Activation on the Medial Wall During Remembered Sequences of Reaching Movements in Monkeys
Nathalie Picard1 and
Peter L. Strick1, 2
1 Research Service, Veterans Administration Medical Center, Syracuse; and 2 Departments of Neurosurgery and Physiology, State University of New York Health Science Center at Syracuse, Syracuse, New York 13210
 |
ABSTRACT |
Picard, Nathalie and Peter L. Strick. Activation on the medial wall during remembered sequences of reaching movements in monkeys. J. Neurophysiol. 77: 2197-2201, 1997. We used the 2-deoxyglucose (2DG) method to map activation in the motor areas on the medial wall of the hemisphere. One group of monkeys licked juice delivered at variable time intervals (LICK task). For these animals, the motor areas on the medial wall displayed restricted activation. 2DG uptake was limited largely to the face representation of the supplementary motor area (SMA). Additional labeling was present more rostrally in the banks of the cingulate sulcus. A second group of animals performed remembered sequences of reaching movements (REM task) for juice rewards. Activation related to licking also was present in this group. In addition, separate, discrete activations were found on the superior frontal gyrus and in the cingulate sulcus during the REM task. The most intense and extensive 2DG labeling was located in the dorsal bank of the cingulate sulcus, coincident with the dorsal cingulate motor area (CMAd). Weaker activations were present in the arm area of the SMA and in the pre-SMA. There was no significant 2DG incorporation in the ventral bank of the cingulate sulcus where the ventral cingulate motor area is located. Our findings suggest that the CMAd is involved more than any other medial area in the preparation for and/or execution of highly practiced, remembered sequences of movements. Overall, our results indicate that the attributes of motor control are not represented equally across the motor areas on the medial wall.
 |
INTRODUCTION |
In recent years, there has been considerable interest in the contribution of the medial wall of the hemisphere to the generation and control of motor behavior (e.g., Dum and Strick 1991
; He et al. 1995
; Luppino et al. 1991
; Matsuzaka et al. 1992
; Shima et al. 1991
; Tanji 1994
). For example, functional imaging studies in humans have revealed intense medial wall activation in a variety of motor tasks (for references and review, see Picard and Strick 1996
). In the past, activation almost anywhere on the medial wall was attributed to the supplementary motor area (SMA). However, it is now clear that the medial wall contains several motor fields. In macaques, the SMA and three cingulate motor areas [rostral (CMAr), dorsal (CMAd), and ventral (CMAv)] project directly to the spinal cord (Dum and Strick 1991
, 1996
; He et al. 1995
). An additional motor field, the pre-SMA, lies rostral to and is distinct from the SMA proper (Luppino et al. 1991
; Matelli et al. 1991
; Matsuzaka et al. 1992
). These five motor fields appear to be present on the medial wall in humans (Picard and Strick 1996
). Given this anatomic framework, we have begun a series of experiments to determine the specific contributions of each motor area to aspects of motor function. In this report, we present evidence that the motor areas on the medial wall are activated differentially when monkeys perform remembered sequences of pointing movements.
 |
METHODS |
Four monkeys (Macaca nemestrina, 5-10.5 kg) were used for these experiments. Two animals performed remembered sequences of reaching movements (REM task) (Mushiake and Strick 1993
) whereas the other two performed a licking task (LICK task). The monkeys faced a panel with five touch pads. In the REM task, they initiated a trial by placing their right hand on a hold key for 0.4-1.3 s. Then, three of five light emitting diodes (LEDs) located over each touch pad were illuminated sequentially. After a delay of 1.25-2.15 s (instruction period), a "go" signal sounded. The monkeys had to release the hold key and contact each of the three touch pads in the instructed order to receive juice rewards. The LEDs remained illuminated until the end of the trial. Eight predetermined sequences were varied pseudorandomly between trials. Monkeys were given considerable practice on the REM task until they performed >90% of the trials correctly and interbutton intervals were relatively brief and regular.
In the LICK task, the monkeys sat in the primate chair and licked juice delivered at variable time intervals (3.2-4.8 s). This approximated the rate of reward delivery for animals performing the REM task. The animals also were presented visual and auditory signals that were comparable with those of the REM task. However, these signals were meaningless and the monkeys did not respond to them. Aside from licking, no movement was required.
