Regional and cellular localization of the CXCl12/SDF-1 chemokine receptor CXCR7 in the developing and adult rat brain

The chemokine stromal cell-derived factor-1 (SDF-1) regulates neuronal development via the chemokine receptor CXCR4. In the adult brain the SDF-1/CXCR4 system was implicated in neurogenesis, neuromodulation, brain inflammation, tumor growth, and HIV encephalopathy. Until the recent identification of RDC1/CXCR7 as the second SDF-1 receptor, CXCR4 was considered to be the only receptor for SDF-1. Here we provide the first map of CXCR7 mRNA expression in the embryonic and adult rat brain. At embryonic stages, CXCR7 and CXCR4 were codistributed in the germinative zone of the ganglionic eminences, caudate putamen, and along the routes of GABAergic precursors migrating toward the cortex. In the cortex, CXCR7 was identified in GABAergic precursors and in some reelin-expressing Cajal-Retzius cells. Unlike CXCR4, CXCR7 was abundant in neurons forming the cortical plate and sparse in the developing dentate gyrus and cerebellar external germinal layer. In the adult brain, CXCR7 was expressed by blood vessels, pyramidal cells in CA3, and mature dentate gyrus granule cells, which is reminiscent of the SDF-1 pattern. CXCR7 and CXCR4 overlapped in the wall of the four ventricles. Further neuronal structures expressing CXCR7 comprised the olfactory bulb, accumbens shell, supraoptic and ventromedial hypothalamic nuclei, medial thalamus, and brain stem motor nuclei. Also, GLAST-expressing astrocytes showed signals for CXCR7. Thus, CXCR4 and CXCR7 may cooperate or act independently in SDF-1-dependent neuronal development. In mature neurons and blood vessels CXCR7 appears to be the preponderant SDF-1-receptor.     

Evidence for tangential migration disturbances in human lissencephaly resulting from a defect in LIS1, DCX and ARX genes

During corticogenesis, neurons adopt different migration pathways to reach their final position. The precursors of pyramidal neurons migrate radially, whereas most of the GABA-containing interneurons are generated in the ventral telencephalon and migrate tangentially into the neocortex. Then, they use a radial migration mode to establish themselves in an inside-out manner in the neocortex, similarly to pyramidal neurons. In humans, the most severe defects in radial migration result in lissencephaly. Lately, a few studies suggested that lissencephaly was also associated with tangential neuronal migration deficits. In the present report, we investigated potential anomalies of this migration mode in three agyric/pachygyric syndromes due to defects in the LIS1, DCX and ARX genes. Immunohistochemistry was performed on paraffin-embedded supra- and infratentorial structures using calretinin, calbindin and parvalbumin antisera. The results were compared with age-matched control brain tissue. In the Miller–Dieker syndrome, GABAergic neurons were found both in upper layers of the cortex and in heterotopic positions in the intermediate zone and in ganglionic eminences. In the DCX mutant brain, few interneurons were dispersed in the cortical plate, with a massive accumulation in the intermediate zone and subventricular zone as well as in the ganglionic eminences. In the ARX-mutated brain, the cortical plate contained almost exclusively pyramidal cells and was devoid of interneurons. The ganglionic eminences and basal ganglia were poorly cellular, suggesting an interneuron production and/or differentiation defect. These data argue for different mechanisms of telencephalic tangential migration impairment in these three agyric/pachygyric syndromes.     

Stromal cell-derived factor 1alpha mediates neural progenitor cell motility after focal cerebral ischemia.

In the adult rodent, stroke induces an increase in endogenous neural progenitor cell (NPC) proliferation in the subventricular zone (SVZ) and neuroblasts migrate towards the ischemic boundary. We investigated the role of stromal cell-derived factor 1alpha (SDF-1alpha) in mediating NPC migration after stroke. We found that cultured NPCs harvested from the normal adult SVZ, when they were overlaid onto stroke brain slices, exhibited significantly (P<0.01) increased migration (67.2+/-25.2 microm) compared with the migration on normal brain slices (29.5+/-29.5 microm). Immunohistochemistry showed that CXCR 4, a receptor of SDF-1alpha, is expressed in the NPCs and migrating neuroblasts in stroke brain. Blocking SDF-1alpha by a neutralizing antibody against CXCR 4 significantly attenuated stroke-enhanced NPC migration. ELISA analysis revealed that SDF-1alpha levels significantly increased (P<0.01) in the stroke hemisphere (43.6+/-6.5 pg/mg) when compared with the normal brain (25.2+/-1.9 pg/mg). Blind-well chamber assays showed that SDF-1alpha enhanced NPC migration in a dose-dependent manner with maximum migration at a dose of 500 ng/mL. In addition, SDF-1alpha induced directionally selective migration. These findings show that SDF-1alpha generated in the stroke hemisphere may guide NPC migration towards the ischemic boundary via binding to its receptor CXCR 4 in the NPC. Thus, our data indicate that SDF-1alpha/CXCR 4 is important for mediating specific migration of NPCs to the site of ischemic damaged neurons.     

