cell concentration and laminar thickness in the frontal cortex of psychotic patients; studies on...

CELL CONCENTRATION AND LAMINAR THICKNESS IN THE FRONTAL CORTEX O F PSYCHOTIC

PATIENTS; STUDIES ON CORTEX REMOVED AT OPERATION

LEWIS P. ROWLAND' AND FRED A. METTLER Department of Neurology, College of Physicians and Surgeons,

Columbia University

INTRODUCTION

During the past 50 years numerous papers have appeared reporting the presence of significant histopathologic alteration in the cerebral cortex in schizophrenia. Almost as many papers have been published which deny the significance of these changes, and the issue has remained moot. Most recently, Wolf and Cowen ('49) have taken a skeptical attitude towards the importance of observed cellular alteration, while Winkelman and Book ('49) feel that the changes noted are present in such a degree as to be of importance. Fo r a fuller discussion of both points of view, these papers and their references may be consulted.

Proponents of the hypothesis that schizophrenia is at- tended by cortical pathology have reported a series of cellular changes which finally results in a dropping-out of cells. However, most of the observers who have reported cell loss have done so on the basis of subjective impressions, reporting both focal acelluler areas and diffuse loss of cells. The presence of changes in the frontal lobes has been most often reported, but other parts of the cerebrum have been impli- cated as well. There has been no agreement as to the subdivi- sion of the frontal lobe most consistently and most severely affected, nor has there been agreement as to which cortical

James Hudson Brown M~morial Junior Fellow

256

THE JOURNAL. OF CONPARATIYE NEUROLOGY, YOL. 90, NO. 3 JUNE 1949

256 LEWIS P. ROWLAND AND FRED A. METTLER

layers are affected, although involvement of the third layer has been most often reported.

Only 4 authors have actually counted the number of cells in the schizophrenic cortex and their results are inconclusive and contradictory.

Dunlap ( ’24, ’28) and his associates counted, independ- ently, specimens from the first frontal convolution of 8 young schizophrenics and 6 controls after sudden death. They reported that “in spite of considerable variation in the cell counts in the individual cases, the general averages obtained” by the three independent observers “are very close,” and they found no significant difference between the schizophrenics and the controls.

Bouman (’28) reviewed the literature up to that time and made cell counts. He concluded that the frontal and temporal lobes were most often affected by a loss of cells which extended over all layers, though not to the same degree. I n three cases, the third layer was most often involved, but not in all.

Hechst (’33) also did cell counts and reported cell loss in all layers ; the most severely affected was, again, the third. He found the greatest loss in the prefrontal region, the in- fraparietal region, and in some areas of the first temporal convolution.

Hechst criticized Bouman for including glial cells in his counts, and counted only neurocytes in his own work. It is difficult to determine how much difference this made in their results because Bouman did not state his figures exactly, but presented results in a graphic form which hinders any attempt at comparison. Hechst compared his own results for schizophrenic brains with the normal values given by Economo and Koskinas ( ’25).

Most recently, Ferrero ( ’47) made quantitative studies on area 9 of Brodmann (area F D of Economo and Koskinas) in 67 cases, including 9 schizophrenics. He could not confirm the reported cell loss and felt that the counts fell within the limits of individual variation.

NUMBER O F CELLS IN FRONTAL CORTEX 257

Thus, quantitative studies have led to no further elucida- tion of the problem as to whether or not there is significant loss of cells in the schizophrenic cortex. Bouman and Heclist found a diminished number of cells, while Dunlap and Fer- rero did not. Discrepancies have also been noted in similar studies on the cortex in mental deficiency. Inaba ('27) and Hammarberg (1895) report a diminished number of cells in this condition, but this is denied by Norman ( '38). A re- view of the methods which have been used by various authors to enumerate the cells of the cerebral cortex may explain some of these inconsistencies.

CELL COUNTS ON THE CEREBRAL CORTEX

&ie t h o ds Three basic methods have been utilized to count cells in

the cortex: the ocular micrometric, the projective, and the photographic.

The use of an ocular micrometer has been employed most often and was used in the studies of Mayer ( '12), Berger ( '21), Berry and Norman ( '33), Bonin ( '38), Bok and van E r p Taalman Kip ('39), Conel ('39, '41, '47), Lashley and Clark ( '46) and Ferrero ( '47). I n this method a glass plate ruled off into measured squares is placed in the microscope ocular and the number of cells appearing within a prescribed area is calculated.

The technique of projecting the slide to be counted upon a surface previously ruled into squares has been used by Santha ( '33), Hechst ( '33) and Bianchi ( '42, '43). This method is probably comparable to the use of an ocular micrometer.

