Abstract

GABAergic interneurons perform crucial roles in cortical development and function. These roles are executed by a diversity of interneuron subtypes, and abnormal function of particular subtypes has been implicated in a variety of neuropsychiatric diseases. However, little is known about the mechanisms that generate interneuron diversity. This paper discusses the potential origins of interneuron subtypes. Evidence is reviewed that suggests bipolar calretinin expressing interneurons may have distinct origins from those that express parvalbumin or somatostatin. In addition, evidence is presented that migratory cells from the subcortical subventricular zone (SVZ) do not proliferate after migration into the cortical SVZ.

Introduction

The cerebral cortex functions in memory and higher-order cognitive processing. These functions are served by essentially two classes of neurons, excitatory projection neurons and inhibitory interneurons. The interneurons, which comprise 20% to 30% of cortical neurons, serve a vital role in modulating cortical output and appear also to affect the postnatal development of cortical circuitry (Jones, 1993; Huang et al., 1999; Fagiolini and Hensch, 2000). These functions are conducted by a remarkable diversity of distinct subtypes that are distinguished by axonal and dendritic morphology (Ramon y Cajal, 1911; Jones, 1975; Lund and Lewis, 1993), chemical markers (DeFelipe, 1993; Kubota et al., 1994; Gonchar and Burkhalter, 1997), connectivity and physiology (Cauli et al., 1997; Kawaguchi and Kubota, 1997; Gupta et al., 2000).

Although the distinct functions of interneuron subtypes are just beginning to be unraveled, both neurological and psychiatric diseases have been linked to abnormalities in interneuron function. Some forms of epilepsy may be related to abnormal number or function of chandelier cells (DeFelipe, 1999). Neuropeptide Y (NPY), expressed by subgroups of interneurons in the hippocampus and neocortex, has been linked to both anti-seizure and anxiolytic effects (Baraban, 2002; Heilig and Thorsell, 2002). Chandelier cell dysfunction has also been implicated schizophrenia (Woo et al., 1998; Volk et al., 2002), and other interneuron abnormalities have been described in bipolar disorder (Benes and Berretta, 2001). Unfortunately, these studies cannot easily distinguish whether the interneuron defects are a cause rather than an effect of the disease.

Despite the relevance to cortical function and dysfunction, little is known about the generation of cortical interneuron diversity. Part of the difficulty may be due to fact that, at first glance, organizing this diversity appears rather forbidding. It has been known for some time that immunochemically detected parvalbumin, somatostatin, and calretinin is present in largely non-overlapping subgroups of interneurons in rodent neocortex (Rogers, 1992; Kubota et al., 1994). These subgroups together comprise >80% of cortical interneurons (Gonchar and Burkhalter, 1997), but the importance of these subgroups in terms of understanding the development of interneuron diversity is complicated by the fact that they cut across morphologically defined subtypes. For example, parvalbumin, present in ∼50% of interneurons in the neocortex of adult rats, is expressed in both Chandelier cells and in some basket cells. On the other hand, some physiological and connectivity characteristics appear to also separate into these same subgroups. Thus, RT-PCR analysis of interneurons characterized as fast spiking, regular spiking or irregular-spiking reveals expression of either parvalbumin, somatostatin or calretinin, respectively (Cauli et al., 2000). Similar findings have also been reported using immunohistochemical methods after electrophysiological recordings (Kawaguchi and Kubota, 1997). Connectivity also appears to distinguish between these neurochemically defined subgroups, as calretinin- and VIP-expressing cells appear to have a greater propensity for innervating other interneurons in both cortex and hippocampus (Gulyas et al., 1996; Defelipe et al., 1999; Gonchar and Burkhalter, 1999). As shall be discussed below, the calretinin-expressing subgroup, as opposed to those expressing parvalbumin or somatostatin, also appear to differ in terms of their origins.

Sources of Cortical Interneurons

Recent fate-mapping experiments using transgenic mice have confirmed the long-held view that cortical projection neurons derive from the dorsal, ‘pallial’ portion of the telencephalon (Iwasato et al., 2000; Gorski et al., 2002). By contrast, in rodents [for reviews see (Parnavelas, 2000; Marin and Rubenstein, 2001)] ferrets (Anderson et al., 2002) and humans (Letinic et al., 2002) many cortical interneurons derive from the ventral (subcortical, pallidal) telencephalon in the anlage of the basal ganglia. Cell tracing and cell transplantation experiments in vitro (Lavdas et al., 1999; Anderson et al., 2001; Polleux et al., 2002) and in vivo (Wichterle et al., 1999, 2001) suggest that the medial ganglionic eminence (MGE) is the primary source of cortical interneurons in rodents.

The fact that many cortical interneurons derive from the subcortical telencephalon suggests two general possibilities for the generation of interneuron diversity. First, interneuron diversity could be established in proliferative zone(s) where they originate. This scenario appears to be the case for other subtypes of neurons, including those in the spinal cord (Jessell, 2000) retina (Livesey and Cepko, 2001), and projection neurons of the cerebral cortex (McConnell, 1995). Alternatively (or in addition), interneuron subtypes could be differentiated from multipotential GABAergic ‘proto’-interneurons by the actions of local cues within the cerebral cortex (Mione et al., 1994).

Clearly, the proper differentiation of interneurons depends upon factors within the cortex (Gotz and Bolz, 1994). Some of these include activity (Antonopoulos et al., 1992; Obst et al., 1998), neurotrophins such as BDNF (Marty et al., 1997; Huang et al., 1999) and cytokines such as leukemia inhibitory factor (LIF) (Wahle et al., 2000). The specificity for these effects on any particular subtype of interneuron has not been well established, although differences in exposure or sensitivity to LIF may explain transient versus permanent expression of NPY in parvalbumin or somatostatin expressing subgroups, respectively (Wahle et al., 2000). Further exploration of epigenetic factors in the differentiation of interneurons are likely to find additional subtype specific influences, but the concept that important steps in the generation of interneuron diversity occur at the cell’s origin is supported by recent evidence that spatial and/or temporal differences exist in the genesis of interneuron subtypes (Table 1).