Modulation of network-driven, GABA-mediated giant depolarizing potentials by SDF-1alpha in the developing hippocampus

Chemokine stromal cell-derived factor-1 (SDF-1, or CXCL12) plays an important role in brain development and functioning. Whole-cell patch clamp recordings were conducted on CA3 neurons in hippocampal slices prepared from neonatal rats between postnatal days 2 and 6 to study the modulatory effects of SDF-1alpha on network-driven, gamma-aminobutyric-acid-mediated giant depolarizing potentials (GDPs), a hallmark of the developing hippocampus. We found that SDF-1alpha, the only natural ligand for chemokine CXC motif receptor 4 (CXCR4), decreased GDP firing without significant effects on neuronal passive membrane properties in neonatal hippocampal neurons. The SDF-1alpha-mediated decrease in GDP firing was blocked by T140, a CXCR4 receptor antagonist, suggesting that SDF-1alpha modulates GDP firing via CXCR4. We also showed that endogenous SDF-1 exerts a tonic inhibitory action on GDPs in the developing hippocampus. As SDF-1/CXCR4 are highly expressed in the developing brain and GDPs are involved in activity-dependent synapse formation and functioning, the inhibitory action of SDF-1alpha on GDPs may reflect a potential mechanism for chemokine regulation of neural development in early neonatal life.     

The SDF-1/CXCR4 pathway and the development of the cerebellar system

Mice deficient for the chemokine receptor CXCR4 show premature translocation of granule cell neuroblasts from their germinal zone into the nascent cerebellum [Y.-R. Zuo et al. (1998) Nature, 393, 595-599]. Here, we used CXCR4-null mice to analyse the early development of cerebellar cortical inhibitory interneurons and pontine neurons which, in the adult, are synaptically integrated with granule cells. Cortical inhibitory interneuronal precursors normally invade the cerebellar anlage of CXCR4-deficient mice, but their dispersal is impeded by dislocated foci of proliferating granule cells, from which they are excluded. This is reminiscent of the strict exclusion of inhibitory interneuronal precursors from the superficial external granule cell layer. As inhibitory interneuronal precursors readily mingle with post-mitotic granule cells both in wild-type and CXCR4-null mice, these findings indicate that the developmentally regulated interactions between granule and inhibitory interneuronal precursors are independent of SDF-1/CXCR4 signalling. In contrast, the transit of pontine neurons from the rhombic lip through the anterior extramural stream to the basilar pons is disrupted in CXCR4-deficient animals. Migrating pontine neurons express CXCR4, and in CXCR4-null animals these cells are found displaced deep into the brainstem. Consequently, nascent pontine nuclei in CXCR4-deficient animals are hypoplastic. Moreover, they fail to express plexin D1, suggesting that SDF-1/CXCR4 signalling may also impinge on axon guidance critical to the orderly formation of granule cell mossy fibre afferents.     

Complex effects of stromal cell-derived factor-1 alpha on melanin-concentrating hormone neuron excitability

Stromal cell-derived factor 1alpha (SDF-1alpha), a chemoattractant for leucocytes and neurons, and its receptor, CXCR4 are expressed in subsets of neurons of specific brain areas. In rat lateral hypothalamic area (LHA) we show, using immunocytochemistry, that CXCR4 is localized within melanin-concentrating hormone (MCH)-expressing neurons, mainly involved in feeding behaviour regulation. We investigated whether SDF-1alpha may control MCH neuronal activity. Patch-clamp recordings in rat LHA slices revealed multiple effects of SDF-1alpha on the membrane potential of MCH neurons, indirect through glutamate/GABA release and direct through GIRK current activation. Moreover, SDF-1alpha at 0.1-1 nM decreased peak and discharge frequency of action potential evoked by current pulses. These effects were further confirmed in voltage-clamp experiments, SDF-1alpha depressing both potassium and sodium currents. At 10 nM, however, SDF-1alpha increased peak and discharge frequency of action potential evoked by current pulses. Using a specific CXCR4 antagonist, we demonstrated that only the depressing effect on AP discharge was mediated through CXCR4 while the opposite effect was indirect. Together, our studies reveal for the first time a direct effect of SDF-1alpha on voltage-dependent membrane currents of neurons in brain slices and suggest that this chemokine may regulate MCH neuron activity.     