The third method, that of counting from photographs, is open to serious criticism. This is the more important, because it was the method used by Economo and Koskinas ( '25). These authors stated at that time that their figures could serve as normal values henceforth and have been so regarded by many workers since then. However, as Agduhr ('41) has pointed out, cells which overlap each other in a piece of


tissue will appear as one in a photograph. Furthermore, it is difficult t o count all the cells in a section, since they actually lie in several optical planes while a photograph focuses on a single plane. I n addition, at the magnification used by Economo and Koskinas it is alniost impossible to distinguish clearly between small neurons and glial cells, a distinction which is often difficult to make even when the cells are examined with an oil immersion lens. Finally, Agduhr pointed out that cells lying at the surface may appear in more than one section, and that counting these cell-fragments as whole cells makes the count too high, as will be further discussed below. These criticisms also apply to the work of Bouman.

Two other methods, essentially comparable to the use of an ocular micrometer, have been used by single investigators. Hammarberg (1895) projected upon the slide to be studied the image of a micrometer etched 011 a glass plate (Abbe's apparatus). In this technique the microscope lenses could be changed without disturbing the micrometer. Dunlap ( '24), on the other hand, merely counted the number of cells per high power field.

Preparatiofi of specimens

Some of the discrepancies in the results of different investigators are probably due to the fact that no constant method of preparation has been employed. Thus, paraffin, celloidin and frozen sections have all been used by one or another of the authors mentioned. As fixatives, formaldehyde, formol and alcohol have been used in varying concentrations. That these agents may have different and variable effects on the cell picture has been shown by Sarkissow ('30a, '30b), Rose ('29, '35) and others. A method of correcting for shrinkage has been suggested by Sugita ( 'B), but this was not used in any of the above studies.

Cells counted

Another source of variation has been the inconstant criteyia for cells to be included in the counts. Hammarberg (1895),

N U M B E R O F CELLS I N FRONTAL CORTEX 259

Berger ('21)' Berry and Norman ('33) and Cone1 (personal communication) counted only those neurons which con- tained nucleoli. Lashley and Clark (personal communication) counted those with nuclei. Economo and Koskinas ( '25) ' counted all nerve cells and cell-fra,gments. Mayer ( ' l a ) included the glia in his estimations, as did Bouman. Some of the authors, including Hechst and Ferrero, make no mention of their criteria.

The kind of cell counted assumes greater importance when one considers the error entailed in counting cell-fragments which lie at either surface of the section, their bodies lying in more than one section. Because of the presence of such cells, it is erroneous to assume that the number of cells per unit area in a 30 p section is twice the number in a 15 p section, as has been shown by Bok and van E r p Taalman Kip ( '39), from whose paper the following is quoted:

NUMBER O F CELLS PER CORTEX COLUMN

Section

30 P 15 P

Theoretical column of 15 @

Mouse Rat Cavia. Rabbit

1105 707 1209 801 1221 771 914 563

398 408 450 351

The figures in each case represent the mean number of cells in 5 columns of equal width counted from the pia to the white matter. In cutting the blocks of tissue at 30p7 a pair of sections at 15 was introduced at constant intervals. The last of the 3 0 p series and the second 15 p section were examined, the first 15 ~1 section being discarded because the inicrotome was less accurate immediately after a change in thickness. The figures given are for homologous areas in different animals. One can see that the cell count in the 3 0 ~ sections is much less than twice the figure derived f o r the 15 p sections. Thus, it would be a mistake to multiply the number of cells in a square with sides of 0.1 mm in a 10 p section by 10 to arrive at the number of cells per (0.1 m ~ n ) ~ . However,


this is essentially what has been done in many of the studies mentioned above, in which 10 v, 25 p and 50 sections have been used. Thus, while most counts are too high, the margin of error varies.

Bok and van E r p Taalman Kip therefore express the number of cells as the difference between the figures for sections of different thickness, further explanation for this being given in their paper.

For the same reason, Economo and Koskinas proposed that their figures be reduced by 33% for cells up to 12 p, 25% f o r cells up to 6 p, and 15-20% for smaller cells, if one would derive precise estimations of cell content. They felt, however, that the uncorrected figures might serve as a basis for relative comparisons.

Agduhr ( '41), however, feels that this reduction method is too gross and inaccurate, and has developed his own formula based upon cell size. Similar formulae, which are siwlilar in principle, but do not agree in particulars, have been conceived by Abercrombie ('46) and FIoderus ( '44). Back- man ('46) prefers the method of Floderus as relatively simple and accurate.

Bianclii ( '42), on the other hand, feels that the use of these formulae may be avoided by counting nucleoli only in 30-40 squares measuring 0.1 mm on a side, if one chooses sections at intervals such that it is unlikely that the same cell will appear twice. Jones ('37) agrees that the source of error arising from split nucleoli is slight. Hammarberg (1895) and Berger ( '21) counted only nucleoli on 5 consecutive 2 0 p slides o r 10 consecutive 1 O v slides.