Temporal Differences in the Generation of Interneuron Subtypes

Cortical interneurons, like the projection neurons, are generally born in an inside-out order with respect to their location within the cortical lamina, and contemporaneously with pyramidal neurons that inhabit the same layer (Miller, 1985; Fairén et al., 1986; Peduzzi, 1988). However, a substantial subpopulation of interneurons, those expressing somatostatin, appear to be born over a relatively narrow timeframe that coincides with genesis of the deeper cortical layers, regardless of the eventual laminar location of the somatostatin-expressing cell (Cavanagh and Parnavelas, 1988). Thus, the majority of somatostatin-expressing interneurons in layers II/III are born prior to those expressing vasoactive intestinal protein (VIP) (Cavanagh and Parnavelas, 1989). Since VIP is largely expressed in bipolar interneurons that colabel with calretinin (Kubota et al., 1994; Porter et al., 1998), this result suggests that the somatostatin and VIP/calretinin-expressing subgroups have distinct temporal origins. Future studies in experimental animals with relatively long periods of neurogenesis in the cortex, such as ferrets, would be highly valuable for examining the issue of interneuron fate and birthdate in detail.

Spatial Differences in the Generation of Interneuron Subtypes

Several studies have presented evidence that spatial differences exist in the origins of some interneuron subtypes. First, genetically labeled cells from the MGE, transplanted back into the MGE of host embryos in vivo, were recently shown to give rise to large numbers of morphologically defined subtypes of interneurons. These included Chandelier, bitufted and basket cells (Wichterle et al., 2001). Colabeling of the MGE-derived cells with somatostatin, parvalbumin and calretinin, which label largely non-overlapping subgroups of cortical interneurons (Kubota et al., 1994; Gonchar and Burkhalter, 1997), revealed interesting results. While at least 70% of the MGE-derived cortical cells expressed somatostatin or parvalbumin, less than 3% expressed calretinin. Since calretinin expressing interneurons are thought to account for ∼17% of cortical interneurons (Gonchar and Burkhalter, 1997), these results suggest that this subgroup of interneurons derives from a spatially or temporally distinct source from interneurons that express somatostatin or parvalbumin.

Similar results were recently found by focally injecting tritiated thymidine into the ventral–lateral wall of the lateral ventricle, or into the neocortical proliferative zone, in neonatal ferrets (Anderson et al., 2002). At this age the pups are still generating neurons destined for the superficial cortical layers, so that the fate of cells born within the cortical or subcortical proliferative zones could be assessed at 6 weeks of age. Some of the subcortically labeled cells gave rise to cortical interneurons expressing parvalbumin or somatostatin, but not calretinin. The cortical injections gave rise to pyramidal neurons, and very small number of apparently GABA-expressing cells, but virtually no neurons expressing somatostatin, parvalbumin or calretinin.

Further evidence that interneuron subtypes may have distinct sources comes from the analysis of mice lacking the homeobox transcription factor Nkx2.1. In these mutants, which die at birth, the normal MGE has been replaced by tissue with a lateral ganglionic eminence (LGE)-like character (Sussel et al., 1999). The early (pre-E14.5), MGE to cortex migration appears to be absent in these animals (Anderson et al., 2001), and the animals lack 50% of cortical GABA-expressing cells at E18.5 (Sussel et al., 1999). Interestingly, later (E14.5–E16.5) subcortical to cortex migration, which in wild-type mice appears to contain cells born in both the MGE and LGE, appears to be intact (Anderson et al., 2001). At these stages the Nkx2.1 mutants lack markers of MGE-derived cells, including Lhx6 and Lhx7. However, although only half of the cortical GABAergic neurons are missing in these mutants, they appear to be missing all of those that express NPY and NOS (Anderson et al., 2001).

Consistent with the findings in tissue sections, primary cultures of cortical cells from E18.5 Nkx2.1 mutants also lack NPY and NOS, and somatostatin expressing cells after 2 weeks in vitro. However, preliminary results suggest that calretinin expressing bipolar neurons do grow out of these mutant cultures (Q. Xu and S.A. Anderson, in preparation). These results further support the notion that somatostatin-expressing multipolar interneurons derive from the MGE, whereas the calretinin-expressing bipolar interneurons do not.

Another mutant in which calretinin-expressing cortical interneurons are differentially affected relative to other subtypes is the flathead rat (Sarkisian et al., 1999, 2001). These animals harbor a recessive mutation in the gene encoding Citron-K (Sarkisian et al., 2002). Neural progenitors in the telencephali of homozygous mutants display cytokinetic abnormalities and apoptosis that particularly affects the ganglionic eminences. In the cortex, interneurons appear to be affected more than projection neurons, but calretinin-expressing interneurons are spared relative to those that express parvalbumin. Although the cause of this difference is unclear, the results again are consistent with the hypothesis that interneuron subtypes are molecularly distinct at the progenitor stage.

Potential Origins of Calretinin-expressing Interneurons

In sum, several lines of evidence indicate that calretinin-expressing bipolar interneurons arise through distinct developmental programs from those that express somatostatin or parvalbumin. What is the basis for this difference? The relative inability to label calretinin-expressing neurons by fate-mapping cortical cells that derived from the MGE in vivo (Wichterle et al., 2001; Anderson et al., 2002) suggests that these cells derive from a different place or time during development. In addition, the apparent sparing of these interneurons in forebrains lacking a normal MGE (Xu and Anderson, in preparation) suggests that they do not derive from the MGE at all. So what might be the source(s) of calretinin expressing cortical interneurons? Reasonable possibilities include the LGE, from which newly postmitotic cells do appear to migrate into the cortex in wild type as well as Nkx2.1 mutants (Anderson et al., 2001). Alternatively, they may derive from anterior subventricular zone (SVZ) of the LGE, which along with the rostral migratory stream (RMS) gives rise to interneurons of the olfactory bulb, some of which also express calretinin. Immunohistochemical staining for DLX1, which labels interneuron precursors in the RMS, appears to also label cells migrating out of the RMS region and into the cortex just prior to reaching the bulb (Anderson et al., 1999). An earlier migration from the retrobulbar area into layer I of the cortex has also been described for calretinin and GABA-expressing cells (Meyer et al., 1998). However, efforts to identify a robust RMS or retrobulbar migration into the overlying cortex in vitro have been unsuccessful (S.A. Anderson, unpublished data).