Enhanced expression of the CXCl12/SDF-1 chemokine receptor CXCR7 after cerebral ischemia in the rat brain

Expression patterns of the second SDF-1 receptor RDC1/CXCR7 were examined after focal ischemia in rats using in situ hybridization. CXCR7 mRNA was identified in the ventricle walls as well as neuronal, astroglial, and vascular cells. After ischemia, intact cortical regions showed a rapid, 4 days-lasting increase in neuronal CXCR7 expression. In the ischemic tissue CXCR7 expression was scarce and associated with blood vessels. Between days 2 and 10 after ischemia-onset, SDF-1 expression increased strongly in the peri-infarct and infarct region, which was accompanied by the appearance of numerous CXCR4-expressing but not CXCR7-expressing cells. These patterns suggest that SDF-1 may influence vascular, astroglial, and neuronal functions via CXCR7 and mediate cell recruitment to ischemic brain areas via CXCR4.     

趋化因子SDF-1体外趋化骨髓基质细胞迁移的实验研究

目的探讨趋化因子SDF-1在体外对骨髓基质细胞的迁移作用。方法采用全骨髓法培养成年Wistar大鼠骨髓基质细胞。取第五代骨髓基质细胞行免疫荧光鉴定；后通过细胞免疫荧光及RT-PCR的方法检测骨髓基质细胞表达趋化因子受体CXCR4的情况．利用Boyden小室法探讨趋化因子SDF-1对骨髓基质细胞的体外趋化作用及其特异性。结果第五代骨髓基质细胞都表达间充质干细胞标记物Vimentin、Laminin及Fibronectin；细胞免疫荧光及RT-PCR结果证实骨髓基质细胞表达趋化因子受体CXCR4；趋化因子SDF-1（5、50、500ng／mL）体外可趋化骨髓基质细胞迁移，抗SDF-1多克隆抗体可对抗其趋化迁移作用。结论SDF-1／CXCR4通路参与骨髓基质细胞体外迁移，为进一步研究骨髓基质细胞的迁移机制提供了理论依据。     

Role of the alpha-chemokine stromal cell-derived factor (SDF-1) in the developing and mature central nervous system

alpha-chemokines, which control the activation and directed migration of leukocytes, participate in the inflammatory processes in host defense response. One of the alpha-chemokines, CXCL12 or stromal cell-derived factor 1 (SDF-1), not only regulates cell growth and migration of hematopoietic stem cells but may also play a central role in brain development as we discuss here. SDF-1 indeed activates the CXCR4 receptor expressed in a variety of neural cells, and this signaling results in diverse biological effects. It enhances migration and proliferation of cerebellar granule cells, chemoattracts microglia, and stimulates cytokine production and glutamate release by astrocytes. Moreover, it elicits postsynaptic currents in Purkinje cells, triggers migration of cortical neuron progenitors, and produces pain by directly exciting nociceptive neurons. By modulating cell signaling and survival during neuroinflammation, SDF-1 may also play a role in the pathogenesis of brain tumors, experimental allergic encephalitis, and the nervous system dysfunction associated with acquired immunodeficiency syndrome.     

Developmental characterization of Zswim5 expression in the progenitor domains and tangential migration pathways of cortical interneurons in the mouse forebrain

