According to the neurodevelopmental hypothesis of schizophrenia, structural brain abnormalities may be one manifestation of a dysconnectivity syndrome in frontotemporolimbic circuits (1–3). The prefrontal region is proposed to be of considerable importance for the pathogenesis of schizophrenia, on the basis of empirical evidence from both neuroimaging (4–8) and postmortem (9–11) studies. The results of magnetic resonance imaging (MRI) studies of frontal volume in schizophrenia are not entirely consistent: there have been reports of both significantly lower volume (5, 7, 12) and no differences (13–15) relative to comparison subjects. In addition, volumetric measures do not generally provide information regarding mechanism. Assessing gyrification as an alternative strategy may, however, provide evidence in postmortem samples for disturbances of development. The central sulcus is observed at about the 20th week of gestation, and the primary fissures are observed near the 26th week (16). Gyrification occurs after the completion of neuronal migration. One model proposes a mechanism related to differential laminar growth in the cortex (17). The gyrification index was proposed by Zilles et al. (18) as a quantitative assessment of gyrification. This measure is calculated as the ratio of the length of the inner perimeter of the cortex (including the depths of the sulci) to the length of the outer perimeter. Gyrification becomes progressively more complex during gestation. The gyrification index reaches a stable plateau value soon after birth and appears to be stable thereafter, even in the presence of atrophic processes affecting gray and white matter at a later age (19). One MRI study (20) applied the gyrification index technique to investigate the left hemisphere in a small group of patients with schizophrenia. A significantly lower than normal mean gyrification index was observed. The gyrification index technique has been most extensively studied and validated in postmortem studies of normal brains, and to our knowledge there have been no postmortem studies of the gyrification index in schizophrenia. The present study was designed to test the hypothesis that disturbances in gyrification could be detected in the prefrontal region of postmortem brains from individuals with schizophrenia.
The prefrontal region was studied in postmortem tissues from 24 patients with schizophrenia and 24 age- and sex-matched comparison subjects, all collected since 1985. Before autopsy, informed consent was obtained from the nearest relative or from the responsible authority in cases of patients under legal care. These procedures were approved by the local ethics committee of the Heinrich-Heine Universität Düsseldorf, where the brains were obtained. Patients with severe systemic illness (metastatic cancer, arteriosclerosis with myocardial infarction or stroke) were excluded from the study. The diagnosis of schizophrenia was established according to DSM-III-R and ICD-9 criteria, on the basis of chart review. All diagnoses of schizophrenia satisfied both the DSM-III-R and ICD-9 criteria.
The anterior boundary of the prefrontal region was defined as the frontal pole, and the posterior boundary was defined as the genu of the corpus callosum. After formalin fixation, the prefrontal region was removed from the hemispheres by using a cut orthogonal to a reference line joining the temporal and occipital poles. The tissue was then embedded in paraffin. Serial sections 20 µm in thickness were then cut, beginning with the level of the genu of the corpus callosum. Modified Heidenhain-Woelcke staining was performed on every 50th slice. We have reported further details previously (21).
Gyrification Index Measurement
The gyrification index, or ratio of the inner to the outer contour of the cortex (18), was measured by tracing an inner contour defined by the complete pial surface of the brain and by tracing an outer contour according to tangential lines connecting the crests of all gyral curvatures (F1). The lengths of these contours were determined as the numbers of pixels in the traced lines by means of an image analysis system (Optimas 6.0, Windows 95). The ratio of the length of the inner contour to the length of the outer contour was calculated in each slice for all subjects. Data for the gyrification index were obtained bilaterally from five different slices of the prefrontal region in 48 brains, without knowledge of diagnosis. From the frontal pole, every 100th slice (interslice distance, 2 mm) was examined. Measurements from the most anterior two slices were subsequently excluded, since tangential cuts of gyri appeared to disproportionately alter the gyrification index. The mean gyrification index of slices 3, 4, and 5, covering the prefrontal region, was calculated. Test-retest and interrater reliabilities were above 95% as assessed by t tests for paired samples.
