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An aberrant crossed visual corticotectal pathway in albino rats
Brain Research, 112 (1976) 37-44 © Elsevier Scientific Publishing Company, Amsterdam - Printed in The Netherlands
AN ABERRANT ALBINO RATS
CROSSED VISUAL C O R T I C O T E C T A L P A T H W A Y
37
IN
M. J. MUSTARI and R. D. LUND Department of Biological Structure, University of Washington School of Medicine, Seattle, Wash. 98195 (u.s.A.)
(Accepted December 31st, 1975)
SUMMARY This study investigates the dynamic nature of the developing corticotectal pathway arising in the visual cortex. Special attention is given to the interaction occurring between the corticotectal pathways of each side of the brain and between corticotectal and retinotectal terminations. Normally the visual cortex of rats projects only to the ipsilateral superior colliculus. If one visual cortex is removed at birth, the remaining visual cortex subsequently shows a bilateral projection to the superior colliculus. The aberrant corticotectal pathway is heavier if the cortical ablation is accompanied by eye removal at birth but eye enucleation alone is not a sufficient stimulus for production of a crossed corticotectal projection. The aberrant crossed pathway shows a topographic order which appears to correspond to that of the normal ipsilateral corticotectal pathway. The pathway differs from the aberrant projections from the retina in that lesions done as late as 20 days postnatal still result in an aberrant crossed corticotectal pathway. This is similar to the aberrant crossed cortical projections from sensorimotor cortex. The pathway would appear to arise as a result of lack of competition from corticotectal axons normally present contralaterally or from attraction of denervated corticotectal sites. While denervated retinotectal sites stimulate sprouting of the corticotectal axons once in the deafferented colliculus, they do not stimulate crossing of the corticotectal projection.
INTRODUCTION A lot of work has been devoted to the question of rigidity and plasticity of retinotectal connections in non-mammalian vertebrates1,3,4,7, 24. From these studies a number of factors such as special affinity and fiber-fiber interactions have been invoked to explain the various patterns of ordered projection of the retina upon the tectum. It appears that similar rules may apply also to the mammalian retinotectal
38 system 17,19,2a. The m a m m a l i a n system does, however, have an a d d i t i o n a l a d v a n t a g e over the n o n - m a m m a l i a n in t h a t besides the p a t h w a y f r o m the retina, the u p p e r layers o f the s u p e r i o r colliculus also receive a p r o j e c t i o n f r o m the visual cortex which is r e t i n o t o p i c a l l y mapped11,12. I t is o f some interest, since it is possible to disturb the o r d e r l y m a p o f the r e t i n a to the colliculus w i t h o u t u p s e t t i n g the c o r t i c o t e c t a l m a p 17 to k n o w m o r e a b o u t the p l a s t i c i t y d e m o n s t r a b l e b y the c o r t i c o t e c t a l p a t h w a y a n d the degree o f i n t e r a c t i o n o f these t w o c o n v e r g e n t p a t h w a y s . The p r e s e n t study is r e s t r i c t e d to the q u e s t i o n o f i n t e r t e c t a l crossing o f corticotectal axons arising in the visual cortex, the o r d e r o f this c r o s s e d p a t h w a y a n d its r e l a t i o n to the r e t i n o t e c t a l t e r m i n a t i o n . A p r e l i m i n a r y r e p o r t o f s o m e o f the results has been given elsewhere 20. MATERIALS AND METHODS Eighty-five a l b i n o rats were used in the study. The v a r i o u s e x p e r i m e n t s u n d e r t a k e n are o u t l i n e d in T a b l e I. I n all b u t the c o n t r o l series, lesions o f the left visual c o r t e x were m a d e b y a s p i r a t i o n at times between fetal d a y 16 a n d p o s t n a t a l d a y 20. Such lesions were i n t e n d e d to involve the whole visual cortex. M o s t e x t e n d e d b e y o n d the limits o f the p r i m a r y visual cortex, a n d resulted in t o t a l loss o f the d o r s a l lateral geniculate b o d y on t h a t side. A few lesions were s u b t o t a l a n d p a r t o f the geniculate r e m a i n e d intact. A f t e r survival o f 1-2 m o n t h s , e i t h e r t o t a l o r s u b t o t a l lesions were m a d e o f the r i g h t visual c o r t e x a n d the a n i m a l s allowed to survive a further 3 - 4 days when t h e y were TABLE I First lesion
Second lesion No. attimals
Age
2 3 monthspostnatal
Crossed Corticotectal* * (%)
6 5 5 17 17 6 6 6 6
Birth Fetal day 16 Fetal day 19 Birth Birth 5 days postnatal 10 days postnatal 15 days postnatal 20 days postnatal
Right v.c. (total or subtotal) 10 Right v.c. (total) 10 Right v.c. (total) 10 Right v.c. (total or subtotal) 50 Right v.c. (total or subtotal) 50 Right v.c. (total) 50 Right v.c. (total) 50 Right v.c. (total) 10 Right v.c. (total) 0
Birth Birth Adult
Right v.c. (total) Right v.c. (total)
Experimental series'
Left visual cortex Left visual cortex Left visual cortex Left v.c. ÷ right eye Left v.c. + both eyes Left visual cortex* Left visual cortex* Left visual cortex* Left visual cortex* Control series
Right eye Both eyes Left visual cortex
2 2 5
* In these animals bilateral eye enucleation at birth. ** Per cent expressed as maximum density of crossed compared to uncrossed.
