Developmenta I Studies of Thalamocortical and Commissural Connections in the Rat Somatic Sensory Cortex S. P. WISE AND E. G. JONES Department ofAnatomy and Neurobiology, Washington University School of Medicine, St. Louis, Missouri 631 I0

ABSTRACT Autoradiographic, axonal degeneration, and horseradish peroxidase fiber tracing methods were employed to investigate the organization, development and potential plasticity of the thalamocortical projection to the somatic sensory cortex of the rat. In the adult animal, thalamocortical terminals are concentrated primarily in layers I and IV and in the upper part of layer VI. Fibers terminating in layers IV and VI arise from a different thalamic region than those terminating in layer I. Discrete clusters of fibers and terminals 250450 p m wide are distributed only to the parts of the SI cortex containing dense aggregates of layer IV granule cells and not to the intervening, less granular and commissurally connected zones. At birth, thalamocortical fibers have invaded the deep part of the developing SI cortex and are concentrated in the upper part of layer VI. Between the age of two and three days, a n additional concentration of fibers appears in the part of the cortical plate which will become layer IV. Layer IV is clearly recognizable by three days of age and the dense granule cell aggregates appear in i t no more than one day later. The ingrowth of commissural fibers (Wise and Jones, ’76) lags behind t h a t of thalamic fibers. The mature commissural fiber pattern is not established until the age of seven days. After removal of the developing thalamocortical system by thalamotomy in newborn rats, subsequent investigation of the commissural system in the adult showed t h a t no commissural fibers or terminals had invaded either laminae or zones of the cortex deprived of thalamic input. Similarly, commissurotomy at birth was not followed by sprouting of thalamic fibers into zones or laminae deprived of commissural connections. The connectional specificity observed in these neocortical fiber systems contrasts markedly with the plasticity of connections reported in allocortical systems. Removal of thalamocortical afferents before they attain their definitive distribution does not radically effect the overall development of the dense granule cell aggregates in layer IV. Within the aggregates, however, subsidiary features such a s the “barrels” fail to appear. This finding suggests t h a t certain elements of cortical architecture such as the dense granule cell aggregates are independent of thalamic afferents while others, such as the barrels, result from the interaction of the developing thalamocortical fibers and/or terminals with maturing neurons. Thalamocortical fibers projecting into the first somatic sensory cortex (SI) of the rat arise from the ventrobasal complex (VB) and the central lateral nucleus (Jones and Leavitt, ’74) and terminate in the cortex in a somato‘76; (Emmers’ ’65; SaPorta and Kruger, ’77). The SI cortex of the rat consists of areas containing dense aggreJ. COMP. NEUR. (1978) 178: 187-208.

gations of granule cells in layer IV, with intervening and surrounding less granular areas. Within many of the larger aggregates, subsidiary (“barrel”) formations (Welker, ’76) are

’ Supported by grant numbers NS10526 and EY00092 from the National Institutes of Health, U.S. Public Health Service. We thank MS. Bertha McClure. Mr. James Fleshman. and Mr. Ronald Steiner for their excellent technical assistance 187

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seen. Only the granular areas are activated by peripheral somesthetic stimuli in anesthetized animals (Welker, '76) ; the less granular areas contain the commissural and other corticocortical fibers (Ryugo and Killackey, '75; Wise and Jones, '76). Previous studies have established the time course of development and the laminar, topographic, and columnar organization of the cpmmissural system of the rat SI cortex (Wise and Jones, '76). The present investigations were undertaken: (1)to obtain similar information concerning the thalamocortical system, (2) to investigate the spatial relationships of the commissural and thalamocortical systems in the normal rat SI cortex, (3) following the demonstration of considerable plasticity in the pattern of connectivity of certain afferent fibers to the allocortex (Westrum, '75; Lynch and Cotman, '76; Zimmer, '76) to evaluate the degree of connectional plasticity or specificity of the neocortical connections; (4) to determine the influence of the developing thalamocortical system upon certain architectural features of the SI cortex. Some of these data have been previously presented in preliminary or abstract form (Wise, '75, '76, '77). MATERIALS AND METHODS

A total of 123 albino rats of the Wistar strain were used in these experiments. A variety of experiments were performed.

Organization of thalamocortical fibers In eight adult rats, injections of radioactive amino acids were made stereotaxically into the ventrobasal complex. Each injection consisted of 0.1-0.2 p1 of a 50 pCiIp1 solution of 5[3H1-l-proline(specific activity 17 Ci/mmole) and 4 3 -[3H1-l-leucine(specific activity 46 Ci/ mmole) and was made through a 31-gauge needle attached to a 1-pl syringe or with air pressure through glass micropipettes having tip diameters of 25-50 pm. After a 1-or 2-day survival period the animals were perfused through the heart with 10%-buffered formalin. The relevant hemispheres from four of the brains were then flattened under pressure (Welker, '76; Wise and Jones, '761, sectioned on a freezing microtome a t 30 pm, mounted on glass slides and dipped in Kodak NTB-2 emulsion for autoradiography (Cowan et al., '72). The other brains were embedded in Paraplast, sectioned a t 20 p m in the frontal plane and also prepared for autoradiography. After a n exposure period of two weeks, the autoradio-

graphs were developed in Kodak D-19 a t 1 7 T , fixed in Kodak Ektaflo and stained through the emulsion with 0.25% thionin.

The spatial relation of thalamocortical and comrnissural fibers In two rats, the ventrobasal complex was lesioned stereotaxically from behind. Immediately thereafter, the corpus callosum was sectioned. The animals were killed after five days by perfusion with formalin. Thirty-micron thick sections were later cut on a freezing microtome and alternate sections were stained by the Wiitanen ('69) method for degenerating fibers and terminals and with thionin. The pattern of thalamocortical degeneration could then be compared to the pattern of granule cell aggregates in layer IV of SI (Welker, '76) and to the distribution of commissural fibers (Wise and Jones, '76). Development of thalamocortical and corticothalamic fibers The development of thalamocortical fibers was investigated by autoradiography in 56 rats of age zero to eight days. In these animals 0.05-0.1 pl injections of 25-50 pCiIp1 [3Hl-proline and leucine were made 1.5 mm anterior and 1.5 mm lateral to lambda and 3.7 mm deep to the cortical surface. The development of corticothalamic fibers was investigated in 16 rats, zero to eight days old and adult, in which 0.05-0.1 p l of the same mixture of proline and leucine was injected into the SI cortex. After a 1-day survival the animals were perfused through the heart with 10% buffered formolsaline or with a solution for formol (10%)acetic acid (5%) in 80% ethanol. All brains were embedded in Paraplast and prepared for autoradiography in the manner described above. A further 18, 1- or 2-day-old animals were injected with 0.05-0.1 p l of a 500 pg/pl solution of horseradish peroxidase (Type VI, Sigma). After a 1-day survival period they were perfused with 1.0-2.0%paraformaldehyde and 1.5-2.5% glutaraldehyde in 0.1 M phosphate buffer. Fixative solutions containing 1%paraformaldehyde and 2.5% glutaraldehyde were found to yield optimal results. These brains were removed, soaked overnight in 30% sucrose in phosphate buffer, then sectioned at 50-100 p m on a freezing microtome, incubated in 0.05% 3,3' diaminobenzidine tetrahydrochloride containing 0.01% hydrogen peroxide (LaVail et al., '73) and counterstained with 0.25% thionin.