Another subcortical source of distinct subtypes of cortical interneurons may lie within the caudal telencephalon. An interneuron migration from ventral to dorsal telencephalon that runs in a caudal-dorsal direction has recently been described in chick (Anderson et al., 1997b; Cobos et al., 2001). In rodents, the caudal ganglionic eminence (CGE) gives rise to neuronal migrations that are distinct from those of the LGE or MGE, and also gives rise to cortical interneurons (Nery et al., 2002). It is not clear whether the CGE gives rise to particular subclasses of cortical interneurons.

Alternatively, calretinin expressing bipolar interneurons could originate from the cortex itself. Recent evidence from explant cultures of human embryos suggests that a substantial number of cortical interneurons arise from the cortical SVZ (Letinic et al., 2002). This may represent a species difference between rodents and humans, although several lines of evidence suggest that the late gestation or early postnatal cortical SVZ of rodents could conceivably give rise to interneurons. First, mice lacking both transcription factors Dlx1 and Dlx2, which also lack migration from the ganglionic eminences into the cortex in explant cultures, nonetheless have only a 75% reduction of cortical GABA-expressing cells at birth (Anderson et al., 1997b, 2001). It remains unclear whether this belies the presence of a ‘leaky’ phenotype with regard to subcortical to cortical migration, migration of interneurons from a subcortical area other than the ganglionic eminences, or the generation of interneurons in the Dlx1/2 mutant cortex itself. In support of the ‘leaky’ hypothesis, a few cells expressing Lhx6, a lim-homeodomain transcription factor that is expressed in cells migrating from the MGE to the cortex (Grigoriou et al., 1998; Sussel et al., 1999), are present in tissue sections from the Dlx1/2 mutant cortex (Anderson et al., 2001).

Another, perhaps stronger line of evidence supporting a cortical source of calretinin expressing interneurons comes from cortical cultures prepared from E14 to E16 rats. These cultures give rise to calretinin expressing interneurons that proliferate in response to Fgf2 and that appear to be distinct from Cajal–Retzius cells (Pappas and Parnavelas, 1998). Interestingly, this effect did not occur for calbindin-expressing interneurons, which label a different subpopulation in postnatal cortex (Defelipe et al., 1999). Moreover, calbindin labels apparent interneuron precursors migrating from the ganglionic eminences (Anderson et al., 1997a), and thus may be restricted to a subcortically derived interneuron subgroup. The implication is that cortical precursors in the rodent, perhaps later in gestation, could generate the calretinin-expressing interneurons.

Studies whose results weigh against this possibility include the recent finding that injections of the S-phase marker [3H]thymidine into the cortical proliferative zone in the neonatal ferret, whose neocortex is at a similar developmentally age as the E15 mouse, do not label calretinin-expressing interneurons in vivo (Anderson et al., 2002). However, this age may be too early for labeling these cells. Another relevant study is that of the cortical phenotype of the Fgf2 mutant (Raballo et al., 2000). These animals live into adulthood but have a dramatic reduction in projection neuron production in the rostral and lateral cortex with a corresponding increase in proportion of GABA-expressing cells (Korada et al., 2002). Calbindin and parvalbumin expressing interneuron subtypes were also increased in proportion, although an affect on calretinin expressing subtypes was not reported.

Virtually All Cortical Interneurons Appear to Express Dlx Genes

In summary, evidence suggests that parvalbumin and somatostatin expressing interneurons derive from the MGE, while the origin(s) of the calretinin expressing subgroup is less clear. On the other hand, all three groups co-localize lacZ in Dlx5/6 enhancer mice (Stuhmer et al., 2002). This enhancer appears to direct β-galactosidase (β-gal) expression within all of the Dlx-expressing interneurons migrating from the ganglionic eminences into the cortex. Although the β-gal message, like that of Dlx5 and 6, becomes nearly undetectable during the second postnatal week, functional β-gal protein appears to persist into adulthood. In adult neocortex, nearly all of the β-gal labeled cells co-label for GABA. This appears to include all the major interneuron subtypes, since over 95% of the GABA expressing cells also express β-gal, including those that express calbindin, parvalbumin or calretinin. Somatostatin was not tested, but two molecules that are largely expressed in somatostatin containing subpopulations, nitric oxide synthase and NPY, also showed nearly 100% co-labeling with the transgenic marker.

The most plausible interpretation of this result is that, in rodents, nearly all cortical interneurons derive from Dlx-expressing proliferative zones of the ventral telencephalon. But there is another possibility. After E14.5 in the mouse, many Dlx-expressing cells appear to migrate from the subcortical SVZ directly into the SVZ of the neocortex. Some of these cells express the postmitotic neuronal marker Tuj1, and, at E16.5, are not proliferating based on their lack of labeling with antibodies against proliferating cell nuclear antigen (PCNA) (Anderson et al., 2001). Slice transplant experiments of E14.5 GE, pulsed with BrdU for 4 h before fixation at 2 days in vitro (DIV), also found that GE cells did not proliferate after migration into the cortex (Polleux et al., 2002).

Surprisingly, although DLX1 expressing cells in the cortical SVZ were negative for PCNA at E16.5, at P0 many of them colabel with PCNA and thus appear to be proliferating (Anderson et al., 2001). So what might be the source of proliferating Dlx-expressing cells in the neonatal cortical SVZ? One possibility, as has been proposed to be the case in humans (Letinic et al., 2002), is that Mash1 expression in a minority of cortical progenitors induces Dlx genes. Another possibility is that, as occurs in the RMS, cells fated to be interneurons may be able to migrate while maintaining the capacity to proliferate. A logical source of this migration would be the MGE or LGE.