Area measurements of gray and white matter were determined, without knowledge of diagnosis, by means of planimetric analysis (21) of the same slices used for measurement of the gyrification index. The ratio of gray to white matter was calculated by dividing the gray matter area by the white matter area. Asymmetry coefficients for the total area measurements were obtained according to the formula 2 × (right–left) / (right + left) of Galaburda et al. (22). A standard postmortem shrinkage factor of 2.25 was applied to all measurements to compensate for the effects of dehydration before paraffin embedding (21). Test-retest and interrater reliabilities for these measurements were above 95% as assessed by t tests for paired samples.
Differences between the demographic characteristics of the schizophrenic and comparison groups were assessed by using two-sided t tests. The dependent variables were the gyrification index (left and right), measurements of gray matter area (left and right) and white matter area (left and right), the ratio of gray to white matter (left and right), and the asymmetry coefficient. First, correlative calculations (Pearson, two-sided) were used to investigate possible effects of age, postmortem time, fixation time, and brain weight on the dependent variables. Second, multivariate analysis of covariance (MANCOVA) was used as the initial statistical procedure with the factors diagnosis and sex. Because the dependent variables could depend on the area measurements, total area (total right area plus total left area) was used as a covariate in all MANCOVA calculations. Potentially confounding variables detected in the correlations were included as additional covariates. Third, significant findings from the multivariate analyses were followed by univariate analyses and subsequent t tests for unpaired samples for the appropriate dependent variables.
Descriptive statistics and confidence intervals for variables that might influence the measurements are shown in t1. At first hospitalization the female schizophrenic patients were significantly older than the male patients (t=–2.80, df=21, p=0.01). The brains of the male patients were significantly heavier than the brains of the female patients (t=4.09, df=18, p=0.001). The mean duration of illness for the male patients was longer than that for the female patients; however, this difference was not statistically significant. No significant differences between the diagnostic groups in mean age at death, postmortem delay, or fixation time were observed.
Correlations between the demographic data and the dependent variables revealed significant associations of the left-sided gray and white matter areas to brain weight (gray matter: r=0.34, df=38, p=0.03; white matter: r=0.32, df=38, p=0.04) and of the right-sided gray and white matter areas to age (gray matter: r=–0.33, df=46, p=0.02; white matter: r=–0.29, df=46, p=0.05). No other correlations between the dependent variables and the potentially confounding variables were found.
Descriptive statistics on gyrification, including 95% confidence intervals, are listed in t2. MANCOVA statistics are shown in t3. For gyrification, only on the right side was a significant diagnosis-by-sex interaction observed, and there was no significant main effect of diagnosis or sex. Analysis of variance also indicated a significant diagnosis-by-sex interaction only on the right side. Subsequent t tests for unpaired samples were performed for the male and female subgroups separately. The male schizophrenic patients differed significantly from the male comparison subjects (t=–2.98, df=20, p=0.007), whereas the female diagnostic groups did not differ significantly. The mean difference was 7%, indicating a relative hypergyria of the right prefrontal region in the male schizophrenic patients (F2).
Descriptive statistics, including 95% confidence intervals, on gray and white matter areas, the ratio of gray to white matter, and asymmetry coefficients are listed in t2. MANCOVA statistics are shown in t3. The significant finding of an influence of sex on left gray and white matter area was not corroborated in the univariate analysis. No significant main effect of diagnosis or sex was observed for any of the other measures.
The main finding of the present study of the prefrontal region in schizophrenia is the significantly greater than normal gyrification on the right side of brains from male schizophrenic patients. However, especially in the interpretation of the presented negative results, the small number of subjects has to be taken into consideration.