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RAT THALAMOCORTICAL DEVELOPMENT Abbreviations CL. Central lateral nucleus of the thalamus CP, Upper part of the cortical plate P, Cerebral peduncle PO, Medial posterior nucleus of the thalamus

SI, First somatic sensory area of the cerebral cortex SII, Second somatic sensory area of the cortex VB, Ventrobasal complex of the thalamus WM, Subcortical white matter I.VI, Layers of the cerebral cortex

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depth ( p i Fig. 1 Grain density counts over traverses of the SI cortex after injections of PHI-proline and L3H1-leucinein VB and a survival of one day. Top: the pattern of labeling of thalamocortical fibers and terminals in a normal adult animal. Bottom: the grain distribution after a VB injection in a n adult rat whose corpus callosum had been sectioned a t birth (fig. 19). Note t hat the laminar distribution of grains is virtually identical in the normal and commissurotomized animals. Stipple, background labeling.

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Effects o f neonatal lesions on afferent fiber systems and on the SI cortex In nine rats, a lesion of SI was made a t one day of age, by removal of the pia mater. In three other l-day-old rats, the corpus callosum was sectioned. After the animals had reached maturity (>4 months), an injection of 13Hl-leucineand proline was made into the thalamic ventrobasal complex (on the side opposite the cortical lesion or opposite the hemisphere retracted during the commissurotomy). Thereafter, the distribution of thalamocortical fibers and terminals was identified by autoradiography as described above. In seven l-day-old rats the thalamus was damaged unilaterally by passing a 25-gauge needle anteriorly in the horizontal plane through the superior colliculus. In five of these rats, after they reached maturity, a single injection of 1 p1 of [3H1-proline and leucine was made into the SI cortex on the side opposite the lesion. After a 2-day survival, the distribution of commissural fibers was studied autoradiographically in sections cut in the frontal plane a t 20 pm. Brains from the other two thalamotomized rats were sectioned on a freezing microtome a t 3 0 p m in the frontal plane. RESULTS

The first somatic sensory cortex (SI) of the rat consists of a molecular layer (layer I) and five cellular layers. Of these, the supragranular layers (layers I1 and 111) often appear fused. Layers V and VI are the main source of efferent fibers (Wise and Jones, ’77) and contain the largest pyramidal cells of the SI cortex. These are situated in the deep part of layer V. The granular layer, layer IV, consists of a number of densely packed aggregations of granule cells which include the barrel fields (Welker and Woolsey, ’74) and which each contain the representation of a specific body part (Welker, ’76). These aggregations are surrounded by zones of cortex in which layer IV is much less distinct. The laminar organization of the second somatic sensory cortex (SII) is similar to that of SI except that the infragranular layers are more prominent and the internal granular layer is nowhere as distinct as in the granule cell aggregates of SI.

Organization o f the thalamocortical fibers Injections of tritiated amino acids which

heavily label cells in the ventrobasal complex (VB) lead to autoradiographic labeling, indicated by a n increase in the density of silver grains over background levels, in both the SI and SII cortex. The laminar organization of thalamocortical fibers and terminals is identical in both areas. After injections confined to VB, the labeling of the SI and SII cortex is most heavily concentrated a t two distinct depths within the cortex (fig. 1).The major peak in grain density (fig. 1) is found over the internal granular layer. This band of labeling typically extends for up to 1 0 0 p m superficial to the upper boundary of layer IV and into layer 111. Beneath this band of labeling a separate, smaller peak in grain density is found a t the border of layers V and VI. Most of the grains are in the upper third of layer VI (fig. 1) but there is an extension into layer V which is most clearly seen in neonatal animals. The layers with the peaks in grain density probably represent those in which the majority of thalamocortical fibers terminate. Sparser accumulations of silver grains occupy large parts of the infragranular layers but these are interpreted as representing labeled thalamocortical fibers. Identical observations have been made using degeneration methods after lesions of the thalamic ventrobasal complex (figs. 2, 3). The thalamocortical fibers emanating from VB a r e not d i s t r i b u t e d homogeneously throughout the SI cortex. First, there is a spatial organization such that only the parts of SI which contain the aggregations of granule cells (including the barrel fields of Welker and Woolsey, ’74) receive thalamocortical fibers and terminals. The intervening and surrounding less granular zones generally have only background levels of labeling as seen in autoradiographic material (figs. 14, 15). Second, the thalamocortical fibers entering a large granule cell aggregate are organized in disjunctive, column-like clusters, each 250450 p m in width. In parts of the ventrolateral aspect of SI these column-like clusters abut each other directly and the density of labeling is so high t h a t each individual cluster can only be distinguished from its neighbor with difficulty. In the barrel fields, however, and also in the dorsomedial parts of SI, the distance between each of the thalamocortical fibers and terminal clusters is greater and the columnlike bundles are discrete and separate (figs. 2, 3). Where the granule cell aggregates have within them distinct “barrels” (Welker and

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Figs. 2 , 3 Dark- and brightfield photomicrographs of immediately adjacent 30 fim frontal sections through the SI cortex. Figure 2 shows in darkfield, degenerating thalamocortical fibers and terminals stained by Wiitanen's method five days after a lesion of VB. The degeneration appears light in darkfield. Note the disjunctive clusters of thalamocortical terminals in both layers IV andVI, of the barrel field (ventrolaterally)as well as in more dorsal parts of t he cortex. Figure 3 is the alternate section stained with thionin. Arrows mark the same blood vessels in each section. Note that the thalamocortical fibers innervate only those patches of SI cortex containing dense aggregates of granule cells in layer IV, and not the surrounding, less granular parts of SI. X 159

Woolsey, '74; Welker, '761, with a central zone of low neuronal density and a peripheral zone of high neuronal density, the thalamocortical terminals are concentrated in the central zone (cf. Killackey, '73; Killackey and Leshin, '75; Caviness et al., '76). The column-like organization of thalamocortical fibers and terminals is most obvious in material stained for degeneration by the Wiitanen ('69) method after lesions of VB (figs. 2-51. With this method, the distinction between fibers and terminals is greater than in the autoradiographic material and the areas of high thalamocortical terminal concentration can be clearly observed. The labeling of the SI thalamocortical system by autoradiography or degeneration shows that the thalamocortical terminal ramifications in both layers IV and VI conform t o the column-like pattern and the labeling in the two layers is in register, i.e., a zone of increased density in layer VI lies immediately deep to one in layer IV (figs. 2, 3).

After injections of L3H1-aminoacids, which also spread to involve parts of the thalamus medial to VB, there is heavy labeling of the superficial half of the molecular layer of SI and SII. This band of labeling is never heavy when an injection is confined to VB (fig. 1). Labeling of the outer part of layer I is often observed in the absence of large peaks in grain density in the deeper, cellular layers. I t appears therefore, t h a t the thalamocortical fibers which are found in layer I arise from a part of the thalamus distinct from those which terminate in the other layers. No further identification of the source of these fibers could be made. Their developmental history will be considered with those arising from VB.

