Electronic Supplementary Material
Subjects
Five of the subjects had been wild-caught and housed in captivity since their capture, while the remaining seven subjects were born and raised in captivity. All subjects lived in social groups ranging from two to thirteen individuals at Yerkes National Primate Research Center and were housed according to institutional guidelines. Brains were collected postmortem, with the cause of death being unrelated to the current study.
Behavioral measurements
In each trial of the manual gesturing task, the experimenter approached the chimpanzee’s home cage carrying food. A piece of food was offered until this elicited an open hand begging gesture from the chimpanzee. All other manual responses (e.g., banging or clapping) were ignored as gestures. The hand used to request food was recorded for each trial and subsequently handedness data on the gesture task was obtained for 10 of the 12 subjects. A minimum of 30 responses were obtained for each individual for each task and a handedness index (HI) was derived for each subject per task by subtracting the number of left-handed responses from the number of right-handed responses and dividing by the total number of responses: HI = (R-L)/[(R+L)/2]. Positive values reflect right-hand preference and negative values represent left-hand preference (Table 1). The absolute value of the HI corresponds to the consistency of directional hand preference. Subjects with z scores greater than 1.95 or less than -1.95 were classified as right- and left-handed, respectively. Subjects with z scores between -1.95 and 1.95 were classified as having no preference. Neuroanatomical measurements were performed blind to the HI scores of the subjects.
Area identification
Adjacent areas were differentiated from area Tpt on the basis of cytoarchitectonic criteria. Area Tpt is characterized cytoarchitectonically by a more homogenous cell density throughout layers II to VI in both macaques and humans (Sweet et al. 2005). The borders of layer IV on either side of layer III and layer V are known to encroach and obscure the layer IV border due to the infiltration of pyramidal cells, creating a broad, central cellular band (Fullerton & Pandya, 2007; Sweet et al. 2005; Galaburda & Pandya, 2005). Area Tpt merges caudally with the inferior parietal lobule, which also has a well developed layer II, deeply stained medium sized pyramidal neurons in the lower tier of layer III, and a layer V and VI with small to medium sized pyramidal neurons (Fullerton & Pandya., 2007).
Shrinkage correction
The presectioning mass for each block was documented and converted to volumetric units by dividing by the specific gravity for brain tissue (1.036 g/cc)
(Gompertz, 1902). In the few cases where presectioning block mass was unavailable, presectioning volumes were estimated by outlining the extent of the block on postmortem MRI scans using Analyze 7.0 software. Using digitally scanned images of each Nissl-stained series we estimated the postprocessing volumes. The outline of each section was determined by thresholding using Image J software (NIH) and the area for each section was calculated. The summed area of sections was converted to volume by multiplying by the distance between measured sections (i.e., for a 1:10 series of 40 µm-thick sections the distance between sections is 400 µm). To obtain a correction factor unique to each block, the presectioning block volume was divided by the postprocessing volume. All shrinkage correction factors were checked to ensure that they were in range of previously published reports before being applied to the postprocessed blocks.
Cortical area volumes and neuron counts
Volumetric data were collected with the Cavalieri method using a 500 μm point counting grid in the StereoInvestigator software (MBF Bioscience, Williston, VT). In each hemisphere, 8 to 20 sections were analyzed for area Tpt at intervals of 400 or 800 μm between sections. As required by the Cavalieri method, sampled sections from each specimen were chosen at standard intervals for each area and the starting section was picked randomly from the first interval. The coefficient of error (Gundersen et al. 1999, m = 1) averaged 0.07 ± 0.02 across both hemispheres and specimens (Table 2); such low values indicate that the precision of the volume estimates was high and that the sampling parameters were sufficient. Each cortical area’s volume was multiplied by the appropriate correction factor to obtain a value representing the pre-shrinkage volume.