0 0 0
39 perfused with buffered p a r a f o r m a l d e h y d e a n d subsequently p r e p a r e d for staining with the F i n k - H e i m e r technique for d e g e n e r a t i n g axons a n d terminals. The s u p e r i o r colliculi o f some b r a i n s were p r e p a r e d for electron m i c r o s c o p y a n d s t u d i e d with a J E O L 100B electron m i c r o s c o p e for identification o f d e g e n e r a t i n g a x o n terminals. RESULTS The c o r t i c o t e c t a l p a t h w a y f r o m the visual cortex to the s u p e r i o r colliculus as described previously for the rat12,13 ends p r e d o m i n a n t l y in the deeper p a r t o f the s t r a t u m g r i s e u m superficiale. Sparse d e g e n e r a t i o n granules are also f o u n d in the strat u m zonale a n d s t r a t u m o p t i c u m a n d occasional d e g e n e r a t i o n granules occur in the u p p e r p a r t o f the s t r a t u m g r i s e u m superficiale following a visual cortex lesion (Fig. la). I f the c o n t r a l a t e r a l eye is r e m o v e d at b i r t h the s t r a t u m griseum superficiale
Fig. 1. a: Fink-Heimer stained preparation from the right superior colliculus of a normal adult rat showing ipsilateral corticotectal degeneration following a right visual cortex lesion. Note sparse degeneration in the upper half of the stratum griseum superficiale. Bar, 100 t~m. b: Fink-Heimer stained preparation showing ipsilateral corticotectal degeneration in the right superior colliculus of an adult rat that had the left visual cortex and both eyes removed at birth, and the right visual cortex lesioned as an adult. Note that the corticotectal degeneration extends to the surface of the superior colliculus. Magnification as in a.
40 shrinks in volume by a little over 50 ~o~4,2~. Subsequently, the corticotectal pathway can be seen projecting through the entire depth of the stratum griseum superficiale as described previously 15 (Fig. lb). The over-all density of the pathway is greater even taking shrinkage into account, than in animals with the eye intact, although it is never as dense as the regular retinotectal pathway. In all the normal animals as well as those with both eyes removed, there is no indication of any crossed corticotectal pathway. After removal of the left visual cortex at birth, an aberrant crossed corticotectal pathway can be demonstrated by subsequent removal of the right visual cortex. The fibers cross over the midline close to the surface and distribute within the deeper part of the stratum griseum superficiale. The crossed pathway is most dense medially and very sparse or absent laterally. To obtain a very approximate figure for the size of the crossed compared with the regular uncrossed pathway, the density of degeneration granules was counted on each side. In these animals the crossed pathway, at its heaviest, was about 1 0 ~ of the regular uncrossed pathway. Since the degenerative stains used show both terminal and preterminal degeneration, it is clear that this figure reflects only the relative size of the pathways which may or may not correlate with their relative synaptic densities. A similar experiment performed in animals in which the initial cortical lesion was made on fetal days 16 and 19, show an identical pattern to those lesions at birth in terms of distribution and density of the crossed corticotectal pathway when tested as adults. If both the left cortex and right eye are removed at birth, the crossed corticotectal arising in the right cortex becomes more prominent. While the ipsilateral corticotectal pathway shows a normal distribution with respect to depth, the crossed corticotectal pathway projects throughout the stratum griseum superficiale with even density (Fig. 2a). The crossing bundle of fibers is more prominent than in the previous case and degeneration can be found right to the lateral border of the colliculus. When both eyes and one cortex are removed at birth the ipsilateral corticotectal pathway projects to the surface of the colliculus as described above and the crossed corticotectal pathway appears generally similar to that described in animals with unilateral cortex and contralateral eye removal at birth. In order to determine whether these aberrant fibers do indeed make synaptic connections some of this group of animals were studied with the electron microscope after lesioning the remaining cortex. Degenerating axodendritic synapses were found throughout the depth of the stratum griseum superficiale of the colliculus contralateral to the lesioned cortex (Fig. 2b). Similar but heavier terminal degeneration was found ipsilaterally. To test whether there was any topographic order to the crossed corticotectal pathway a series of animals was prepared in which both eyes and one visual cortex were removed at birth. Subsequently, small lesions were made in the remaining visual cortex and the degeneration due to these lesions was studied in the superior colliculus. Ipsilateral degeneration was always in the predicted position. Contralateral degeneration tended to show a mirror topography. For example, lesions which produced a local patch of degeneration close to the midline on the ipsilateral side gave
41
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:
Fig. 2. a: Fink-Heimer stained preparation showing crossed corticotectal degeneration extending to the surface of the left superior colliculus in an adult rat that had the right eye and left visual cortex removed at birth, and the right visual cortex lesioned as an adult. Note degenerating abzrrant crossed corticotectal pathway. Bar, 50/~m. b: electron micrograph taken from a section through the stratum griseum superficiale of the left superior colliculus of an adult rat that had both eyes and the left visual cortex removed at birth and the right visual cortex lesioned as an adult. Note the dense degenerating crossed corticotectal terminal making an axodendritic contact. a c i r c u m s c r i b e d p a t c h also close to the midline contralaterally. Similarly, lesions p r o ducing a n t e r i o r or p o s t e r i o r d e g e n e r a t i o n ipsilaterally show similar patches c o n t r a laterally. Lesions p r o d u c i n g lateral d e g e n e r a t i o n ipsilaterally give a very sparse p r o j e c t i o n c o n t r a l a t e r a l l y . W h e n this can be localized, it is situated in a lateral position. H o w e v e r , in a n u m b e r o f cases it a p p e a r s s o m e w h a t diffuse. Such diffuseness c o u l d reflect staining o f b o t h passage fibers a n d t e r m i n a l s r a t h e r t h a n necessarily i n d i c a t i n g a diffuse t e r m i n a t i o n . To assess the effect o f t i m i n g o f the initial c o r t i c a l lesion on the f o r m a t i o n o f the a b e r r a n t crossed c o r t i c o t e c t a l p a t h w a y , bilateral eye enucleations were p e r f o r m e d on a set o f a n i m a l s at birth, a n d subsequently, a u n i l a t e r a l visual c o r t i c a l lesion was m a d e on p o s t n a t a l days 5, 10, 15 a n d 20. A f t e r 2.5 m o n t h s , animals f r o m each g r o u p were t a k e n w i t h o u t further surgery a n d stained with the F i n k - H e i m e r m e t h o d to test for residual d e g e n e r a t i o n froi"n the initial lesions. N o residual d e g e n e r a t i o n was
