Organization of the Anterior Limb of the Internal Capsule in the Rat.

Dysfunction of the orbitofrontal (OFC) and anterior cingulate (ACC) cortices has been linked with several psychiatric disorders, including obsessive-compulsive disorder, major depressive disorder, posttraumatic stress disorder, and addiction. These conditions are also associated with abnormalities in the anterior limb of the internal capsule, the white matter (WM) bundle carrying ascending and descending fibers from the OFC and ACC. Furthermore, deep-brain stimulation (DBS) for psychiatric disorders targets these fibers. Experiments in rats provide essential information on the mechanisms of normal and abnormal brain anatomy, including WM composition and perturbations. However, whereas descending prefrontal cortex (PFC) fibers in primates form a well defined and topographic anterior limb of the internal capsule, the specific locations and organization of these fibers in rats is unknown. We address this gap by analyzing descending fibers from injections of an anterograde tracer in the rat ACC and OFC. Our results show that the descending PFC fibers in the rat form WM fascicles embedded within the striatum. These bundles are arranged topographically and contain projections, not only to the striatum, but also to the thalamus and brainstem. They can therefore be viewed as the rat homolog of the primate anterior limb of the internal capsule. Furthermore, mapping these projections allows us to identify the fibers likely to be affected by experimental manipulations of the striatum and the anterior limb of the internal capsule. These results are therefore essential for translating abnormalities of human WM and effects of DBS to rodent models.SIGNIFICANCE STATEMENT Psychiatric diseases are linked to abnormalities in specific white matter (WM) pathways, and the efficacy of deep-brain stimulation relies upon activation of WM. Experiments in rodents are necessary for studying the mechanisms of brain function. However, the translation of results between primates and rodents is hindered by the fact that the organization of descending WM in rodents is poorly understood. This is especially relevant for the prefrontal cortex, abnormal connectivity of which is central to psychiatric disorders. We address this gap by studying the organization of descending rodent prefrontal pathways. These fibers course through a subcortical structure, the striatum, and share important organization principles with primate WM. These results allow us to model primate WM effectively in the rodent.

Human and monkey ventral prefrontal fibers use the same organizational principles to reach their targets: tracing versus tractography.

This article is a comparative study of white matter projections from ventral prefrontal cortex (vPFC) between human and macaque brains. We test whether the organizational rules that vPFC connections follow in macaques are preserved in humans. These rules concern the trajectories of some of the white matter projections from vPFC and how the position of regions in the vPFC dictate the trajectories of their projections in the white matter. To address this question, we present a novel approach that combines direct tracer measurements of entire white matter trajectories in macaque monkeys with diffusion MRI tractography of both macaques and humans. The approach allows us to provide explicit validation of diffusion tractography and transfer tractography strategies across species to test the extent to which inferences from macaques can be applied to human neuroanatomy. Apart from one exception, we found a remarkable overlap between the two techniques in the macaque. Furthermore, the organizational principles followed by vPFC tracts in macaques are preserved in humans.

Thalamocortical tract between anterior thalamic nuclei and cingulate gyrus in the human brain: diffusion tensor tractography study.

Most portions of the Papez circuit have been identified by diffusion tensor tractography (DTT). However, no DTT study on the proportion of the Papez circuit between the anterior thalamic nuclei and cingulate gyrus has been reported. We attempted to reconstruct the thalamocortical tract between the anterior thalamic nuclei and cingulate gyrus using DTT. All the reconstructed thalamocortical tracts originated from the anterior thalamic nuclei, ascended through the genu of the internal capsule, the anterior limb of the internal capsule, and the white matter around the anterior horn of the lateral ventricle in the anterior and lateral direction, and then terminated at the anterior cingulate gyrus. In terms of FA, MD, and tract volume, no significant differences were observed between hemispheres (p > 0.05). We reconstructed the thalamocortical tract between the anterior thalamic nucleus and cingulate gyrus in the human brain using DTT. We believe that the methodology and results of this study will be helpful to researchers investigating the Papez circuit.