There are further considerations in the criteria for choosing cells. I n commenting upon the discrepancies in the calculation of the total number of cells in the cerebrum, Economo and Koskinas state that Berger ( '21), and Thompson (1889), as well as Hammarberg, must have counted only those cells showing the characteristic size and shape for the particular layer counted, omitting smaller and atypical cells, and that the criteria of size to be included varied with the author.

NUMBER O F CELLS I N FRONTAL CORTEX 261

They state that if one subtracts the figure for granule cells from their own figure for total cell content, one arrives a t a number approximately the mean of the other authors.

One final variable has been the difficulty mentioned by several investigators in differentiating small neurocytes from glial cells in Nissl preparations.

Number of counts: selection of loci

I t has been the impression of most students of cytoarchitecture that the width of the various cortical layers varies in different positions on the wall, lip and crown of a gyrus and in the floor of a sulcus. This has been most thoroughly discussed by Bok ( '29J. One would expect'this to have an effect on the cell counts within a single layer a t different loci, but this assumption may not be true, according to the unpublished observations of H. Idema (quoted by Bok and van Erp Taal- man Kip, '39). Nevertheless, Hammarberg, Berger, Ferrero, Bok and van E r p Taalman Kip, Berry and Norman, and Conel have confined their measurements to the crowns of gyri. Some of the others counted random loci, while others make no mention of what sites were selected.

The number of loci in each layer which are counted per slide and the number of slides per area have varied from observer to observer. Often no information in this regard is given at all. Perhaps the most disconcerting silence is that of Economo and Koskinas. It is assumed that they used 6 brains in their study, but they may have used more for cell counts. The exact number is not conspicuously stated. X o indication is given as to how many counts from how many slides their mean figures represent, nor are the plates in their atlas always taken from the same brain. The importance of individual variation has been emphasized by Lashley and Clark ('46) and Lashley ('47), but has apparently been in- adequately considered by most workers, including those who purport to establish normal values.

Hammarberg and Bianchi specifically state that their methods have given generally constant results. Conel (personal


communication) counts 10 adjacent squares in a horizontal row in 6-10 slides for each layer, because the “cells are not scattered uniformly throughout any on’e layer.” Ferrero (personal communication) used a similar method, but with fewer slides. Norman (’38) has analyzed his figures for statistical variation, as has Bonin (’38) to some extent. Lashley and Clark found reliable differences of 25% from homologous loci in the same brain. Few others have considered this matter.

Thus, a variety of methods of sectioning, preparing and counting cells in the cerebral cortex have been used, some of which are inadequate and some have not been reported in detail. For this reason one must use caution in comparing the results of different authors. Of the papers which have dealt specifically with the enumeration of cells in schizophrenia, Bouman’s work is subject to the same criticism as that of Economo and Koskinas, both on account of his use of photographs and inadequate regard for the possibility of individual variation, as well as for the omission of detailed description of his methods. These latter criticisms apply to Hechst, and to a lesser extent to Ferrero. Dunlap (’28) admitted that he could not distinguish between the different areas of the frontal lobes and was not certain that his sections from different cases are from homologous areas. Further- more, he was not able to differentiate the various cortical layers and therefore merely counted at a measured distance from the pia. This method seems t o be subject to error because different layers may acutally lie at the same distance from the pia in different brains o r even in different parts of the same brain.

MATERIALS AND METHODS

The material used in the present investigation has already been described from both the points of view of anatomy (Mettler, ’49) and pathology (Wolf and Cowen, ’49). Detailed studies of the clinical condition of the patients may also be

NUMBER O F CELLS I N FRONTAL CORTEX 263

found in the same volume, “Some problems of the human frontal lobe.”

The tissue examined had been surgically ablated from 23 patients, including 22 schizophrenics and one manic-depres- sive. There are no “normal” controls, but it was felt that if cell loss is actually an integral part of the histopathology of schizophrenia, then we might expect to find some correlation between the severity of the cellular diminution, if any, and the severity o r duration of the psychosis, the age of the patient or the kind of treatment previously received. Lest there be any bias on the part of the observers, such data were not consulted until all tissue examinations had been completed.

The cortical tissue used for these studies was fixed in 10% formalin (the exact time elapsing between ablation and fixation in each case is given in Mettler, ’49), dehydrated in alcohol, embedded in paraffin, and cut at l o p . Every 10th section was stained with cresyl violet and, of the slides so prepared, every 5th section was examined, except when such a section was too distorted o r when it had been used for other studies and was therefore no longer available. I n such circumstances the next consecutive slide was taken. I n a few instances the series of slides examined is grossly irregular, because of the lack of suitable tissue for counting at the usual intervals.

A binocular microscope equipped with an ocular net-micrometer and a mechanical stage was used. Counts were made under oil immersion (objective 1.8 mm, 97 x ) with a 10 X ocular. At this magnification each side of the large micrometer square measured 0.1 mm and was subdivided into squares with sides of 0.025mm, as calibrated from a Leitz Objektmikrometer with units of 0.01 mm.