Migratory Cells from the Ganglionic Eminences Do Not Appear to Proliferate in the Cortical SVZ In Vitro

To test the hypothesis that the neonatal cortical SVZ is partially ‘seeded’ by a late-gestation migration from the ganglionic eminences, we used a slice transplantation assay (Anderson et al., 2001), modified by the use of GFP-expressing donor tissues (Polleux et al., 2002). Ganglionic eminence (GE) proliferative zone, from E15.5, E17.5 or E18.5 donor mice in which all cells express green fluorescent protein (GFP, a kind gift from Andras Nagy) (Hadjantonakis et al., 1998) were transplanted homotopically and isochronically into slices from wild-type mice (Fig. 1a). The slices were maintained on polycarbonate filters floating on medium containing Neurobasal/B27 (Gibco, Rockville, MD). After 44 h of culture, BrdU was added to the medium for 4 h to label cells in S-phase (Miller and Nowakowski, 1988). The slices were then fixed in 4% paraformaldehyde, cryoprotected in 20% sucrose, embedded in OCT and sectioned at 10 μm on a cryostat. The ability of GFP-expressing donor cells from the GE to proliferate within the cortex was assessed by BrdU and GFP immunodetection and epifluorescence. The cortical proliferative zone was identified by the presence of BrdU labeled cells. Only those 10 μm hemisections containing at least 50 GFP+ cells and 50 BrdU+ cells within this zone were counted. Cells were counted in at least three sections from each of three separate transplants.

As shown previously, a large number of cells from the subcortical donor tissue migrated into the cortex of the host slice (Fig. 1b,c) (Anderson et al., 2001; Polleux et al., 2002). Although many of these cells migrated into the cortical proliferative zone (PZ), and although a large number of the cortical PZ cells incorporated BrdU, double labeled cells were extremely rare at any of the three ages (Fig. 1c). Summed totals (double labeled/all GFP+ cells in the cortical PZ from all counted sections) reveal 1/551 from slices started at E15.5, 0/475 started at E17.5, and 3/683 started at E18.5.

To determine whether the mitogenic properties of Fgf2 may be necessary for the proliferation of migratory GFP-expressing cells 10 ng/ml of Fgf2 was added to the medium. Again, migration of GFP-labeled GE cells into the cortical PZ was robust, as was proliferation in the cortical PZ. However, the percentage of GFP expressing cells that incorporated BrdU within the cortical proliferative zone remained extremely small (E15.5 + 2DIV, 4/734; E17.5 + 2DIV, 0/560, E18.5 + 2DIV, 0/518).

To confirm that DLX1-expressing cells do proliferate in these slices, we tested DLX1 and BrdU double labeling in the cortical PZ of the non-transplanted hemisphere. In the cortical proliferative zone of sections of E17.5 + 2DIV there were 54 DLX1/BrdU co-labeled cells out of 416 that expressed Dlx1 (13%, data not shown). This percentage is consistent with the previous, unquantified finding that roughly half of DLX1 expressing cells in the cortical SVZ of tissue sections prepared at P0 also express PCNA (Anderson et al., 2001), a marker that is expressed throughout the cell cycle and shortly beyond.

These findings are consistent with a previous report that used similar methods at earlier time points (Polleux et al., 2002). They suggest that some proliferative, DLX expressing cells in the cortical SVZ of rodents derive from truly cortical, ‘pallial’ progenitors, as has been proposed to occur in humans (Letinic et al., 2002). However, their interpretation must be tempered by the many limitations of explant cultures, and by the possibility that proliferating DLX-expressing cells could migrate into the cortical SVZ from Dlx expressing regions other than the ganglionic eminences, such as the RMS. Regardless of the initially pallial or pallidal origin of the mitotic Dlx expressing cell in the cortical SVZ, it would be interesting to determine whether they selectively give rise to calretinin-expressing interneuron subtypes. Indeed, they might not give rise to neurons at all, as Dlx genes also appear to be expressed in progenitors of cortical oligodendrocytes (He et al., 2001; Marshall and Goldman, 2002) and astrocytes (Marshall and Goldman, 2002).

In summary, although the issue is far from settled, converging lines of evidence suggest that spatial and temporal origins distinguish between major neurochemically and morphologically defined interneuron subtypes. Specifically, parvalbumin (Pv) expressing Chandelier cells (and probably also Pv-expressing basket cells), as well as somatostatin expressing interneurons, appear to derive from the MGE. Small, bipolar, calretinin expressing interneurons appear not to derive from the MGE, and some evidence suggests that they may undergo their final mitosis within the perinatal SVZ of the cortex itself. In relation to a recent report that the majority of interneurons in humans derive from the cortical SVZ (Letinic et al., 2002), it is interesting to note that in the cortex of humans and other primates a far higher percentage of interneurons express calretinin than in rodents (Conde et al., 1994; Gabbot and Bacon, 1996; Gabbot et al., 1997; Gonchar and Burkhalter, 1997; Kawaguchi and Kubota, 1997).

The presence of spatial and temporal differences in the origins of distinct interneuron subtypes does not constitute proof that differences in factors present at these origins accounts for later phenotypic differences. Differential influences encountered during migration, or based upon the timing or location of their entry into the cortical plate, may also direct the initial expression of sub-phenotypes. Still, identification of the sources of interneuron subtypes will be a crucial step for studies of the factors that may determine their fate. Gaining knowledge of these factors should have important therapeutic benefits by shedding light on potential mechanisms of diseases that involve interneuron deficits. In addition, the directed use of factors that dictate the fates of interneuron subtypes could be useful for therapeutic cell replacement strategies for diseases such as epilepsy.