42 found. Second lesions were made on similar animals involving the entire remaining visual cortex. They show a heavy crossed corticotectal projection in animals when the initial cortical lesion was made on postnatal days 5 and 10 that differed little in density or distribution from the crossed corticotectal projections observed in animals that received cortical lesions and eye enucleation at birth. Those animals in which the initial cortical lesions were made on day 15 show a crossed distribution similar to the previous animals but this is of reduced density, being about 20 ~ of the amount found in 5- and 10-day cases. Those animals with initial cortical lesions made on postnatal day 20 show very light crossed degeneration and this can be only found near the midline. DISCUSSION The corticotectal pathway from the visual area like that from the sensorimotor cortex 1° shows the capacity for giving a crossed projection in the absence of a corticotectal pathway from the opposite side. This occurs even though there is not a natural commissure joining the appropriate laminae. Electron microscopy shows that the aberrant pathway forms synaptic contacts in the contralateral colliculus. Whether these contacts can affect the physiology of the postsynaptic cells or whether they are suppressed by other inputs remains to be determined. The crossed projection is not the result of a random process and is subject to certain constraints. For example, it ends only in laminae appropriate for the visual corticotectal pathway and shows a pattern of topographic order which relates to the retinotectal map although not to the map of the visual field. As in most other systems in which a major reorganization of a pathway Occurs 14,16,20,2a, the aberrant pathway shows progressively less development, the older the animal at the time of the initial lesion. Thus by 20 days very little crossed degeneration is found. This is comparable in timing for the crossed corticotectal pathway from the sensorimotor cortex 9 but later than the aberrant ipsilateral retinotectal pathway (10-12 day) induced by unilateral eye lesions a4. It would be expected, therefore, that the restriction of the capacity to sprout reflects some component of the individual pathway rather than a stabilization within the tectum itself, such as of the astrocytic component. The relationship between the corticotectal and retinotectal pathways is particularly interesting. First, as discussed previously, if the eyes are removed at birth the corticotectal pathway, either regular or crossed appears to sprout to innervate the whole stratum griseum superficiale. Such apparent sprouting could be artifactual resulting from loss of the upper part of the stratum griseum superficiale into which optic axons have their heaviest projection. This would seem unlikely since Golgi studies (unpublished) indicate that the cell types characteristic of the upper part of the stratum griseum superficiale s are still present after neonatal eye enucleation. Further, the degree of shrinkage after enucleation is less than would be accounted for by a total loss of the upper half of the stratum griseum superficiale and the increased density of the corticotectal pathway is less than could be accounted for by shrinkage alone. This implies that corticotectal fibers have taken over sites made available by
43 retinal deafferentation. The second point concerning the interrelation of retinotectal and corticotectal pathways relates to the fact that although optic deafferentation may result in an increased depth of distribution of corticotectal fibers within one colliculus, the control studies show that it does not on its own stimulate an aberrant crossing of corticotectal fibers. The underlying mechanism resulting in the formation of the crossed corticotectal pathway is unclear. Certainly it is not an indiscriminate response to deafferentation (or delayed innervation) ofcorticotectal sites. Alternatively, it could reflect an inhibitory interaction normally present between corticotectal axons from each side of the brain. Thus, if the axons on one side are absent during a critical stage of development, those of the other side cross the midline and innervate the area normally served by the absent axons. A similar pattern of mutual inhibition has been indicated in vitro 2 and could be used to explain several other studies concerning in vivo development and plasticity 5,6,Is,z2. It is surprising perhaps that the aberrant pathway occurs even though its normal ipsilateral synaptic connection appears present and undisturbed by the surgery. Furthermore, the stimulus for crossing is effective over considerable distances as from the lateral side of one colliculus to the lateral side of the other. In order to understand more fully the mechanisms controlling the pathway, it would seem important to know how it develops and how this development relates to the normal development of the corticotectal pathway. In particular it would be interesting to know how the topographic order establishes itself. Is it instantaneous with fibers growing directly to appropriate sites or is it progressive in the manner indicated in the moving retinotectal map and compression in fish where fibers gradually form their connections? (see Gaze3; Gaze and Hope 4 for summary). One useful feature of this system is the relatively late time at which sprouting becomes reduced and the similarity of the sprouting after lesions made at 15 days postnatal and at fetal day 16 (when cells generating the pathway are just being formed). This means that the plasticity of the pathway can be more extensively and easily studied than in the retinotectal system where early neonatal and fetal times are necessary in rat and hamster to show plastic phenomenon14,19,2a, 23. A further use of study of the corticotectal system will be to see whether the rules of development derived largely from study of the retinotectal pathway are indeed applicable to other pathways, particularly to ones arising in the cerebral cortex. Particularly interesting are questions concerning the effects of partial visual cortical and superior collicular lesions and how they may compare with similar retinal and tectal manipulations in fish. Do the same sorts of compressions and expansions occur ? Further, can intertectal crossing of corticotectal fibers be induced by tectal lesions as is the case for retinal fibers2~? Thus it is hoped that this system will be one which is amenable to many of the experimental regimens to which the retinotectal systems have been subjected. Its special interests lie in the fact that it is a pathway of cerebral origin rather than one arising from primary afferents; it can be manipulated late in development; and it interacts with the retinotectal system.