GABAergic interneurons play an essential role in modulating cortical networks. The progenitor domains of cortical interneurons are localized in developing ventral forebrain, including the medial ganglionic eminence (MGE), caudal ganglionic eminence (CGE), preoptic area (POA), and preoptic hypothalamic border domain (POH). Here, we characterized the expression pattern of Zswim5, an MGE-enriched gene in the mouse forebrain. At E11.5–E13.5, prominent Zswim5 expression was detected in the subventricular zone (SVZ) of MGE, POA, and POH, but not CGE of ventral telencephalon where progenitors of cortical interneurons resided. At E15.5 and E17.5, Zswim5 expression remained in the MGE/pallidum primordium and ventral germinal zone. Zswim5 mRNA was markedly decreased after birth and was absent in the adult forebrain. Interestingly, the Zswim5 expression pattern resembled the tangential migration pathways of cortical interneurons. Zswim5-positive cells in the MGE appeared to migrate from the MGE through the SVZ of LGE to overlying neocortex. Indeed, Zswim5 was co-localized with Nkx2.1 and Lhx6, markers of progenitors and migratory cortical interneurons. Double labeling showed that Ascl1/Mash1-positive cells co-expressed Zswim5. Zswim5 expressing cells contained none or at most low levels of Ki67 but co-expressed Tuj1 in the SVZ of MGE. These results suggest that Zswim5 is immediately upregulated as progenitors exiting cell cycle become postmitotic. Given that recent studies have elucidated that the cell fate of cortical interneurons is determined shortly after becoming postmitotic, the timing of Zswim5 expression in early postmitotic interneurons suggests a potential role of Zswim5 in regulation of neurogenesis and tangential migration of cortical interneurons.     

Chemokine receptor expression by neural progenitor cells in neurogenic regions of mouse brain

We previously demonstrated that chemokine receptors are expressed by neural progenitors grown as cultured neurospheres. To examine the significance of these findings for neural progenitor function in vivo, we investigated whether chemokine receptors were expressed by cells having the characteristics of neural progenitors in neurogenic regions of the postnatal brain. Using in situ hybridization we demonstrated the expression of CCR1, CCR2, CCR5, CXCR3, and CXCR4 chemokine receptors by cells in the dentate gyrus (DG), subventricular zone of the lateral ventricle, and olfactory bulb. The pattern of expression for all of these receptors was similar, including regions where neural progenitors normally reside. In addition, we attempted to colocalize chemokine receptors with markers for neural progenitors. In order to do this we used nestin-EGFP and TLX-LacZ transgenic mice, as well as labeling for Ki67, a marker for dividing cells. In all three areas of the brain we demonstrated colocalization of chemokine receptors with these three markers in populations of cells. Expression of chemokine receptors by neural progenitors was further confirmed using CXCR4-EGFP BAC transgenic mice. Expression of CXCR4 in the DG included cells that expressed nestin and GFAP as well as cells that appeared to be immature granule neurons expressing PSA-NCAM, calretinin, and Prox-1. CXCR4-expressing cells in the DG were found in close proximity to immature granule neurons that expressed the chemokine SDF-1/CXCL12. Cells expressing CXCR4 frequently coexpressed CCR2 receptors. These data support the hypothesis that chemokine receptors are important in regulating the migration of progenitor cells in postnatal brain.     

Stromal-derived factor 1 signalling regulates radial and tangential migration in the developing cerebral cortex

Stromal-derived factor 1 (SDF-1), a known chemoattractant, and its receptor CXCR4 are widely expressed in the developing and adult cerebral cortex. Recent studies have highlighted potential roles for SDF-1 during early cortical development. In view of the current findings, our histological analysis has revealed a distinct pattern of SDF-1 expression in the developing cerebral cortex at a time when cell proliferation and migration are at peak. To determine the role of chemokine signalling during early cortical development, embryonic rat brain slices were exposed to a medium containing secreted SDF-1 to perturb the endogenous levels of chemokine. Alternatively, brain slices were treated with 40 muM of T140 or AMD3100, known antagonists of CXCR4. Using these experimental approaches, we demonstrate that chemokine signalling is imperative for the maintenance of the early cortical plate. In addition, we provide evidence that both neurogenesis and radial migration are concomitantly regulated by this signalling system. Conversely, interneurons, although not dependent on SDF-1 signalling to transgress the telencephalic boundary, require the chemokine to maintain their tangential migration. Collectively, our results demonstrate that SDF-1 with its distinct pattern of expression is essential and uniquely positioned to regulate key developmental events that underlie the formation of the cerebral cortex.     

