarchitectonic boundaries for that subject.

    RESULTS
Abstract
Introduction
Methods
Results
Discussion
References

Characteristics of the responses to different modes of same-site skin stimulation

SI HINDLIMB REGION. The two temporal series of average difference images shown in the top and middle rows of Fig. 1 were obtained from the anterior parietal hindlimb region contralateral to stimuli delivered to a site on a subject's right foot. The panels in the top row show the distribution of changes in reflectance evoked at different times after onset of 25-Hz flutter stimulation (time, in seconds after stimulus onset, is indicated by the number over each panel). The parameters of the 25-Hz flutter stimulus used were 400 µm peak-to-peak amplitude; duration 7 s; probe diameter 2 mm; intertrial interval 60 s. In the absence of sinusoidal motion the probe indented the skin by 0.5 mm. In contrast, the images in the panels in the middle row of Fig. 1 were obtained from the same subject with skin-heating stimulation (32-48-32°C ramps; 1 ramp/2.4 s; 6 ramps per trial; intertrial interval 180 s). A stationary contact thermode advanced 0.5 mm into the skin was used to deliver the skin-heating stimuli to the same site studied with 25-Hz flutter stimulation.

Clearly (see Fig. 1), the responses of the SI hindlimb region to the 25-Hz flutter and skin-heating stimuli have different temporal and spatial characteristics. First, the response to flutter became evident and attained maximal intensity earlier than the response evoked by skin heating. Second, whereas the response to skin heating increased slowly and progressively in both intensity (i.e., exhibited "slow temporal summation") and spatial extent (i.e., exhibited "spatial radiation") during the series of six skin heating ramps delivered in each trial, the spatial extent of the response of 25-Hz cutaneous flutter remained relatively constant during the 7-s period during which the flutter stimulus was applied (e.g., note that the responses obtained at 2.4 and at 6.3 s after trial onset are virtually identical). Third, the responses to flutter and skin heating occupy very different sites within the anterior parietal hindlimb region. At all times after trial onset the signal evoked by flutter remained localized within area 3b (see top images in Fig. 1 obtained at 2.4-6.3 s), whereas at all times after the relatively delayed appearance of the response to skin heating the most intense part of the signal occupied area 3a (see images in middle row of Fig. 1 between 3.7-6.3 s).

Unfortunately, a substantial part of area 3a in the subject that provided the data in Fig. 1 was not in exposed cortex but occupied the depths of the anterior bank of the central sulcus (see cortical surface and map of cytoarchitectonic boundaries in Fig. 1, bottom left panels). As a result, the full spatial extent of the area 3a response to skin heating in this subject could not be ascertained. Despite this limitation, the data illustrated in Fig. 1 clearly suggest (see threshold responses evoked by both modes of stimulation in panel at bottom right) that the responses evoked by same-site 25-Hz flutter or noxious skin heating occupy essentially nonoverlapping territories within the part of the SI hindlimb cortex that was accessible for study with the IOS imaging method.

SI FORELIMB REGION. The temporal series of average difference images in Fig. 2 allow direct comparison of the spatial and temporal attributes of the responses evoked in the anterior parietal forelimb region of a squirrel monkey by 25-Hz flutter and by skin-heating stimulation of the same site on the volar hand (the layout of Fig. 2 is identical to that of Fig. 1; see legend to Fig. 2 for additional details about the stimulus conditions that were used). As was the case for responses recorded from the hindlimb region, the signals evoked in the contralateral anterior parietal forelimb region by same-site 25-Hz flutter and by skin-heating stimulation exhibited very different spatiotemporal characteristics. That is, with 25-Hz flutter stimulation a reflectance decrease appeared early and attained maximal intensity relatively quickly (within 2.6-4.0 s after stimulus onset), and at all the times at which imaging data were obtained after onset of flutter the reflectance decrease occupied a spatially restricted region that included neighboring parts of areas 3b and 1 (see images in Fig. 2, top row).


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FIG. 2. Temporal evolution of the response of anterior parietal forelimb region to 25-Hz flutter (images in top row) and to noxious heating (48°; images in middle row) of the same site (filled circle on figurine at bottom right) on palmar surface of the contralateral hand. All images were obtained from the same squirrel monkey subject. Figure format same as Fig. 1.