44 ACKNOWLEDGEMENTS We thank Ren6e Wise and Joy Baisinger for valuable technical help and Doris Ringer and Vicki Neff for editorial assistance. Supported by U.S.P.H.S. Grants EY-00596 and GM-00136 from the National Institutes of Health. REFERENCES 1 Cowan, W. M., Studies on the development of the avian visual system. In D. S. Pease (Ed.), Cellular Aspects o f Neural Growth and Differentiation, Univ. Calif. Press, Los Angeles, Calif., 1971, pp. 177-222. 2 Dunn, G. A., Mutual contact inhibition of extension of chick sensory nerve fibers in vitro, aT. comp. Neurol., 143 (1971) 491 507. 3 Gaze, R. M., Neuronal specificity, Brit. reed. Bull., 30 (1974) 116-121. 4 Gaze, R. M. and Hope, R. A., The formation of continuously ordered mappings. In M. Corner and D. Swaab (Eds.), Perspectives in Brain Research, Progress hz Brain Research, Vol. 45, Elsevier, Amsterdam, 1976, in press. 5 Gottlieb, D. I. and Cowan, W. M., Evidence for a temporal factor in the occupation of available synaptic sites during development of the dentate gyrus, Brain Research, 41 (1972) 452-456. 6 Guillery, R. W., Binocular competition in the control of geniculate cell growth, J. comp. Neurol., 144 (1972) 117-129. 7 Hunt, R. K. and Jacobsen, M., Neuronal specificity revisited, Curr. Top. Develop. Biol., 8 (1974) 203-259. 8 Langer, T. P. and Lund, R. D., The upper layers of the superior colliculus of the rat: a Golgi study, J. comp. NeuroL, 158 (1974) 405-436. 9 Leong, S. K., Unpublished observations. 10 Leong, S. K. and Lund, R. D., Anomalous bilateral corticofugal pathways in albino rats after neonatal lesions, Brain Research, 62 (1973) 218-221. 11 Lund, R. D., Terminal distribution in the superior colliculus of fibers originating in the visual cortex, Nature (Lond.), 204 (1964) 1283-1285. 12 Lund, R. D., The occipitotectal pathway of the rat, J. Anat. (Lond.), 11 (1966) 51-62. 13 Lund, R . D . , Synaptic patterns of the superficial layers of the superior colliculus of the rat, J. comp. Net~rol., 135 (1969) 179-201. 14 Lund, R. D., Cunningham, T. J. and Lund, J. S., Modified optic projections after unilatera! eye removal in young rats, Brain Behav. Evol., 8 (1973) 51-72. 15 Lund, R. D. and Lund, J. S., Modifications of synaptic patterns in the superior colliculus of the rat during development and following deafferentation, Vision Res., 11, Suppl. 3 (1971) 281 298. 16 Lund, R. D. and Lund, J. S., Synaptic adjustment after deafferentation of the superior colliculus of the rat, Science, 171 (1971) 804-807. 17 Lund, R. D. and Lurid, J. S., Plasticity in the developing visual system: the effects of retinal lesions made in young rats, J. comp. Neurol., (1976) in press. 18 Lynch, G., Mosko, S., Parks, T. and Cotman, L., Hyperdevelopment of the dentate gyrus commissural system after entorhinal lesions in immature rats, Brain Research, 50 (1973) 174-178. 19 Miller, B. F. and Lund, R . D . , The pattern of retinotectal connections in albino rats can be modified by fetal surgery, Brain Research, 91 (1975) 119-125. 20 Mustari, M. J., An aberrant contralateral visual corticotectal pathway in albino rats, Anat. Rec., 181 (1975) 433 (Abstract). 21 Schneider, G. E., Mechanisms of functional recovery following lesions of visual cortex or superior colliculus in neonate and adult hamsters, Brain Behav. Evol., 3 (1970) 295-323. 22 Schneider, G. E., Competition for terminal space in formation of abnormal retinotectal connections, and a functional consequence, Anat. Rec., 169 (1971) 420. 23 Schneider, G. E. and Jhaveri, S. R., Neuroanatomical correlates of spared or altered function after brain lesions in the newborn hamster. In P. G. Stein, J. J. Rosen and N. Butters (Eds.), Plasticity and Recovery o f Function hi the Central Nervou~ System, Academic Press, New York, 1974, pp. 65-109. 24 Sperry, R.W., Chemoaffinity in the orderly growth of nerve fiber patterns and connections, Proe. nat. Acad. Sci. (Wash.), 50 (1963) 703-710. 25 Tsang, Y., Visual centers in blinded rats, J. eomp. NeuroL, 66 (1937) 211-261.