Role of the atypical cadherin Celsr3 during development of the internal capsule.

The development of axonal tracts requires interactions between growth cones and the environment. Major bundles, particularly in the internal capsule, are completely defective in mice with constitutive mutation of Celsr3. In order to understand better how Celsr3 controls axonal tract formation, we generated a conditional allele that allowed inactivation of Celsr3 in different sectors of the forebrain. Effects of Celsr3 inactivation specifically in the telencephalon, in the ventral forebrain, or in the cortex, demonstrate essential roles for the gene, both in the neurons that project their axons to subcerebral targets such as the spinal cord, as well as in cells that guide projecting axons through the ventral forebrain. These observations provide unequivocal in vivo evidence that heterotypic interactions between axons and guidepost cells govern axonal path formation in mammals, and that Celsr3 plays a key role in this process. In absence of cortico-subcortical connections, mice can survive up to P20, allowing studies of behavior and cortical maturation. Mutant mice with defective corticospinal tracts survive normally and provide a model to evaluate in vivo the role of this tract in motor function in rodents.

Brain fiber tract plasticity in experimental spinal cord injury: diffusion tensor imaging.

Diffusion tensor imaging (DTI) and immunohistochemistry were performed in spinal cord injured rats to understand the basis for activation of multiple regions in the brain observed in functional magnetic resonance imaging (fMRI) studies. The measured fractional anisotropy (FA), a scalar measure of diffusion anisotropy, along the region encompassing corticospinal tracts (CST) indicates significant differences between control and injured groups in the 3 to 4 mm area posterior to bregma that correspond to internal capsule and cerebral peduncle. Additionally, DTI-based tractography in injured animals showed increased number of fibers that extend towards the cortex terminating in the regions that were activated in fMRI. Both the internal capsule and cerebral peduncle demonstrated an increase in GFAP-immunoreactivity compared to control animals. GAP-43 expression also indicates plasticity in the internal capsule. These studies suggest that the previously observed multiple regions of activation in spinal cord injury are, at least in part, due to the formation of new fibers.

Early forebrain wiring: genetic dissection using conditional Celsr3 mutant mice.

Development of axonal tracts requires interactions between growth cones and the environment. Tracts such as the anterior commissure and internal capsule are defective in mice with null mutation of Celsr3. We generated a conditional Celsr3 allele, allowing regional inactivation. Inactivation in telencephalon, ventral forebrain, or cortex demonstrated essential roles for Celsr3 in neurons that project axons to the anterior commissure and subcerebral targets, as well as in cells that guide axons through the internal capsule. When Celsr3 was inactivated in cortex, subcerebral projections failed to grow, yet corticothalamic axons developed normally, indicating that besides guidepost cells, additional Celsr3-independent cues can assist their progression. These observations provide in vivo evidence that Celsr3-mediated interactions between axons and guidepost cells govern axonal tract formation in mammals.

Substantial migration of SVZ cells to the cortex results in the generation of new neurons in the excitotoxically damaged immature rat brain.

Mammalian SVZ progenitors continuously generate new neurons in the olfactory bulb. After injury, changes in SVZ cell number suggest injury-induced migration. Studies that trace the migration of SVZ precursors into neurodegenerating areas are lacking. Previously, we showed a decrease in BrdU+SVZ cells following excitotoxic damage to the immature rat cortex. Here, we demonstrate that NMDA-induced injury forces endogenous Cell Tracker Green (CTG) labeled VZ/SVZ precursors out of the SVZ into the neurodegenerating cortex. CTG+/Nestin+/Filamin A+ precursors are closely associated with vimentin+/GFAP+/GLAST+ filaments and express both chemokine receptor CXCR4 and Robo1. In the cortex, SVZ-derived progenitors show a progressive expression of developing, migrating and mature neurons and glial markers. CTG+/GFAP+ astrocytes greatly outnumber CTG+/MAP2+/NeuN+ neurons. SVZ-derived progenitors differentiate into both tbr1+ cortical glutamatergic neurons and calretinin+ interneurons. But, there is little integration of these neurons into the existing circuitry, as seen by Fluorogold retrograde tracing from the internal capsule.