Table 1

Summary of mutants with prominent abnormalities of cortical interneuron genesis or migration

Mutant (gene) Max. age analyzed Cortical interneuron phenotype Other telencephalic phenotypes 
References (limited to those addressing the interneuron phenotype): 1, Anderson et al., 1997b; 2, Pleasure et al., 2000; 3. Sussel et al., 1999; 4. Casarosa et al., 1999; 5. Chapouton et al., 1999; 6, Monaghan et al., 1997; 7, Sarkisian et al., 2001; 8, Powell et al., 2001; 9, Polleux et al., 2002; 10, Shinozaki et al., 2002; 11, Muzio et al., 2002; 12, Kitamura et al., 2002
Ncx, neocortex; Hp, hippocampus; V–D, ventral–dorsal.; calret., calretinin; somat, somatostatin; calbin., calbindin; Pv, parvalbumin; ERCx, entorhinal cortex; NOS, nitric oxide synthase; GAD, glutamate decarboxylase. 
Dlx1/Dlx21 P0 Ncx: 75% ⇓ GABA+ striatal dysgenesis 
 Hp2: ∼100% ⇓ GABA+ ⇓ ß GABA+ cells in olfactory bulb  
Nkx2.13 E19.5 50% ⇓ GABA+ normal MGE replaced by  
 no NPY, somat., NOS ‘LGE-like’ tissue  
Mash14 P0 ⇓ GABA+ dysgenesis of striatum and 
  ventral pallidum  
PAX65 P0 1x–2x ⇑ GABA+ abnormal cortical migration, patterning 
 ⇑ V–D migration and thalamic connectivity  
Tailless6 adult ⇓ calret. +,somat.+ ⇓ rhinencephalon, ⇓dentate 
  normal calbin.+, Pv+, NOS+  
Flathead7 2 wks in ERCx, ∼70% ⇓ GABA+ 40% ⇓ Ctx size 
(Citron K) in S1, ⇓somat.+, Pv+ ⇑ cell death in GE > Ctx  
 in S1 normal Calret.+ Abnormal neuronal cytokinesis  
u-PAR8 P0 20–65% ⇓ calbindin ⇑ cell packing in neocortex 
TrkB9 E15 30% ⇓ calbindin  
 ⇓ V–D migration   
Emx1/Emx210 E18.5 ⇓ GABA, ⇓ V–D migration cortical lamination defects  
Emx2/Pax611 E16.5 ⇑ ⇑ GAD cortex re-specified as basal ganglia 
ARX12 E12.5 ⇓ V–D migration Pax6–/– like patterning defect of LGE 
 P3 no NPY, NOS ⇓ Cortical proliferation 
Mutant (gene) Max. age analyzed Cortical interneuron phenotype Other telencephalic phenotypes 
References (limited to those addressing the interneuron phenotype): 1, Anderson et al., 1997b; 2, Pleasure et al., 2000; 3. Sussel et al., 1999; 4. Casarosa et al., 1999; 5. Chapouton et al., 1999; 6, Monaghan et al., 1997; 7, Sarkisian et al., 2001; 8, Powell et al., 2001; 9, Polleux et al., 2002; 10, Shinozaki et al., 2002; 11, Muzio et al., 2002; 12, Kitamura et al., 2002
Ncx, neocortex; Hp, hippocampus; V–D, ventral–dorsal.; calret., calretinin; somat, somatostatin; calbin., calbindin; Pv, parvalbumin; ERCx, entorhinal cortex; NOS, nitric oxide synthase; GAD, glutamate decarboxylase. 
Dlx1/Dlx21 P0 Ncx: 75% ⇓ GABA+ striatal dysgenesis 
 Hp2: ∼100% ⇓ GABA+ ⇓ ß GABA+ cells in olfactory bulb  
Nkx2.13 E19.5 50% ⇓ GABA+ normal MGE replaced by  
 no NPY, somat., NOS ‘LGE-like’ tissue  
Mash14 P0 ⇓ GABA+ dysgenesis of striatum and 
  ventral pallidum  
PAX65 P0 1x–2x ⇑ GABA+ abnormal cortical migration, patterning 
 ⇑ V–D migration and thalamic connectivity  
Tailless6 adult ⇓ calret. +,somat.+ ⇓ rhinencephalon, ⇓dentate 
  normal calbin.+, Pv+, NOS+  
Flathead7 2 wks in ERCx, ∼70% ⇓ GABA+ 40% ⇓ Ctx size 
(Citron K) in S1, ⇓somat.+, Pv+ ⇑ cell death in GE > Ctx  
 in S1 normal Calret.+ Abnormal neuronal cytokinesis  
u-PAR8 P0 20–65% ⇓ calbindin ⇑ cell packing in neocortex 
TrkB9 E15 30% ⇓ calbindin  
 ⇓ V–D migration   
Emx1/Emx210 E18.5 ⇓ GABA, ⇓ V–D migration cortical lamination defects  
Emx2/Pax611 E16.5 ⇑ ⇑ GAD cortex re-specified as basal ganglia 
ARX12 E12.5 ⇓ V–D migration Pax6–/– like patterning defect of LGE 
 P3 no NPY, NOS ⇓ Cortical proliferation 
Figure 1.

Migratory cells from the subcortical telencephalon do not proliferate in the cortical SVZ. (A) Dark field photomicrograph of an E18.5 telencephalic explant shortly after homotopic transplantation of subcortical tissue from a GFP-expressing donor. The donor tissue is pseudocolored green. (B) Epifluorescence photomicrograph (red = BrdU, green = GFP) of a 10 μm section of the same slice (boxed area). BrdU was added to the culture medium 4 h before fixation at 2 days in vitro. (C) Higher magnification view of the boxed area in (B). Co-labeling of BrdU and GFP in cells that migrated from the donor tissue into the cortex of the host explant is not present. Scale bars: 500 μm in (A), 100 μm in (B). Ncx, neocortex; Se, septal area; St, striatum.

Figure 1.

Migratory cells from the subcortical telencephalon do not proliferate in the cortical SVZ. (A) Dark field photomicrograph of an E18.5 telencephalic explant shortly after homotopic transplantation of subcortical tissue from a GFP-expressing donor. The donor tissue is pseudocolored green. (B) Epifluorescence photomicrograph (red = BrdU, green = GFP) of a 10 μm section of the same slice (boxed area). BrdU was added to the culture medium 4 h before fixation at 2 days in vitro. (C) Higher magnification view of the boxed area in (B). Co-labeling of BrdU and GFP in cells that migrated from the donor tissue into the cortex of the host explant is not present. Scale bars: 500 μm in (A), 100 μm in (B). Ncx, neocortex; Se, septal area; St, striatum.

