Agenesis of the putamen and globus pallidus caused by recessive mutations in the homeobox gene GSX2.

Basal ganglia are subcortical grey nuclei that play essential roles in controlling voluntary movements, cognition and emotion. While basal ganglia dysfunction is observed in many neurodegenerative or metabolic disorders, congenital malformations are rare. In particular, dysplastic basal ganglia are part of the malformative spectrum of tubulinopathies and X-linked lissencephaly with abnormal genitalia, but neurodevelopmental syndromes characterized by basal ganglia agenesis are not known to date. We ascertained two unrelated children (both female) presenting with spastic tetraparesis, severe generalized dystonia and intellectual impairment, sharing a unique brain malformation characterized by agenesis of putamina and globi pallidi, dysgenesis of the caudate nuclei, olfactory bulbs hypoplasia, and anomaly of the diencephalic-mesencephalic junction with abnormal corticospinal tract course. Whole-exome sequencing identified two novel homozygous variants, c.26C>A; p.(S9*) and c.752A>G; p.(Q251R) in the GSX2 gene, a member of the family of homeobox transcription factors, which are key regulators of embryonic development. GSX2 is highly expressed in neural progenitors of the lateral and median ganglionic eminences, two protrusions of the ventral telencephalon from which the basal ganglia and olfactory tubercles originate, where it promotes neurogenesis while negatively regulating oligodendrogenesis. The truncating variant resulted in complete loss of protein expression, while the missense variant affected a highly conserved residue of the homeobox domain, was consistently predicted as pathogenic by bioinformatic tools, resulted in reduced protein expression and caused impaired structural stability of the homeobox domain and weaker interaction with DNA according to molecular dynamic simulations. Moreover, the nuclear localization of the mutant protein in transfected cells was significantly reduced compared to the wild-type protein. Expression studies on both patients' fibroblasts demonstrated reduced expression of GSX2 itself, likely due to altered transcriptional self-regulation, as well as significant expression changes of related genes such as ASCL1 and PAX6. Whole transcriptome analysis revealed a global deregulation in genes implicated in apoptosis and immunity, two broad pathways known to be involved in brain development. This is the first report of the clinical phenotype and molecular basis associated to basal ganglia agenesis in humans.

Distribution of brain iron accrual in adolescence: Evidence from cross-sectional and longitudinal analysis.

To track iron accumulation and location in the brain across adolescence, we repurposed diffusion tensor imaging (DTI) and functional magnetic resonance imaging (fMRI) data acquired in 513 adolescents and validated iron estimates with quantitative susceptibility mapping (QSM) in 104 of these subjects. DTI and fMRI data were acquired longitudinally over 1 year in 245 male and 268 female, no-to-low alcohol-consuming adolescents (12-21 years at baseline) from the National Consortium on Alcohol and NeuroDevelopment in Adolescence (NCANDA) study. Brain region average signal values were calculated for susceptibility to nonheme iron deposition: pallidum, putamen, dentate nucleus, red nucleus, and substantia nigra. To estimate nonheme iron, the corpus callosum signal (robust to iron effects) was divided by regional signals to generate estimated R2 (edwR2 for DTI) and R2 * (eR2 * for fMRI). Longitudinal iron deposition was measured using the normalized signal change across time for each subject. Validation using baseline QSM, derived from susceptibility-weighted imaging, was performed on 46 male and 58 female participants. Normalized iron deposition estimates from DTI and fMRI correlated with age in most regions; both estimates indicated less iron in boys than girls. QSM results correlated highly with DTI and fMRI results (adjusted R2 = 0.643 for DTI, 0.578 for fMRI). Cross-sectional and longitudinal analyses indicated an initial rapid increase in iron, notably in the putamen and red nucleus, that slowed with age. DTI and fMRI data can be repurposed for identifying regional brain iron deposition in developing adolescents as validated with high correspondence with QSM.

Spatiotemporal variations of magnetic susceptibility in the deep gray matter nuclei from 1 month to 6 years: A quantitative susceptibility mapping study.