Conventional procedures for qualitative 2-deoxyglucose (2DG) were followed (e.g., Juliano et al. 1981
). On the day of the experiment, an intravenous catheter was inserted in the leg for 2DG and drug injection. After stable task performance was achieved, monkeys received 60-100 µCi/Kg of 14C-2-deoxyglucose (55 mCi/mmol; American Radiolabeled Chemicals). The monkeys continued the task for 45 min. They then were killed with sodium pentobarbital (40 mg/kg) and perfused with saline and fixative. The brains were blocked, frozen, and stored at
70°C until sectioning.
Autoradiographs of coronal brain sections (25- or 30-µm thick) were digitized at a resolution of 55-62 µm by 45-51 µm. For each pixel, gray scale values were converted to 14C tissue concentration equivalents based on 14C standards (Amersham). We measured the average 14C values in the middle cortical layers. To do this, we drew a line through the cortex midway between the cortical surface and the white matter. 14C values of four pixels, two on each side of this line, were averaged. These averaged values were then used to generate a flattened map of activation on the medial wall using sections spaced 90 or 100 µm apart. The methods for unfolding the medial wall and generating a flattened map have been described in detail previously (Dum and Strick 1991
). Serial sections every 300 or 400 µm were thionin-stained for cytoarchitectonic analysis. We then sampled 2 mm2 regions of interest on the maps. Background was defined as the amount of 2DG labeling in a portion of the leg representation of primary motor cortex (M1) on the precentral gyrus. 2DG uptake >3 SD from the background mean was considered significant.
In all animals, the highest 2DG incorporation in sensorimotor cortex was located in lateral portions of the primary somatic sensory cortex (SI) on the precentral gyrus (orofacial representation in areas 3a/3b) (e.g., Pritchard et al. 1986
). To compare activations among animals, the range of activation was defined as the difference between activation in face SI and background for each monkey. Mean 14C values in specific regions of interest were transformed into percentages of this range. Finally, to enhance visual presentation, 14C values in functional maps were interpolated linearly between sections and slightly smoothed (3 data points average in the y axis, 5 points Gaussian filter in the x axis).
We compared the distribution of 2DG labeling on the medial wall with the distribution of neurons projecting to lower cervical segments of the spinal cord (C7-T1) (He et al. 1995
). To obtain a map of corticospinal neurons that was comparable with the 2DG maps, we replotted data from animal H2 of He et al. (1995)
using a contour program (Transform, Spyglass). The number of corticospinal neurons in 200-µm-wide bins through cortical layer V of coronal sections spaced 200 µm apart was entered into Transform. The data was interpolated linearly between sections, smoothed (3 × 3 average) and then color-coded according to density.
 |
RESULTS |
The global uptake of 2DG in the brain varied between animals. However, the location and relative intensity of labeling in the motor areas of the medial wall were similar for the two animals in each group after normalization of 14C values (see METHODS). Representative results from one subject in each task group are presented here. This presentation will be limited to activations present in the motor areas of the medial wall contralateral to the moving arm.
Licking control
Only two foci on the medial wall had significant 2DG labeling during the LICK task (Table 1). One was located on the medial portion of the superior frontal gyrus at approximately the rostro-caudal level of the genu of the arcuate sulcus (Fig. 1A). This focus of activation is in a region that corresponds to the face/neck representation of the SMA (Luppino et al. 1991
; Mitz and Wise 1987
; Muakkassa and Strick 1979
). The second site of activation was located in the cingulate sulcus and includes the face representation of the CMAr (Fig. 1A) (He et al. 1995
; Luppino et al. 1991
; Muakkassa and Strick 1979
; Picard and Strick 1996
). In the caudal portion of the cingulate sulcus, 2DG uptake was not significantly different from that of background (Table 1). This reflects the absence of a substantial representation of the face in this region of the cingulate sulcus (Luppino et al. 1991
; Morecraft and Van Hoesen 1992
; Morecraft et al. 1996
; Muakkassa and Strick 1979
). Some activation on the superior frontal gyrus extended rostral to the face SMA. This labeling is actually an extension of activation in the supplementary eye field, which is located predominantly on the lateral surface of the hemisphere (not shown) (Schlag and Schlag-Rey 1987
). Similarly, activation extended rostral to the CMAr into regions of prefrontal cortex.