Segregation of thalamocortical and commissural fibers It has already been demonstrated (Wise and Jones, '76) that commissural fibers entering the rat SI are distributed in a series of column-

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Figs. 4,5 Higher power dark- and brightfield photomicrographs of adjacent sections from the same brain from which figures 2 and 3 were taken. The same blood vessels are marked (X). The Wiitanen stained section (fig. 4) is slightly shrunken relative to the thionin stained section (fig. 5). Nevertheless, the three thalamocortical terminal clusters can be seen to be concentrated within the dense aggregates of layer IV granule cells. Figure 4 also shows part of t h e deeper, infragranular concentration of thalamocortical terminals. X 60.

like bundles primarily to the relatively agranular zones of cortex lying between and around the granular cell aggregates. Within the column-like bundles, terminal ramifications are

concentrated in two major zones: in layers II11 and V-VI. This columnar and laminar distribution has been confirmed in the present set of experiments. In addition, the experi-

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ments in which commissural and thalamocortical fibers were labeled in the same experiment clearly indicate a segregation of the two fiber systems. Figure 6 shows clearly separate bundles of degenerating commissural and thalamocortical fibers that give rise to dense clusters of degenerating terminal ramifications that do not overlap to any extent.

Development of the thalamocortical fibers At birth the cortical plate consists of two parts, a superficial, densely packed zone of highly immature and probably to some extent still migrating cells, and a deeper, less densely packed, and much thicker part in which layers V and VI are already distinguishable (the “subplate” of Rice and Van der Loos, ’77). Unpublished experiments employing the retrograde transport of HRP in 1-day-oldrats have demonstrated that the corticofugal fibers emanating from the SI cortex and projecting to brainstem sites and the spinal cord originate from neuronal somata confined to layer V (S. P. Wise, P. G. H. Clarke and E. G. Jones, unpublished data), as they do in adult rats (Wise and Jones, ’77). This finding supports the designation of this lamina in the neonatal cortical plate, as layer V. The superficial, densely packed zone will be referred to as the upper cortical plate. Superficial to the upper cortical plate is the marginal zone or immature molecular layer. Deep to the cortical plate, as a whole, is the intermediate zone which consists mostly a t this stage of the immature subcortical white matter. For the purposes of describing fiber development, rats in their first postnatal day are termed 0 day old and those in their second day 1day old. The recorded age of the animal, unless otherwise specified, is the age a t perfusion. In all cases, preliminary data on the time course of thalamocortical fiber development based on age measurements starting simply a t birth were verified with animals of known gestational age (starting a t 21-22 days postcoitum). No differences in the pattern of thalamocortical development are observed between SI and SII, but the descriptions and illustrations will be of the development of these fibers in SI. In rats in which VB and adjacent thalamic nuclei were injected a t zero and 1day old and which were killed within 1 2 to 24 hours, the pattern of thalamocortical labeling in SI is quite unlike that in the adult. Injections of

Fig. 6 A darkfield photomicrograph of a 30-pm frontal section from the dorsomedial part of SI stained by the Wiitanen method and taken from a rat five days subsequent to a lesion of VB and section of the corpus callosum. The degenerating bundles of thalamocortical (right) and commissural (left) fibers, can be distinguished on t h e basis of their characteristic laminar organization. The fibers from these two afferent fiber systems are also clearly distributed to different, though adjacent, regions of the SI cortex. Surface of brain indicated by arrows. X 60.

the thalamus in these animals lead to a concentration of transported label over the outer part of layer VI and over the outer part of layer I but labeling over the intervening upper cortical plate does not greatly exceed background levels (figs. 7, 8, 16). The labeling of the subcortical white matter is very heavy relative to that of the gray matter (figs. 7, 8, 161, suggesting the presence of fibers which have not penetrated the cortex. In 1- to 2-day-old rats killed 24 hours after injecting horseradish peroxidase (HRP) into the presumptive SI cortex and underlying white matter, there is heavy retrograde labeling of cells in the ipsilateral ventrobasal complex as well as in the central lateral nu-

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Figs. 7 , 8 Dark- and brightfield photomicrographs from a 20-pm thick autoradiographic section taken from an animal which was perfused a t one day of age, 18 hours after a n injection of PHI-proline and PHI-leucine into th e thalamus. Accumulated labeling can be seen to be confined to the subcortical white matter and to layer VI of th e developing cortex. Layer V is present but no granular or supragranular layers can be distinguished a t this age (fig. 8). Arrow marks t he pial surface (see fig. 16).Thionin counterstain. X 70.

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Fig. 9 Darkfield and brightfield (inset) photomicrographs from the same part of a 50-gm frontal section through the thalamus. The darkfield photomicrographs show neurons labeled by retrograde axonal transport of horseradish peroxidase. The animal was two days old when perfused, 24 hours after a n injection of the enzyme into the presumptive SI cortex. Note the large number of retrogradely labeled thalamocortical neurons in VB and the central lateral nucleus (CL). The large number of labeled thalamic cell somata suggests that even a t two days of age, most of the thalamocortical cells have fibers in the cortex or subcortical white matter. Note also the clustering of neurons in the arcuate nucleus of VB which projects to the barrel fields of SI. Thionin counterstain. Figure 9, x 65; inset X 25.

cleus, both of which are recognizable a t this age. Virtually every neuron in VB is found to be heavily labeled with granules of HRP reaction product (fig. 9) and clustered aggregations of cells are clearly seen in the arcuate nucleus (fig. 9). A similar pattern of labeling is observed in all animals injected with HRP a t subsequent ages including adults. Injections of [3H1-aminoacids made into the thalamus late on the third day of postnatal life (2-day-oldanimals) and a t the age of three days lead t o a more adult like pattern of labeling in the cortex. In these cases, label in the presumptive SI is concentrated in three zones: in the upper part of layer VI with extension into the deep part of layer V; in the outer part of layer I; in the deepest aspect of the upper cortical plate (figs. 10, 11, 16). Since this part of the cortical plate contains, a t this age, the cells which form layer IV in the mature animal (Berry and Rogers, '65; Rice and Van der LOOS,'771, the thalamocortical fiber distribution in these 2- to 3-day-old animals has at-

tained a pattern similar to that of the mature rat. There is little evidence, a t this stage of development, for a disjunctive clustering of the thalamocortical fibers. In the subcortical white matter, labeling is heavy but is not much more intense than the labeling in the cortex; in contrast to the high ratio of subcortical to cortical labeling observed in younger animals. There is however, considerable labeling of the deep layers of the cortex which suggests the continued presence of fibers which have yet to reach their ultimate destination in more superficial parts of the cortex (fig. 16). None of the dense aggregates of layer IV cells are readily distinguishable a t three days of age. Layer IV itself is only barely discernible from the rest of the upper cortical plate a t this stage. Between the ages of three and four days, however, the deepest part of the upper cortical plate matures into a clearly recognizable layer IV (DISCUSSION and fig. 13) and shortly thereafter, the aggregates of cells in

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Figs. 10, 11 Dark- and brightfield photomicrographs of the same autoradiograph from an animal perfused very late on its third postnatal day (late 2 days old), after injection of PHI-amino acids into the thalamus 24 hours previously. The fibers have grown into the deepest part of the upper cortical plate (CP) and have adopted a relatively mature distribution pattern (cf. fig. 16: 3 days old). A clear layer IV is not yet discernible but it will form the deep part of the upper cortical plate which already contains thalamocortical fibers. 20-pm thick frontal section, thionin counterstain. X 125.

layer IV become separated from each other to form the pattern of aggregates (some of which contain barrels) seen in the adult (figs. 3, 5, 20). The laminar distribution of thalamocortical fibers a t four days old is virtually identical to that of the adult. After injections of L3H1amino acids which involve VB a t this age, the heaviest concentration of labeling is over layer IV, extending into the deepest part of a now recognizable layer 111. Additional peaks of labeling occur over the outer part of layer I, and over the upper part of layer VI, extending into layer V in the zone beneath the layer of large pyramidal cells (figs. 12, 13). By the age of four days, the labeled thalamocortical fibers are clearly distributed only t o the granule cell aggregates and now form disjunctive, column-like masses approximately 100-400 p m in width. The labeling of the cortex a t short survival periods is now greater than that

of the white matter and this pattern is found in adulthood. A t ages later than four days, the cortex, particularly the supragranular layers, continues t o mature and the upper cortical plate becomes an obvious layer 11. By six to seven days of age, the supragranular layers have begun to achieve an adult-like architecture, though initially they are still much thinner than in the adult. The distribution of thalamocortical fibers, as demonstrated by the density of labeling following injections of PHI-amino acids into the thalamus, does not greatly change after this time (figs. 14-16).As noted above, however, it becomes possible to distinguish in the larger brain the differential thalamic projection to layer I from that to layers IV and VI. Moreover, the proportion of labeling in layer IV increases (fig. 16) during the first postnatal week.