Stromal cell-derived factor 1-mediated CXCR4 signaling in rat and human cortical neural progenitor cells

Stromal cell-derived factor 1 (SDF-1) and the chemokine receptor CXCR4 are highly expressed in the nervous system. Knockout studies have suggested that both SDF-1 and CXCR4 play essential roles in cerebellar, hippocampal, and neocortical neural cell migration during embryogenesis. To extend these observations, CXCR4 signaling events in rat and human neural progenitor cells (NPCs) were examined. Our results show that CXCR4 is expressed in abundance on rat and human NPCs. Moreover, SDF-1alpha induced increased NPCs levels of inositol 1,4,5-triphosphate, extracellular signal-regulated kinases 1/2, Akt, c-Jun N-terminal kinase, and intracellular calcium whereas it diminished cyclic adenosine monophosphate. Finally, SDF-1alpha can induce human NPC chemotaxis in vitro, suggesting that CXCR4 plays a functional role in NPC migration. Both T140, a CXCR4 antagonist, and pertussis toxin (PTX), an inactivator of G protein-coupled receptors, abrogated these events. Ultimately, this study suggested that SDF-1alpha can influence NPC function through CXCR4 and that CXCR4 is functional on NPC.     

HIV-1, chemokines and neurogenesis

HIV-1 infection of the brain results in a large number of behavioural defecits accompanied by diverse neuropathological signs. However,it is not clear how the virus produces these effects or exactly how the neuropathology and behavioural defecits are related. In this article we discuss the possibility that HIV-1 infection may negatively impact the process of neurogenesis in the adult brain and that this may contribute to HIV-1 related effects on the nervous system. We have previously demonstrated that the development of the dentate gyrus during embryogenesis requires signaling by the chemokine SDF-1 via its receptor CXCR4. We demonstrated that neural progenitor cells that give rise to dentate granule neurons express CXCR4 and other chemokine receptors and migrate into the nascent dentate gyrus along SDF-1 gradients. Animals deficient in CXCR4 receptors exhibit a malformed dentate gyrus in which the migration of neural progenitors is stalled. In the adult, neurogenesis continues in the dentate gyrus. Adult neural progenitor cells existing in the subgranlar zone, that produce granule neurons, express CXCR4 and other chemokine receptors, and granule neurons express SDF-1 suggesting that SDF-1/CXCR4 signaling is also important in adult neurogenesis. Because the cellular receptors for HIV-1 include chemokine receptors such as CXCR4 and CCR5 it is possible that the virus may interfere with SDF-1/CXCR4 signaling in the brain including disruption of the formation of new granule neurons in the adult brain.     

Intracellular CXCR4 signaling, neuronal apoptosis and neuropathogenic mechanisms of HIV-1-associated dementia.

The mechanism(s) by which HIV-1 affects neural injury in HIV-1-associated dementia (HAD) remains unknown. To ascertain the role that cellular and viral macrophage products play in HAD neurotoxicity, we explored one potential route for neuronal demise, CXCR4. CXCR4, expressed on lymphocytes and neurons, is both a part of neural development and a co-receptor for HIV-1. Its ligand, stromal cell-derived factor-1alpha (SDF-1alpha), affects neuronal viability. GTP binding protein (G-protein) linked signaling after neuronal exposure to SDF-1alpha, virus-infected monocyte-derived macrophage (MDM) secretory products, and virus was determined. In both human and rat neurons, CXCR4 was expressed at high levels. SDF-1alpha/beta was detected predominantly in astrocytes and at low levels in MDM. SDF-1beta/beta was expressed in HAD brain tissue and upregulated in astrocytes exposed to virus infected and/or immune activated MDM conditioned media (fluids). HIV-1-infected MDM secretions, virus and SDF-1beta induced a G inhibitory (Gi) protein-linked decrease in cyclic AMP (cAMP) and increase inositol 1,4, 5-trisphosphate (IP3) and intracellular calcium. Such effects were partially blocked by antibodies to CXCR4 or removal of virus from MDM fluids. Changes in G-protein-coupled signaling correlated, but were not directly linked, to increased neuronal synaptic transmission, Caspase 3 activation and apoptosis. These data, taken together, suggest that CXCR4-mediated signal transduction may be a potential mechanism for neuronal dysfunction during HAD.     