Counts were made only at the crown of a gyrus and never beyond a point on the curvature of approximately 15” from the midline. Five counts were made for each layer on each of 5 slides from a portion of the section where cortical archi- tecture had not been distorted by the operative procedure.


I n counting the cells of the second layer (Brodmann terminology), the slide was adjusted so that the upper micrometer line lay just within the upper border of the layer. For layer 111, the cross-lines of the micrometer were placed at the mid-point between the lower border of layer I1 and the upper border of layer IV under low power, the objective then being changed to oil. This would correspond to I I Ib of Economo and Koskinas. Selection of the locus in layer IV was similar to that of layer 11, except that its borders are not as regular as the latter and care was taken to remain within the central part of the layer. A similar procedure was followed for layer VI, where counts were done just within its upper border, usually well-demarcated from layer V. The mid-point of layer V was chosen in a manner similar to that f o r layer 111.

In this way one proceeded down a line from layer I1 to layer VI along the direction of the radial striations, counting at the intervals described and not moving the field in a horizontal direction except when a large vessel came into view, in which case the next adjacent field was chosen. With this exception, bias was more or less eliminated in the selection of fields to be counted because of the relatively mechanical procedure.

The selection of cells to be counted posed a problem. At first it was decided to count only those cells with nucleoli. Two factors militated against this otherwise desirable technique. First, it was soon noted that in almost all cases the cells in the second layer were sclerotic or otherwise distorted, so that cell structure was obscured. (Such phenomena are often seen in so-called normal material and are not regarded as specific findings in psychoses, see Wolf and Cowen, '49.) One could not count merely those cells with nucleoli here. Furthermore, such cells were not infrequently found in layer I11 and some provision had to be made for counting them. Second, it was noted that many cells containing clearly dis- cernible nucleoli had poorly shaped nuclear- and cell-mem- branes, many resembling cell-shadows or cell-fra,gments. On

N U M B E R O F CELLS I N FRONTAL CORTEX 265

the other hand, some well-formed cells, with a symmetrical, uninterrupted nuclear membrane and well-developed cell body possessed no nucleolus. If the former were to be included in the counts it was felt that the latter should be, too. Thus the cells actually counted included all well-formed cells with or without nucleoli and all cells with nucleoli, however poorly formed. Sclerotic cells were included if the cell membrane, although malformed, was complete. Cell-fragments, cell-shadows and glial cells were not counted, nor were cells which bordered on the edge of the micrometer square, unless they possessed a clear nucleolus which touched either the upper or righthand lines. Some difficulty was noted in dis- tinguishing very small neurocytes with scanty cytoplasm from glial cells. Such cells were counted only if they possessed a well-defined iucleolus. Care was taken to count all the cells in all of the optical planes included in the section.

Because the sections are 10 p thick these counts would have to be multiplied by 10 to arrive at the number of cell per (0.1mm)3. The dangers of such a procedure have already been discussed and our figures a re therefore expressed as the number of cells per (0.1 mm)2 in a section of 10 p. Al- though this prevents accurate comparison with the results of other workers, it should nevertheless suffice for the comparison of counts within this series. Moreover, one must be cautious in comparing the results of different investigators because of the variables mentioned.

The thickness of the various layers was measured by means of a straightline ocular micrometer, the units of which equalled .015mm a t the magnification used in this aspect of the study (430 x). One measurement was made on each of 5 slides, passing in a direction from the pial surface to the white matter at the crown of a gyrus. I n flat gyri, selection of the locus to be measured was random.

Cell size, unfortunately, was not measured due to limita- tions of time, so that correction for cells lying at the surfaces has not been made. However, the present methods ought to

266 L E W I S P. ROWLAND AND FRED A. METTLER

give some indication of the trends within the series in a relative manner.

OBSERVATIONS AND DISCUSSION

Tables 1, 2, 3 and 4 give the average number of cells per .0001 mm3 [that is (0.1 mm)2 of a section 10 p thick] in the various layers of different frontal lobe areas in each of the cases studied. Each figure given represents the mean of 25 counts for each h e m i s p h e r e 4 counts on each of 5 slides. Standard deviations have been calculated according to the method described by Hill ( ’48) for the derivation of standard deviation in small series. Table 5 summarizes these two data.

Despite the variables mentioned above, most other authors have obtained cell counts (per .001mm3) which seem to fall into the same general range, as indicated in table 6. If our own figures were to be multiplied by 10 in order to make them comparable, the figures for the present series would be much higher than any of the previous estimates, almost twice those of Economo and Koskinas. The explanation for this dis- crepancy lies partly in the discussion by Bok and van Erp

EXPLANATION OF TABLES

Tables 1-4 Average number of cells per .0001 mm’ (that is, 0.1 mma of a section 10 p thick), i n the various layers of different Brodmann areas in each of the cases available for study of those areas.

