Quantitative data on cell loss and cellular atrophy of intralaminar nuclei following cortical and subcortical lesions

Brain Research, 89 (1975) 43-59

43

© Elsevier Scientific Publishing Company, Amsterdam - Printed in The Netherlands

Q U A N T I T A T I V E D A T A O N C E L L LOSS A N D C E L L U L A R A T R O P H Y OF I N T R A L A M I N A R N U C L E I F O L L O W I N G C O R T I C A L A N D S U B C O R T I C A L LESIONS

G. MACCHI, A. QUATTRINI, P. CHINZARI, G. M A R C H E S I AND G. CAPOCCHI

Neurological Clinic, Catholic University, Rome (Italy) (Accepted December 2nd, 1974)

SUMMARY

Morphological changes in the intralaminar nuclei centralis medialis, paracentralis and centralis lateralis of the thalamus of adult cats after cortical excisions have been determined by means of a quantitative method. The number and size of the remaining neurons on the operated side have been compared with those of the normal side. The differences between the normal and the operated side have been compared to those found between the two sides in the control animals. The most important result is the demonstration that after cortical ablations the intralaminar nuclei show not only chromatolytic or atrophic changes of their cells but also a true cell loss. These reactions are qualitatively similar to those observed in the specific nuclei of the thalamus, the only difference being a quantitative one. As a consequence it can be suggested that some intralaminar nuclei project to certain areas of the cerebral cortex which also receive projections from one or other specific thalamic nucleus. A large essential connection of the intralaminar nuclei, in particular the nucleus centralis medialis, with subcortical structures is confirmed.

INTRODUCTION

The projections of the intralaminar nuclei to the cortex are of great relevance since these nuclei have been considered part of the 'non-specific thalamic system' which, on the basis of physiological effects elicited by its stimulation, must be thought to differ from the 'specific thalamic system '6. The anterior intralaminar nuclei include the nucleus paracentralis (PC) and the nucleus centralis lateralis (CL), both contained within the lamina medullaris interna, and the nucleus centralis medialis (CE). From a topographical point of view, the latter nucleus belongs to the midline group of nuclei but it can be considered

44 also as 'intralaminar' because of its close association with the lamina medullaris interna and the other intralaminar nuclei. There exists considerable disagreement among various authors with regard to the histological changes observed in the intralaminar nuclei following hemidecortication. Lashley 9 in rat, Bodian 3 in opossum, Rose and Woolsey22 in rabbit, Walker e,~','~ in primates, and Powell is in man describe only slight reactions in the nucleus paracentralis and nucleus centralis lateralis. In contrast, according to Combs 4 in rat, Waller 27 and Peacock and Combs in cat and monkey 1~,17 hemidecortication affects the neurons of the same nuclei. According to Nashold, Hanbery and Olszewski 14 the nuclei paracentralis and centralis lateralis project in a diffuse manner to the cortex, while Akimoto, Negishi and Yamada 2 and Adrianov ~ observed retrograde reaction in these nuclei following fronto-parietal ablations. In contrast to the nuclei paracentralis and centralis lateralis, the nucleus centralis medialis has been reported not to display any cell changes after cortical ablations 4,11A6,~7. The data obtained by means of the retrograde degeneration method after lesions of the basal ganglia, in particular of the head of caudate nucleus, have induced many authors to emphasize the subcortical connections of the intralaminar nuclei~,19,z4. Others 11 have suggested the existence of a projection of the nuclei centralis lateralis and paracentralis to the basal ganglia and to the fronto-parietal cortex, but could not determine whether the neuronal changes in the intralaminar nuclei represented a retrograde or a transneuronal effect. Observations with the anterograde degeneration technique have tended to confirm projections of the intralaminar nuclei to the putamen, the claustrum, the orbital cortex, the suprasylvian gyrus, the limbic cortex and the rhinencephalon~L In more recent work Powell and Cowan z° believed that they found a true retrograde reaction only in animals with lesion of the internal capsule, whereas they interpreted the cell changes in the intralaminar nuclei after cortical ablations (atrophy, chromatolysis, gliosis) as transneuronal. In contrast, recent data by Murray 13 demonstrate cell loss in the intralaminar nuclei following ablation of orbito-frontal, limbic and parieto-occipital cortical areas. In view of the disagreement among various authors in evaluating the alterations in the intralaminar nuclei after cortical ablations, the morphological changes of the intralaminar nuclei after experimental lesions have been determined in the present study by means of a quantitative method. In particular, the number and size of the neurons remaining on the operated side have been compared with the normal side. This approach provides a numerical estimate of the cell changes and, at the same time, avoids the use of imprecise terms such as 'light' or 'heavy' degeneration which have often been used in the literature.

Anatomy of the centralis medialis, paracentralis, and centralis lateralis nuclei We have traced the anatomical limits of the intralaminar centralis medialis, paracentralis and centralis lateralis nuclei according to the terminology suggested by Rose 21 and followed by Murray 13. In tracing the limits with neighboring areas we have been guided by cell structure characteristics and by the connections with the internal medullary lamina.