In contrast to the temporal properties of the response to cutaneous flutter, skin-heating stimulation (see images in Fig. 2, middle row) evoked a reflectance decrease in the forelimb region of areas 3a and 4 that did not become evident until 3.4 s after trial onset, and from that point until the end of the trial (6 skin-heating ramps were applied during each trial at a frequency of 0.42 Hz) both the intensity and spatial extent of the stimulus-evoked signal increased progressively. Similar results were obtained from the other two subjects in whom the SI forelimb region was studied in the same way.

Time course of stimulus-evoked activity in areas 3b/1 and area 3a

The available difference images in Figs. 1 and 2 clearly show that the time course and cytoarchitectonic location of the anterior parietal reflectance decrease---the dark region(s) in each image in Figs. 1 and 2---evoked by skin heating are very different from those evoked by cutaneous flutter. It is far more difficult, however, to discern the effects, if any, of skin stimulation on the reflectance of regions outside the zones of maximal reflectance decrease. For this purpose an approach to quantification of the imaging data was developed.

Figure 3 identifies the approach and illustrates how it was used to quantify the temporal evolution of the anterior parietal responses to cutaneous flutter and skin-heating stimulation, with the imaging data obtained from the experiment shown in Fig. 2. First, an average difference image was generated from the data acquired at the time after stimulus onset when the response to each condition of skin stimulation was maximal; the images in Fig. 3, top, second and third images from left, were generated in this way. Second, the 1 × 1 mm field (boxel) that exhibited the largest intrinsic signal was identified in each image generated in step 1 (the rectangles labeled 3a and 3b in Fig. 3, top right, show the locations of the two boxels). The 1 × 1 mm boxel in which the reflectance decrease was largest during 25-Hz flutter stimulation was in area 3b, and the boxel in which the reflectance decrease was largest during skin heating stimulation was in area 3a. Third, the average pixel value within each of the two 1 × 1 mm boxels was determined for each of the temporal series of difference images generated from the experimental data. Fourth, the series of values obtained in the manner indicated in the preceding step was plotted as a function of time after stimulus onset to reveal the temporal evolution of the response evoked within each boxel by each mode of skin stimulation. The temporal evolutions of area 3b and area 3a responses to 25-Hz flutter and skin-heating stimulation revealed by use of this four-step approach are shown by the plots in Fig. 3, middle and bottom. During 25-Hz cutaneous flutter stimulation (Fig. 3, bottom plot) average reflectance decreased rapidly within the area 3b boxel (the 1 × 1 mm region that responded maximally to flutter), whereas average reflectance simultaneously underwent a more modest decrease within the area 3a boxel (the 1 × 1 mm region that responded maximally to skin heating). A very different pattern of anterior parietal reflectance change occurred in response to skin heating, i.e., average reflectance in the area 3a boxel decreased, and simultaneously it increased in the area 3b boxel.


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FIG. 3. Approach to quantification of intrinsic signals evoked in the forelimb region of anterior parietal cortex of a squirrel monkey by same-site 25-Hz flutter and noxious skin heating. Plots were generated with the same intrinsic optical signal data used to generate the difference images of Fig. 2. Top panels, left-to-right: reference image showing surface vascular pattern and central sulcus; response to 25-Hz flutter; response to noxious skin heating; locus in each image of a boxel that exhibited the largest average reflectance decrease in response to flutter (3b) or skin heating (3a). Boxel 3a is in area 3a; Boxel 3b is in area 3b. Middle: time course of signal measured within area 3a boxel (bullet ) and area 3b boxel (black-square) during noxious skin heating. Stimulus time line shown below X-axis. Bottom: time course of signal measured within area 3a boxel (bullet ) and in area 3b boxel (black-square) during 25-Hz flutter stimulation. Stimulus time line shown below X-axis.

Application of the paired boxel analysis method to the data obtained from the other three subjects in which the SI forelimb area was studied (Fig. 4) revealed that average reflectance in the area 3a boxel always underwent a large decrease during skin heating, and average reflectance in the area 3b boxel always underwent a large decrease during 25-Hz flutter. In contrast, for both the skin heating and 25-Hz flutter conditions there was considerable subject-to-subject variation in the direction, magnitude, and time course of the average reflectance change in the nonmaximally activated boxel.


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FIG. 4. Time course of area 3a (solid lines) and area 3b (dotted lines) responses to 25-Hz skin flutter stimulation (plots in right column) or noxious skin heating (plots in left column) in 3 additional subjects (see Fig. 3). Plots in each row are from same subject; plots in different rows from different subjects. SD, stimulus duration; ITI, intertrial interval. Two different conditions of noxious skin heating were used---top left is plot for data obtained with 20 presentations (trials) of a single skin heating ramp (37-48-37°C; ramp duration 3.5 s; ITI 90 s); middle and bottom left plots are for data obtained in 20 trials each consisting of 6 2.4-s skin-heating ramps.