BACKGROUND: Quantitative susceptibility mapping (QSM) is emerging as a technique that quantifies the paramagnetic nonheme iron in brain tissue. Brain iron quantification during early development provides insights into the underlying mechanism of brain maturation. PURPOSE: To quantify the spatiotemporal variations of brain iron-related magnetic susceptibility in deep gray matter nuclei during early development by using QSM. STUDY TYPE: Retrospective. SUBJECTS: Eighty-seven infants and children aged 1 month to 6 years. FIELD STRENGTH/SEQUENCE: Enhanced T2 *-weighted angiography using a 3D gradient-echo sequence at 3.0T. ASSESSMENT: QSM was calculated by modified sophisticated harmonic artifact reduction for phase data and sparse linear equations and sparse least squares-based algorithm. Means of susceptibility in deep gray matter nuclei (caudate nucleus, putamen, globus pallidus, thalamus) relative to that in splenium of corpus callosum were measured. STATISTICAL TESTS: Relationships of mean susceptibility with age and referenced iron concentration were tested by Pearson correlation. Differences of mean susceptibility between the selected nuclei in each age group were compared by one-way analysis of variance (ANOVA) and Fisher's Linear Significant Difference (LSD) test. RESULTS: Positive correlations of susceptibility with both referenced iron concentration and age were found (P < 0.0001); particularly, globus pallidus showed the highest correlation with age (correlation coefficient, 0.882; slope, 1.203; P < 0.001) and greatest susceptibility (P < 0.05) among the selected nuclei. DATA CONCLUSION: QSM allows the feasible quantification of iron deposition in deep gray matter nuclei in infants and young children, which exhibited gradual accumulation at different speeds. The fastest and highest iron accumulation was observed in the globus pallidus with increasing age during early development. LEVEL OF EVIDENCE: 4 Technical Efficacy:Stage 2 J. Magn. Reson. Imaging 2018.

Validation of a brain MRI relaxometry protocol to measure effects of preterm birth at a flexible postnatal age.

BACKGROUND: Magnetic resonance imaging (MRI) is a useful tool to study brain growth and organization in preterm neonates for clinical and research purposes, but its practicality can be limited by time and medical constraints. The aim of this study was to determine if MRI relaxometry of the deep nuclei, as opposed to white matter, would reflect the influence of gestational age at birth on structures essential to motor development, regardless of postnatal age at the time of imaging. RESULTS: This was a prospective observational study of infants without brain injury on conventional neuroimaging who were cared for in the neonatal intensive care unit (NICU) at Vanderbilt. Infants were studied using MRI relaxometry within a 2-month window of postmenstrual term age. In 45 infants, white matter MRI T1 relaxation times were influenced by both gestational age and postnatal age at imaging time (R(2) = 0.19 for gestational age vs. R(2) = 0.34 adjusting for both gestational age and age at imaging; all p < 0.01). Similar results were obtained with T2 relaxation times. In contrast, globus pallidus T1 reflected gestational age but was minimally affected by postnatal age (R(2) = 0.50 vs. 0.57, p < 0.001). CONCLUSIONS: The results obtained using this imaging protocol are consistent with the slow maturation of the globus pallidus, essential to normal development of complex motor programs into adulthood. Globus pallidus MRI relaxometry measures the impact of gestational age at birth on brain development independent of postnatal age in preterm infants and should prove useful for predictive modeling in a flexible time-window around postmenstrual term age.

Brain growth rate abnormalities visualized in adolescents with autism.

Autism spectrum disorder is a heterogeneous disorder of brain development with wide ranging cognitive deficits. Typically diagnosed before age 3, autism spectrum disorder is behaviorally defined but patients are thought to have protracted alterations in brain maturation. With longitudinal magnetic resonance imaging (MRI), we mapped an anomalous developmental trajectory of the brains of autistic compared with those of typically developing children and adolescents. Using tensor-based morphometry, we created 3D maps visualizing regional tissue growth rates based on longitudinal brain MRI scans of 13 autistic and seven typically developing boys (mean age/interscan interval: autism 12.0 ± 2.3 years/2.9 ± 0.9 years; control 12.3 ± 2.4/2.8 ± 0.8). The typically developing boys demonstrated strong whole brain white matter growth during this period, but the autistic boys showed abnormally slowed white matter development (P = 0.03, corrected), especially in the parietal (P = 0.008), temporal (P = 0.03), and occipital lobes (P = 0.02). We also visualized abnormal overgrowth in autism in gray matter structures such as the putamen and anterior cingulate cortex. Our findings reveal aberrant growth rates in brain regions implicated in social impairment, communication deficits and repetitive behaviors in autism, suggesting that growth rate abnormalities persist into adolescence. Tensor-based morphometry revealed persisting growth rate anomalies long after diagnosis, which has implications for evaluation of therapeutic effects.