45 The centralis medialis nucleus has quite clear cell structure characteristics; the whole of its cranio-caudal extension, from a transverse cranial plane, going through the rostral part of the medio-dorsal nucleus to the caudal transverse plane through the middle part of the ventralis posterior nucleus. The cell structure characteristics set the nucleus centralis medialis apart from the rhomboideus nucleus, laying dorsally, as well as from the reuniens and submedius nuclei, which are ventrally located. There is a much higher concentration of cell bodies than in the nuclei reuniens and submedius; the cell shape is more uniform than in the nucleus rhomboideus and the cytoplasm coloring is more intense than in the other two nuclei. The limits with the adjacent paracentralis nucleus are less clear-cut; we have therefore set them where the elongate cell bodies of the nucleus paracentralis start to prevail, their main axis being parallel to the ventrodorsal course of the fibers of the internal medullary lamina. The paracentralis nucleus lies rostrally on a transverse plane going through the rostral part of the medio-dorsal nucleus and continues in a caudal direction, always included in the internal medullary lamina until the transverse plane going through the middle third of the ventralis posterior nucleus. It borders dorso-medially with the medio-dorsal nucleus, and ventrally with the ventralis lateralis and ventralis posterior. As regards cell structure, the paracentralis nucleus differs from the bordering intralaminar nuclei and from the surrounding thalamic nuclei because of the prevalent spindle-like shape of its cell bodies. The limit between the paracentralis and the centralis lateralis nuclei lies where the cell bodies start to assume a polymorphous aspect; their concentration decreases and their volume tends to grow. The borders of the centralis lateralis nucleus with the antero-ventral nucleus (AV), laying dorsally in the more rostral sections and with the lateralis dorsalis nucleus (LD), laying dorsally in the medio-caudal area, are well defined. This is due to the more homogeneous characteristics of the neuronal thalamic population and to the higher cellular density in the above-mentioned nuclei, as compared to the nucleus centralis lateralis. The same may be said for the medialis dorsalis (DM), and lateralis posterior (LP) nuclei, which border, respectively, medially and laterally, upon the centralis lateralis nucleus. MATERIAL AND METHODS

In the present study 14 adult cats were used. Seven of these sustained extensive cortical ablations (PG 78, PG 115, PG 116, PG 91, PG 56, PG 83, P G 73); 1 h a d a deep lesion involving the internal capsule and the basal ganglia (GA 95); 4 cats received a restricted lesion of either the primary acoustic (PG 16) or the somatic (PG 30) and visual cortex (PG 81, PG 82), and 2 cats were not operated (C1, C2). The 4 cats with lesions of the primary sensory cortex and the 2 unoperated animals were used as controls. The experimental animals were operated under aseptic conditions and after 18 days were perfused with 1 0 ~ formalin solution. The cortex and

46 thalamus were then embedded in paraffin, and serial sections cut at 20 l~,m were stained with cresyl violet. In all cases 1 of every 3 sections was used for counting and measuring cells along the entire craniocaudal extent of the intralaminar nuclei. Two parameters have been considered for quantitative evaluation: (a) the number of neurons was compared in corresponding areas of the intralaminar nuclei ipsi- and contralateral to the lesion. This provided a quantitative determination of the loss of cells on the operated side, (b) the average size of the cell bodies remaining in the intralaminar nuclei of the operated side was compared with that of the normal side. By this means it was possible to determine the degree of atrophy of the remaining cell bodies on the operated side. The calculation of the number of cells has been made by means of a graticule inserted into a × 8 eyepiece of the microscope; only cells displaying the nucleus and nucleolus were counted. The difference in number of cells between the normal and the operated side was determined by considering equal to 100 the total number of cells counted in each of the nuclei of the non-operated side. In each section the cells were counted in several fields according to the extent of the nucleus in question. Ten fields were counted in the nuclei paracentratis and centralis lateralis and 8 fields in the nucleus centralis medialis in order to examine the whole cross-sectional area of the nucleus. Each level was sampled by two examiners and the average taken. The average perikaryal size was calculated from a sample of 200 randomly chosen cells in each case. The cross-sectional area of each cell body was projected and drawn at high magnification ; the area of every cell was then measured with a planimeter. The difference in cell size between the normal and the operated side was obtained by considering equal to 100 the total surface of a corresponding number of cells measured on the normal side. The difference between the findings on the normal and experimental side was compared to that established between the two sides of the control animals. Furthermore the results have undergone statistical analysis with the Student's t-test for dependent phenomena with observations which may be coupled. It was impossible to undertake statistical analysis for experiment G A 95 since it was the only example of lesions of striatum and internal capsule. In this case however the differences between the two sides were so great that a statistical confirmation was not needed. RESULTS