The gyrification index appears to be a reliable measure of gyrification (18, 19), which is stable throughout adult life. The gyrification index did not show any significant relationship with either age or brain weight in the present study group. This confirms that the gyrification index is an ontogenetic stable feature, at least in two different groups of subjects, i.e., the present study group and the group worked up for the anatomical study (19). The difference in the gyrification indexes for the right prefrontal region in the schizophrenic and comparison subjects thus suggests the presence of a developmental abnormality affecting this region in male schizophrenic patients. In the only other available study of the gyrification index in schizophrenia (20), which used MRI, nine right-handed male schizophrenic patients had a significantly lower mean gyrification index for the entire left hemisphere than did comparison subjects (F=5.56, p<0.03). There was no interaction between diagnostic group and cortical region. However, the results of that study are difficult to compare with the present findings for several reasons. First, only the left hemisphere was assessed in the MRI study, and different findings could have been observed on the right side. Second, another issue concerns the differences in resolution between magnetic resonance images acquired as 1.5-mm slices and then reformatted for analysis to 2.0 mm, in the MRI study, and 20-µm-thick postmortem sections, in our study. Third, the existing validation of the significance of the gyrification index is limited to postmortem studies (18, 23). Other investigations of the prefrontal region in schizophrenia demonstrated wide cerebral fissures in the frontal area (4, 24) and an abnormal frontal gyral pattern in a single schizophrenic patient (25). A three-dimensional MRI study could not demonstrate any relevant abnormalities in the frontal lobe (26).
Although abnormalities of the gyrification index are likely to reflect disturbances in processes of brain development, the mechanism of cortical folding is still unclear. A difference in growth "pressure" between cytoarchitectonic layers in the neocortex has been proposed (17). Reports of disorganization of the architecture and distribution of neurons in prefrontal regions in schizophrenia could be consistent with this possibility (27–29). Another proposed mechanism of gyrification is given by a tension-based theory of morphogenesis (30). According to this model, differences in mechanical tension along axons, dendrites, or glial processes connecting different brain regions result in cortical folding. If this model is correct, patients with schizophrenia and abnormalities of gyrification might be expected to have abnormalities of functional neural connectivity that could be demonstrated with imaging techniques such as positron emission tomography or functional MRI.
No abnormalities in gray or white matter area in the prefrontal region were observed in the postmortem brain slices we examined for gyrification in schizophrenia. Previous MRI studies of total prefrontal volume in schizophrenia have shown lower than normal volume (5, 7, 31–35) or no difference from comparison subjects (13–15, 25). The results of MRI studies used to differentiate between gray and white matter volumes have been similarly inconsistent (7, 15, 31, 35, 36). Using the plane-cut method of sectioning the frontal lobe at the frontal border of the corpus callosum, Breier et al. (35) found a significantly lower than normal volume in schizophrenia, which was pronounced in the white matter. In contrast, Zipursky et al. (31) described widespread low volumes in the gray matter. Similarly, Schlaepfer et al. (7) found significantly lower than normal volumes in the gray matter of all heteromodal association areas. Suddath et al. (15) did not find any volume differences when using the plane-cut method. Low volume in the prefrontal region has been correlated with deficits in neuropsychological performance (6, 8, 26, 27).
The present findings of disturbed gyrification in schizophrenia support the concept of a neurodevelopmental component in the mechanism of illness. These abnormalities of cerebral structure could be markers of developmentally impaired connectivity between cortical regions in schizophrenia. Whether these gyrification disturbances also affect the clinical presentation of schizophrenic patients remains to be established through in vivo experiments.
Presented at the 9th Biennial Winter Workshop for Schizophrenia Research, Davos, Switzerland, Feb. 7–14, 1998. Received July 17, 1998; revision received July 13, 1999; accepted July 21, 1999. From the Department of Psychiatry, Friedrich Wilhelms University of Bonn. Address reprint requests to Dr. Vogeley, Department of Psychiatry, Friedrich Wilhelms University of Bonn, Sigmund-Freud-Strasse 25, 53105 Bonn, Germany; email@example.com (e-mail).
Brain Slice of Prefrontal Region in Normal Man Showing Tracings of Cortex Contours Used to Determine Gyrification Indexa
aThe gyrification index is the ratio of the inner contour (white line) to the outer contour (black line). The index was measured with automatized image analysis after manual tracing of the contours and was calculated according to the method of Zilles et al. (18).
Gyrification Indexa in the Prefrontal Region of Postmortem Brains From Male and Female Schizophrenic Patients and Normal Comparison Subjects
aRatio of inner and outer surface contours of the cortex.
bThe red horizontal lines represent mean values, and the shaded boxes are 95% confidence intervals.