Estrogen receptor alpha, G-protein coupled estrogen receptor 1, and aromatase: Developmental, sex, and region-specific differences across the rat caudate-putamen, nucleus accumbens core and shell.

Sex steroid hormones such as 17ß-estradiol (estradiol) regulate neuronal function by binding to estrogen receptors (ERs), including ERα and GPER1, and through differential production via the enzyme aromatase. ERs and aromatase are expressed across the nervous system, including in the striatal brain regions. These regions, comprising the nucleus accumbens core, shell, and caudate-putamen, are instrumental for a wide-range of functions and disorders that show sex differences in phenotype and/or incidence. Sex-specific estrogen action is an integral component for generating these sex differences. A distinctive feature of the striatal regions is that in adulthood neurons exclusively express membrane but not nuclear ERs. This long-standing finding dominates models of estrogen action in striatal regions. However, the developmental etiology of ER and aromatase cellular expression in female and male striatum is unknown. This omission in knowledge is important to address, as developmental stage influences cellular estrogenic mechanisms. Thus, ERα, GPER1, and aromatase cellular immunoreactivity was assessed in perinatal, prepubertal, and adult female and male rats. We tested the hypothesis that ERα, GPER1, and aromatase exhibits sex, region, and age-specific differences, including nuclear expression. ERα exhibits nuclear expression in all three striatal regions before adulthood and disappears in a region- and sex-specific time-course. Cellular GPER1 expression decreases during development in a region- but not sex-specific time-course, resulting in extranuclear expression by adulthood. Somatic aromatase expression presents at prepuberty and increases by adulthood in a region- but not sex-specific time-course. These data indicate that developmental period exerts critical sex-specific influences on striatal cellular estrogenic mechanisms.

Developmental regulation of nerve growth factor and its receptor in the rat caudate-putamen.

In prior studies, nerve growth factor (NGF) administration induced a robust, selective increase in the neurochemical differentiation of caudate-putamen cholinergic neurons. In this study, expression of NGF and its receptor was examined to determine whether endogenous NGF might serve as a neurotrophic factor for these neurons. The temporal pattern of NGF gene expression and the levels of NGF mRNA and protein were distinct from those found in other brain regions. NGF and high-affinity NGF binding were present during cholinergic neurochemical differentiation and persisted into adult-hood. An increase in NGF binding during the third postnatal week was correlated with increasing choline acetyltransferase activity. The data are consistent with a role for endogenous NGF in the development and, possibly, the maintenance of caudate-putamen cholinergic neurons.

Early setting of grammatical processing in the bilingual brain.

The existence of a "critical period" for language acquisition is controversial. Bilingual subjects with variable age of acquisition (AOA) and proficiency level (PL) constitute a suitable model to study this issue. We used functional magnetic resonance imaging to investigate the effects of AOA and PL on neural correlates of grammatical and semantic judgments in Italian-German bilinguals who learned the second language at different ages and had different proficiency levels. While the pattern of brain activity for semantic judgment was largely dependent on PL, AOA mainly affected the cortical representation of grammatical processes. These findings support the view that both AOA and PL affect the neural substrates of second language processing, with a differential effect on grammar and semantics.

Gray matter differences between pediatric obsessive-compulsive disorder patients and high-risk siblings: a preliminary voxel-based morphometry study.