(1) Control animals (see Table I and Fig. 1) (A) In the 2 unoperated animals (C1 and C2) the number of cells on one side, as compared to the other was found to differ by a maximum of 5 . 1 2 ~ for the nucleus centralis lateralis, 1.84% for the nucleus paracentralis, and 3.52 % for the nucleus centralis medialis. As for cell size variation the difference was found not to exceed 2.40 ~. (B) In the 4 cats with ablations in the somatic sensory areas $1 and $2 ( P G 30), the primary acoustic area (PG 16) and the occipital visual cortices (PG 81 and P G 82),

47

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Fig. 1. S u m m a r y of the quantitative findings on the control animals. A b o v e : the cortical lesions of 4 cats operated in sensory cortical areas of the right side whose ablation does not p r o v o k e a n y microscopical change in the i n t r a l a m i n a r nuclei. Below: each c o l u m n represents the quantitative data for n u m b e r of cells c o u n t e d a n d the m e a n cell d i m e n s i o n of the right side of the i n t r a l a m i n a r nuclei. Ca a n d C2 are n o n - o p e r a t e d controls. T h e variations in m i n u s or in plus reported with the white space in the c o l u m n s have been calculated by considering both the total n u m b e r of cells c o u n t e d a n d the average perikaryal size of the left side in the i n t r a l a m i n a r nuclei equal to 100. These abbreviations apply to this a n d the following figures. A M : n. anterior medialis; A V : n. anterior ventralis; C E : n. centralis medialis; C L : n. centralis lateralis; c l a u s t r u m ; D M : n. medialis dorsalis; G L : n. geniculatus lateralis; I: insula; L D : n. lateralis dorsalis; LP: n. lateralis posterior; P: p u t a m e n ; PC: n. paracentralis; P U : n. pulvinar; R: n. reticularis; R E : n. reuniens; T O : t u b e r c u l u m olf a c t o r i u m ; V A : n. ventralis anterior; V P L : n. ventralis postero-lateralis; V P M : n. ventralis posteromedialis.

48 TABLE I CONTROL ANIMALS

Numerical values in the right side of each intralaminar nucleus when compared with the lef! side values considered equal to 100.

PG PG PG PG C1 C2

16 30 81 82

Cell number in the right side (%)

Cell size in the right side (%)

CL

PC

CE

CL

PC

CE

~.52 --1.96 ~ 1.24 --0.52 75.12 -~ 1.86

--4.75 +0.63 --I.35 40.85 + 1.84 --0.70

--3.42 ~0.84 --3.05 4.68 --2.40 + 3.52

--5.12 +2.07 --5.44 --1.51 --2.40 ~0.44

--2.46 --2.99 2-4.90 --0.50 --0.40 +0.24

--5.16 --2.33 -t-0.45 --1.49 --0.51 +0.29

of the right hemisphere, no change in the neuronal or glial population is detectable in the intralaminar nuclei. The quantitative data indicate that in these cases the difference in the number of cells between the normal and the operated side does not exceed 4.75 ~ in the nucleus paracentralis, or 3.42 ~o in the nucleus centralis medialis. Moreover, the lower figures do not always refer to the operated side. In fact, in cases P G 30 and P G 82 a slightly higher number of cells was counted in the nucleus paracentralis of the operated side and in case P G 81 the same is true of the nucleus centralis lateralis. With regard to the size of the remaining cells, the variations reach a maximum o f - - 5 . 1 6 ~ on the operated side of the nucleus centralis medialis in experiment PG 16 and --5.44 ~ on the operated side of the nucleus centralis lateralis in the experiment P G 81. (2) Experimental lesions (see Table II and Fig. 2) Prefrontal lesion. In the case with ablation of the gyrus proreus (PG 78) retro-

grade degeneration is found throughout the rostro-caudal extent of the specific thalarnic nucleus medialis dorsalis. In the nucleus centralis lateralis the number of cells on the operated side differs by --7.83 ~ from that on the normal side. This is a difference slightly in excess of the maximal variation between the two sides in the control animals (5.12~). More marked are the corresponding results for nuclei paracentralis ( - - 1 1 . 6 0 ~ ) and centralis medialis (--13.12 ~). The mean cell size of the neurons in the operated side of P G 78 show a reduction of respectively 10.12~ and 10.50 ~/o in the nuclei paracentralis and centralis medialis. Lesions o f the motor a n d p r e m o t o r areas. In these experiments (PG 115 and PG 116) the cortical destruction also involves the subjacent white matter and therefore some fiber bundles to gyrus proreus and to the superior part of cingular gyrus are interrupted. This may explain the appearance of a retrograde reaction in the nuclei ventralis lateralis and ventralis anterior, the pars paralamellaris of the nucleus medialis dorsalis, and the nucleus anterior medialis. Outside these specific thalamic nuclei cell loss is evident in the nucleus paracentralis ( - - 2 2 . 9 5 ~ ) and less consistently