This work was supported by grants from NARSAD, and the NIMH (K08-MH01620-05) to S.A.

References

Anderson SA, Eisenstat DD, Shi L, Rubenstein JL (
1997
) Interneuron migration from basal forebrain to neocortex: dependence on Dlx genes.
Science
 
278
:
474
–476.
Anderson SA, Qiu M, Bulfone A, Eisenstat DD, Meneses J, Pedersen R, Rubenstein JL (
1997
) Mutations of the homeobox genes Dlx-1 and Dlx-2 disrupt the striatal subventricular zone and differentiation of late born striatal neurons.
Neuron
 
19
:
27
–37.
Anderson S, Mione M, Yun K, Rubenstein JLR (
1999
) Differential origins of projection and local circuit neurons: role of Dlx genes in neocortical interneuronogenesis.
Cereb Cortex
 
9
:
646
–654.
Anderson SA, Marin O, Horn C, Jennings K, Rubenstein JL (
2001
) Distinct cortical migrations from the medial and lateral ganglionic eminences.
Development
 
128
:
353
–363.
Anderson SA, Kaznowski CE, Horn C, Rubenstein JL, McConnell SK (
2002
) Distinct origins of neocortical projection neurons and interneurons in vivo.
Cereb Cortex
 
12
:
702
–709.
Antonopoulos J, Papadopoulos GC, Michaloudi H, Cavanagh ME, Parnavelas JG (
1992
) Postnatal development of neuropeptide Y-containing neurons in the visual cortex of normal- and dark-reared rats.
Neurosci Lett
 
145
:
75
–78.
Baraban SC (
2002
) Antiepileptic actions of neuropeptide y in the mouse hippocampus require y5 receptors.
Epilepsia
 
43
Suppl 5:
9
–13.
Benes FM, Berretta S (
2001
) GABAergic interneurons: implications for understanding schizophrenia and bipolar disorder.
Neuropsychopharmacology
 
25
:
1
–27.
Casarosa S, Fode C, Guillemot F (
1999
) Mash1 regulates neurogenesis in the ventral telencephalon.
Development
 
126
:
525
–534.
Cauli B, Audinat E, Lambolez B, Angulo MC, Ropert N, Tsuzuki K, Hestrin S, Rossier J (
1997
) Molecular and physiological diversity of cortical nonpyramidal cells.
J Neurosci
 
17
:
3894
–3906.
Cauli B, Porter JT, Tsuzuki K, Lambolez B, Rossier J, Quenet B, Audinat E (
2000
) Classification of fusiform neocortical interneurons based on unsupervised clustering.
Proc Natl Acad Sci USA
 
97
:
6144
–6149.
Cavanagh ME, Parnavelas JG (
1988
) Development of somatostatin immunoreactive neurons in the rat occipital cortex: a combined immunocytochemical–autoradiographic study.
J Comp Neurol
 
268
:
1
–12.
Cavanagh ME, Parnavelas JG (
1989
) Development of vasoactive-intestinal-polypeptide-immunoreactive neurons in the rat occipital cortex: a combined immunohistochemical–autoradiographic study.
J Comp Neurol
 
284
:
637
–645.
Chapouton P, Gärtner A, Götz M (
1999
) The role of Pax6 in restricting cell migration between developing cortex and basal ganglia.
Development
 
126
:
5569
–5579.
Cobos I, Puelles L, Martinez S (
2001
) The avian telencephalic subpallium originates inhibitory neurons that invade tangentially the pallium.
Dev Biol
 
239
:
30
–45.
Conde, F, Lund, JS, Jacobowitz DM, Baimbridge KG, Lewis DA (
1994
) Local circuit neurons immunoreactive for calretinin, calbindin D-28k or parvalbumin in monkey prefrontal cortex: distribution and morphology
J Comp Neurol
 
341
95
–116.
DeFelipe J (
1993
) Neocortical neuronal diversity: chemical heterogeneity revealed by colocalization studies of classic neurotransmitters, neuro-peptides, calcium-binding proteins, and cell surface molecules.
Cereb Cortex
 
3
:
273
–289.
DeFelipe J (
1999
) Chandelier cells and epilepsy.
Brain
 
122
:
1807
–1822.
Defelipe J, Gonzalez-Albo MC, Del Rio MR, Elston GN (
1999
) Distribution and patterns of connectivity of interneurons containing calbindin, calretinin, and parvalbumin in visual areas of the occipital and temporal lobes of the macaque monkey.
J Comp Neurol
 
412
:
515
–526.
Fagiolini M, Hensch TK (
2000
) Inhibitory threshold for critical-period activation in primary visual cortex.
Nature
 
404
:
183
–186.
Fairén A, Cobas A, Fonseca M (
1986
) Times of generation of glutamic acid decarboxylase immunoreactive neurons in mouse somatosensory cortex.
J Comp Neurol
 
251
:
67
–83.
Gabbott PL, Bacon SJ (
1996
) Local circuit neurons in the medial prefrontal cortex in the monkey: II. Quantitative areal and laminar distributions.
J Comp Neurol
 
364
:
609
–636.
Gabbott PL, Jays PR, Bacon SJ (
1997
) Calretinin neurons in human medial prefrontal cortex (areas 24a,b,c, 32′, and 25).
J Comp Neurol
 
381
:
389
–410.
Gonchar Y, Burkhalter A (
1997
) Three distinct families of GABAergic neurons in rat visual cortex.
Cereb Cortex
 
7
:
347
–358.
Gonchar Y, Burkhalter A (
1999
) Connectivity of GABAergic calretinin-immunoreactive neurons in rat primary visual cortex.
Cereb Cortex
 