AN ABERRANT ALBINO RATS
CROSSED VISUAL C O R T I C O T E C T A L P A T H W A Y
37
IN
M. J. MUSTARI and R. D. LUND Department of Biological Structure, University of Washington School of Medicine, Seattle, Wash. 98195 (u.s.A.)
(Accepted December 31st, 1975)
SUMMARY This study investigates the dynamic nature of the developing corticotectal pathway arising in the visual cortex. Special attention is given to the interaction occurring between the corticotectal pathways of each side of the brain and between corticotectal and retinotectal terminations. Normally the visual cortex of rats projects only to the ipsilateral superior colliculus. If one visual cortex is removed at birth, the remaining visual cortex subsequently shows a bilateral projection to the superior colliculus. The aberrant corticotectal pathway is heavier if the cortical ablation is accompanied by eye removal at birth but eye enucleation alone is not a sufficient stimulus for production of a crossed corticotectal projection. The aberrant crossed pathway shows a topographic order which appears to correspond to that of the normal ipsilateral corticotectal pathway. The pathway differs from the aberrant projections from the retina in that lesions done as late as 20 days postnatal still result in an aberrant crossed corticotectal pathway. This is similar to the aberrant crossed cortical projections from sensorimotor cortex. The pathway would appear to arise as a result of lack of competition from corticotectal axons normally present contralaterally or from attraction of denervated corticotectal sites. While denervated retinotectal sites stimulate sprouting of the corticotectal axons once in the deafferented colliculus, they do not stimulate crossing of the corticotectal projection.
INTRODUCTION A lot of work has been devoted to the question of rigidity and plasticity of retinotectal connections in non-mammalian vertebrates1,3,4,7, 24. From these studies a number of factors such as special affinity and fiber-fiber interactions have been invoked to explain the various patterns of ordered projection of the retina upon the tectum. It appears that similar rules may apply also to the mammalian retinotectal
38 system 17,19,2a. The m a m m a l i a n system does, however, have an a d d i t i o n a l a d v a n t a g e over the n o n - m a m m a l i a n in t h a t besides the p a t h w a y f r o m the retina, the u p p e r layers o f the s u p e r i o r colliculus also receive a p r o j e c t i o n f r o m the visual cortex which is r e t i n o t o p i c a l l y mapped11,12. I t is o f some interest, since it is possible to disturb the o r d e r l y m a p o f the r e t i n a to the colliculus w i t h o u t u p s e t t i n g the c o r t i c o t e c t a l m a p 17 to k n o w m o r e a b o u t the p l a s t i c i t y d e m o n s t r a b l e b y the c o r t i c o t e c t a l p a t h w a y a n d the degree o f i n t e r a c t i o n o f these t w o c o n v e r g e n t p a t h w a y s . The p r e s e n t study is r e s t r i c t e d to the q u e s t i o n o f i n t e r t e c t a l crossing o f corticotectal axons arising in the visual cortex, the o r d e r o f this c r o s s e d p a t h w a y a n d its r e l a t i o n to the r e t i n o t e c t a l t e r m i n a t i o n . A p r e l i m i n a r y r e p o r t o f s o m e o f the results has been given elsewhere 20. MATERIALS AND METHODS Eighty-five a l b i n o rats were used in the study. The v a r i o u s e x p e r i m e n t s u n d e r t a k e n are o u t l i n e d in T a b l e I. I n all b u t the c o n t r o l series, lesions o f the left visual c o r t e x were m a d e b y a s p i r a t i o n at times between fetal d a y 16 a n d p o s t n a t a l d a y 20. Such lesions were i n t e n d e d to involve the whole visual cortex. M o s t e x t e n d e d b e y o n d the limits o f the p r i m a r y visual cortex, a n d resulted in t o t a l loss o f the d o r s a l lateral geniculate b o d y on t h a t side. A few lesions were s u b t o t a l a n d p a r t o f the geniculate r e m a i n e d intact. A f t e r survival o f 1-2 m o n t h s , e i t h e r t o t a l o r s u b t o t a l lesions were m a d e o f the r i g h t visual c o r t e x a n d the a n i m a l s allowed to survive a further 3 - 4 days when t h e y were TABLE I First lesion
Second lesion No. attimals
Age
2 3 monthspostnatal
Crossed Corticotectal* * (%)
6 5 5 17 17 6 6 6 6
Birth Fetal day 16 Fetal day 19 Birth Birth 5 days postnatal 10 days postnatal 15 days postnatal 20 days postnatal
Right v.c. (total or subtotal) 10 Right v.c. (total) 10 Right v.c. (total) 10 Right v.c. (total or subtotal) 50 Right v.c. (total or subtotal) 50 Right v.c. (total) 50 Right v.c. (total) 50 Right v.c. (total) 10 Right v.c. (total) 0
Birth Birth Adult
Right v.c. (total) Right v.c. (total)
Experimental series'
Left visual cortex Left visual cortex Left visual cortex Left v.c. ÷ right eye Left v.c. + both eyes Left visual cortex* Left visual cortex* Left visual cortex* Left visual cortex* Control series
Right eye Both eyes Left visual cortex
2 2 5
* In these animals bilateral eye enucleation at birth. ** Per cent expressed as maximum density of crossed compared to uncrossed.