49

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Fig. 2. Summary of the quantitative findings on the experimental animals. Above: the cortical lesions of each experiment and two transverse sections of the thalamus are schematically presented. Below: each column corresponds to one of the intralaminar nuclei considered. The white space in the column indicates the percentage of cell loss and cell atrophy in the operated side with respect to the normal side in which the total number of cells counted and the average size of remaining cell bodies have been considered equal to 100.

in the nucleus centralis lateralis ( - - 1 3 . 1 3 ~ in P G 116 a n d - - 1 4 . 8 9 ~ in P G 115) a n d centralis medialis ( - - 9 . 5 3 ~ in P G 116 a n d - - 1 1 . 1 7 ~ in P G 115). Also, as far as the size of the r e m a i n i n g cells is concerned, the most consistent difference between the operated a n d n o r m a l side is f o u n d in the nucleus paracentralis (--29.21 ~ in PG 116 a n d - - 1 4 . 5 0 ~ in P G 115); in the nucleus centralis lateralis it is significant (--19.50 ~ ) only in case P G 116, while in the nucleus centralis medialis it does n o t reach significance in either case ( P G 115 or P G 116). Lesion of the posterior parietal area. In the cat P G 56 the cortical lesion, which is

50 TABLE II EXPERIMENTAL ANIMALS

Percentage of cell loss and cell atrophy for each experiment. Cell loss in the operated side (%) Cell atrophy in the operatedside (%)

PG 78 PG 115 PG 116 PG 91 PG 56 PG 83 PG 73 GA 95

CL

PC

- - 7.83 14.89 13.13 27.65 - 19.30 20.58 22.12 36.77

--11.60 --22.95 --23.00 29.20 -- 1.81 --11.57 --29.18 --36.23

CE

13.12 11.17 9.53 8.13 5.00 7.40 10.56 23.78

CL

- - 1.50 5.22 19.50 28.62 26.50 - 30.50 - 29.75 29.50

PC

10.12 14.50 29.21 23.50 4.79 10.65 40.50 30.50

CE

10.50 3.86 0.47 3.50 1.50 4.50 3.42 --24.12

centered on the anterior part of the suprasylvian gyrus, also destroys the subjacent white matter and thus, some projection fibers to and from the supracingular, lateral and ectosylvian gyri. Retrograde degeneration of specific nuclei of the thalamus occurs mainly in the lateralis posterior and lateralis dorsalis nuclei but some retrogradely affected cells also appear in the nucleus ventralis anterior dorsalis, the anteroventral part of the nucleus geniculatus lateralis, the intermedial part of the posterior complex and even the more rostral sectors of the nucleus geniculatus medialis. The cell loss in the intralaminar nuclei of the operated side is significant only in the nucleus centralis lateralis ( 19.30~, Fig. 3C), while in the nucleus paracentralis and the nucleus centralis medialis the difference between the normal and the operated side is no greater than in control animals ( 1.81 ~ and 5.0~). The average cell size of the remaining population shows a conspicuous diminution in the nucleus centralis lateralis ( - - 2 6 . 5 0 ~ ) on the operated side but not in the other intralaminar nuclei (---4.79 ~ and 1.50~). Lesion o f the fronto-parietal region. In cat P G 91 the cortical ablation destroys the sigmoidal gyrus and the rostral part of the suprasylvian and lateral gyri. Retrograde degeneration can be ascertained in the nucleus ventralis posterior, the nucleus ventralis lateralis, the ventral part of the nucleus ventralis anterior, the anterior part of the nucleus lateralis posterior, the anteroventral border of the nucleus geniculatus lateralis, and to a lesser extent the anterior and medial part of the PO complex. The cell loss in the intralaminar nuclei of the operated side is highly significant in the nucleus centralis lateralis ( 27.65 ~ ) and the nucleus paracentralis ( - 2 9 . 2 0 ~ ) . By contrast there is a less appreciable change in the nucleus centralis medialis (--8.13 ~o)Comparable changes are found in the cell size of the remaining neurons in the nucleus paracentralis ( 23.50~) and the nucleus centralis lateralis ( - - 2 8 . 6 2 ~ ) in contrast with the nucleus centralis medialis ( 3.50~). Lesion o f the parieto-temporo-occipital region. In cat PG 83 the ablation largely destroys the cortex and the underlying white matter of the parietal posterior region, of the anterior suprasylvian gyrus, of the $2 area of the anterior ectosytvian gyrus and

51

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Fig. 3. Histological view of CL in various experiments: GA 95 (normal side in A and operated side in AD, P G 73 (normal side in B and operated side in B1), and PG 56 (normal side in C and operated side in Ca). Magnification approximately × 120.