Neuroimaging studies have identified alterations in frontostriatal circuitry in obsessive-compulsive disorder (OCD). Voxel-based morphometry (VBM) allows for the assessment of differences in gray matter density across the whole brain. VBM has not previously been used to examine regional gray matter density in pediatric OCD patients and the siblings of pediatric OCD patients. Volumetric magnetic resonance imaging (MRI) studies were conducted in 10 psychotropic naïve pediatric patients with OCD, 10 unaffected siblings of pediatric patients with OCD, and 10 healthy controls. VBM analysis was conducted using SPM2. Statistical comparisons were performed with the general linear model, implementing small volume random field corrections for a priori regions of interest (anterior cingulate cortex or ACC, striatum and thalamus). VBM analysis revealed significantly lower gray matter density in OCD patients compared to healthy in the left ACC and bilateral medial superior frontal gyrus (SFG). Furthermore, a small volume correction was used to identify a significantly greater gray matter density in the right putamen in OCD patients as compared to unaffected siblings of OCD patients. These findings in patients, siblings, and healthy controls, although preliminary, suggest the presence of gray matter structural differences between affected subjects and healthy controls as well as between affected subjects and individuals at risk for OCD.

Selection and Validation of Reference Genes for RT-PCR Expression Analysis of Candidate Genes Involved in Morphine-Induced Conditioned Place Preference Mice.

The expression of reference genes should be constitutively stable under the experimental conditions, so determining stable reference genes is critical for obtaining reliable results in gene expression studies. Morphine addiction persistently influences neurotransmitters and signal transduction systems, which may negatively alter behavioral responses at the cellular levels and interfere the expression of reference genes. In order to research morphine dependence, animal models are commonly used in physiology, pathology, and therapeutics field since human trials have many limitations. Therefore, it is necessary to select stable reference genes in standardized animal model. The objective of this study is to find out a set of optimal reference genes to standardize the gene expression of morphine-induced conditioned place preference (CPP) mice. During the process, eight reference genes were chosen. Then, the stability of their expression in two different brain tissues (Caudate Putamen and Hippocampus) was tested in two developmental stages (puberty and adult) under two treatments (physiological saline as control and morphine). Based on two algorithm-based methods (geNorm and NormFinder), which can rank and assess the stability of expression of eight reference genes, thereby quantifying the transcriptional levels of these genes by high sensitive, specific, and accurate real-time quantitative reverse transcription PCR (RT-qPCR) assays.

Ventricular shape and relative position abnormalities in preterm neonates.

Recent neuroimaging findings have highlighted the impact of premature birth on subcortical development and morphological changes in the deep grey nuclei and ventricular system. To help characterize subcortical microstructural changes in preterm neonates, we recently implemented a multivariate tensor-based method (mTBM). This method allows to precisely measure local surface deformation of brain structures in infants. Here, we investigated ventricular abnormalities and their spatial relationships with surrounding subcortical structures in preterm neonates. We performed regional group comparisons on the surface morphometry and relative position of the lateral ventricles between 19 full-term and 17 preterm born neonates at term-equivalent age. Furthermore, a relative pose analysis was used to detect individual differences in translation, rotation, and scale of a given brain structure with respect to an average. Our mTBM results revealed broad areas of alterations on the frontal horn and body of the left ventricle, and narrower areas of differences on the temporal horn of the right ventricle. A significant shift in the rotation of the left ventricle was also found in preterm neonates. Furthermore, we located significant correlations between morphology and pose parameters of the lateral ventricles and that of the putamen and thalamus. These results show that regional abnormalities on the surface and pose of the ventricles are also associated with alterations on the putamen and thalamus. The complementarity of the information provided by the surface and pose analysis may help to identify abnormal white and grey matter growth, hinting toward a pattern of neural and cellular dysmaturation.

Thalamic alterations in preterm neonates and their relation to ventral striatum disturbances revealed by a combined shape and pose analysis.