0 0 0
39 perfused with buffered p a r a f o r m a l d e h y d e a n d subsequently p r e p a r e d for staining with the F i n k - H e i m e r technique for d e g e n e r a t i n g axons a n d terminals. The s u p e r i o r colliculi o f some b r a i n s were p r e p a r e d for electron m i c r o s c o p y a n d s t u d i e d with a J E O L 100B electron m i c r o s c o p e for identification o f d e g e n e r a t i n g a x o n terminals. RESULTS The c o r t i c o t e c t a l p a t h w a y f r o m the visual cortex to the s u p e r i o r colliculus as described previously for the rat12,13 ends p r e d o m i n a n t l y in the deeper p a r t o f the s t r a t u m g r i s e u m superficiale. Sparse d e g e n e r a t i o n granules are also f o u n d in the strat u m zonale a n d s t r a t u m o p t i c u m a n d occasional d e g e n e r a t i o n granules occur in the u p p e r p a r t o f the s t r a t u m g r i s e u m superficiale following a visual cortex lesion (Fig. la). I f the c o n t r a l a t e r a l eye is r e m o v e d at b i r t h the s t r a t u m griseum superficiale
Fig. 1. a: Fink-Heimer stained preparation from the right superior colliculus of a normal adult rat showing ipsilateral corticotectal degeneration following a right visual cortex lesion. Note sparse degeneration in the upper half of the stratum griseum superficiale. Bar, 100 t~m. b: Fink-Heimer stained preparation showing ipsilateral corticotectal degeneration in the right superior colliculus of an adult rat that had the left visual cortex and both eyes removed at birth, and the right visual cortex lesioned as an adult. Note that the corticotectal degeneration extends to the surface of the superior colliculus. Magnification as in a.
40 shrinks in volume by a little over 50 ~o~4,2~. Subsequently, the corticotectal pathway can be seen projecting through the entire depth of the stratum griseum superficiale as described previously 15 (Fig. lb). The over-all density of the pathway is greater even taking shrinkage into account, than in animals with the eye intact, although it is never as dense as the regular retinotectal pathway. In all the normal animals as well as those with both eyes removed, there is no indication of any crossed corticotectal pathway. After removal of the left visual cortex at birth, an aberrant crossed corticotectal pathway can be demonstrated by subsequent removal of the right visual cortex. The fibers cross over the midline close to the surface and distribute within the deeper part of the stratum griseum superficiale. The crossed pathway is most dense medially and very sparse or absent laterally. To obtain a very approximate figure for the size of the crossed compared with the regular uncrossed pathway, the density of degeneration granules was counted on each side. In these animals the crossed pathway, at its heaviest, was about 1 0 ~ of the regular uncrossed pathway. Since the degenerative stains used show both terminal and preterminal degeneration, it is clear that this figure reflects only the relative size of the pathways which may or may not correlate with their relative synaptic densities. A similar experiment performed in animals in which the initial cortical lesion was made on fetal days 16 and 19, show an identical pattern to those lesions at birth in terms of distribution and density of the crossed corticotectal pathway when tested as adults. If both the left cortex and right eye are removed at birth, the crossed corticotectal arising in the right cortex becomes more prominent. While the ipsilateral corticotectal pathway shows a normal distribution with respect to depth, the crossed corticotectal pathway projects throughout the stratum griseum superficiale with even density (Fig. 2a). The crossing bundle of fibers is more prominent than in the previous case and degeneration can be found right to the lateral border of the colliculus. When both eyes and one cortex are removed at birth the ipsilateral corticotectal pathway projects to the surface of the colliculus as described above and the crossed corticotectal pathway appears generally similar to that described in animals with unilateral cortex and contralateral eye removal at birth. In order to determine whether these aberrant fibers do indeed make synaptic connections some of this group of animals were studied with the electron microscope after lesioning the remaining cortex. Degenerating axodendritic synapses were found throughout the depth of the stratum griseum superficiale of the colliculus contralateral to the lesioned cortex (Fig. 2b). Similar but heavier terminal degeneration was found ipsilaterally. To test whether there was any topographic order to the crossed corticotectal pathway a series of animals was prepared in which both eyes and one visual cortex were removed at birth. Subsequently, small lesions were made in the remaining visual cortex and the degeneration due to these lesions was studied in the superior colliculus. Ipsilateral degeneration was always in the predicted position. Contralateral degeneration tended to show a mirror topography. For example, lesions which produced a local patch of degeneration close to the midline on the ipsilateral side gave
41
, i¸ : i f : / :
:
Fig. 2. a: Fink-Heimer stained preparation showing crossed corticotectal degeneration extending to the surface of the left superior colliculus in an adult rat that had the right eye and left visual cortex removed at birth, and the right visual cortex lesioned as an adult. Note degenerating abzrrant crossed corticotectal pathway. Bar, 50/~m. b: electron micrograph taken from a section through the stratum griseum superficiale of the left superior colliculus of an adult rat that had both eyes and the left visual cortex removed at birth and the right visual cortex lesioned as an adult. Note the dense degenerating crossed corticotectal terminal making an axodendritic contact. a c i r c u m s c r i b e d p a t c h also close to the midline contralaterally. Similarly, lesions p r o ducing a n t e r i o r or p o s t e r i o r d e g e n e r a t i o n ipsilaterally show similar patches c o n t r a laterally. Lesions p r o d u c i n g lateral d e g e n e r a t i o n ipsilaterally give a very sparse p r o j e c t i o n c o n t r a l a t e r a l l y . W h e n this can be localized, it is situated in a lateral position. H o w e v e r , in a n u m b e r o f cases it a p p e a r s s o m e w h a t diffuse. Such diffuseness c o u l d reflect staining o f b o t h passage fibers a n d t e r m i n a l s r a t h e r t h a n necessarily i n d i c a t i n g a diffuse t e r m i n a t i o n . To assess the effect o f t i m i n g o f the initial c o r t i c a l lesion on the f o r m a t i o n o f the a b e r r a n t crossed c o r t i c o t e c t a l p a t h w a y , bilateral eye enucleations were p e r f o r m e d on a set o f a n i m a l s at birth, a n d subsequently, a u n i l a t e r a l visual c o r t i c a l lesion was m a d e on p o s t n a t a l days 5, 10, 15 a n d 20. A f t e r 2.5 m o n t h s , animals f r o m each g r o u p were t a k e n w i t h o u t further surgery a n d stained with the F i n k - H e i m e r m e t h o d to test for residual d e g e n e r a t i o n froi"n the initial lesions. N o residual d e g e n e r a t i o n was
