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Seven monkeys (Macaca fuscata) were used. Before the start of the behavioral training, four animals underwent section of the splenium of the CC and the AC and served at first as the posterior-split group. After the behavioral experiments, one monkey was immediately injected with tracers and dedicated to histological analyses (15). Three of the posterior-split monkeys underwent further section of the remaining anterior part of the CC and then served as the full-split group. Three animals served as unoperated controls. Surgery was carried out with sodium pentobarbital anesthesia (25 mg per kilogram of body weight per hour, intravenously) under sterile conditions. In the posterior-split operation, at least one-third of CC from the posterior end of the splenium was aspirated. The lateral ventricle was entered at the level of the interventricular foramen, and AC was exposed and cauterized (24). In the full-split operation, the callosal lesion was extended anteriorly from the level of the AC to the rostrum of the CC.
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to obtain coronal and sagittal MRIs in all the posterior-split monkeys
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At the end of the experiment, the animals were perfused with 4% paraformaldehyde in phosphate buffer (pH 7.4). Adjacent series of sections (50 μm) were stained with cresyl violet or stained for myelin with the modified Gallyas silver technique. Two full-split, one posterior-split, and one unoperated control monkey were injected with the retrograde fluorescent tracers FB and DY 14 to 16 days before perfusion. With a 1-μl Hamilton syringe, nine sites surrounding the anterior middle temporal sulcus in the inferotemporal cortex were injected with DY (2%, 0.25 to 0.5 μl), and 12 to 14 sites in the ventrolateral, dorsolateral, and lateral orbital prefrontal areas were injected with FB (3%, 0.25 to 0.5 μl). For these animals, a series of sections were examined by computerized microscopy (Zeiss, KS-400) for the presence of fluorescently labeled cells.
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Figures 3A and 4A illustrate the behavioral tasks. Procedures for the visual stimulus-stimulus association task were as described in detail in (2, 23, 24), except that in this study the animals must maintain fixation during parafoveal presentation of the visual stimuli and respond with saccade. Visual stimuli were monochrome Fourier descriptors extending approximately 2° X 2°. While the monkey fixated within 0.4° to 0.6° of a spot on a computer monitor, one cue (for 0.7s) and two choice stimuli (0.5 to 1.0 s) were sequentially presented 2.5° lateral to the fixation spot. In the INTRA condition (Fig. 3A), all the stimuli were presented to the identical visual hemifield. In the INTER condition (Fig. 4A), choices were presented to the opposite side of the cue. When the fixation spot was turned off, the animal saccaded to one of the choices. The animal was rewarded when he correctly selected the choice stimulus instructed by the cue. Incorrect trials were followed with additional correction trials, which were not included in the data analysis. If the monkey failed to maintain fixation, the trial was aborted. Eye position was monitored with the scleral search coil method [S. J. Judge, B. J. Richmond, F. C. Chu, Vision Res. 20, 535 (1980)]. For one control animal, a measurement system with a charge-coupled device camera was also used [K. Matsuda, Neurosci. Res. 20, S270 (1996)].
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Figures 3A and 4A illustrate the behavioral tasks. Procedures for the visual stimulus-stimulus association task were as described in detail in (2, 23, 24), except that in this study the animals must maintain fixation during parafoveal presentation of the visual stimuli and respond with saccade. Visual stimuli were monochrome Fourier descriptors extending approximately 2° X 2°. While the monkey fixated within 0.4° to 0.6° of a spot on a computer monitor, one cue (for 0.7s) and two choice stimuli (0.5 to 1.0 s) were sequentially presented 2.5° lateral to the fixation spot. In the INTRA condition (Fig. 3A), all the stimuli were presented to the identical visual hemifield. In the INTER condition (Fig. 4A), choices were presented to the opposite side of the cue. When the fixation spot was turned off, the animal saccaded to one of the choices. The animal was rewarded when he correctly selected the choice stimulus instructed by the cue. Incorrect trials were followed with additional correction trials, which were not included in the data analysis. If the monkey failed to maintain fixation, the trial was aborted. Eye position was monitored with the scleral search coil method [S. J. Judge, B. J. Richmond, F. C. Chu, Vision Res. 20, 535 (1980)]. For one control animal, a measurement system with a charge-coupled device camera was also used [K. Matsuda, Neurosci. Res. 20, S270 (1996)].