Finding the neuroanatomical correlates of prematurity is vital to understanding which structures are affected, and to designing efficient prevention and treatment strategies. Converging results reveal that thalamic abnormalities are important indicators of prematurity. However, little is known about the localization of the abnormalities within the subnuclei of the thalamus, or on the association of altered thalamic development with other deep gray matter disturbances. Here, we aim to investigate the effect of prematurity on the thalamus and the putamen in the neonatal brain, and further investigate the associated abnormalities between these two structures. Using brain structural magnetic resonance imaging, we perform a novel combined shape and pose analysis of the thalamus and putamen between 17 preterm (41.12 ± 5.08 weeks) and 19 term-born (45.51 ± 5.40 weeks) neonates at term equivalent age. We also perform a set of correlation analyses between the thalamus and the putamen, based on the surface and pose results. We locate significant alterations on specific surface regions such as the anterior and ventral anterior (VA) thalamic nuclei, and significant relative pose changes of the left thalamus and the right putamen. In addition, we detect significant association between the thalamus and the putamen for both surface and pose parameters. The regions that are significantly associated include the VA, and the anterior and inferior putamen. We detect statistically significant surface deformations and pose changes on the thalamus and putamen, and for the first time, demonstrate the feasibility of using relative pose parameters as indicators for prematurity in neonates. Our methods show that regional abnormalities of the thalamus are associated with alterations of the putamen, possibly due to disturbed development of shared pre-frontal connectivity. More specifically, the significantly correlated regions in these two structures point to frontal-subcortical pathways including the dorsolateral prefrontal-subcortical circuit, the lateral orbitofrontal-subcortical circuit, the motor circuit, and the oculomotor circuit. These findings reveal new insight into potential subcortical structural covariates for poor neurodevelopmental outcomes in the preterm population.

Catecholamines inhibit lipid peroxidation in young, aged, and Alzheimer's disease brain.

Some catecholamines and indolamines inhibit lipid peroxidation. Recent studies indicate that catecholaminergic inhibition of lipid peroxidation may be receptor mediated in vivo and in cell cultures. Because oxidative stress is one of the hypothesized pathogenic mechanisms for neurodegenerative diseases, including Alzheimer's disease (AD), we hypothesized that catecholaminergic and indolaminergic inhibition of lipid peroxidation would be altered in AD as compared to age-matched non-AD. To test this hypothesis we studied the effect of a variety of neurotransmitters and their antagonists on ascorbate-stimulated lipid peroxidation in membrane fragment preparations derived from postmortem human brain. In this in vitro system, the inhibition of lipid peroxidation by dopamine and serotonin did not appear to be receptor mediated. Further, our findings indicate that there is no apparent effect of age or AD on the inhibition of lipid peroxidation by catecholaminergic and indolaminergic agents.

Glycoconjugate boundaries during early postnatal development of the neostriatal mosaic.

The dispositions of galactosyl-containing glycoconjugates were studied during postnatal development of the caudate putamen in mice. The binding of the lectin peanut agglutinin, which has an affinity for galactosyl B-1,3 N-acetylgalactosamine residues, was compared to acetylcholinesterase staining and tyrosine hydroxylase immunoreactivity in the immature and adult neostriatum. The binding of peanut agglutinin conjugated to horseradish peroxidase, in sections that were processed for peroxidase histochemistry, was extremely pronounced in the neostriatum through the first postnatal week and constituted ringlike or polygonally shaped structures, which, overall, produced a variegated mosaic. These structures consist of outer rims of dense lectin-associated reaction product surrounding lightly labeled centers. Lectin delineations of the neostriatal mosaic are no longer visible in the second postnatal week. When adjacent sections were processed for lectin binding or acetylcholinesterase histochemistry, the dense lectin binding sites represented borders of acetylcholinesterase-rich and -poor zones. The distribution of dense patches of tyrosine hydroxylase immunoreactive fibers and terminals also coincides with the acetylcholinesterase-rich zones during the same times, and thus the glycoconjugate-delineated boundaries can also be directly compared with the distribution of nigrostriatal dopaminergic projections. The findings presented here represent the first demonstration of a probe that recognizes apparent borders of neostriatal compartments during a limited period of development. They are consistent with previous observations made on transient glycoconjugate "hidden boundaries" during development of other central nervous system structures, including the somatosensory cortical barrel field, and thalamic and brainstem nuclei (Cooper and Steindler, '86a,b; Steindler and Cooper, in press). In those studies, glia were shown to be the major source of glycoconjugate-associated patterns, and thus, glia and glycoconjugates that they synthesize during pattern formation events may be involved in the formation and stabilization of neurochemically distinct components of the neostriatal mosaic.