9
:
683
–696.
Gorski JA, Talley T, Qiu M, Puelles L, Rubenstein JL, Jones KR (
2002
) Cortical excitatory neurons and glia, but not GABAergic neurons, are produced in the Emx1-expressing lineage.
J Neurosci
 
22
:
6309
–6314.
Gotz M, Bolz J (
1994
) Differentiation of transmitter phenotypes in rat cerebral cortex.
Eur J Neurosci
 
6
:
18
–32.
Grigoriou M, Tucker AS, Sharpe PT, Pachnis V (
1998
) Expression and regulation of Lhx6 and Lhx7, a novel subfamily of LIM homeodomain encoding genes, suggests a role in mammalian head development.
Development
 
125
:
2063
–2074.
Gulyas AI, Hajos N, Freund TF (
1996
) Interneurons containing calretinin are specialized to control other interneurons in the rat hippocampus.
J Neurosci
 
16
:
3397
–3411.
Gupta A, Wang Y, Markram H (
2000
) Organizing principles for a diversity of GABAergic interneurons and synapses in the neocortex.
Science
 
287
:
273
–278.
Hadjantonakis AK, Gertsenstein M, Ikawa M, Okabe M, Nagy A (
1998
) Generating green fluorescent mice by germline transmission of green fluorescent ES cells.
Mech Dev
 
76
:
79
–90.
He W, Ingraham C, Rising L, Goderie S, Temple S (
2001
) Multipotent stem cells from the mouse basal forebrain contribute GABAergic neurons and oligodendrocytes to the cerebral cortex during embryogenesis.
J Neurosci
 
21
:
8854
–8862.
Heilig M, Thorsell A (
2002
) Brain neuropeptide Y (NPY) in stress and alcohol dependence.
Rev Neurosci
 
13
:
85
–94.
Huang ZJ, Kirkwood A, Pizzorusso T, Porciatti V, Morales B, Bear MF, Maffei L, Tonegawa S (
1999
) BDNF regulates the maturation of inhibition and the critical period of plasticity in mouse visual cortex.
Cell
 
98
:
739
–755.
Iwasato T, Datwani A, Wolf AM, Nishiyama H, Taguchi Y, Tonegawa S, Knopfel T, Erzurumlu RS, Itohara S (
2000
) Cortex-restricted disruption of NMDAR1 impairs neuronal patterns in the barrel cortex.
Nature
 
406
:
726
–731.
Jessell TM (
2000
) Neuronal specification in the spinal cord: inductive signals and transcriptional codes.
Nat Rev Genet
 
1
:
20
–29.
Jones EG (
1975
) Varieties and distribution of non-pyramidal cells in the somatic sensory cortex of the squirrel monkey.
J Comp Neurol
 
160
:
205
–267.
Jones EG (
1993
) GABAergic neurons and their role in cortical plasticity in primates.
Cereb Cortex
 
3
:
361
–372.
Kawaguchi Y, Kubota Y (
1997
) GABAergic cell subtypes and their synaptic connections in rat frontal cortex.
Cereb Cortex
 
7
:
476
–486.
Kitamura K, Yanazawa M, Sugiyama N, Dobyns WB, Yokoyama M, Morohashi K (
2002
). Mutation of ARX causes abnormal development of forebrain and testes in mice and X-linked lissencephaly with abnormal genitalia in humans
Nat Genet
 
32
:
359
–369.
Korada S, Zheng W, Basilico C, Schwartz ML, Vaccarino FM (
2002
) Fibroblast growth factor 2 is necessary for the growth of glutamate projection neurons in the anterior neocortex.
J Neurosci
 
22
:
863
–875.
Kubota Y, Hattori R, Yui Y (
1994
) Three distinct subpopulations of GABAergic neurons in rat frontal agranular cortex.
Brain Res
 
649
:
159
–173.
Lavdas AA, Grigoriou M, Pachnis V, Parnavelas JG (
1999
) The medial ganglionic eminence gives rise to a population of early neurons in the developing cerebral cortex.
J Neurosci
 
19
:
7881
–7888.
Letinic K, Zoncu R, Rakic P (
2002
) Origin of GABAergic neurons in the human neocortex.
Nature
 
417
:
645
–649.
Livesey FJ, Cepko CL (
2001
) Vertebrate neural cell-fate determination: lessons from the retina.
Nat Rev Neurosci
 
2
:
109
–118.
Lund JS, Lewis DA (
1993
) Local circuit neurons of developing and mature macaque prefrontal cortex: Golgi and immunocytochemical characteristics.
J Comp Neurol
 
328
:
282
–312.
Marin O, Rubenstein JL (
2001
) A long, remarkable journey: tangential migration in the telencephalon.
Nat Rev Neurosci
 
2
:
780
–790.
Marshall CA, Goldman JE (
2002
) Subpallial dlx2-expressing cells give rise to astrocytes and oligodendrocytes in the cerebral cortex and white matter.
J Neurosci
 
15
:
9821
–9830.
Marty S, Berzaghi Mda P, Berninger B (
1997
) Neurotrophins and activity-dependent plasticity of cortical interneurons.
Trends Neurosci
 
20
:
198
–202.
McConnell SK (
1995
) Strategies for the generation of neuronal diversity in the developing central nervous system.
J Neurosci
 
15
:
6987
–6998.
Meyer G, Soria JM, Martínez-Galán JR, Martín-Clemente B, Fairén A (
1998
) Different origins and developmental histories of transient neurons in the marginal zone of the fetal and neonatal rat cortex.
J Comp Neurol
 
397
:
493
–518.
Miller MW (
1985
) Cogeneration of retrogradely labeled corticocortical projection and GABA-immunoreactive local circuit neurons in cerebral cortex.
Brain Res
 
355
:
187
–192.
Miller MW, Nowakowski RS (
1988
) Use of bromodeoxyuridine-immunohistochemistry to examine the proliferation, migration, and time of origin of cells in the central nervous system.
Brain Res
 
457
:
44
–52.
Mione MC, Danevic C, Boardman P, Harris B, Parnavelas JG (
1994
) Lineage analysis reveals neurotransmitter (GABA or glutamate) but not calcium-binding protein homogeneity in clonally related cortical neurons.
J Neurosci
 