Table 5 Summary of results obtained by counting the cells of the frontal lobes of psychotic patients.

Number of cells per .001 mm3 of frontal cortex according t o Brodmann terminology, as determined in six previous studies. The figures of Economo and Koskinas, Hammarberg, Berry and Norman, and Bianchi are for the ‘ ‘normal ’ ’ human; Hechst and Ferrero ’s results are for schizophrenics.

Overall thickness of frontal cortex and thickness of specific layers for the various Brodmann areas (table 7 ) and area 9 alone (table 8) a re determined by the present investigators on biopsy material obtained from psychotic patients (tables 7 and 8) , by Economo and Koskinas on post-mortem material from “normal” individuals (tables 7 and 8), and by various other authors (table 8).

Table 6

Tables 7 and 8

NUMBER O F C E L L S I N F R O N T A L CORTEX 267

Taalman Kip already cited, but the figures of the remaining authors a re subject to this source of error, though perhaps not to the same degree, and are still lower than ours. Failure to account for split cells is, therefore, probably only a partial explanation.

TABLE 1

Number of cells per do01 n i d

( A R E A 6 ) 2~

i 24L R

( A R E A 8 )

7L R

8 R

25L R

27L R

M E A N

R A N G E L-_= STAND DEV.

2

9.76

11.00 12.72

14.32 16.36

12.83

9.76 - 16.36

2.63

9.68 14.04

11.96 12.52

12.32

12.44 14.04

10.60 12.68

12 25

968- 14.04

1.442

___-

__-- __-

~-

3

3 36

4 32 5 44

4 0 0 4 12

4 25

3 3 6 - 5 44

0 241

3 12 4 68

3 24 4 96

4 16

4 6 0 4 72

3 84 4 8 0

4 23

3 12- 496

727

_ _ _ ~ ____

___ -

4

- - - - -

8 16 10.64

I I 12 I3 24

II 32

13.40 13.76

7 20 9 24

10 90

7 2 0 - 13 76

2 347 =__=

5

3.04

3.56 4 92

4 96 4.60

4.22

3.04 - 4.96

0.268

4.72 4.92

4.0 0 5.00

4.68

5.08 5.84

3.44 4.92

4.73

3.44- 5.84

J I I

6

4 24

5 44 6 48

6 16 5 92

5 65

4 24- 6 48

0 279

6 0 4 7 36

6 16 7 24

6 24

744 8 04

4 64 652

6 6 3

6 0 4 - 8 0 4

I022 __ ____

Measurements for cortical thickness (table 7) show that our specimens are also significantly thinner than those of previous investigators, a s compared in table 8. This suggests that the present biopsy specimens are thinner and more concentrated than those of previous studies, all of which studies were made OH post-mortem materid. If a certain amount of post-mortem swelling occurs, one would expect cerebral

268 LEWIS P. ROWLASD A S D F R E D A. METTLER

12.24

16.16 18.72

12.84

tissue taken froiii cadavers to be broader aiiil less concentrated. Such a tendency was postulated by Rose ('as), but his work canriot be considered coiiclusive. Weil ( '45) points out the importance of the time elapsing between death and autopsy iii any consideration of post-mortem swelling and

3 8 0

5 20 6 84

4 48

TABLE 2

Number of cells per do01 mrn8

10.92 14.48

13.96 13.28

14 00 13.84

( A R E A 9) 3L R

4L

7L R

8 R

13L R

4.16 4 32

4 40 5.44

5.32 4.68

22L R

25L R

2 7L R

31L R

32L R

MEAN

RANGE

, STAND. DEV.

2

15 44 13 08

1184

I4 04 I5 32

12 08

I4 52 15 80

I I 28 15 08

1504 I4 04

1436 12.96

1456 13 44

15 52 13.84

I4 01

1128- 15 80

1.371

3

5 40 4 9 6

3 64

4 04 5 00

3 64

5 0 8 4 64

3 72 5 04

4 60 5 24

5 60 4 36

5 96 6 8 0

4 72 5.5 6

489

3 64- 6 80

0 844 _ _

1

415 12.60 1232 I 447322

13.76 1 5 6 0 13.92 4.68

13.28 5.08 15.52 I 6.28

15.80 15.76

10 92- 3 8 0 - 18.72

1.610 0.791

6

6.88 7.24

6.96

7.32 8 28

7.32

832 7.64

6.60 7.00

736 7.40

7.00 6.08

7.28 8.04

8.12 828

7.39

6.08- 8.32

0.910

this pheiioiiieiioii is also alluded to by Ehrnrooth ( '29). Strecker ('29) fouiid evidence to suggest that the swelling occurs p a ~ t l y at the expense of the cerebrospinal fluid, which decreases in amount in proportion to the time after death. However, Reichardt ( '19)? who studied extensively the phe- iiomeiia associated with hraiii swelling, was iiot convinced that swelling coiisistciitlF occui's after cleatli, altliough he

NUMBER OF CELLS IN FRONTAL CORTES 269

1 STAND. OEV. -

described it in a variety of pathological conditions. Camerer ( '48) expressed the opinion that post-mortem swelling regu- larly occurs and tends to interfere with estimates of the effects which are produced by fixatives and other fluids.