Correlation mapping: comprehensive approach to evaluation of the functional relationship between areas 3b/1 and area 3a during skin heating and cutaneous flutter stimulation

The results shown in Figs. 3 and 4 indicate that a skin-heating stimulus consistently causes activation (indicated by a decrease in reflectance) of area 3a and that 25-Hz cutaneous flutter consistently activates area 3b and/or area 1, and that the effects of either mode of stimulation on anterior parietal territories that neighbor the maximally activated region were neither large nor consistent from one subject to the next. On the other hand, the relatively small changes in average reflectance detected by the paired boxel method (the method illustrated in Figs. 3 and 4) at locations outside the maximally responding anterior parietal region were more difficult to interpret, that is, it was not clear that the intrinsic signal that occurred at sites outside the maximally responding region were meaningful or were merely attributable to random variation ("noise").

Uncertainly about the meaning of variation in average reflectance at off-focus anterior parietal sites prompted us to formulate and use a more powerful and comprehensive method for quantitatively evaluating the imaging data---the method of correlation mapping. The specific hypothesis we sought to test with this method was that changes in average reflectance evoked by either skin heating or 25-Hz flutter skin stimulation not only are different in the regions of areas 3a and 3b/1 that receive their input from the stimulated skin site but are opposing (i.e., the changes in reflectance evoked in cytoarchitectonic areas 3a and 3b/1 by each mode of skin stimulation are negatively correlated). The main strengths of the correlation mapping approach (relative to the analysis of boxel pairs) are that it 1) provides an outcome (a correlation map) constructed from all the information in an image, 2) provides objective measures (correlation values) of associations between the time-dependent reflectance changes at loci within the region of maximal activity and changes in reflectance at loci outside the region of maximal activation, 3) enables assessment of the statistical significance of the correlation values in different anterior parietal regions, and 4) enables objective delineation of anterior parietal regions functionally related to each stimulation mode based on the value and sign of the correlation with the time-dependent behavior of the region activated maximally by skin stimulation.

A correlation map was generated by calculating for each subject/stimulus condition the correlation (ranging between -1 for negative correlation and +1 for positive correlation) between the temporal series of reflectance values at each pixel location and the temporal series of average reflectance values determined for the boxel at the region of maximal activation (see METHODS for details). The results (Fig. 5) indicate that repetitive skin stimulation with a skin-heating stimulus evokes a spatially extensive and complex pattern of reflectance changes at "off-focus" anterior parietal locations, which develops in parallel with the response of the maximally responding region in area 3a. More specifically, for all four subjects studied with the skin heating stimulus, the time course of the changes in average reflectance observed at every locus within a large, mostly continuous zone (red coding in each panel in the middle column of Fig. 5 defines anterior parietal loci with correlation values between +0.5 and +0.8) was positively correlated with the time course of the response of the region that responded with the maximal reflectance decrease (thresholded region designated with black code in each panel on the right). Thus, at all locations within the red-colored zone in each of the middle panels of Fig. 5, average reflectance decreased after onset of skin heating with a time course similar to that observed in the region of maximal activation in area 3a. Also evident in all the maps of Fig. 5 are extensive, largely continuous regions comprised of loci whose average reflectance increased with a time course similar to that of the reflectance decrease that occurred in the maximally activated region (blue-colored regions in the middle panels of Fig. 5 define loci with correlation values between -0.5 and -0.8).


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FIG. 5. Correlation maps generated from imaging data evoked by 25-Hz flutter (panels in left column) and skin heating (middle column). Each row of maps was obtained from a different subject (n = 4); maps in top 3 rows were obtained from 3 subjects stimulated on forelimb skin site; maps in bottom row from single subject stimulated on hindlimb site. Panels in column on right show the threshold responses to both modes of same-site stimulation (gray shading, region of maximal response to 25-Hz flutter; black shading, region of maximal response to skin heating). Color scale at left indicates value of the correlation coefficient. For each map the correlation (estimated by the correlation coefficient, r) between the changes in reflectance over time at each pixel, and the time course of the average reflectance decrease within the region of maximal response for each stimulus condition was computed (the regions of maximal response were defined as the area 3b and area 3a boxels shown in Figs. 3 and 4). For each map, the value of |r| that had to be exceeded to be regarded as significantly different from zero (P < 0.05): for maps in left column---top (0.340); second from top (0.553); third from top (0.482); bottom (0.340); for maps in middle column---top (0.270); second from top (0.260); third from top (0.390); bottom (0.230).