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Figs. 12, 13 Dark- and brightfield photomicrographs of the same autoradiograph showing labeled thalamocortical fiber ramifications in the SI cortex of a 4-day-old rat after injection of PHI-proline and leucine in the thalamus 24 hours previously. By this age a clear layer IV is seen, as well as a n immature layer 111, but a densely packed remnant of the upper cortical plate remains (fig. 13).The labeling is concentrated over layer IV (extending into the deep part of layer 111). and over layer VI (extending into the deep part of layer V). There is also some labeling of layer I, and heavily labeled fibers appear in the subcortical white matter. Thionin counterstain. X 80.

Development of corticothalamic fibers Injections of PHI-amino acids into the cortex of O-day-oldanimals perfused a t one day of age, show that fibers, at this early stage, have reached the ventrobasal (fig. 17), posterior, reticular, central lateral and other nuclear complexes in the thalamus. In comparison with similar experiments conducted in adult animals, the density of labeled corticothalamic fiber ramifications in the thalamic neuropil of the neonate does not appear to be as great. Conversely, a relatively larger proportion of labeling is observed over fiber bundles within the external medullary lamina and in the ventrobasal complex than in the adult. The distribution of the corticothalamic labeling

changes little over the course of post-natal development and the relative proportions of fiber and neuropil labeling assume their adult pattern within the first two or three days. A more complete description of the distribution of corticothalamic fibers emanating from the SI of the mature rat has been published elsewhere (Wise and Jones, '77).

Influence of thalamocortical and commissural fibers on cortical cytoarchitecture In the brains of adult animals, thalamotomized a t birth, some aspects of cortical cytoarchitecture in SI, such as the dense, granule cell aggregates are easily seen, especially in the 30-pm thick sections. In a typical case most of the ventrobasal

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Figs. 14, 15 Dark- and brightfield photomicrographs taken from a 20-pm frontal section of t h e brain of a r a t perfused when six days old after injection of PHI-proline and PHI-leucine into the thalamus 24 hours previously. At this age all layers of the SI cortex are identifiable and the labeled thalamocortical fibers have adopted an adult-like distribution pattern (fig. 16). Note the disjunctive clustering of grains in layer IV (see also figs. 2-5) and the laminar concentrations in layers IV and VI. Arrows mark same blood vessel. Thionin counterstain. X 65.

complex and many other thalamic nuclei are clearly absent on the operated side. In the case illustrated (fig. 181,only the most dorsolateral aspect of VB, a large part of the lateral posterior nucleus, and a few of the midline nuclei, are present on the affected side, while the contralateral thalamus is largely unaffected. Incidental damage includes significant destruction of the striatum, dilation of the lateral ventricle with displacement of the hippocampal formation, and a nearly complete section of the internal capsule. The latter is reflected in a considerable reduction in the size of the cerebral peduncle and pyramidal tract on the side ipsilateral to the lesion (fig. 21). In the cortex there is shrinkage of the layerV pyramidal cells in all areas, particularly the sensory and motor. In those areas of SI topographically related to the lesioned part of VB,

the total combined thickness of the supragranular and granular layers is significantly reduced (13%; p < 0.001, Kolmogorov-Smirnov test) and the layers appear thinner and more palely staining upon qualitative examination (figs. 20, 21). The aggregates of layer IV granule cells on the lesioned side of the brain, do not appear as dense as on the contralateral (normal) side. However, they are roughly equivalent in position t o the normal, their horizontal extent is the same and they are separated from each other by relatively less granular cortex with about the same spacing as normal. Subsidiary cytoarchitectural features of the layer IV aggregates, on the lesioned side, particularly the barrels with clear centers, are not seen, though they are obvious in the control hemisphere (figs. 20, 21). In the case illustrated, a

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Fig. 16 Grain density counts of traverses through t h e SI cortex subsequent to VB injections and autoradiographic demonstration of thalamocortical fibers and terminals a t various postnatal ages. The highly immature pattern of grains seen a t one day, in which there is a high concentration of grains in the subcortical white matter and a secondary peak a t the border of layers V and VI (figs. 10, ll),gives way to a much more adult-like laminar distribution by three days of age (figs. 12,131.The proportion of labeling in layer IV increases over the first postnatal week. Bv nine davs of aee the thalamocortical fiber distribution is indistinguishable from that of the adult (cf. fig. 1). "

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few barrels are observed in the caudal aspects of SI on the lesioned side, in a part of the cortex where the reduction of thickness in layer IV is also minimal. The preservation of this part of SI is considered t o be due to the maintenance of some normal thalamic connections with the intact dorsolateral part of VB (Davidson, '65; Emmers, '65). In other cases with thalamic lesions in which all or part of VB had been destroyed a t one day of age, the same observations were made. In each case, the part of SI topagraphically related t o the part of VB removed has attenuated; granule cell aggregates are

present but no barrels appear within them. In all cases, the contralateral, control hemisphere shows dense granule cell aggregates of normal thickness and with a normal complement of barrels. In complementary experiments, the corpus callosum was sectioned a t zero or one day of age and the cortex subsequently examined a t maturity (fig. 19). In these cases there is incidental damage to the cortex of one and sometimes both sides, but in all cases the dense, granule cell aggregates and the barrels within those that normally contain them, appear unaffected, in spite of the failure of the com-

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Fig. 17 Darkfield and brightfield (inset) photomicrographs taken from a 20-rm thick frontal section of the thalamus of a rat perfused at 1 day old 12 hours after an injection of PHI-amino acids in the presumptive SI cortex. This experiment demonstrates that before the thalamocortical fibers have established a mature organizational pattern (fig. 16), corticothalamic fibers, as demonstrated by transported label, are present in the ventrobasal complex. Thionin counterstain. Figure 17, X 70; inset, x 25.

missural fibers t o penetrate the intervening less granular areas.