Ultrastructural localization of delta-opioid receptors in the rat caudate-putamen nucleus during postnatal development: relation to synaptogenesis.

During development, delta-opioid receptors (DORs) in the rat caudate-putamen nucleus (CPN) appear later than mu-opioid receptors (MORs), whose developmental pattern specifically relates to synaptogenesis. We used electron microscopic immunocytochemistry to determine whether there are also age-related changes in subcellular localization of DORs in the rat CPN. Sections from postnatal day (P) 0-P30 and adult dorsomedial CPN were immunogold-silver labeled to examine the plasmalemmal and cytoplasmic distribution of these receptors. In addition, immunoperoxidase labeling was used to determine the numerical density of synapses relative to DOR-labeled profiles. Immunolabeling for DOR was undetectable at P0, light at P5, and dense from P10 onward. The labeling during P5-P10 was mainly localized in somatodendritic profiles but also was readily seen in axon terminals, most of which formed asymmetric synapses with dendrites. From P15, a few immunogold particles were seen in contact with postsynaptic densities in spines, and the proportion of these particles significantly increased in P30 and adult CPN. Other particles were localized in the cytoplasm of dendrites and terminals without significant age-related changes. Stereological analysis showed that compared with labeled dendritic shafts and spines, labeled axon terminals have a closer correlation with synapse formation. These results are in marked contrast with MORs, which show an age-related increase in association with dendritic plasma membrane and a good correlation in the developmental pattern of MOR-labeled spines with synapse formation (Wang et al. [2003] Neuroscience 118:695-708). Together, our results suggest receptor-type specific roles for endogenous opioids acting at both pre- and postsynaptic sides in the developing CPN.

Early postnatal development of the monkey neostriatum: a Golgi and ultrastructural study.

Paired specimens of the neostriatum were taken from monkeys at zero (newborn), one, two, four, eight, and 16 weeks of age, and prepared for Golgi impregnations and electron microscopy. Light microscopy shows that in the first postnatal week, the structure contains the five neuronal types and four categories of afferent axons described in the adult, as well as some cells too undifferentiated to classify. Most neurons exhibit immature dendritic features, including local enlargements, terminal growth cones with filopodia, and filiform processes. In spiny type I cells, various levels of maturity may coexist in regions of a single dendrite, in different dendrites of the same neuron, and among individual cells. Spine density increases progressively with age, but the relative distribution of spine types remains about the same. Spiny type II neurons show some decline in spine density, and generally mature sooner than spiny type I cells. The long axons of spiny neurons have varicosities which disappear at about eight weeks. In younger animals (newborn and one week), the dendrites of aspiny neurons (types I, II, and III) may have a "spiny" appearance, exhibiting many spine-like and filiform processes. Concurrently, the short axons vary in degree of arborization from very immature to well developed. Electron microscopy corroborates the developmental features recognized in the Golgi material: dendritic and axonal growth cones, filopodia and varicosities, as well as various stages of maturation in somata and dendrites. Degenerating elements, mostly of an axonal nature, are seen up to eight weeks. The synapses which reach maturity at birth are of the asymmetric axospinous type, in which the axonal profile contains small round vesicles, and of the symmetric axodendritic class, with the presynaptic elements having pleomorphic vesicles. Some synapses are slower to mature and appear at one to eight postnatal weeks. These include those made by profiles with pleomorphic vesicles, forming either symmetric contacts with somata and axon initial segments, or asymmetric contacts with spines. The same applies to the asymmetric axodendritic synapses made by elements containing small round vesicles. Finally, profiles containing large round or flat vesicles are the latest to participate in mature synapses formation. Findings indicate that a considerable degree of qualitative and quantitative change takes place in the monkey neostriatal neuropil during early postnatal development, especially in the first eight-week period.

Species-specific patterns of glycoprotein expression in the developing rodent caudoputamen: association of 5'-nucleotidase activity with dopamine islands and striosomes in rat, but with extrastriosomal matrix in mouse.