The correlation maps of Fig. 5 demonstrate that the anterior parietal region (red-yellow region) in which the change in average reflectance is correlated positively with the reflectance change in the area 3a region that responded maximally to either skin heating (middle panels) or the area 3b region that responded maximally to 25-Hz flutter (left panels) is much larger than the region that underwent the maximal stimulus-evoked reflectance decrease (black and gray regions in right panels). The maps in Fig. 5 also reveal that the region in which the change in average reflectance is correlated negatively with the reflectance change in the area that responded maximally to skin heating only partially approximates the location of the region that responded maximally to 25-Hz flutter (it typically is much larger than the region of maximal response to 25-Hz flutter and, in 3 of the 4 subjects, is almost exclusively located within area 1). In summary, the results obtained by correlation mapping are generally consistent with the suggestion (of the "raw" IOS imaging observations) that extensive regions in areas 3b/1 and area 3a in SI hindlimb or forelimb cortex participate simultaneously in the SI global response to repetitive delivery of a 25-Hz or skin-heating stimulus to a discrete skin site. They also indicate that the spatiotemporal distribution of influences evoked within SI by skin-heating and 25-Hz flutter stimulation is very different, i.e., skin heating evokes strong activation of area 3a and simultaneous suppression/inhibition of anterior parietal regions located posterior to area 3a, whereas 25-Hz flutter evokes vigorous activation of areas 3b/1 that is accompanied by weak or no significant suppression/inhibition of area 3a.

    DISCUSSION
Abstract
Introduction
Methods
Results
Discussion
References

A role for area 3a in pain perception?

The available evidence indicates that it may not be the activity of area 3b or area 1 but area 3a activity that most closely parallels the intensity of the pain experience evoked by repetitive stimulation of the nociceptors activated by skin heating. This conclusion is consistent with the IOS imaging data presented in a previous report (Tommerdahl et al. 1996) in which it was demonstrated that the area 3a optical response to 0.33/s tapping of the skin with a probe maintained at 48-52°, like the second pain percept of humans (Price et al. 1977, 1994; Vierck et al. 1997), increases progressively (exhibits slow temporal summation), grows in spatial extent (exhibits spatial radiation), and remains evident (exhibits persistence) for a substantial time (15-30 s) after stimulus termination. Additional support for a role for area 3a in signaling the intensity of skin nociceptor activation is provided by the finding that the optical response of area 3a to noxious skin heating survives interruption of the dorsal column pathway (Tommerdahl and Whitsel 1995) and by the discovery that layer IV of area 3a but not layer IV in areas 3b and 1 receives a prominent projection from the thalamic neurons that selectively receive the output of the lamina I neurons of the spinal cord dorsal horn (Craig 1991, 1995a,b, 1996; Craig and Kniffki 1995).

The observation that in response to skin heating a reflectance increase occurs in area 1 (and sometimes also in area 3b) and at the same time reflectance decreases in area 3a leads us to anticipate that the neuroelectrical activity of area 1 neurons (and in some instances also area 3b neurons) is actively suppressed/inhibited by skin heating. This possibility remains to be evaluated in neurophysiological experiments in which the spike discharge responses of area 3b/1 neurons to cutaneous flutter stimulation are recorded before and during same-site heating, but the demonstration of area 3b/1 neuron suppression/inhibition by skin heating in neurophysiological recording experiments is fully consistent with a number of human psychophysical observations. For example, 1) the ability of humans to detect a cutaneous flutter stimulus is decreased during application of a tonic noxious skin heating stimulus, 2) the elevation in threshold for detection of cutaneous flutter during skin heating becomes more evident as the intensity of the pain experience evoked by the skin heating stimulus is increased (Apkarian 1995; Apkarian et al. 1992, 1994), and 3) the magnitude of the elevation of the threshold for detection of flutter increases as the distance between the skin heating and flutter stimuli is decreased (Bolanowski et al. 1998). The observation that in three of four subjects the activity in area 1 was significantly suppressed/inhibited while the activity in area 3b was either marginally or not suppressed/inhibited (in the fourth subject the activity of both areas 1 and 3b was suppressed/inhibited) was not predicted, but in retrospect seems consistent with a well-known feature of the organization of anterior parietal interareal connections. That is, the main target of corticocortical connections arising in area 3b is area 1 (Bunton and Fabri, 1995); thus suppression/inhibition of area 1 would be the most economical way to reduce the impact on other cortical territories of influences arising in both areas 3b and 1.