Nitric oxide donor up-regulation of SDF1/CXCR4 and Ang1/Tie2 promotes neuroblast cell migration after stroke

We tested the hypothesis that a nitric oxide donor, DETA-NONOate, up-regulates stromal cell-derived factor-1 (SDF1) and angiopoietin 1 (Ang1) in the ischemic brain and their respective receptors chemokine CXC motif receptor 4 (CXCR4) and Tie2 in the subventricular zone (SVZ) and thereby promote SVZ neuroblast cell migration after stroke. C57BL/6J mice were subjected to middle cerebral artery occlusion (MCAo), and 24 hr later DETA-NONOate (0.4 mg/kg) or phosphate-buffered solution was intravenously administered. Mice were sacrificed at 14 days for histological assessment or sacrificed at 3 days for analysis by real-time polymerase chain reaction and migration after MCAo. To elucidate whether SDF1/CXCR4 and Ang1/Tie2 pathways mediate DETA-NONOate-induced SVZ migration after stroke, SDF1alpha, Ang1 peptide, a specific antagonist of CXCR4 (AMD3100), and a neutralizing antibody of Tie2 (anti-Tie2) were used in vitro. DETA-NONOate significantly increased the percentage area of doublecortin (DCX, a marker of migrating neuroblasts)-immunoreactive cells in the SVZ and ischemic boundary zone. DETA-NONOate significantly increased the expression of SDF1 and Ang1 in the ischemic border and up-regulated CXCR4 and Tie2 in the SVZ compared with MCAo control. DCX-positive cell migration from SVZ explants was significantly increased in the DETA-NONOate treatment group compared with MCAo-alone animals. In vitro, SDF1alpha and Ang1 significantly increased SVZ explants cell migration. In addition, inhibition of CXCR4 or Tie2 significantly attenuated DETA-NONOate-induced SVZ cell migration. Our data indicate that treatment of stroke with a nitric oxide donor up-regulates SDF1/CXCR4 and Ang1/Tie2 pathways and thereby likely increases SVZ neuroblast cell migration.     

SDF-1alpha is expressed in astrocytes and neurons in the AIDS dementia complex: an in vivo and in vitro study

Recent in vitro studies suggest that the alpha chemokine stromal-derived factor-1alpha (SDF-1alpha) and its receptor CXCR-4 may contribute to neuronal apoptosis in HIV infection of the brain. The cellular and regional expression of this chemokine and its relationship to the AIDS dementia complex (ADC), however, have remained undetermined. Using immunohistochemistry and semiquantitative RT-PCR, we examined the expression of SDF-1alpha in the frontal cortex (FC), the adjacent deep white matter (DWM). and the basal ganglia (BG) of 17 patients with ADC and 5 normal controls, and the FC and temporal cortex of 6 patients with Alzheimer disease (AD). Additionally, SDF-1alpha expression was studied in 3 different neuronal cultures: differentiated SK-N-MC cells, primary human fetal neuronal, and mouse hippocampal cultures. SDF-1alpha staining was predominantly localized to astrocytes in all 3 groups in the gray matter of the FC and the BG, often in the vicinity of cortical and basal ganglia neurons, but was generally absent in the DWM. Further, the number of positive neurons was significantly greater in the BG of AIDS subjects with advanced brain disease compared to subjects with lesser disease (p = 0.029). All cultures showed prominent SDF-1alpha staining of neurons within the cytoplasm and in neurites, whereas preferential expression in GABA-ergic neurons was found in hippocampal cultures. This is the first study to show that SDF-1alpha is constitutively expressed in astrocytes of the deep and cortical gray matter as well as in neurons of the human brain. Its increased expression in basal ganglia neurons of patients with advanced HIV CNS disease suggests it may also contribute to pathogenesis.     

Aristaless-related homeobox gene disruption leads to abnormal distribution of GABAergic interneurons in human neocortex: evidence based on a case of X-linked lissencephaly with abnormal genitalia (XLAG)

X-linked lissencephaly with abnormal genitalia (XLAG) is a rare disorder caused by mutations in the aristaless-related homeobox (ARX) gene, located on Xp22.13. Arx-null mice show loss of tangential migration of GABAergic interneurons, presumably being related to caudal ganglionic eminence tangential migration. In the present study, we investigated a subpopulation of GABAergic interneurons in the brain of an infant with XLAG, who had a novel nonsense mutation of the ARX gene, compared with those of age-matched normal controls and Miller-Dieker syndrome. We performed immunocytochemistry for interneuron and migration markers. We found that glutamic acid decarboxylase (GAD)- and calretinin (CR)-containing cells were significantly reduced in the neocortex and located in the white matter and neocortical subventricular zone, while neuropeptide Y- or cholecystokinin-containing cells were normally distributed. Moreover, in the neocortical subventricular region, the GAD- and CR-containing cells expressed the radial migration marker Mash-1 as well as nestin. Our findings suggest that ARX protein controls not only the tangential migration of GABAergic interneurons from the ganglionic eminence, but also may serve to induce radial migration from the neocortical subventricular zone.