42 found. Second lesions were made on similar animals involving the entire remaining visual cortex. They show a heavy crossed corticotectal projection in animals when the initial cortical lesion was made on postnatal days 5 and 10 that differed little in density or distribution from the crossed corticotectal projections observed in animals that received cortical lesions and eye enucleation at birth. Those animals in which the initial cortical lesions were made on day 15 show a crossed distribution similar to the previous animals but this is of reduced density, being about 20 ~ of the amount found in 5- and 10-day cases. Those animals with initial cortical lesions made on postnatal day 20 show very light crossed degeneration and this can be only found near the midline. DISCUSSION The corticotectal pathway from the visual area like that from the sensorimotor cortex 1° shows the capacity for giving a crossed projection in the absence of a corticotectal pathway from the opposite side. This occurs even though there is not a natural commissure joining the appropriate laminae. Electron microscopy shows that the aberrant pathway forms synaptic contacts in the contralateral colliculus. Whether these contacts can affect the physiology of the postsynaptic cells or whether they are suppressed by other inputs remains to be determined. The crossed projection is not the result of a random process and is subject to certain constraints. For example, it ends only in laminae appropriate for the visual corticotectal pathway and shows a pattern of topographic order which relates to the retinotectal map although not to the map of the visual field. As in most other systems in which a major reorganization of a pathway Occurs 14,16,20,2a, the aberrant pathway shows progressively less development, the older the animal at the time of the initial lesion. Thus by 20 days very little crossed degeneration is found. This is comparable in timing for the crossed corticotectal pathway from the sensorimotor cortex 9 but later than the aberrant ipsilateral retinotectal pathway (10-12 day) induced by unilateral eye lesions a4. It would be expected, therefore, that the restriction of the capacity to sprout reflects some component of the individual pathway rather than a stabilization within the tectum itself, such as of the astrocytic component. The relationship between the corticotectal and retinotectal pathways is particularly interesting. First, as discussed previously, if the eyes are removed at birth the corticotectal pathway, either regular or crossed appears to sprout to innervate the whole stratum griseum superficiale. Such apparent sprouting could be artifactual resulting from loss of the upper part of the stratum griseum superficiale into which optic axons have their heaviest projection. This would seem unlikely since Golgi studies (unpublished) indicate that the cell types characteristic of the upper part of the stratum griseum superficiale s are still present after neonatal eye enucleation. Further, the degree of shrinkage after enucleation is less than would be accounted for by a total loss of the upper half of the stratum griseum superficiale and the increased density of the corticotectal pathway is less than could be accounted for by shrinkage alone. This implies that corticotectal fibers have taken over sites made available by
43 retinal deafferentation. The second point concerning the interrelation of retinotectal and corticotectal pathways relates to the fact that although optic deafferentation may result in an increased depth of distribution of corticotectal fibers within one colliculus, the control studies show that it does not on its own stimulate an aberrant crossing of corticotectal fibers. The underlying mechanism resulting in the formation of the crossed corticotectal pathway is unclear. Certainly it is not an indiscriminate response to deafferentation (or delayed innervation) ofcorticotectal sites. Alternatively, it could reflect an inhibitory interaction normally present between corticotectal axons from each side of the brain. Thus, if the axons on one side are absent during a critical stage of development, those of the other side cross the midline and innervate the area normally served by the absent axons. A similar pattern of mutual inhibition has been indicated in vitro 2 and could be used to explain several other studies concerning in vivo development and plasticity 5,6,Is,z2. It is surprising perhaps that the aberrant pathway occurs even though its normal ipsilateral synaptic connection appears present and undisturbed by the surgery. Furthermore, the stimulus for crossing is effective over considerable distances as from the lateral side of one colliculus to the lateral side of the other. In order to understand more fully the mechanisms controlling the pathway, it would seem important to know how it develops and how this development relates to the normal development of the corticotectal pathway. In particular it would be interesting to know how the topographic order establishes itself. Is it instantaneous with fibers growing directly to appropriate sites or is it progressive in the manner indicated in the moving retinotectal map and compression in fish where fibers gradually form their connections? (see Gaze3; Gaze and Hope 4 for summary). One useful feature of this system is the relatively late time at which sprouting becomes reduced and the similarity of the sprouting after lesions made at 15 days postnatal and at fetal day 16 (when cells generating the pathway are just being formed). This means that the plasticity of the pathway can be more extensively and easily studied than in the retinotectal system where early neonatal and fetal times are necessary in rat and hamster to show plastic phenomenon14,19,2a, 23. A further use of study of the corticotectal system will be to see whether the rules of development derived largely from study of the retinotectal pathway are indeed applicable to other pathways, particularly to ones arising in the cerebral cortex. Particularly interesting are questions concerning the effects of partial visual cortical and superior collicular lesions and how they may compare with similar retinal and tectal manipulations in fish. Do the same sorts of compressions and expansions occur ? Further, can intertectal crossing of corticotectal fibers be induced by tectal lesions as is the case for retinal fibers2~? Thus it is hoped that this system will be one which is amenable to many of the experimental regimens to which the retinotectal systems have been subjected. Its special interests lie in the fact that it is a pathway of cerebral origin rather than one arising from primary afferents; it can be manipulated late in development; and it interacts with the retinotectal system.