Developmental differences in white matter architecture between boys and girls.

Previous studies have found developmental differences between males and females in brain structure. During childhood and adolescence, relative white matter volume increases faster in boys than in girls. Sex differences in the development of white matter microstructure were investigated in a cohort of normal children ages 5-18 in a cross-sectional diffusion tensor imaging (DTI) study. Greater fractional anisotropy (FA) in boys was shown in associative white matter regions (including the frontal lobes), while greater FA in girls was shown in the splenium of the corpus callosum. Greater mean diffusivity (MD) in boys was shown in the corticospinal tract and in frontal white matter in the right hemisphere; greater MD in girls was shown in occipito-parietal regions and the most superior aspect of the corticospinal tract in the right hemisphere. Significant sex-age interactions on FA and MD were also shown. Girls displayed a greater rate of fiber density increase with age when compared with boys in associative regions (reflected in MD values). However, girls displayed a trend toward increased organization with age (reflected in FA values) only in the right hemisphere, while boys displayed this trend only in the left hemisphere. These results indicate differing developmental trajectories in white matter for boys and girls and the importance of taking sex into account in developmental DTI studies. The results also may have implications for the study of the relationship of brain architecture with intelligence.

NADPH-diaphorase-positive cells in the thalamic nuclei and internal capsule in humans.

The nuclei of the dorsal thalamus and reticular nucleus in humans were found to contain separated NADPH-diaphorase (NADPH-d)-positive neurons. Staining of NADPH-d-positive neurons and all their processes, along with previous studies of neurons in the nuclei of the dorsal thalamus based on the Golgi method, allowed the type of these cells to be identified as sparsely branched. The main, densely branched, efferent neurons did not contain NADPH-d. NADPH-d-positive neurons included reticular cells and cells of one of the types of short-axon interneurons. The internal capsule contained large numbers of NADPH-d-positive reticular neurons. NADPH-d-positive neurons were found in contact with vessels. Thus, NADPH-d-positive cells of the dorsal thalamus, reticular nucleus, and internal capsule were evolutionarily more ancient and less structurally complex cells.

Morphological study of the perireticular nucleus in human fetal brains.

Abstract The perireticular nucleus consists of scattered neurons that are located in the internal capsule. The presence of perireticular neurons in the rat, ferret, cat and human has been described previously. Evidence suggests that the perireticular neurons in various species decrease in number with increasing gestation, but in humans this finding has not been supported by quantitative data. This study aimed to investigate (1) the morphology of the human fetal perireticular neurons, (2) the average number of perireticular neurons within the anterior and posterior crus of the internal capsule per unit area, and (3) the magnitude and the stage of neuronal loss in the human perireticular nucleus subsequent to maturation. Nissl-stained sections of the internal capsule of human fetal brains of 24, 26.5, 32, 35, 37 and 39 weeks of gestation showed a number of clearly distinguishable large perireticular and small microglia cells. A regular increase of both perireticular and microglial cells was observed up to 32 weeks of gestation, after which a dramatic reduction in the number of both perireticular and microglia cells was observed. The average number of perireticular and the microglia cells per unit area, located within the posterior crus, was more than in the anterior crus of the internal capsule. In the adult, no perireticular neurons were detected within the internal capsule. The results show that perireticular neurons are not restricted to the region lateral to the thalamus and medial to the globus pallidus (posterior crus) but are also present at the region lateral to the caudate nucleus and medial to the globus pallidus (anterior crus).

[NADPH-diaphorase positive cells of the human thalamic nuclei and internal capsule]. / NADPH-diaforazo-pozitivnye kletki iader talamusa i vnytrennei kapsuly.