The optical fractionator parameters employed in neuron counting were a 50x50 μm counting frame and a 1200x1200 μm scan grid (on the same sections used for volumetric estimates). A standard disector depth of 7 μm, with a 3 µm guard zone at the top was set in all cases. Section thickness was measured at every 5th site during counting. Neurons were counted in layers II to VI if a clear nucleolus came into focus within the permitted boundaries of the counting frame, according to the principles of the optical fractionator method (West et al. 1991). The calculated coefficients of error (Schmitz & Hof, 2000) were within the range of ≤0.10 for each cortical area in each specimen (Table 3). Neuron density was calculated as the ratio of total neuron number over the shrinkage-corrected cortical area volume.
Validation and inter-observer variability
The volume of each cortical area was calculated and subsequently mapped back onto the accompanying MRIs. Intraclass correlation coefficients (ICC) were calculated to assess the impact of inter-observer variability on both the absolute volume and the reported AQs. These results indicate strong agreement between observers for both absolute volume (ICC = 0.98; P < 0.001) and AQ (ICC = 0.98; P < 0.0001). This indicates that the subjective judgment of area boundaries delineated by these two observers covaries in a systematic fashion, suggesting that the quantitative measures of regional volumes and neuron numbers presented here are reliable.
The transfer of area boundaries onto the associated MRIs represents a further challenge to an assessment of the repeatability of these methods. Analysis of the correlation between histologically-derived volumes and the volumes mapped onto MRI space revealed excellent congruency (r = 0.97; P < 0.001). Area volumes mapped onto the MRIs were on average 23% lower than estimates of volumes obtained directly from the histological sections.
Finally, when each cortical area was mapped back onto the MRIs and the percentage of spatial overlap between the volumes delineated by Observer 1 and Observer 2 was calculated, a mean percentage overlap of 65 ± 10% was obtained. Thus, a portion of the interindividual variation in the location of cortical areas can be attributed to differences among observers in the subjective definition of boundaries. Therefore, our probability maps should cautiously be considered underestimates.
References
1. Fullerton, B.C. & Pandya, D.N. 2007 Architectonic analysis of the auitory related areas of the superior temporal region in the human brain. J Comp Neurol 504, 470-498.
2. Galaburda, A. & Pandya, D. 1983 The intrinsic architechtonic and connectional organization of the superior temporal region of the rhesus monkey. J Comp Neurol 221, 169-184.
3. Gompertz, R.H.C. 1902 Specific gravity of the brain. J Physiol 27, 459-462.
4. Gundersen, H.J.G., Jensen, E.B., Kieû, K. & Nielsen, J. 1999 The efficiency of systematic sampling in stereology-reconsidered. J Microsc 193, 199-211.
5. Schmitz, C. & Hof, P.R. 2000 Recommendations for straightforward and rigorous methods of counting neurons based on computer simulation. J Chem Neuroanat 20, 93-114.
6. Sweet, R.A., Dorph-Petersen, K. & Lewis, D.A. 2005 Mapping auditory core, lateral belt, and parabelt cortices in the human superior temporal gyrus. J Comp Neurol 491, 270-289.
7. West, M.J., Slomianka, L. & Gundersen, H.J.G. 1991 Unbiased stereological estimation of the total number of neurons in the subdivisions of the rat hippocampus using the optical fractionator. Anat Rec 231, 482-497.