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The visual stimulus presented to one visual hemifield (16) must be lateralized to the contralateral hemisphere in the current paradigm, because each visual hemifield is both anatomically and functionally represented in the contralateral hemisphere except for a strip of 1° or narrower, if any, at the vertical meridian [J. Stone, J. Leicester, S. M. Sherman, J. Comp. Neurol. 150, 333 (1973); M. Sugishita, C. R. Hamilton, I. Sakuma, I. Hemmi, Neuropsychologia 32, 399 (1994); R. Fendrich and M. S. Gazzaniga, ibid. 27, 273 (1989)]. Restriction of visual input to one hemisphere is further confirmed by our own findings that learning of visual long-term memory was not transferred in posterior-split animals and that full-split animals could not perform the INTER task above chance (see text).
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The visual stimulus presented to one visual hemifield (16) must be lateralized to the contralateral hemisphere in the current paradigm, because each visual hemifield is both anatomically and functionally represented in the contralateral hemisphere except for a strip of 1° or narrower, if any, at the vertical meridian [J. Stone, J. Leicester, S. M. Sherman, J. Comp. Neurol. 150, 333 (1973); M. Sugishita, C. R. Hamilton, I. Sakuma, I. Hemmi, Neuropsychologia 32, 399 (1994); R. Fendrich and M. S. Gazzaniga, ibid. 27, 273 (1989)]. Restriction of visual input to one hemisphere is further confirmed by our own findings that learning of visual long-term memory was not transferred in posterior-split animals and that full-split animals could not perform the INTER task above chance (see text).
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The visual stimulus presented to one visual hemifield (16) must be lateralized to the contralateral hemisphere in the current paradigm, because each visual hemifield is both anatomically and functionally represented in the contralateral hemisphere except for a strip of 1° or narrower, if any, at the vertical meridian [J. Stone, J. Leicester, S. M. Sherman, J. Comp. Neurol. 150, 333 (1973); M. Sugishita, C. R. Hamilton, I. Sakuma, I. Hemmi, Neuropsychologia 32, 399 (1994); R. Fendrich and M. S. Gazzaniga, ibid. 27, 273 (1989)]. Restriction of visual input to one hemisphere is further confirmed by our own findings that learning of visual long-term memory was not transferred in posterior-split animals and that full-split animals could not perform the INTER task above chance (see text).
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Before measurement of learning transfer, acquisition of at least three stimulus sets was required for each left and right INTRA condition. A transfer test was then carried out with four new stimulus sets in a daily 200-trial session for 1 to 3 days until a criterion level was reached. The criterion was defined as 80% or more correct responses for two consecutive 20-trial blocks within a session. The averaged number of trials for individual animals to reach the criterion was statistically analyzed by a repeated-measures ANOVA and post hoc t tests.
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The saving score for each stimulus set was calculated as follows: saving score = (TC1 - TC2)/(TC1 + TCZ) × 100; TC1 and TC2, trials to criterion for the first and second hemisphere, respectively. The range of the score is from -100% to +100%, where 100% indicates perfect transfer and 0% means no transfer.
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After completion of the first experiment, the posterior-split animals were shaped into the INTER condition and trained with the same four stimulus sets that were used for the learning transfer test (18). It took 285 ± 105 trials (mean ± SE; N = 3) to reach the criterion in the INTER condition. In performance test, the averaged score over two consecutive 100-trial sessions was recorded for each of the four stimulus sets per animal on three conditions: left INTRA, right INTRA, and INTER. After the full-split surgery, performance for these same stimulus sets was again tested for every condition. For the INTER condition, however, blocks of 10 INTER trials and of 5 INTRA trials were alternated to maintain motivation. The averaged score for individual animals in each condition was statistically analyzed with repeated-measures ANOVA and post hoc t tests.
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Haxby, J.V.1
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note
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Experiments were conducted in accordance with the NIH Guide for the Care and Use of Laboratory Animals and with the regulations of the University of Tokyo School of Medicine. This work was supported by grants from Research Fellowships of the Japan Society for the Promotion of Science for Young Scientists to I.H. and T.I. and by a grant-in-aid for Specially Promoted Research from the Ministry for Education, Science, and Culture of Japan (07102006) to Y.M. We thank S. Konishi and M. Kameyama for collaboration; H. Niki for helpful discussion; C. Hamada for statistical assistance; T. Kitamura, J. Yamada, M. Yukie, and M. Yoshida for histological advice; and T. Kirino for encouragement.
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