Connectional specificity of thalamocortical and commissural fibers Several experiments were performed to evaluate the potential of either commissural or thalamocortical fibers to form aberrant connections following the destruction of the other set of fibers early in development. In mature animals in which the commissural system had been removed by hemidecortication or commissurotomy (fig. 19) a t zero or one day of age, the distribution of thalamocortical fibers was demonstrated by injection of PHI-proline and PHI-leucine into VB. In all of these cases, the distribution of thalamocortical fibers in SI and SII was identical to that seen in normal, mature animals (fig. 1).After injections of VB and adjacent nuclei, the labeling is concentrated in the outer part of layer VI extending into layer V, in layer IV extending into layer 111, and in the superficial

part of layer I. The labeling of thalamocortical fibers remains restricted to those areas of cortex containing the dense aggregates of granule cells, as in normal animals, even though the commissural fibers have never entered the cortex and are absent from the adjacent, less granular zones of SI. In parallel experiments, a thalamotomy at one day of age (fig. 18) was followed by investigation of the distribution of commissural fibers and terminals in the mature animal. Here, also no discrepancy could be detected between the pattern of organization of the commissural fibers in comparison with normal rats. The commissural fibers, in the experimental cases in which VB and other thalamic nuclei had been destroyed terminate in discrete patches in the relatively agranular parts of the SI cortex, between and surrounding the dense, granule cell aggregates. The patches are 2 0 0 - 3 0 0 ~ min width and labeled fibers and terminals are concentrated primarily in the supragranular layers (fig. 22).

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Fig. 18 Photomicrograph of a 30-pm thick frontal section of the brain of an adult rat that sustained a unilateral thalamotomy (right) a t birth. Only a small portion of the lateral thalamic complex remains intact on the lesioned side. Frozen section, thionin stain. X 10. Fig. 19 Photomicrograph of a 20 pm thick, frontal section of the brain of an adult rat whose corpus callosum had been sectioned a t birth. The right hemisphere has been damanged by retraction during surgery. Injections of tritiated amino acids in the thalamus of the undamaged side reveal no abnormalities of the thalamocortical fiber distribution (fig. 1). Paraffin section, thionin stain. X 10. DISCUSSION

Laminar organization The terminal ramifications of thalamocortical fibers in the somatic sensory cortex of the rat are concentrated a t three depths: (i) a t the junction of layers V and VI, mostly in layer VI, (ii) in layer IV with an extension into the

deepest part of layer 111; (iii) in the superficial part of layer I. These findings are comparable to those that have recently been reported with autoradiographic and electron microscopic studies in other species and in other areas of the rat cortex (Drager, '74; Strick and Sterling, '74; Rosenquist et al., '74; Benevento and Rezak, '75; Jones, '75a; Ribak and Peters, '75;

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Figs. 20,21 Photomicrographs taken from the two sides of a frontal section of the brain of an adult r a t which had sustained a thalamotomy a t birth (fig. 18). The SI cortex on the normal side (fig. 20)shows some of the dense aggregates of granule cells in layer IV. The edges of these aggregates are marked by thin arrows. Within one of the aggregates, "barrels" can be seen (open arrows). The SI cortex from the lesioned side (fig. 21) is significantly reduced in thickness, and the cells of the infragranular layers are markedly shrunken but the aggregations of layer IV granule cells, the edges of which are marked by thin arrows, remain. However, no barrels are present in the lower aggregate corresponding to that shown in figure 20. Thirty-micron thick frozen sections, thionin stain. x 40.

Robson and Hall, '75; Wise, '75; Jones and Burton, '76; LeVay and Gilbert, '76; Peters and Feldman, '76; Rakic, '76, '77). The observation that a thalamic projection to the superficial part of layer I arises from a nucleus of the thalamus other than VB, accords well with previous investigations of thalamic inputs to this layer (Benevento and Rezak, '75; Jones, '75a; Jones and Burton, '76). The identification of the cells of origin of this projection remains uncertain. The likely origin seems to be in the intralaminar nuclei (Jones and Leavitt, '74) but other nuclei such as the ventromedial complex (Herkenham, '76) are also possible.

Topographic and spatial organization The terminal ramifications of thalamocortical fibers in the mature rat SI are restricted to those zones of cortex which contain the dense granule cell aggregates described by Welker ('76) and Welker and Woolsey ('74). This findFig. 22 Grain density count of a traverse through t h e SI cortex of 4 month old rat, thalamotomized a t birth (fig. 18) and injected with PHI-amino acids in the SI cortex of the contralateral (undamaged) side. Twenty-four-hour survival. As demonstrated autoradiographically, commissural fibers projecting from the normal hemisphere into the hemisphere deprived of thalamic input, have a laminar distribution pattern typical of normal animals. The inset shows the grain distribution of labeled commissural fibers in normal animals (Wise and Jones, '76: fig. 2).

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ing has been reported previously in abstract form by Ryugo and Killackey ('75; see also Killackey and Belford, '76). The commissural fibers do not penetrate the granule cell aggregates (Ryugo and Killackey, '75; Wise and Jones, '761, and as shown in the present study, there appears to be little or no spatial overlap between the two afferent systems. In the rat the relatively agranular zones also contain the cells that give rise to the commissural fibers (Wise and Jones, '76; see also York and Caviness, '75, in the mouse). Therefore, there appears to be a complete segregation of thalamic and commissurally related cortex. Whether the relatively agranular areas of SI between and surrounding the granule cell aggregates are truly "athalamic" or receive thalamic input from some as yet unidentified thalamic nucleus is not known but according to Welker ('761, the less granular zones between the aggregates cannot be activated by stimulation of the periphery in deeply anesthetized animals. No similar "athalamic" areas seem to be present in the SI cortex of cats or monkeys. In these species, the distribution of bundles of commissural fibers is also disjunctive but where commissural bundles are present they overlap bundles of thalamocortical fibers to a considerable extent (Jones et al., '75; Wise and Jones, unpublished observations). Columnar organization In the sensory areas of the cortex, the similarities in receptive field and submodality properties of cells in vertically oriented columns extending through all layers are thought to be based in the first instance upon the thalamocortical input (Mountcastle, '57; Powell and Mountcastle, '59; Hubel and Wiesel, '62, '72, '74a,b; Abeles and Goldstein, '70; Wiesel et al., '74; Jones, '75a,b; LeVay et al., '75). Columns of this type have been found to have a discrete cylindrical or slab-like organization. They measure about 500 pm in width in primates and are somewhat narrower in rats (Armstrong-James, '75; Wise and Jones, '76) though the total width of the terminal aborization of single thalamocortical fibers as demonstrated by the Golgi technique is similar in both rodents (Lorente de NO, '49) and monkeys (Jones, '75a). In the present study, it has been possible to demonstrate the same type of column-like organization with axonal degeneration and autoradiographic techniques. The 250-450 p m wide clusters of thalamocortical ramifications in layer IV, seem to

represent the terminations of bundles containing many fibers. As in the layer IV clusters, in the infragranular layers there is a similar but less dense, secondary patch of apparent thalamocortical terminations. This cluster has similar dimensions and is in register with a cluster in the overlying layer IV. The column-like clusters of thalamocortical fibers with their bilaminar termination are found in relation to each of the large aggregations of granule cells that make up layer IV. They are most discrete and most clearly separated from one another in the aggregations which represent the mystacial and other facial vibrissae (see also Killackey and Leshin, '75; Caviness et al., '76) and in the dorsomedial cortex which represents limbs and trunk. Each cluster is interpreted as forming the basis of a n electrophysiologically defined column (see also Axelrad et al., '76; Durham and Woolsey, 77). Development At about the time of birth, thalamocortical fibers have penetrated the somatic sensory cortex and accumulate in the upper part of layer VI. At this time, corticothalamic fibers which in the adult arise from layers V and VI (Wise and Jones, '77) have already entered the ventrobasal complex (see also Wise et al., '77 in the cat). At present, it is uncertain whether the peak of thalamocortical labeling seen in layer VI in our experiments a t these earliest stages represent fibers terminating there, or growing fibers which ultimately will terminate in more superficial laminae. The heavy concentration of label in the white matter beneath SI suggests the presence of many growing fibers that have not yet entered the cortex. Although the retrograde labeling experiments would imply that the axons ofall the thalamocortical relay neurons in VB have already reached this general region by the time of birth, no evidence has been obtained in the present study to show whether they accumulate there over a long period of time before invading the cortex. A relatively protracted "waiting period" of this type has been demonstrated in thalamocortical fiber development in the striate cortex of the rat (Lund and Mustari, '77) and monkey (Rakic, '76, '77); in the development of the commissural system of the rat SI cortex (Wise and Jones, '76); in the development of thalamocortical fibers of the cat sensory motor cortex (Wise et al., '77); and in the development of retinotectal fibers in