Explaiiation of the higher count in our material on the basis of post-niortem swelling in the material used in pre-

TABLE 3

Ntimber of cells per .0001 nu&

LAYER

AREA 10) 7~ R

13R

21L R

22L R

27R

32R

33L R

36 R 4 4L

49L R

MEAN

RANGE

2

14.36 13.40

15.92

16.36 15.52

13.72 14.80

13 12

12.48

13.88 16.16

12 84

17.08

14.00 1328

14.46

12.48 I708

1.594

3

4.92 6.04

4 .OO

4.8 4 4 84

4.3 6 4 40

3.88

4.7 2

4 44 5.76

4 52

5.60

4.32 4.36

4.74

3.88- 6.04

0.609

4

15.60 1784

I3 48

15 00 13 48

15.44 14.28

1460

15.00

I3 92 14.40

12.88

I5 56

16.48 16.24

I4 95

12.88- 17.84

1.290

5

5 48 6 04

3 96

5.20 5 04

4 60 4 76

4 0 0

6.80

4 40 4 84

5 00

5 44

5.04 5 16

5.05

3.96- 6 80

0.233

6

7.04 8 44

7. 12

7.92 7.3 2

7.52 7.32

6.56

8.20

7.80 6.96

7.24

8.08

7.84 7.84

7.55

6.56- 8.44

0.254

vions studies is complicated by other factors, not the least of which is the difficulty in defiriiiig the boundaries between the various larers and between the gray matter and the white. Furthermore, embedding in paraffin may cause more shrinkage than celloidin (Stowell, '41). It should also be noted that according to Sarkissow ( '30a) the shrinkage of different brains in the siiiiic enibedding inaterial will he different ; even

270 LEWIS P. ROWLANI) A K D FRED A. METTLER

7.92 7.72

6.84

7.51

6.84- 8.48

0.585

6.96

7. I2

7.04

7.80 6.24

4.68

6.16

7.12 6.84

6.47

4.68 - 7.80

1.100

6.72 7.04

6.08 6.48

6.44

7.76

6.75

6.08- 7.76 0.637

_.__

AREA II) 1 8 L

R

1

3 6 L R

42L R

49R

MEAN

RANGE

STAND. DEV.

AREA 4 4 ) 33L

47L

ME AN

(AREA 45) 13L R

I9R

33L

36 L R

MEAN

RANGE

STAND. DEV. (AREA 46) 4L

R 22L

R

36R

49L

M E A N

RANGE

STAND. DEV.

TABLE 4

Xirmber of rd l s per .@0@1 mnl'

2

13.84 12.96

t4.64 13.80

13.56 I I .20

12.12

13.13

I I .20 -. 14.64

1.258

15.0

16.24

15.62

____

12 80 14.56

9.20

I3 88

I5 20 15 28

1349

9 2- 15 28

2 297

13 04 11.96

I I 4 0 10.96

15 4 4

I4 00

12 80

10 96- 15 44 1712

3

4 04 4 60

4 32 4 80

3 9 6 4 0 4

4 4 4

4 31

3 96- 4 80

0 371

5 24

4 60

4 92

5 12 4 36

3 20

4 84

4 24 4 84

443

3 20- 5 12

0 721

5 12 3 64

3 36 3 88

4 36

4 64

4 17

3 36- 5 12 0 632

-~ -__

~ _ _

4

14 52 14 32

1268 12 80

I3 52 13 08

13 00

I3 42

1268- I4 52

0 717

15 24

10 52

12 88

12 24 12 36

10 36

II 56

14 12 12 60

12 21

10 36- 14 12

I124

I3 44 14 44

12 16 12 48

13 40

16 56

I3 77

12 16- 16 56 I536

___

5

5.64 5.80

5.44 5.4 8

5.6 4 5.36

4.76

5 45

4.76 - 5.80

0.249

4.88

4.20

4.54

4.60 4.32

3.00

3.88

4.28 4.28

4.06

3.00 - 4.60

0.5 74

5.00 4.92

4.04 5.20

4.40

6.96

5.09

4.04- 6.96 0.996

____ .____

-___

NUMBER O F CELLS IN' FRONTAL CORTEX 271

different par ts of the same brain may shrink to different deg Orees.

Kevertheless, one additional factor suggests that pre- mortem tissue is more compact than the same tissue after death. Most of the other authors have sectioned their material perpendicular to the surface and to the longitudinal axis of the gyrus, in the manner prescribed by Economo and

TABLE 5

Iluntber of cells per .0001 mn? ia carious areas of t k e human frontal lobe

STAND. DEV

_____ ~

MEAN I 2 1 44 RANGE

I 1 STAND. DEV. 1 I

MEAN

6 1 45 RANGE

STAND. DEV

MEAN I 1 6 1 46 RANGE

I STAND. DEV . ..