52 of the entire temporal and occipital regions. The retrograde degeneration in the specific nuclei of the thalamus extends to the nucleus ventralis anterior pars dorsalis, the nucleus lateralis dorsalis and lateralis posterior, the pulvinar, the nuclei geniculatus lateralis and medialis and the PO complex. The destruction of the subcortical white substance determines also some retrograde reaction in the nucleus anterior ventralis. Cell loss is evident in the nucleus centralis lateralis (--20.58 ~o): but much less so in the nucleus paracentralis (--11.57~) and inconspicuous in the nucleus centralis medialis (--7.40 0/,~). The average size of the remaining neurons is remarkably reduced in the nucleus centralis lateralis of the operated side (--30.50~) while in the nucleus paracentralis it has declined by 10.05% and in the nucleus centralis medialis by an insignificant 4.5 ~.

Fig. 4. Experiment GA 95. On the left side one can see the heavy reduction in the number of cells in the nuclei centralis medialis and paracentralis of the lesioned hemisphere, Cresyl violet staining. Magnification approximately < 40.

53

Lesion of the fronto-parieto-temporo-occipital region. In cat PG 73 there is a subtotal hemidecortication sparing only part of the gyrus proreus and of the medial aspect of the occipital cortex. The retrograde degeneration of the thalamus on the affected side is marked in all of the specific nuclei except part of the nucleus medialis dorsalis and nucleus geniculatus lateralis. The retrograde reaction is also present in PO complex and in the nucleus geniculatus medialis pars magnocellularis but absent in the nucleus geniculatus medialis caudalis. The cell loss of the intralaminar nuclei, on the operated side, reaches a high degree in the nuclei paracentralis (--29.18 ~) and centralis lateralis (--22.12 ~) and it is only moderately appreciable in the nucleus centralis medialis (--10.56~). The cell size of the remaining neurons likewise has been reduced markedly in the nuclei paracentralis (--40.50 ~) and centralis lateralis

Fig. 5. Experiment PG 73. On the right side are represented the nuclei centralis medialis and paracentralis of the affected side. Cresyl violet staining. Magnification approximately × 40.

54 (--29.75 ~o) ipsilateral to the lesion in contrast to the n o r m a l values o b t a i n e d for the nucleus centralis medialis (Figs. 4 and 5).

Lesion o f the internal capsule and basal ganglia. In cat G A 95 the lesion extends i n depth far e n o u g h to destroy the white matter of the semioval center in the frontal region a n d the anterior part of the internal capsule a n d basal ganglia. I n this case several specific nuclei belonging to the anterior, ventralis lateralis, medialis-dorsalis a n d partially lateratis posterior a n d ventralis posterior groups of the t h a l a m u s as well as the midline nuclei a n d the p a r a f a s c i c u l a r - c e n t r o m e d i a n complex exhibit a m a r k e d retrograde degeneration of their cells. Differences in the n u m b e r of cells between the u n o p e r a t e d a n d operated side reach the highest level e n c o u n t e r e d in the present study: 36.23 ~ for the nucleus paracentralis as well as 36.77 0(, for the nucleus centralis lateralis. In the nucleus centralis medialis cell loss a m o u n t s to 2 3 . 7 8 ~ o n the operated side, a percentage which is twice as high as that in the most extensive cortical ablations. Cell a t r o p h y is marked in the paracentralis ( - - 3 0 . 5 0 ~ ) a n d centralis lateralis ( - - 2 9 . 5 0 ~ ) nuclei but the greatest increase in cell size loss with respect to the most extensive cortical lesion is shown by the nucleus centralis medialis (--24.12 ~ ) (Figs. 3 and 4). TABLE II1 SUMMARYOF THE CELLLOSSIN EXPERIMENTALAND CONTROLANIMALS The numbers indicate the total number of cells counted in the normal (N) and operated (P) side in the experiments, in the right (R) or left (L) side in the controls.

CL

PC

N

P

CE

N

P

N

P

1340 1829 1574 t813 1382 1704 2084

1184 1295 1212 1397 1357 " 1507 1475

1314 1943 1522 1979 2lt0 2231 2190

1141 1787 1377 1767 2009 2045 1951

Experimental anima~ PG PG PG PG PG PG PG

78 91 116 115 56 83 73

1483 1718 1623 1854 2054 1526 2161

•

1367 1242 1410 1578 1556 1212 1683

t - 6.83 P < 0.001

CL

t - 4.36 P < 0.001

PC

L

R

t - - 9.97 P < 0.001

CE

L

R

L

R

1732 1260 1940 1411 1944 1494

1650 1268 t914 1424 1981 1483

1933 1199 2422 1335 1993 1511

1867 1189 2349 1326 1945 1566

Control anima~ PG 16 PG 30 PG81 PG82 C1 C2 •

1759 1582 1687 1349 2095 2066 t = 1.01 Pn.s.

1750 1551 1699 1342 2208 . 2105 '~

t = 0.66 Pn.s.

• , t = test for dependent phenomena.

t ~1.41 P n.s.