RAT THALAMOCORTICAL DEVELOPMENT

the chick (Crossland et al., '74). It is likely therefore, that such a stage occurs in the rat SI thalamocortical system, and that it does so prenatally. The thalamocortical fibers grow into the upper cortical plate between the ages of two and three postnatal days. This timing seems to coincide with the first appearance of cortical evoked potentials in response t o peripheral or thalamic stimulation (Verley and Axelrad, '75) and with the appearance of histochemically demonstrable succinate dehydrogenase activity (Killackey and Belford, '76). I t is not yet known, however, whether synapses are already formed a t this stage. In the visual cortex of the rat (Lund and Mustari, '771, synapses start to form as soon as the thalamic fibers begin to enter the cortex. The invading thalamocortical fibers seem to accumulate only in the deepest part of the upper cortical plate and a t no stage are they distributed uniformly throughout the upper cortical plate. Berry and Rogers ('65) have shown that neurons "born" on the eighteenth day of gestation are segregated in the deepest aspect of the upper cortical plate by two to three days of age (24 days post-conception). Here, they will form layer IV of the mature cortex. This suggests that the cells upon which the arriving thalamocortical fibers will terminate aggregate first in layer IV and that the fibers grow directly t o meet them. There is no clear evidence of an initial diffuseness of laminar organization followed by reorganization, even though the deep part of the upper cortical plate does not become obvious as layer IV until about a day after the arrival of the fibers. Thereafter, layer IV becomes discontinuous and forms the dense, granule cell aggregates typical of the adult internal granular layer (at 4 days old). The column-like pattern of thalamocortical fiber distribution is apparently established very early in development for it can be clearly observed in 4-day-old animals. The early development of a columnar pattern is comparable to the development of the precursors of ocular dominance columns in the monkey striate cortex (Rakic, '76, '77) before birth and possibly to the disjunctive patches of corticocortical fibers seen in the frontal cortex of immature monkeys (Goldman and Nauta, '77). Physiological studies have shown that the columnar properties of the rat SI cortex are established by six days of age, but no data are available on younger animals (Armstrong-

205

James, '75). I t is interesting that the small aggregations of VB cells demonstrated with the HRP method, (similar to the "barreloids" of Van der Loos, '76) and which may form the basis of the clustering of the thalamocortical fibers, are present a t or shortly after birth (fig. 9).This finding would seem t o imply that the secondary aggregation of the fibers in the cortex could be to some extent affected by extrinsic factors such as bundling of incoming fibers, and not simply by factors within the maturing cortex itself. Influence of afferent fiber systems on cortical cytoarchitecture Removal of the thalamic afferents to the cortex before they penetrate into the developing granular layer prevents the formation of the barrels, but not of the layer IV granule cell aggregates in which the barrels normally lie. The aggregates, though reduced in thickness, acquire in the absence of thalamic afferents, a similar horizontal extent to that seen in the normal rat. These findings are consistent with the hypothesis that the thalamocortical fibers and terminals interact during development with the immature neurons of the cortex, particularly those of layer IV, and are necessary for the formation of the definitive barrels. The findings of several other studies are consistent with this view. Subsequent t o removal of vibrissae in neonatal mice (Van der Loos and Woolsey, '73; Weller and Johnson, '75) and rats (Killackey et al., '76) the barrels related t o those vibrissae fail to appear in their definitive form. The effect of removal of a vibrissa on the ultimate appearance of a barrel is greatest in very young animals and becomes less with age (Woolsey and Wann, '76). On the fifth postnatal day, when the barrels first become evident in the mouse (Rice and Van der Loos, '771, removal of vibrissae is almost ineffective in altering cortical architecture. Thus, the presence of thalamocortical fibers seems to be unnecessary for the formation of basic cortical lamination and cytoarchitecture, as reflected in the large granule cell aggregates, but the establishment of thalamocortical connections is vital for the appearance of those special subsidiary units of cortical cytoarchitecture that are unique to rodents and a few species belonging to other orders. Connectional specificity The time course of thalamocortical fiber de-

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velopment in the rat SI cortex shows that these fibers have established an adult-like pattern in the highly immature cortex. They have also established this pattern before the fibers of the commissural system penetrate the cortical plate in large numbers. The commissural fibers enter the cortex a t five days of age and only adopt a relatively mature pattern of organi%ation by seven days of age (Wise and Jones, ’76; see also Mires e t al., ‘75). The two fiber systems not only have a different time course of postnatal development but are also distributed largely to different zones and laminae in SI. I t was therefore of interest to determine whether the laminar or spatial organization of either of these fiber systems could be modified by removal of the other system early in postnatal life. Somewhat surprisingly, no experimental manipulation resulted in the formation of aberrant connections by either fiber system. The fibers grow specifically into those zones and laminae of the SI cortex in which they are normally found. These neocortical fiber systems are, therefore, clearly different from comparable systems in the allocortex. In the dentate gyrus similar experiments have shown that commissural fibers have the potential to sprout and occupy laminae and synaptic sites denervated by removal of other afferent fibers normally emanating from the entorhinal cortex (Lynch, Mosko, Parks and Cotman, ’73; Lynch and Cotman, ’75). Similar examples of sprouting of one afferent fiber system into a deafferented zone have been reported for other connections of the hippocampal formation (Lynch and Cotman, ’75; Zimmer, ’76). Although i t probably does not occur in all systems, neuroplasticity of this type has been demonstrated in many other sites (Bernstein and Goodman, ’73; Kerr, ’75). Generally, it has been found that developing or immature neuronal systems are more “plastic” and therefore form aberrant connections more readily than mature systems after experimental manipulation (Lynch, Stanfield and Cotman, ’73; Lund and Miller, ’75; Leong and Lund, ’73). We have not been able to obtain evidence that this occurs in the neocortex. Also of relevance to our findings is the evidence that different fiber systems projecting into the same terminal region compete for available synaptic sites in a manner which gives the earliest arriving fibers an advantage (Gottlieb and Cowan, ’72). Therefore, the finding that the thalamocorti-