Isolated NADPH-diaphorase (NADPH-d)-positive neurons were demonstrated in the nuclei of human dorsal thalamus and nucleus reticularis. Staining of NADPH-d-positive neurons with all their processes and preceding study of neurons of dorsal thalamus using Golgi method enabled the identification of their types and their determination as sparsely-branched cells. Main types of efferent densely-branched neurons had no demonstrable NADPH-d activity. NADPH-d-positive neurons were represented by reticular neurons and by one type of short-axon interneurons. Capsula interna contains numerous NADPH-d-positive reticular neurons. NADPH-d-positive cells forming contacts with blood vessels were found. Thus, NADPH-d-positive cells of dorsal thalamus, reticular nucleus and capsula interna appear to be evolutionally more ancient and structurally less complex.

The thalamic reticular and perireticular nuclei in developing rabbits: patterns of parvalbumin expression.

Due to its strategic position, the thalamic reticular nucleus (TRN) plays an important role within the thalamo-cortical circuits. The perireticular thalamic nucleus (PRN) is a smaller group of cells, which is associated with the TRN and lies among the fibres of the internal capsule (IC). Studies of nuclei in rodents and carnivores have been conducted employing a number of different tools. The use of calcium-binding proteins is one example. It needs to be noted that rabbits have been regarded as intermediate between rodents and carnivores in relation to local GABAergic circuits. In the present study, sections from rabbits at different ages (prenatal, postnatal and adult) were examined to determine the parvalbumin (PV) expression in the developing TRN and PRN. In the TRN, there is one wave of PV expression during development, from caudal parts of the nucleus towards the rostral pole. At E22 there is already an incipient PV expression. In the adult stage, the TRN is completely positive to PV. The present study clearly indicates the presence of the PRN in the developing rabbit. The first PV positive cells were visible at E24, meanwhile the immunoreactivity was at its maximum at early postnatal stages (P0-P8). Two different types of perireticular cells in the IC were identified and the changes concerning neuronal morphology and orientation were described. The comparison between these results and previous data obtained in rats, ferrets or cats suggest that rabbits could represent an intermediate stage in the evolution of thalamic circuits and could be considered as useful neurobiological model.

A novel role for p75NTR in subplate growth cone complexity and visual thalamocortical innervation.

In cortical development, subplate axons pioneer the pathway from neocortex to the internal capsule, leading to the proposal that they are required for subsequent area-specific innervation of cortex by thalamic axons. A role for p75 neutrophin receptor (NTR) in area-specific thalamic innervation of cortex is suggested by the observation that p75NTR expression is restricted to subplate neurons in a low-rostral to high-caudal gradient throughout the period of thalamocortical innervation. In vitro, neurotrophin 3 binding to p75NTR increases neurite length and filopodial formation of immunopurified subplate neurons, suggesting a role for p75NTR in subplate growth cone morphology and function in vivo. Consistent with this idea, subplate growth cones have markedly fewer filopodia in mice lacking p75NTR than in wild type mice. Despite this gross morphologic defect, many subplate axons in knock-out mice pioneer the projection to the internal capsule as they do in wild-type mice. However a few subplate axons in the knock-out mice make ectopic projections rostral in the intermediate zone and frontal cortex. Concomitant with the altered morphology of subplate growth cones, mice lacking p75NTR have diminished innervation of visual cortex from the lateral geniculate nucleus, with markedly reduced or absent connections in 48% of knock-out mice. Thalamic projections to auditory and somatosensory cortex are normal, consistent with the gradient of p75NTR expression. Our present results are unusual in that they argue that p75NTR functions in a novel way in subplate neurons, that is, in growth cone morphology and function rather than in axon extension or neuronal survival.

The N-methyl-D-aspartate antagonist CNS 1102 protects cerebral gray and white matter from ischemic injury following temporary focal ischemia in rats.