55 TABLE IV S U M M A R Y OF T H E R E D U C T I O N IN SIZE OF T H E C E L L BODIES IN 2 0 0

CELLS D E S I G N E D IN E X P E R I M E N T A L

A N D C O N T R O L ANIMALS A T H I G H M A G N I F I C A T I O N

The numbers indicate the total surface of cells in the normal (N) and operated (P) side in the experiments or in the right (R) and left (L) side in the controls, measured by means of a planimeter.

CL

PC

N

P

CE

N

P

N

P

7.09 6.54 6.69 8.30 4.49 6.59 7.23

6.37 4.88 4.74 7.89 4.27 5.88 4.30

6.64 6.33 5.34 8.20 4.49 6.18 5.82

5.94 6.10 5.31 7.72 4.42 5.90 5.62

Experimental animals PG PG PG PG PG PG PG

78 91 116 115 56 83 73

7.68 7.28 7.27 9.00 5.83 7.11 7.64

•

7.57 5.19 6.01 8.53 4.30 4.94 5.36

t 4.72 P < 0.001

CL

t ~ 3.54 P < 0.005

PC

L

R

t 3.45 P < 0.005

CE

L

R

L

R

7.52 5.94 5.05 5.73 4.40 6.78

7.33 5.76 5.32 5.76 4.38 6.80

8.18 5.96 5.00 5.70 4.27 6.50

7.70 5.82 5.02 5.61 4.24 6.52

Control animals PG PG PG PG C1 C2

16 30 81 82

10.38 6.64 10.66 8.98 4.83 8.22

•

t

9.83 6.78 10.08 8.93 4.71 8.19 1.79

Pn.s.

t

0.18

Pn.s.

t1.65 Pn.s.

• , t = test for dependent phenomena.

Statistical evaluation o f the results (Tables II1 a n d IV) As regards the n u m b e r o f the r e m a i n i n g cells, the statistical data on one h a n d confirm the histological data for the centralis lateralis a n d paracentralis nuclei (for which the Student's t-test is significant in the operated subjects a n d is non-significant in the controls); on the other h a n d the data would also be favorable for a significance of the results c o n c e r n i n g the centralis medialis nucleus. The S t u d e n t ' s t-test applied to the centralis medialis nucleus has provided a value of 9.97 (13 df) with P ~> 0.001 for all the ablations considered. O n the c o n t r a r y in the control p r e p a r a t i o n s the value was 1.41 with a non-significant P (11 df). As regards the sizes of the r e m a i n i n g cells the results are superimposable, because there is significance in the experimental preparations, a n d non-significance in the controls (see Table IV). DISCUSSION

The most i m p o r t a n t result which emerges f r o m the experimental data is the

56 demonstration, by means of a quantitative method, that after cortical ablations the intralaminar nuclei considered here show not only chromatolytic or atrophic changes of their cells but also a true cell loss. This is particularly evident when we consider that the control material included not only unoperated cats but also cats in which the sensory acoustic, visual and somatic areas have been removed, i.e. cortical areas whose destruction causes neither gtial proliferation nor cell changes, as is well known, in the intralaminar nuclei. In this respect the present quantitative data are in agreement with the findings reported by Murray 13 from a qualitative microscopical study. The cell alterations are comparable with those observed in the specific nuclei of the thalamus, the only difference being a quantitative one TM. It thus appears that appropriately placed cortex lesions cause cell loss and chromatolytic and atrophic changes of some of the remaining neurons in both the intralaminar and specific thalamic nuclei. The statistical analysis of the results has confirmed this comprehensive datum, showing clearly that the results in the controls are not significant. The percentage of cell loss in the intralaminar nuclei is higher after lesions of the internal capsule and striatum than it is in cases of lesion confined to the cortex. Moreover, it is greater in the n. centralis medialis, but the morphology of the cell alterations is the same. Therefore the present data do not support Powell and Cowart's 20 view according to which only the alterations observed in the intralaminar nuclei after lesions in the internal capsule and striatum can be attributed to retrograde effects, while the atrophic and chromatolytic changes of cells in the intralaminar nuclei after cortical ablations would be the consequence of transneuronal changes. The intralaminar nuclei under study react in a quite different way to the various cortical ablations. In some cases, even in those with extensive cortical ablations (PG 83), the nucleus centralis medialis shows a percentage of cell loss and cell atrophy which remains within the limits of the variations observed in the control experiments. However, in cases PG 78, PG 115, 116 and PG 73 in which either the prefrontal area (PG 78) or the motor and premotor cortices (PG 115, PG 116, PG 73) were ablated, cell loss in this nucleus was in the order of 13.12}o~, t1.17~, 9 . 5 3 ~ and t0.56~o respectively. However the reduction average size of the neurons remaining in the nucleus of the operated side is appreciable only in case PG 78 ( - - t 0 . 5 0 ~ ) . These findings suggest a limited dependence of several neurons of the nucleus centralis medialis upon prefrontal as well as motor and premotor areas of the cortex. From a statistical point of view these data provide a significant value, also for the Student's t-test applied to the centralis medialis nucleus. The nucleus centralis lateralis, by contrast, displays a marked cell loss in all of the cortical ablations, except for case PG 78 with a lesion of the gyrus proteus (--7.83 ~). The most severe retrograde degeneration is found in the nucleus in cases with fronto-parietal or posterior parietal ablations (PG 91, PG 56). If the destruction includes also the temporo-occipital-cortical areas (PG 83, PG 73) the amount of cell loss does not increase significantly. The extirpation of the motor and premotor areas alone (PG 115, PG 116) induces a moderate cell loss, whereas the lesion confined to the somato-sensory cortex (PG 30) is without effect. With regard to the average cell size of the remaining neurons in the nucleus