The idea that a region in human cerebral cortex in a location corresponding to that occupied by area 3a plays an important role in pain perception is not new. In particular, the post-World War studies of large numbers of patients with localized brain injuries limited to cerebral cortex led clinical investigators to the concept that damage of a region in the depths of the posterior wall of the central sulcus (the position occupied by much of area 3a) is accompanied by a prominent and selective loss of cutaneous pain sensibility. Conversely, lesions that involve much of anterior parietal cortex but spare the tissue in the immediate vicinity of the central sulcus are associated with hyperpathia (Kleiset 1934; Marshall 1951; Russell 1945). Interestingly, these early clinical investigators raised the possibility that these effects of parietal cortex lesions on pain perception might be explained in terms of disturbed interactions among parietal territories that in healthy subjects were mutually inhibitory; Peele (1944) regarded this interpretation as fully consistent with the results obtained in a study of the effects of parietal cortical lesions in monkeys. It is unclear why the results reported by these early studies were neglected because they were reviewed in detail by Perl (1984). Our observations in squirrel monkey that a skin-heating stimulus vigorously and consistently activates area 3a and simultaneously suppresses/inhibits the activity in those parts of areas 3b/1 that represent the stimulated skin area lead us to recommend reconsideration of the proposal that dynamic anterior parietal interareal inhibitory interactions contribute importantly to pain information processing in cerebral cortex.

At first glance, the demonstration by this and our previous study (Tommerdahl et al. 1996) that area 3a in squirrel monkey cortex is activated by a condition of skin-heating stimulation that evokes well-characterized pain experiences in humans appears at odds with the recent evidence provided by PET and fMRI human brain imaging studies. For example, it was reported that a transient and modestly painful skin-heating stimulus that activates multiple neocortical regions also activates a territory that, on the basis of its hemispheric location, was regarded as SI (Apkarian 1995; Apkarian et al. 1992; Bushnell et al. 1995; Casey et al. 1994; Coghill et al. 1994; Derbyshire et al. 1994; Jones et al. 1991; Talbot et al. 1991). Moreover, in subjects in whom a prolonged (tonic) noxious skin-heating stimulus evoked moderate to strong pain, SI activity not only did not increase but was consistently suppressed/inhibited to a level below that present before stimulation (Apkarian et al. 1995; see also Jones and Derbyshire 1994). The variety of reports indicating that anterior parietal and ventrobasal thalamic activity is abnormally low in patients experiencing severe chronic pain also were viewed as consistent with the observation that SI activity in normal subjects is suppressed/inhibited during moderate to strong stimulus-evoked pain (for recent reviews of clinical pain imaging studies see Apkarian et al. 1995; Derbyshire et al. 1994; Jones and Derbyshire 1994). Given observations such as these, it is not surprising that SI was considered to play only a minor and clinically irrelevant role in pain encoding. Perhaps the most influential result in this regard is that measures of SI activity obtained in human PET and fMRI studies have not exhibited the monotonically accelerating relationship between response magnitude and intensity of the skin-heating stimulation expected of a region that encodes pain intensity (one that signals an increase in the intensity of perceived pain magnitude by an increase in activity).

A straightforward way to account for the discrepancy between our IOS imaging results and the findings reported by the human PET and fMRI studies becomes evident when it is recognized that the activity of each different anterior parietal cytoarchitectonic areas was evaluated independently in the present study, whereas the published human imaging investigations pool the activity in area 3a with the activity of the other anterior parietal fields (areas 3b, 1, and 2). When viewed in this context, the results obtained in this study raise the possibility that, under certain conditions of skin stimulation, an across-area measure of SI activity gives a grossly inaccurate estimate of the actual state of anterior parietal activation. An across-area activity measure would, at least in squirrel monkey, yield a value for activity substantially less than that actually evoked in area 3a by a skin-heating stimulus. That in squirrel monkey the activity of area 3a consistently is accompanied by suppression/inhibition of the activity in area 1 (and less frequently in area 3b as well; Fig. 5) implies that the view of activity provided by an across-area measure of SI activity could be highly unrepresentative of that actually present. The estimate of the SI activity evoked by skin heating would thus be the algebraic sum of the stimulus-evoked increase in area 3a activity and the suppressed/inhibited activity levels in areas 3b and area 1. Our conclusion is therefore that an across-area measure of anterior parietal activity in squirrel monkey would grossly underestimate the extent to which anterior parietal cortex participates in the processing of information evoked by noxious skin heating. Furthermore, because the negative correlation between area 3a and area 3b/1 activity appears to become increasingly more prominent as the intensity of the skin-heating stimulus is increased, it is anticipated that the extent to which an across-area activity measure underestimates the area 3a response increases as the intensity of the noxious skin-heating stimulus is increased. This would account for the finding of the PET and fMRI human imaging studies that SI activity evoked by noxious skin heating fails to increase monotonically with stimulus strength in the manner expected for a cortical region, which, if it were involved importantly in pain information processing, should vary its activity in parallel with the intensity of the stimulus-evoked pain experience.