cal and commissural systems tend to be dissociated and innervate adjacent patches and different laminae of the SI cortex suggested the possibility that in the absence of competition from one set of fibers the remaining system might grow into the inappropriate zones andlor laminae. This occurred in neither of the two types of experiments carried out. One explanation for the failure of thalamic fibers to sprout into the agranular zones and t h e laminae normally occupied by commissural fibers is that the thalamic fibers clearly arrive in advance of the commissural fibers (Wise and Jones, ’76). They may, therefore, have irreversibly established their appropriate number of synapses before synaptic sites become available in the zones and laminae that are to become commissurally connected, or may have been prevented from doing so by a third competing system. The possibility that regrowth of the commissural fibers (through some route other than the corpus callosum), prevents thalamocortical fibers sprouting, has been ruled out by autoradiographic experiments on split-brain rats in which no labeled fibers could be traced to the opposite side after injections of SI (unpublished results). In the case of the later developing commissural system, removal of thalamic fibers before they establish connections, should leave potential thalamocortical synaptic sites vacant, though it is possible t h a t in the two to three day lag between the arrival of the two systems, thalamocortical synaptic sites, not colonized by thalamic terminals may have failed to appear, appeared and disappeared, or become occupied by a third fiber system. The third “competing system” might be derived from an axonal system intrinsic to the cortex. However, the developmental history and even the mature organization of associational fiber systems in the rat SI cortex is scarcely known (but see Ryugo and Killackey, ’75). It is also possible, in the rat, t h a t the afferent fibers are restricted to a radial orientation, and that limitations on their horizontal growth prevents them from invading cortical zones in which they are not normally found. In the cat SI cortex, where, in the commissurally connected parts of the representation, thalamocortical and commissural fiber bundles are not dissociated from one another but overlap considerably (Wise and Jones, unpublished), neonatal removal of the commissural system does not result in any alteration in the

RAT THALAMOCORTICAL DEVELOPMENT

laminar distribution of thalamocortical fibers and terminals (Wise, ’76). In this case radial sprouting of thalamic fibers clearly does not occur. This, together with the work on the rat, would seem to imply that the thalamocortical and commissural fiber systems each form connections with specific classes of cells (or specific parts of cells) in a particular layer of the cortex and that they are not able to form aberrant connections on inappropriate cells or inappropriate parts of cells. LITERATURE CITED Abeles, M., and M. H. Goldstein 1970 Functional architecture in cat primary auditory cortex. Columnar organization and organization according to depth. J. Neurophysiol., 33: 172-187. Armstrong-James, M. 1975 The functional status and columnar organization of single cells responding to cutaneous stimulation in neonatal rat somatosensory cortex SI. J. Physiol. (London), 246: 501-538. Axelrad, H., R. Verley, and E. Farkas 1976 Responses, evoked in mouse and r a t SI cortex by vihrissa stimulation. Neurosci. Letters, 3: 265-274. Benevento, L. A., and M. Rezak with the assistance of J. Bos 1975 Extrageniculate projections to layers VI and I of striate cortex (area 17) in the rhesus monkey (Macaca mulatta). Brain Res., 96: 51-55. Bernstein, J. J., and D. C. Goodman (eds.) 1973 Neuromorphological plasticity. Brain Behav. and Evol., 8: 4-164. Berry, M., and A. W. Rogers 1965 The migration of neuroblasts in the developing cerebral cortex. J. Anat. (London), 99: 691-709. Caviness, V. S., Jr., D. 0. Frost and N. L. Hayes 1976 Barrels in somatosensory cortex of normal and reeler mutant mice. Neurosci Letter, 3: 7-14. Cowan, W. M., D. I. Gottlieb, A. E. Hendrickson, J. L. Price and T. A. Woolsey 1972 The autoradiographic demonstration of axonal connections in t he central nervous system. Brain Res., 37: 21-51. Crossland, W. J., W. M. Cowan, L. A. Rogers and J. P. Kelly 1974 The specification of the retino-tectal projection in the chick. J. Comp. Neur., 155: 127-164. Davidson, N. 1965 The projection of afferent pathways on the thalamus of the rat. J. Comp. Neur., 124: 377-390. Drager, U. C. 1974 Autoradiography of tritiated proline and fucose transported transneuronally from the eye to the visual cortex in pigmented and albino mice. Brain Res., 82: 284-292. Durham, D., and T. A. Woolsey 1977 Barrels and columnar cortical organization: evidence from 2-deoxy-glucose (2DG) experiments. Anat. Rec., 187: 570. Emmers, R. 1965 Organization of the first and second somesthetic regions (91 and SII) in the rat thalamus. J. Comp. Neur., 124: 215-228. Goldman, P. S., and W. J. H. Nauta 1977 Columnar distribution of cortico-cortical fibers in the frontal association, limbic, and motor cortex of the developing rhesus monkey. Brain Res., 122: 393-413. Gottlieb, D. I., and W. M. Cowan 1972 Evidence for a temporal factor in the occupation of available synaptic sites during the development of the dentate gyrus. Brain Res., 41: 452-456. Herkenham, M. 1976 The nigro-thalamo-cortical connections mediated hy t h e nucleus ventralis medialis thalami: evidence for a wide cortical distribution in the rat. Anat. Rec., 184: 426.

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Hubel, D. H., and T. N. Wiesel 1962 Receptive fields, binocular interaction and functional architecture in the cat’s visual cortex. J. Physiol., 160: 106-154. 1972 Laminar and columnar distribution of geniculo-cortical fibers in the macaque monkey. J. Comp. Neur., 146: 421-450. 1974 Sequence regularity and geometry of orientation columns in the monkey striate cortex. J. Comp. Neur., 158: 267-295. 1974 Uniformity of monkey striate cortex: a parallel relationship between field size, scatter and magnification factor J. Comp. Neur., 158: 296-306. Hubel, D. H., T. N. Wiesel and S. LeVay 1977 Plasticity of ocular dominance columns in monkey striate cortex. Phil. Trans. roy. SOC.Lond. B., 278: 377-409. Jones, E. G. 1975a Lamination and differential distribution of thalamic afferents within the sensory-motor cortex of the squirrel monkey. J. Comp. Neur., 160: 167-204. 1975b Varieties and distribution of non-pyramidal cells in the somatic sensory cortex of the squirrel monkey. J. Comp. Neur., 160: 205-268. Jones, E. G., and H. Burton 1976 Areal differences in the laminar distribution of thalamic afferents in cortical fields of the insular, parietal, and temporal regions of primates. J. Comp. Neur., 168: 197-248, Jones, E. G., H. Burton and R. Porter 1975 Commissural and cortico-cortical “columns” in the somatic sensory cortex of primates. Science, 190: 572-574. Jones, E. G., and R. Y. Leavitt 1974 Retrograde axonal transport and the demonstration of non-specific projections to the cerebral cortex and striatum from thalamic intralaminar nuclei in the rat, cat and rhesus monkey. J. Comp. Neur., 154: 349-378. Jones, E. G., and T. P. S. Powell 1970 An electron microscopic study of the laminar pattern and mode of termination of afferent fibre pathways in the somatic sensory cortex of the cat. Phil. Trans. roy. SOC.Lond. B., 257: 45-62. Kerr, F. W. L. 1975 Neuroplasticity of primary afferents in the neo-natal cat and some results of early deafferentation of the trigeminal spinal nucleus. J. Comp. Neur., 163: 305-328. Killacky, H. P. 1973 Anatomical evidence for cortical subdivisions based on vertically discrete thalamic projections from the ventral posterior nucleus to cortical barrels in the rat. Brain Res., 51: 326-331. Killackey, H. P., G. Belford, R. Ryugo and D. K. Ryugo 1976 Anomalous organization of thalamocortical projections consequent to vibrissae removal in the newborn rat and mouse. Brain Res., 104: 309-315. Killackey, H. P., and S. Leshin 1975 The organization of specific thalamocortical projections to the posteromedial barrel suhfield of the ra t somatic sensory cortex. Brain Rea., 86: 469-472. LaVail, J. H., K. R. Winston and A. Tish 1973 A method based on retrograde intraaxonal transport of protein for identification of cell bodies of origin of axons terminating within the CNS. Brain Res., 58: 470-477. LeVay, S., and C. D. Gilbert 1976 Laminar patterns of the geniculocortical projection in the cat. Brain Res., 113: 1-19. LeVay, S., D. H. Hubel and T. N. Wiesel 1975 The pattern of ocular dominance columns in macaque visual cortex revealed by reduced silver stain. J. Comp. Neur., 159: 559-576. Leong, S. K., and R. D. Lund 1973 Anomalous bilateral corticofugal pathways in albino rats after neonatal lesions. Brain Res., 62: 218.221. Lorente de NO, R. 1949 Cerebral cortex: architectonics, intracortical connections. In: Physiology of the Nervous