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| FIG. 1.
Relative 2-deoxyglucose (2DG) uptake in medial wall during LICK task (A) and REM task (B). Each map is from a single monkey. Local 2DG concentration is color coded as a function of the number of standard deviations above mean background uptake in each animal (see text). Long dashed lines mark the fundus of cingulate sulcus that is "opened up." , level of arcuate genu on lateral surface of hemisphere. Arrow heads mark location of sections shown in Fig. 3. CC, corpus callosum; CgG, cingulate gyrus; CgSv, cingulate sulcus, ventral bank; CgSd, cingulate sulcus, dorsal bank; SFG, superior frontal gyrus. Scale bars = 5 mm.
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Remembered sequence
All the areas that displayed activation during the LICK task also displayed activation during the REM task (Table 1; Fig. 1, A and B). In addition, performance of the REM task was associated with labeling in the arm representations of three of the five motor areas on the medial wall (Table 1; Fig. 1B). Activation also was present bilaterally in medial portions of SI. This area contains the representation of the hindlimb and tail and extends onto caudal portions of the superior frontal gyrus and into adjacent portions of the dorsal bank of the cingulate sulcus (Nelson et al. 1980
; Peele 1942
; Vogt and Pandya 1978
). This area was adequately sampled and found to be activated in one animal that performed the LICK task. Furthermore, medial portions of SI also were activated bilaterally in animals trained to perform reaching movements that are visually guided (unpublished observations). Thus activation in medial portions of SI does not appear to be task specific.
Sites exclusively activated during the REM task are shown in Fig. 2A. By far the most extensive and intense activation during the REM task was located on the dorsal bank of the cingulate sulcus, in the arm area of the CMAd (Table 1; Figs. 2A and 3). The second region containing smaller foci of activation was located in the arm area of the SMA on the superior frontal gyrus. However, in relative terms, the intensity of 2DG labeling in the CMAd was >1.5 times greater than that in the SMA (Table 1). Furthermore, the difference between the activation during the REM task and that during the LICK task was many times greater in the CMAd than in the SMA. The third site of activation was located in a region of the superior frontal gyrus that lies rostral to the genu of the arcuate sulcus. This region lacks substantial direct projections to either the spinal cord (Fig. 2B) or M1 (Dum and Strick 1991
) and corresponds to the pre-SMA (Luppino et al. 1991
; Matsuzaka et al. 1992
). The intensity of activation in the pre-SMA was similar to that of the SMA and, thus, much less than the CMAd (Table 1, Fig. 2A).

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| FIG. 2.
A: motor areas activated exclusively during the REM task. This map was created by removing activations present during the LICK task (Fig. 1A) from activations generated by the REM task (Fig. 1B). B: density map of corticospinal neurons projecting to segments C7-T1 (adapted from He et al. 1995 ). Color scale = >1 neuron (blue) to >4.5 neurons (white) (fractions result from smoothing).
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| FIG. 3.
2DG label in brain sections through the dorsal cingulate motor area (CMAd) during LICK task (left) and REM task (right). Location of each coronal section is indicated in Fig. 1. See Fig. 1 for color scale. SPCS, superior precentral sulcus. Scale bar = 1 mm.
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DISCUSSION |
An unexpected finding of the present study is that a single motor area on the medial wall, the CMAd, was the major site of activation during the REM task. It is unlikely that the intense activation of the CMAd during the REM task is simply due to the involvement of this cortical area in movement execution per se. Savaki et al. (1995)
did not observe 2DG labeling in the CMAd of monkeys that performed simple reaching movements guided by visual cues. Our preliminary observations indicate that 2DG uptake in the CMAd is not as intense when movement sequences identical to those of the present study are performed under visual guidance; other cortical areas are activated more intensely than the CMAd in this condition (unpublished observations). Our findings suggest that the CMAd is involved more than any other medial area in the preparation for and/or execution of highly practiced, remembered sequences of movements. Overall, these observations indicate that the attributes of motor control are not represented equally across the motor areas on the medial wall.