12.83 9.76-

16.36

9.68- 14.04 1.442

3.36- 4'25 5.44 I 241

4.23 10.90 7.20 13.76 2.347

1 ::) 1401 1 I I 28- 3.64- 10.92- 15.80 6.80 18.72

14.46 4.74 14.45 12 48- 3.88- 12 88- 17.08 17.84 1.594 ,609 I .290

1.371 -

3.44 6.04 0.04 1.022

5.02 7.39

6.84 ,791

I I 5.05 7.55 3.96- I ::3:- I 6.80 .233 1 .254 1

272 LEWIS P. EOWLAND A S D FRED A. METTLEF:

4 ECONOMO 8 1 6-8 KOSKINAS

~ --

Koskinas. However, the biopsy tissue in the present investigation was sectioned in a constant frontal plane, regardless of the direction of the gyrus. For this reason man:- of our

TABLE 6

Cell counts of various authors; no. of cclls per .001 wim3

I I -I-. LAYER ' AREA '-.

.

1 HECHST I

BERRY a , NORMAN

.~ -

BlANCHl ' t - -

1

1 s ECONOMO 8 1 K O S K I N A S

t 1 H A M M A R B E R G I t- -

H E C H S T

~-

BlANCHl

1 F E R R E R O 1 13 I

-.

a -30

49 45 42 18 17 I8 57 51 43 17 20 18

--

30 ' 29

15+ ..

79 65 65' 22 22 20 -. 1

~ 15 12

4 0 j

i I I

31 I 18

12-15

70-8

6 0

I 62 7i 17 69 7;

8 5

49

0 - 2 5

, b - 1 5 15-17

.

15 I5 I

I6 14 16 ~ 22 22 20 I5 13 14 21 20 I7

16 I 25

I- ~-

, -__ -

19 20. 22! 26 23 26

0 I -30-401 a2 - 4 0 a-25-40 b - 28 1 b - 18

15 ' 15-20 - 1

19 17 19 122 20 22 18 15 20'20 20 22

I5 30 1 30-35

I5 -17

32 I :: 1 16

sections are oblique and should therefore be broader than one mould expect in a perpendicular section. That they are actually narrower, not broader, strengthens the assumption that this difference is not spnrious.

NUMBER OF CELLS I S F R O S T A L CORTEX 273

I t must be emphasized that the normal range of values for neither cell concentration nor laminar thickness in the frontal lobe is known with accuracy, even in post-mortem tissue. In the current study, considerable overlap from a I*CR

TABLE 7

Average tAicPness of layws ( m n ) for the kirinan f ron ta l lobe in biopsy specimens

Figures in ( ) from Econonio and Koskiiies

itandard devi-l Y, of E.B K. ' ition of total i measurement I

I 1

to area was seen for both cell concentration and laiiiiiiar thickness. Tli~is, for instance, altliougli the average figui*c~ for cell polnilat ion and laniiiiar thickness of the pyramidal layers tends to decrease as one procedes rostrally, in iridi-

374 LEWIS P. ROWLAND A N D FRED A. METTLER

BRODMANN

HAMMARBERG

FERRERO _ _ _ _ _ _ _ _ ~ -

L P R 6 FAM

vidual cases one might find figures for a rostra1 area which exceed the average value for a more posterior locus.

I n addition to the role of individual variation in accounting for this overlap, other variable factors must be considered.

PARAFFIN - 1 - PARAFFIN .30 .20

_. - -~

PARAFFIN 209 1145

-1 ~

PARAFFIN

TABLE 8

1 7 i i c k i i ~ s s i i i nini of area 9 accordircg t o various azithors

E C O N O M O e KOSKINAS I

m .78

- _- I .30 ._

.911

.70

IY

.21

- ___

. 20

.I59 ~.

. I 3

(3.45)

.49 2.22

Average figures for 2.5 coziwts compared with larger sanipk in area 9 1 27 1 iz 1 LEFT ~ 14.36 I 5 .60 1 14.00 , 5.31 1 7.00 1 13.93 5.15 13.80 5 .32 7.00 1 14.56 I :J 13.28 5.08 7 2 0

12 70 I 5.04 6.87

2 5 LEFT 3' 5 0 13 20

I

i 31 1 25 6 3

~ RIGHT I 13.44 I 6.80 i 15.52 1 6.28 I 8.04 1

In selecting areas to be counted, an attempt was made to avoid transitioiial areas. However, this could iiot always be done with certainty, since the identification of the various areas within the granular portion of the frontal lobe is not always obvious. If there is a, gradient as one passes from the frontal pole caudally, with an increasing total cortical