The glycoprotein 5'-nucleotidase is a cell surface phosphatase and represents a new marker for striosomes in the adult rat caudoputamen. We report here on its developmental expression in the rat and mouse striatum, and show an unexpected converse 5'-nucleotidase chemoarchitecture of the caudoputamen in these closely related species. In the rat, 5'-nucleotidase activity was first visible as neuropil staining in tyrosine hydroxylase-positive dopamine islands of the midstriatum on postnatal day 1, and by the end of the first postnatal week, 5'-nucleotidase-positive dopamine islands also appeared rostrally. This compartmental pattern persisted thereafter, so that in adult animals, in all but the caudal caudoputamen, zones of enhanced 5'-nucleotidase staining were restricted to calbindin-D28k-poor striosomes. Weak 5'-nucleotidase activity also emerged in the matrix. In striking contrast, in the mouse striatum, enhanced 5'-nucleotidase activity was preferentially associated with extrastriosomal tissue. Enzymatic reaction first appeared on embryonic day 18, and developed over the first postnatal week into a mosaic pattern in which the matrix was stained but the dopamine islands were unstained. The matrix staining itself was heterogeneous. After the second postnatal week, most of the caudoputamen was stained, and in adult mice only rostral striosomes expressed low 5'-nucleotidase activity. We conclude that in rats, 5'-nucleotidase represents one of the few substances that maintains a preferential dopamine island/striosome distribution during striatal development. In mice, 5'-nucleotidase activity is expressed preferentially in the matrix during development, and its compartmental pattern is gradually lost with maturation, except very rostrally. These findings do not suggest an instructive role of the enzyme in striatal compartment formation in either species, but do suggest the possibility that 5'-nucleotidase contributes to the differentiation of striatal compartments during development.

Heterogeneous development of calbindin-D28K expression in the striatal matrix.

In the present study, we attempted to trace the development of the striatal matrix by analyzing the ontogenetic expression of calbindin-D28K (calbindin), a calcium binding protein selectivity expressed in medium-sized neurons of the matrix compartment of the mature rat's caudoputamen. The localization of calbindin was documented in a series of developing rat brains, as was the compartmental location of these cells relative to tyrosine hydroxylase (TH)-immunostained dopamine islands, sites of future striosomes. Medium-sized striatal neurons appeared in the striatum at embryonic day (E) 20, and from their first appearance, the calbindin-positive neurons had highly heterogeneous distributions. They first formed a latticework of patches and bands in a ventral region of the caudoputamen. By postnatal day (P) 7, this early calbindin-positive lattice had evolved into a mosaic in which circumscript pockets of low calbindin-like immunoreactivity appeared in more extensive calbindin-rich surrounds. With further development, the mosaic gradually encroached on all but the dorsolateral caudoputamen, a district that is calbindin-poor at adulthood. A special lateral branch of the striatal calbindin system was also identified, distinct from the rest of the calbindin-positive mosaic in several developmental characteristics. In the parts of the caudoputamen where the developing calbindin system and dopamine island system were both present, the dopamine islands invariably lay in calbindin-poor zones. Most dopamine islands, however, only filled parts of the corresponding calbindin-poor zones. Moreover, there were some calbindin-poor zones for which TH-positive dopamine islands could not be detected. Thus during development, calbindin was expressed in the extrastriosomal matrix of the striatum, but the matrix could be divided into calbindin-rich and calbindin-poor zones. In the calbindin-rich regions, there were patches of especially intense calbindin expression and zones of weaker expression. These results suggest that there is neurochemical heterogeneity in the striatal matrix during the prolonged developmental period in which the early calbindin-positive lattice expands to form the calbindin-positive matrix of the mature striatum. Surprisingly, calbindin expression in the matrix, although eventually distributed in strictly complementary fashion to striosomes, does not originate as a system complementary to dopamine islands. The prolonged disparity between the borders of dopamine islands and calbindin-poor zones, and the different spatiotemporal schedules of development of the islands and the calbindin gaps suggest instead that the final match between the borders of striosomes and surrounding matrix results from dynamic processes occurring early in postnatal development. Candidate mechanisms for the gradual adjustment of these borders are proposed.