Matsunami and Kawashima (1995)
found 2DG uptake in the cingulate sulcus of monkeys performing wrist movements. It may be significant that their monkeys were making movements to remembered targets. However, it is difficult to compare these results with ours because Matsunami and Kawashima (1995)
do not provide any measure of the intensity of the activation they saw. In addition, the site of cingulate activation in most of their animals does not appear to correspond to the region of the CMAd that was activated in our study.
Activation during the REM task was not limited to the CMAd but was present in other motor areas on the medial wall as well. This observation suggests that, even after extensive practice, the performance of sequential movements requires the active involvement of multiple motor areas. Interestingly, activation was absent from two arm areas on the medial wall
the CMAr and the CMAv. The lack of activation in these areas is significant in two respects. First, the dissociation between the 2DG labeling on the dorsal and ventral banks of the cingulate sulcus provides further support for splitting these regions into two separate areas, the CMAd and CMAv (Dum and Strick 1991
; He et al. 1995
). Second, neuron activity is present in the CMAr and CMAv during simple motor tasks (Shima et al. 1991
), and neurons in the CMAv receive proprioceptive input from the arm and hand (Cadoret and Smith 1995
). The absence of substantial labeling in the CMAr and CMAv indicates that the specific aspects of voluntary movement that are controlled by the CMAr and CMAv were not particularly engaged by the REM task. Thus these results provide further evidence for functional specificity within the motor areas on the medial wall.
It is important to recognize that foci of 2DG uptake may not overlap precisely with foci of cellular activity because metabolic markers primarily reflect synaptic activity(Jueptner and Weiller 1995
). However, prior studies have shown striking correlations between the location of 2DG activation in regions of somatosensory and visual cortex and sites of cellular activity (e.g., Juliano and Whitsel 1987
; Tootell et al. 1988
). Furthermore, there is close correspondence between our sites of 2DG uptake during licking and sequential arm movements and the maps of face and arm representation on the medial wall generated by other techniques (e.g., Dum and Strick 1991
; He et al. 1995
; Luppino et al. 1991
; Shima et al. 1991
). Thus we believe that 2DG uptake in cortex provides a reliable indication of the relative location and intensity of neural activity in the motor areas.
The intense and localized activation of the CMAd during the REM task raises the possibility that the motor functions of the medial wall that have been traditionally attributed to the SMA in human subjects actually may involve the CMAd alone or in combination with the SMA. Thus our observations stress the need for precise localization of activation sites within a well-defined anatomic framework (Picard and Strick 1996
). Concepts about the function of the medial wall in motor control have emphasized the role of the SMA in programming internally generated and sequential movements like those performed during the REM task (e.g., Goldberg 1985
; Halsband et al. 1994
; Mushiake et al. 1991
; Passingham 1987
; Roland et al. 1980
; Tanji 1994
). Although the SMA and pre-SMA were activated during the REM task, overall 2DG incorporation on the superior frontal gyrus was relatively low. The modest activation of the SMA and pre-SMA during the REM task in part may be due to task particulars or overtraining. However, drawing any conclusion about SMA function is made even more difficult by our observation that the simple behavior of licking for juice rewards always led to intense activation in the face representation of the SMA. These observations argue against the notion that the SMA is involved exclusively in complex aspects of motor control.
 |
ACKNOWLEDGEMENTS |
We thank M. Page for the development of computer programs and M. O'Malley-Davis for expert technical assistance.
This work was supported by the Veterans Affairs Medical Research Service and Rehabilitation Research and Development Service, U.S. Public Health Service Grant 24328 (P. L. Strick), and Medical Research Council of Canada (N. Picard).
 |
FOOTNOTES |
Address for reprint requests: P. L. Strick, VA Medical Center, Research Service (151S), 800 Irving Ave., Syracuse, NY 13210.
Received 4 November 1996; accepted in final form 2 January 1997.
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