57 centralis lateralis, the degree of atrophy parallels the severity of cell loss in cases with cortical ablations including the posterior parietal area. These findings suggest a particular dependence of the nucleus centralis lateralis upon the posterior parietal area and, to a lesser extent, the motor and premotor areas in the sigmoid gyrus. In the nucleus paracentralis (PC), cell loss is maximal after combined motor and premotor lesions (PG 115, PG 116) or following extensive ablations including the motor cortex (PG 91, PG 73), while in cases with posterior parietal or parietotemporo-occipital ablations (PG 83, PG 56) the difference in cell number between operated and intact side is insignificant and in several cases (for example PG 56) smaller than the difference observed between right and left side in control preparations. Case PG 78 with a lesion limited to the prefrontal area displays a moderate cell loss in the nucleus paracentralis ( - - 1 1 . 6 0 ~ ) which may be significant again if one considers the small amount of cortex ablated. These results suggest a dependence of the nucleus paracentralis upon the motor and premotor cortex and, to a lesser degree, upon the prefrontal cortex. The cell atrophy in the nucleus paracentralis seems to be correlated with the extent of the ablation of the motor and premotor areas and in more limited degree the prefrontal area. The percentage of cell loss in the intralaminar nuclei is higher after a lesion involving also the internal capsule and striatum. This difference is particularly evident in the nucleus centralis medialis (CE) in which such lesions cause a cell loss twice as great as that observed after the most extensive cortical ablations. These data support the existence of a large essential connection of the intralaminar nuclei, in particular the CE, with the subcortical structures involved by such a deep lesion. Also the marked cell atrophy observed in the nucleus centralis medialis strongly suggests the existence of a great number of collaterals from this nucleus to the basal ganglia. The question now arises as to what meaning should be ascribed to the changes observed in the intralaminar nuclei. Our results are not in contrast, especially for the nucleus centralis medialis, with the classical opinion that the intralaminar nuclei pro-

Fig. 6. Schematic representation of the chief cortical areas whose ablation determines a serious damage to the intralaminar nuclei.

58 ject to the basal ganglia and rhinencephalic area. The anterograde degeneration studies o f N a u t a and Whitlock 15 and the observation o f Scheibel and Scheibel z3 with the Golgi method support this point o f view. On the other hand, the consistent finding of alterations o f the 'retrograde' type in the nuclei paracentralis and centralis lateralis and to a lesser degree, in the nucleus centralis medialis after cortical excisions which involve territories mainly connected with adjacent specific nuclei (lateralis posterior, ventralis lateralis, medialis dorsalis), cannot be underestimated. On this subject we must further consider very positive the fact that the statistical evaluation by means of the Student's t-test has also confirmed the validity o f the results obtained in the experimental preparations c o m p a r e d to the controls. The possible occurrence o f transneuronal degeneration in a thalamic area after cortical lesion has been discussed in a previous publication TM. In this respect it should be pointed out that transneuronal effect might contribute to the observed atrophic and chromatolytic changes, in agreement with the conclusions o f Powell and C o w a n 20. However, in the present experiments with a survival period o f 18 days, it is very unlikely that cell loss might have occurred as a consequence o f transneuronal effectslL Consequently it can be suggested that some intralaminar nuclei project directly to certain areas o f the cerebral cortex to which also some specific thalamic nuclei project (see Fig. 6) as the recent data obtained with horseradish peroxidase method would also demonstrateT, 8. Such dual projections would seem to converge in particular u p o n anterior parts o f the cortex. This could represent the anatomical basis for a complex integration o f specific and non-specific thalamic systems not only at the thalamic but also at the striate and cortical levels.