BACKGROUND AND PURPOSE: Cerebral white matter is as sensitive as gray matter to ischemic injury and is probably amenable to pharmacological intervention. In this study we investigated whether an N-methyl-D-aspartate (NMDA) antagonist, CNS 1102, protects not only cerebral gray matter but also white matter from ischemic injury. METHODS: Ten rats underwent 15 minutes of temporary focal ischemia and were blindly assigned to CNS 1102 intravenous bolus injection (1. 13 mg/kg) followed by intravenous infusion (0.33 mg/kg per hour) for 3.75 hours or to vehicle (n=5 per group) immediately after reperfusion. Seventy-two hours after ischemia, the animals were perfusion fixed for histology. The severity of neuronal necrosis in the cortex and striatum was semiquantitatively analyzed. The Luxol fast blue-periodic acid Schiff stain and Bielschowsky's silver stain were used to measure optical densities (ODs) of myelin and axons, respectively, in the internal capsule of both hemispheres, and the OD ratio was calculated to reflect the severity of white matter damage. RESULTS: Neuronal damage in both the cortex and the striatum was significantly better in the drug-treated group than in the placebo group (P<0.05). The OD ratio of both the axons (0.93+/-0.08 versus 0.61+/-0.18; P<0.01) and the myelin sheath (0.95+/-0.07 versus 0.67+/-0.19; P=0.01) was significantly higher in the CNS 1102 group than in the placebo group. The neurological score was significantly improved in the drug-treated group (P<0.05). CONCLUSIONS: The NMDA receptor antagonist CNS 1102 protects not only cerebral gray matter but also white matter from ischemic injury, most probably by preventing degeneration of white matter structures such as myelin and axons.

Netrin-1 promotes thalamic axon growth and is required for proper development of the thalamocortical projection.

The thalamocortical axon (TCA) projection originates in dorsal thalamus, conveys sensory input to the neocortex, and has a critical role in cortical development. We show that the secreted axon guidance molecule netrin-1 acts in vitro as an attractant and growth promoter for dorsal thalamic axons and is required for the proper development of the TCA projection in vivo. As TCAs approach the hypothalamus, they turn laterally into the ventral telencephalon and extend toward the cortex through a population of netrin-1-expressing cells. DCC and neogenin, receptors implicated in mediating the attractant effects of netrin-1, are expressed in dorsal thalamus, whereas unc5h2 and unc5h3, netrin-1 receptors implicated in repulsion, are not. In vitro, dorsal thalamic axons show biased growth toward a source of netrin-1, which can be abolished by netrin-1-blocking antibodies. Netrin-1 also enhances overall axon outgrowth from explants of dorsal thalamus. The biased growth of dorsal thalamic axons toward the internal capsule zone of ventral telencephalic explants is attenuated, but not significantly, by netrin-1-blocking antibodies, suggesting that it releases another attractant activity for TCAs in addition to netrin-1. Analyses of netrin-1 -/- mice reveal that the TCA projection through the ventral telencephalon is disorganized, their pathway is abnormally restricted, and fewer dorsal thalamic axons reach cortex. These findings demonstrate that netrin-1 promotes the growth of TCAs through the ventral telencephalon and cooperates with other guidance cues to control their pathfinding from dorsal thalamus to cortex.

Expression of MAP1a and MAP1b in the ganglionic eminence and the internal capsule of the human fetal brain.

The expression of microtubule-associated proteins 1a and 1b (MAP1a and 1b) were investigated in two transient structures, the ganglionic eminence (GE) being a prominent part of the telencephalic proliferative zone and the perireticular nucleus (PR) within the internal capsule (IC). Anti-MAP1a immunolabels PR neurons from 18 weeks of gestation (wg) onwards, whereas anti-MAP1b immunolabels long IC fibers between 18 and 22 wg. MAP1b is further present in thalamic fibers that seem to terminate at the medial margin of the GE, in a moderate number of cells of the GE and its medial extension, the gangliothalamic body (GTB). From 26 to 33 wg MAP1b is expressed in short fiber bundles of the IC, a few MAP1b-positive cells are seen in the GE. MAP1a has so far been described to appear in differentiated neurons and to be related to late developmental events. However, the transient PR being involved in axonal guidance as an intermediate target shows a precocious MAP1a-expression. The MAP1b-finding that thalamocortical fibers accumulate at the GE-margin indicates that this region represents an intermediate target for these fibers. The short MAP1b fiber bundles found in the IC are in accordance with cell culture experiments showing that MAP1b is concentrated in distal parts of outgrowing axons.