Do corticocortical interareal inhibitory interactions contribute to anterior parietal information processing?

Although inhibitory interactions between the neural responses to skin heating and innocuous mechanical skin stimuli were described at subcortical levels of the somatosensory projection path (for discussion see Apkarian et al. 1995), we regard the extensive system of horizontal connections that link the different anterior parietal cytoarchitectural areas (Burton and Fabri 1995; DeFelipe et al. 1986) as the most likely neural substrate responsible for the suppressive/inhibitory effects detected in this IOS imaging study. This conclusion is based on the following functional/anatomic observations. First, the afferent drive evoked by cutaneous flutter reaches area 3b and 1 via a route (the spinal dorsal column-dorsal column nucleusvventrobasal thalamus projection path), which until very recently (Dykes and Craig 1998) was regarded as remaining relatively free of interactions with activity that ascends the neuraxis via the spinothalamic pathways (Mountcastle 1984). Second, the spinal dorsal horn is probably not the site of the suspected interactions because dorsal horn neurons are not a significant source of the afferent drive that reaches SI as a result of cutaneous flutter stimulation (Dreyer et al. 1974; Jain et al. 1997; Tommerdahl et al. 1996); however, dorsal horn neurons recently have been shown to be involved in the control of the excitability and of the receptive field size and locus of neurons in the dorsal column nuclei) (Dykes and Craig 1998). Third, the system of corticocortical connections that interlinks the different anterior parietal areas is more massive by far than any set of subcortical connections that might account for the observed relationship among activities of the different anterior parietal fields observed during skin heating stimulation. Taken together, these established and widely accepted aspects of somatosensory nervous system organization make it difficult to adequately explain the highly coordinated, opposing behaviors of areas 3b/1 and area 3a during either cutaneous flutter or noxious skin heating in terms of interactions taking place at any level before cerebral cortex. Insofar as the physiological effects of corticocortical interactions are concerned, although the direct synaptic action of the glutaminergic connections that interrelate the different anterior parietal fields is excitatory, these connections are known to mediate interareal inhibitory effects when they are tonically and strongly activated. The putative mechanism that was suggested to enable glutaminergic corticocortical connections to inhibit the activity of the cell columns in which they terminate was described (Tommerdahl et al. 1996; see also Hirsch and Gilbert 1991). In conclusion, we regard corticocortical inhibitory interactions as the most likely explanation of the suppressive/inhibitory effects detected in this study, but the possibility that both cortical and subcortical inhibitory mechanisms are involved cannot be ruled out.

    ACKNOWLEDGEMENTS

  Expert technical assistance was provided by C. Wong. The authors are indebted to Dr. W. Maixner for advice in the development and use of the apparatus used to deliver controlled mechanical-thermal skin tapping stimuli and Dr. M. Quiberra for help with development and statistical evaluation of IOS correlation maps.

  This research was supported by National Institute of neurological Disorders and Stroke Grants R29 NS-32358 to M. Tommerdahl and RO1 NS-35222 to B. Whitsel and by Whitehall Foundation Grant S93-10 to M. Tommerdahl. K. A. Delemos was supported by National Institute if Dental Research Grant PO1 DE-07509 (W. Maixner, Program Director) and by Whitaker Foundation Special Opportunity Award "Engineering In Systems Neuroscience At UNC-CH".

    FOOTNOTES

  Address for reprint requests: M. Tommerdahl, 155 Medical Research Bulding CB #7545, Dept. of Physiology, University of North Carolina, Chapel Hill, NC 27599.

  Received 9 July 1998; accepted in final form 31 August 1998.

    REFERENCES
Abstract
Introduction
Methods
Results
Discussion
References

0022-3077/98 $5.00 Copyright ©1998 The American Physiological Society