REFERENCES 1 ADRIANOV, O. S., Sur les liaisons et les fonctions des noyaux thalamiques du syst6me 'nonsp6cifique', Acta neurol, belg., 60 (1960) 704-722. 2 AKIMOTO,K., NEGJSm, K., AND YAMADA,K., Studies on thalamo-cortical connection in cat by means of retrograde degeneration method, Folia psychiat, neuroL jap., 10 (1956) 39-82. 3 BODIAN,D., Studies on the diencephalon of the Virginia opossum. III. Thalamo-cortical projections, J. comp. NeuroL, 77 (1942) 535-575. 4 CoMas, C. M., Fibers and cell degeneration in the albino rat brain after hemidecortication, J. comp. Neurol., 90 (1949) 373-394. 5 COWAN,W. M., AND POWELL,T. P. S., The projection of the midline and intralaminar nuclei of the thalamus of the rabbit, J. Neurol. Neurosurg. Psychiat., 18 (1955) 266-279. 6 DEMPSEY,E. W., ANOMORJSON,R. S., The production of rhythmically recurrent cortical potential after localized thalamic stimulation, Amer. J. Physiol., 135 (1942) 293-300. 7 JONES,E. G.. AND LEAVITT, R. Y., Retrograde axonal transport and the demonstration of nonspecific projections to the cerebral cortex and striatum from thalamic intralaminar nuclei in the rat, cat, monkey, J. comp. NeuroL, 154 (1974) 349-378. 8 KUYPERS, H. G J. M.. KIEVIT,J., AND GROEN-KLEVANT,C., Retrograde axonal transport of HRP in rat's forebrain, Brain Research, 67 (1974) 211-218. 9 LASHLEY,K. S.. Thalamo-cortical connections of the rat's brain. J. comp. Neurol., 75 (1941) 67-121. 10 MACCHI,G., Introductory statement about the thalamo-cortical connections. Arch. itaL Biol.. 107 (1969) 547-569. 11 MACCHI,G., E ANO~LERI,F., Anatomia sperimentale dei sistemi a proiezione diffnsa delrencefato. In Atti Convengno di Fisiol. e Clin. dei Sistemi a Proiezione diffusa dell'encefalo. 23-24 novembre, Maccari, Parma, 1957.

59 12 MATHEWS, M. R., Further observation on transneuronal degeneration in the lateral geniculate nucleus of the macaque monkey, J. Anat. (Lond.), 98 (1964) 255-264. 13 MURRAY, M., Degeneration of some intralaminar nuclei after cortical removals in the cat, J. comp. Neurol., 127 (1966) 341-367. 14 NASHOLD, B. S., HANBERY, J., AND OLSZEWSKI, J., Observations on the diffuse thalamic projections, Electroenceph. clin. Neurophysiol., 7 (1955) 609-620. 15 NAUTA, W. J. H., AND WHITLOCK, D. G., An anatomical analysis of the non-specific thalamic projection system. In E. D. ADRIAN, F. BREMER AND A. JASPER (Eds.), Brain Mechanisms and Consciousness, Blackwell, Oxford, 1954, pp. 8l-116. 16 PEACOCK, J. H., AND COMBS, M. C., Retrograde cell degeneration in diencephalic and other structures after hemidecortication of rhesus monkeys, Exp. Neurol., 11 (1965) 367-399. 17 PEACOCK, J. H., AND COMBS, M. C., Retrograde cell degeneration in adult cat after hemidecortication, J. comp. Neurol., 125 (1965) 329-336. 18 POWELL, T. P. S., Residual neurons in the human thalamus following hemidecortication, Brain, 75 (1952) 571-584. 19 POWELL, T. P. S., AND COWAN, W. M., A study of thalamo-striate relations in the monkey, Brain, 79 (1956) 364-390. 20 POWELL, T. P. S., AND COWAN W. M., The interpretation of the degenerative changes in the intralaminar nuclei of the thalamus, J. Neurol. Neurosurg. Psychiat., 30 (1967) 140-153. 21 ROSE, J. E., The thalamus of the sheep cellular and fibrous structure and comparison with pig, rabbit and cat, J. comp. Neurol., 77 (1942) 469-524. 22 ROSE, J. E., AND WOOLSEY, C. N., A study of thalamo-cortical relations in the rabbit, Bull. Johns Hopk. Hosp., 73 (1943) 65-128. 23 SCHEIBEL, E. M., AND SCHEIBEL, A. B., Structural organization of the unspecific thalamic nuclei and their projection toward cortex, Brain Research, 6 (1967) 60 94. 24 STEEENS, R., ET DROOGLEEVER-FORTUYN, J. Contribution ~, l'6tude de la structure et de quelques connexions des noyaux interm6diaires du thalamus chez le lapin, Schweiz. Arch. Neurol. P,~'chiat., 72 (1953) 299-318. 25 WALKER, A. E., The retrograde cell degeneration in the thalamus of Macacus Rhesus following hemidecortication, J. comp. Neurol., 62 (1935) 407-487. 26 WALKER, A. E., The thalamus of chimpanzee. Its nuclear structure normal and following hemidecortication, J. comp. Neurol., 69 (1938) 487-507. 27 WALLER, W. H., The thalamus of the cat after hemidecortication, J. Anat. (Lond.), 72 (1938) 475-487.