44 ACKNOWLEDGEMENTS We thank Ren6e Wise and Joy Baisinger for valuable technical help and Doris Ringer and Vicki Neff for editorial assistance. Supported by U.S.P.H.S. Grants EY-00596 and GM-00136 from the National Institutes of Health. REFERENCES 1 Cowan, W. M., Studies on the development of the avian visual system. In D. S. Pease (Ed.), Cellular Aspects o f Neural Growth and Differentiation, Univ. Calif. Press, Los Angeles, Calif., 1971, pp. 177-222. 2 Dunn, G. A., Mutual contact inhibition of extension of chick sensory nerve fibers in vitro, aT. comp. Neurol., 143 (1971) 491 507. 3 Gaze, R. M., Neuronal specificity, Brit. reed. Bull., 30 (1974) 116-121. 4 Gaze, R. M. and Hope, R. A., The formation of continuously ordered mappings. In M. Corner and D. Swaab (Eds.), Perspectives in Brain Research, Progress hz Brain Research, Vol. 45, Elsevier, Amsterdam, 1976, in press. 5 Gottlieb, D. I. and Cowan, W. M., Evidence for a temporal factor in the occupation of available synaptic sites during development of the dentate gyrus, Brain Research, 41 (1972) 452-456. 6 Guillery, R. W., Binocular competition in the control of geniculate cell growth, J. comp. Neurol., 144 (1972) 117-129. 7 Hunt, R. K. and Jacobsen, M., Neuronal specificity revisited, Curr. Top. Develop. Biol., 8 (1974) 203-259. 8 Langer, T. P. and Lund, R. D., The upper layers of the superior colliculus of the rat: a Golgi study, J. comp. NeuroL, 158 (1974) 405-436. 9 Leong, S. K., Unpublished observations. 10 Leong, S. K. and Lund, R. D., Anomalous bilateral corticofugal pathways in albino rats after neonatal lesions, Brain Research, 62 (1973) 218-221. 11 Lund, R. D., Terminal distribution in the superior colliculus of fibers originating in the visual cortex, Nature (Lond.), 204 (1964) 1283-1285. 12 Lund, R. D., The occipitotectal pathway of the rat, J. Anat. (Lond.), 11 (1966) 51-62. 13 Lund, R . D . , Synaptic patterns of the superficial layers of the superior colliculus of the rat, J. comp. Net~rol., 135 (1969) 179-201. 14 Lund, R. D., Cunningham, T. J. and Lund, J. S., Modified optic projections after unilatera! eye removal in young rats, Brain Behav. Evol., 8 (1973) 51-72. 15 Lund, R. D. and Lund, J. S., Modifications of synaptic patterns in the superior colliculus of the rat during development and following deafferentation, Vision Res., 11, Suppl. 3 (1971) 281 298. 16 Lund, R. D. and Lund, J. S., Synaptic adjustment after deafferentation of the superior colliculus of the rat, Science, 171 (1971) 804-807. 17 Lund, R. D. and Lurid, J. S., Plasticity in the developing visual system: the effects of retinal lesions made in young rats, J. comp. Neurol., (1976) in press. 18 Lynch, G., Mosko, S., Parks, T. and Cotman, L., Hyperdevelopment of the dentate gyrus commissural system after entorhinal lesions in immature rats, Brain Research, 50 (1973) 174-178. 19 Miller, B. F. and Lund, R . D . , The pattern of retinotectal connections in albino rats can be modified by fetal surgery, Brain Research, 91 (1975) 119-125. 20 Mustari, M. J., An aberrant contralateral visual corticotectal pathway in albino rats, Anat. Rec., 181 (1975) 433 (Abstract). 21 Schneider, G. E., Mechanisms of functional recovery following lesions of visual cortex or superior colliculus in neonate and adult hamsters, Brain Behav. Evol., 3 (1970) 295-323. 22 Schneider, G. E., Competition for terminal space in formation of abnormal retinotectal connections, and a functional consequence, Anat. Rec., 169 (1971) 420. 23 Schneider, G. E. and Jhaveri, S. R., Neuroanatomical correlates of spared or altered function after brain lesions in the newborn hamster. In P. G. Stein, J. J. Rosen and N. Butters (Eds.), Plasticity and Recovery o f Function hi the Central Nervou~ System, Academic Press, New York, 1974, pp. 65-109. 24 Sperry, R.W., Chemoaffinity in the orderly growth of nerve fiber patterns and connections, Proe. nat. Acad. Sci. (Wash.), 50 (1963) 703-710. 25 Tsang, Y., Visual centers in blinded rats, J. eomp. NeuroL, 66 (1937) 211-261.