List of Tables
Table 1: Previously published data on handedness(Hopkins, 1995; Hopkins & Cantalupo, 2004; Taglialatela et al, 2006)
and planum temporale asymmetry for the 12 individuals in the current study
Subject /
Tube Task HI
/Gesture Task HI
/ Planum temporale surface area AQ /Planum temporale grey matter volume AQ
CO630 / 1.000 / 0.6 / -0.1 / -0.1CO342 / 0.360 / -0.2 / 0.0 / 0.0
CO406 / 0.340 / -0.2 / -0.2 / 0.1
CO336 / -0.160 / 1.0 / -0.1 / 0.0
CO408 / 1.000 / N/A / 0.0 / -0.2
CO242 / 0.680 / N/A / N/A / 0.2
CO507 / 0.910 / 0.0 / -0.4 / -0.1
CO491 / 0.510 / 0.0 / 0.0 / -0.1
CO423 / 0.350 / 0.6 / -0.2 / -0.2
CO301 / -0.710 / 0.4 / -0.1 / -0.2
CO273 / 0.650 / 0.3 / N/A / 0.0
CO357 / -1.000 / -0.1 / -0.3 / -0.4
Table 2: Stereologic estimates and CE of shrinkage-corrected volumes (mm3) for area Tpt in each individual
Subject / Sex / Age / Whole Brain Vol. / Left Vol. / CE / Right Vol. / CE
CO630 / Female / 13 / 415.4 / 310.5 / 0.10 / 239.2 / 0.13
CO342 / Female / 35 / 348.1 / 375.0 / 0.06 / 535.7 / 0.06
CO406 / Female / 42 / 327.8 / 446.4 / 0.07 / 824.4 / 0.06
CO336 / Female / 44 / 332.9 / 274.5 / 0.09 / 226.6 / 0.05
CO408 / Female / 45 / 312.9 / 474.9 / 0.05 / 263.6 / 0.06
CO242 / Female / 48 / 298.2 / 569.3 / 0.09 / 168.4 / 0.07
CO507 / Male / 17 / 384.0 / 455.4 / 0.08 / 558.7 / 0.1
CO491 / Male / 18 / 364.6 / 257.1 / 0.07 / 193.1 / 0.08
CO423 / Male / 25 / 419.7 / 469.3 / 0.08 / 230.8 / 0.06
CO301 / Male / 35 / 409.3 / 291.1 / 0.08 / 256.3 / 0.12
CO273 / Male / 40 / 341.2 / 388.5 / 0.06 / 244.9 / 0.06
CO357 / Male / 41 / 377.2 / 546.8 / 0.04 / 304.0 / 0.04
Mean / 34 / 360.9 / 404.9 / 337.2
SD / 12 / 40.7 / 105.5 / 197.7
CV / 36 / 11.3 / 26.1 / 58.6
Table 3: Stereologic estimates and CE of neuron density (1000s/mm3) and total neuron number (×106) for area Tpt in each individual
Left / Right
Subject / Sex / Neuron density / Total Neuron No. / CE / Neuron density / Total Neuron No. / CE
CO630 / Female / 30.19 / 9.37 / 0.07 / 25.54 / 6.11 / 0.09
CO342 / Female / 33.37 / 12.51 / 0.06 / 24.01 / 12.87 / 0.08
CO406 / Female / 34.03 / 15.19 / 0.06 / 39.89 / 32.89 / 0.05
CO336 / Female / 53.18 / 14.59 / 0.08 / 25.69 / 5.82 / 0.08
CO408 / Female / 40.27 / 19.12 / 0.06 / 25.61 / 6.75 / 0.06
CO242 / Female / 10.66 / 6.07 / 0.07 / 22.04 / 3.71 / 0.06
CO507 / Male / 34.79 / 15.84 / 0.06 / 26.29 / 14.69 / 0.06
CO491 / Male / 25.53 / 6.56 / 0.07 / 12.94 / 2.49 / 0.14
CO423 / Male / 44.71 / 20.98 / 0.06 / 59.23 / 13.67 / 0.06
CO301 / Male / 34.47 / 10.03 / 0.08 / 24.24 / 6.21 / 0.09
CO273 / Male / 34.49 / 13.39 / 0.07 / 27.42 / 6.72 / 0.05
CO357 / Male / 28.01 / 15.31 / 0.06 / 38.33 / 11.65 / 0.07
Mean / 33.64 / 13.25 / 29.27 / 10.29
SD / 10.39 / 4.60 / 11.74 / 8.17
CV / 30.89 / 34.72 / 40.11 / 79.39
Table 4: Volumes and centroid, x, y, z coordinates for the regions of area Tpt that overlap in at least 5 of 12 individuals
Centroid
Hemisphere / Volume / X / Y / Z
Left / 219 / 105 / 73 / 71
Right / 529 / 24 / 73 / 68