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System. Third ed. J. F. Fulton, ed. Oxford Univ. Press, New York, pp. 274-301. Lund, R. D., and B. F. Miller 1975 Secondary effects of fetal eye damage in rats on intact central optic projections. Brain Res., 92: 279-289. Lund, R. D., and M. J. Mustari 1977 Development of the geniculocortical pathway in rats. J. Conip. Neur., 173: 289-305. Lynch, G . , and C. W. Cotman 1975 The hippocampus as a model for studying anatomical plasticity in the adult brain. In: The Hippocampus. Vol. 1. R. L. lsaacson and K. H. Pribram, eds. i a v e n Press, pp. 123-154. Lynch, G., S. Mosko, T. Parks and C. Cotman 1973 Relocation and hyperdevelopment of the dentate gyrus commissural system after entorhinal lesions in immature rats. Brain Res., 50: 174-178. Lynch, G., B. Stanfield and C. W. Cotman 1973 Developmental differences in post-lesion axonal growth in the hippocampus. Brain Res., 59: 155-158. Mbres, P., J. Mbres and E. Kozakova-Matlova 1975 Development of the interhemispheric response in rats. Life Sci., 5: 5-10. Mountcastle, V. B. 1957 Modality and topographic properties of single neurons of cat’s somatic sensory cortex. J. Neurophysiol., 20: 408-434. Peters, A., and M. Feldman 1976 The projection of the lateral geniculate nucleus to area 17 of the rat cerebral cortex. I. General description. J. Neurocytol., 5: 63-84. Powell, T. P. S., and V. B. Mountcastle 1959 Some aspects of the functional organization of t he cortex of t he postcentral gyrus of the monkey: A correlation of findings obtained in a single unit analysis with cytoarchitecture. Bull. Johns Hopkins Hosp., 105: 133-162. Rakic, P. 1976 Prenatal genesis of connections subserving ocular dominance in the rhesus monkey. Nature, 261: 467-471. 1977 Prenatal development of the visual system in rhesus monkey. Phil. Trans. roy. SOC.Lond. B., 278: 245-260. Ribak, C. E., and A. Peters 1975 An autoradiographic study of the projections from the lateral geniculate body of the rat. Brain Research, 92: 341-368. Rice, F. L., and H. Van der Loos 1977 Development of barrels and barrel field in the somatosensory cortex of the mouse. J. Comp. Neur., 171: 545-560. Robson, J. A., and W. C. Hall 1975 Connections of layer VI in striate cortex of the grey squirrel (Sciurus carolinensis). Brain Res., 93: 133-139. Rosenquist, A. C., S. B. Edwards and L. A. Palmer 1974 An autoradiographic study of t he projections of the dorsal lateral geniculate nucleus and the posterior nucleus in the cat. Brain Res., 80: 71-93. Ryugo, R., and H. P. Killackey 1975 Corticocortical connections of the barrel field of rat somatosensory cortex. Neurosci. Abstr., 1: 126. Saporta, S., and L. Kruger 1977 The organization of thalamocortical relay neurons in the rat ventrobasal complex studied by the retrograde transport of heorseradish peroxidase. J. Comp. Neur., 174: 187-208. Strick, P. L., and P. Sterling 1974 Synaptic termination of afferents from the ventrolateral nucleus of the thalamus in the cat motor cortex. A light and electron microscouic study. J. Comp. Neur., 153: 77.106.

Van der Loos, H. 1976 Barreloids in mouse somatosensory thalamus. Neurosci. Letters, 2: 1-6. Van der Loos, H., and T. A. Woolsey 1973 Somatosensory cortex: structural alterations following early injury to sense organs. Science, 179: 395-398. Verley, R., and H. Axelrad 1975 Postnatal ontogenesis of potentials elicited in the cerebral cortex by afferent stimulation. Neurosci. Letters, 1: 99-104. Welker, C. 1976 Receptive fields of barrels in the somatosensory neocortex of the rat. J. Comp. Neur., 166: 173-190. Welker, C., and T. A. Woolsey 1974 Structure of layer IV in the somatosensory neocortex of the rat: description and comparison with the mouse. J. Comp. Neur., 158: 437-454. Weller, W. L., and J. I. Johnson 1975 Barrels in cerebral cortex altered by receptor disruption in newborn but not five-day-old mice. (Cricetidae and Muridae). Brain Res., 83: 504-508. Westrum, L. E. 1975 Axonal patterns in olfactory cortex after olfactory bulb removal in newborn rats. Exp. Neurol., 47: 442-447. Wiesel, T. N., D. H. Hubel and D. M. K. Lam 1974 Autoradiographic demonstration of ocular dominance columns in the monkey striate cortex by means of transneuronal transport. Brain Res., 79: 273-279. Wiitanen, J. T. 1969 Selective silver impregnation of degenerating axon terminals in the central nervous system of the monkey (Macaca rnulatta). Brain Res., 14: 546-548. Wise, S. P. 1975 The laminar organization of certain afferent and efferent fiber systems in the ra t somatosensory cortex. Brain Res., 90: 139-142. 1976 The development and specificity of connections in the somatic sensory cortex of the rat. Anat. Rec., 184: 565-566. 1977 The thalamocortical system of the ra t somatic sensory cortex: organization, development and connectional specificity. Neurosci. Abstr., 3: 495. Wise, S. P., S. H. C. Hendry and E. G. Jones 1977 Prenatal development of sensory-motor cortical projections in cats. Brain Res., in press. Wise, S. P., and E. G. Jones 1976 The organization and postnatal development of the commissural projection of the somatic sensory cortex of the rat. J. Comp. Neur., 168: 313-343. 1977 Cells of origin and terminal distribution of descending projections of the ra t somatic sensory cortex. J. Comp. Neur., 175: 129-158. Woolsey, T. A,, and H. Van der Loos 1970 The structural organization of layer IV in the somatosensory region (SI) of the mouse cerebral cortex: the description of a cortical field composed of discrete cytoarchitectonic units. Brain Res., 17: 205-242. Woolsey, T. A., and J. R. Wann 1976 Quantitative changes in mouse cortical barrels consequent to vibrissal damage a t different postnatal ages. Anat. Rec., 184: 567-568. Yorke, C. H., and V. S. Caviness 1975 Interhemispheric neocortical connections of the corpus callosum in the normal mouse: a study based on anterograde and retrograde methods. J. Comp. Neur., 164: 233-246. Zimmer, J. 1976 Postlesion remodelling of fiber connections in the hippocampus and fascia dentata: I. Observations on immature rats. EXD. Brain Res.. Suuul.. I : 191-196. I

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