NUMBER OF CELLS I N FRONTAL CORTEX 275

thickness, an increase in width and cell coiiceiitratioii in the pyramidal layers and a decrease in these values for the granular layers, then some variation might be expected if one does not choose analagous sites within iiidividual areas of the cortex for calculation. Similarly, the difference in the amount of curvature of a gyrus may have an effect on the number of cells, even in relatively analogous loci. For all

TABLE 10

Correlation of cell counts g i t h varicxts factors Nwnzber of cells per .001 mnlP by layers

“Figures in parentheses are standard deviations

DURATION (<22 mos )

E S T 14.00 14.04

INSULIN

(+ I 14.72 (-1 14.08

METRAZOL 14.78 14.52

m

5.17 ( .9681* 4.67 ( ,667)

4.92 (1.004) 497 ( ,994)

4.88 ( ,963) 4.90 ( ,655)

4.17 (1.081) 4.75 ( ,736)

5.16 ( .377) 4.74 ( ,895)

4.74 ( .653) 5.04 (l.OOl)

5.34 (1206) 4 6 9 ( ,837)

4.72 ( 689) 476 ( 834)

4.82 [ 715) 4.90 ( ,629)

4 9 0 ( .824) 4 5 6 (.244)

4.83 t.791) 4.60 (.238)

5.16 ( 615) 474 ( ,599)

~

4.92 ( ,791) 4.54 ( .312)

n?

13.66 14.41

14.67 13.48

14.29 13.65

14.03 14.21

14.12 14.07

I4 98 13 18

I4 52 13 61

14.44 15.39

14 73 15.19

14.79 15.13

15.01 14.85

~~

15.28 14.92

15.58 14.22

4.29 (2.896) 1 7.31 4.93 ( ,834) 762 . .

5.31 ( 728) 1 7.60 4.72 f ,763) 7.20

4.72 ( .663) 7.35 5.34 ( ,823) I 7.72

t--

4.84 ( .704) 5.37 ( .709)

6.12 ( .961)

276 LEWIS P. €:OWLAND A S D FEED A. METTLER

this, however, the rostrocaudal transition seems to be very gradual in the present series ; although the trend seems real, there are not very marked differences for these values in iieighboriiig areas.

That a larger sample would prohably not have given sig- iiificaiitly different figures may be seen in table 9, in whicli the figures f o r 25 counts ( 5 slides) are compared with average figures derived from a larger number slides from the same case.

Because the normal values for cell concentration in pre- mortem tissue are not available, the data were analyzed in the following manner:

For areas 9 and 10, the cases were divided into two groups for each of 6 factors. Those who were notably deteriorated were compared with those whose outlook was more favorable ; those with a “psyohiatric grade” of 0-2 were compared with the more favorable group (“psychiatric grade” of 3-5). Those with long periods of institutionalization were compared with those who had spent less time in institutions. Younger patients were compared with older. Finally, comparisons were made between those patients who liad received, respectively, shock, insulin and metrazol therapy and those who had not. For area 9, one additional category was analyzed, those who liad had rapid EEG activity before operation were compared with those wlio had not. For each category the average number of cells was calculated from the average figures for the cases in that group and the results are given in table 10. I t can be seen that there are no significant differences fo r any of the factors considered.

Although the “psychiatric grade” and the duration of institutionalization are relatively ci.ude measures, one might expect to see some difference in the number of cells if cell loss is more severe in the most deteriorated cases or in those of longer duration, as suggested h g Jlislrolczy (’37) and others. The 1)wseiit figures do not lend support to such a belief.

NUMBER OF CELLS IN FRONTAL CORTEX 277

The lack of necessary change with electroshock therapy is in keeping with what is apparently the consensus among workers in this particular field (Windle et al, '45).

SUMMARY

The literature relating to cell counts on the cerebral cortex has been reviewed. Individual variation, variable fissura- tion, different methods of preparing and sectioning tissue, variable reaction of different brains to the same fixative, various methods of counting and variation in the time after death before fixation may lead to variable measurements of cellular concentration and laminar thickness. It is felt that the normal range for these values in the frontal lobe is not known with accuracy. Comparison of the results of different authors must be made with caution.

Examination of the present biopsy material from the brains of psychotic patients suggests that a certain amount of post- mortem swelling has made the tissue examined by previous workers broader and less concentrated.

Comparison of cell counts within this series shows no significant difference between those patients who had severe mental disease and those whose prognosis was somewhat better, nor between those who had been institutionalized for more than 22 months and those who had been for less than 22 months, nor between those who had received shock therapp, insulin or metrazol and those who had not. Finally, there was no significant difference between the older group of' patients and the younger, nor between those who had had rapid electroencephalographic activity pre-operatively and those who had not.

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NUMBER OF C E L L S IN F R O N T A L CORTEX 279

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1935

1930s

1930b

1941


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cell concentration and laminar thickness in the frontal cortex of psychotic patients; studies on...

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