14
:
107
–123.
Monaghan AP, Bock D, Gass P, Schwager A, Wolfer DP, Lipp HP, Schutz G (
1997
) Defective limbic system in mice lacking the tailless gene.
Nature
 
390
:
515
–517.
Muzio L, DiBenedetto B, Stoykova A, Boncinelli E, Gruss P, Mallamaci A (
2002
) Conversion of cerebral cortex into basal ganglia in Emx2(–/–) Pax6(Sey/Sey) double-mutant mice.
Nat Neurosci
 
5
:
737
–745.
Nery S, Fishell G, Corbin JG (
2002
) The caudal ganglionic eminence is a source of distinct cortical and subcortical cell populations
Nat Neurosci
 
5
:
1279
–1287.
Obst K, Bronzel M, Wahle P (
1998
) Visual activity is required to maintain the phenotype of supragranular NPY neurons in rat area 17.
Eur J Neurosci
 
10
:
1422
–1428.
Pappas IS, Parnavelas JG (
1998
) Basic fibroblast growth factor promotes the generation and differentiation of calretinin neurons in the rat cerebral cortex in vitro.
Eur J Neurosci
 
10
:
1436
–1445.
Parnavelas JG (
2000
) The origin and migration of cortical neurones: new vistas.
Trends Neurosci
 
23
:
126
–131.
Peduzzi JD (
1988
) Genesis of GABA-immunoreactive neurons in the ferret visual cortex.
J Neurosci
 
8
:
920
–931.
Pleasure SJ, Anderson S, Hevner R, Bagri A, Marin O, Lowenstein DH, Rubenstein JL (
2000
) Cell migration from the ganglionic eminences is required for the development of hippocampal GABAergic interneurons.
Neuron
 
28
:
727
–740.
Polleux F, Whitford KL, Dijkhuizen PA, Vitalis T, Ghosh A (
2002
) Control of cortical interneuron migration by neurotrophins and PI3-kinase signaling.
Development
 
129
:
3147
–3160.
Porter JT, Cauli B, Staiger JF, Lambolez B, Rossier J, Audinat E (
1998
) Properties of bipolar VIPergic interneurons and their excitation by pyramidal neurons in the rat neocortex.
Eur J Neurosci
 
10
:
3617
–3628.
Powell EM, Mars WM, Levitt P (
2001
) Hepatocyte growth factor/scatter factor is a motogen for interneurons migrating from the ventral to dorsal telencephalon.
Neuron
 
30
:
79
–89.
Raballo R, Rhee J, Lyn-Cook R, Leckman JF, Schwartz ML, Vaccarino FM (
2000
) Basic fibroblast growth factor (Fgf2) is necessary for cell proliferation and neurogenesis in the developing cerebral cortex.
J Neurosci
 
20
:
5012
–5023.
Ramon y Cajal S (1911) Histology of the nervous system. New York: Oxford University Press.
Rogers JH (
1992
) Immunohistochemical markers in rat cortex: co-localization of calretinin and calbindin-D28k with neuropeptides and GABA.
Brain Res
 
587
:
147
–157.
Sarkisian MR, Rattan S, D’Mello SR, LoTurco JJ (
1999
) Characterization of seizures in the flathead rat: a new genetic model of epilepsy in early postnatal development.
Epilepsia
 
40
:
394
–400.
Sarkisian MR, Frenkel M, Li W, Oborski JA, LoTurco JJ (
2001
) Altered interneuron development in the cerebral cortex of the flathead mutant.
Cereb Cortex
 
11
:
734
–743.
Sarkisian MR, Li W, Di Cunto F, D’Mello SR, LoTurco JJ (
2002
) Citronkinase, a protein essential to cytokinesis in neuronal progenitors, is deleted in the flathead mutant rat.
J Neurosci
 
22
:
RC217
.
Shinozaki K, Miyagi T, Yoshida M, Miyata T, Ogawa M, Aizawa S, Suda Y (
2002
) Absence of Cajal–Retzius cells and subplate neurons associated with defects of tangential cell migration from ganglionic eminence in Emx1/2 double mutant cerebral cortex.
Development
 
129
:
3479
–3492.
Stuhmer T, Puelles L, Ekker M, Rubenstein JL (
2002
) Expression from a Dlx gene enhancer marks adult mouse cortical GABAergic neurons.
Cereb Cortex
 
12
:
75
–85.
Sussel L, Marin O, Kimura S, Rubenstein JL (
1999
) Loss of Nkx2.1 homeobox gene function results in a ventral to dorsal molecular respecification within the basal telencephalon: evidence for a transformation of the pallidum into the striatum.
Development
 
126
:
3359
–3370.
Volk DW, Pierri JN, Fritschy JM, Auh S, Sampson AR, Lewis DA (
2002
) Reciprocal alterations in pre- and postsynaptic inhibitory markers at chandelier cell inputs to pyramidal neurons in schizophrenia.
Cereb Cortex
 
12
:
1063
–1070.
Wahle P, Gorba T, Wirth MJ, Obst-Pernberg K (
2000
) Specification of neuropeptide Y phenotype in visual cortical neurons by leukemia inhibitory factor.
Development
 
127
:
1943
–1951.
Wichterle H, Garcia-Verdugo JM, Herrera DG, Alvarez-Buylla A (
1999
) Young neurons from medial ganglionic eminence disperse in adult and embryonic brain.
Nat Neurosci
 
2
:
461
–466.
Wichterle H, Turnbull DH, Nery S, Fishell G, Alvarez-Buylla A (
2001
) In utero fate mapping reveals distinct migratory pathways and fates of neurons born in the mammalian basal forebrain.
Development
 
128
:
3759
–3771.
Woo TU, Whitehead RE, Melchitzky DS, Lewis DA (
1998
) A subclass of prefrontal gamma-aminobutyric acid axon terminals are selectively altered in schizophrenia.
Proc Natl Acad Sci USA
 
95
:
5341
–5346.