Luis Puelles

Pallial and subpallial derivatives in the embryonic chick and mouse telencephalon, traced by the expression of the genes Dlx-2, Emx-1, Nkx-2.1, Pax-6, and Tbr-1

Pallial and subpallial morphological subdivisions of the developing chicken telencephalon were examined by means of gene markers, compared with their expression pattern in the mouse. Nested expression domains of the genes Dlx‐2 and Nkx‐2.1, plus Pax‐6‐expressing migrated cells, are characteristic for the mouse subpallium. The genes Pax‐6, Tbr‐1, and Emx‐1 are expressed in the pallium. The pallio‐subpallial boundary lies at the interface between the Tbr‐1 and Dlx‐2 expression domains. Differences in the expression topography of Tbr‐1 and Emx‐1 suggest the existence of a novel “ventral pallium” subdivision, which is an Emx‐1‐negative pallial territory intercalated between the striatum and the lateral pallium. Its derivatives in the mouse belong to the claustroamygdaloid complex. Chicken genes homologous to these mouse genes are expressed in topologically comparable patterns during development. The avian subpallium, called “paleostriatum,” shows nested Dlx‐2 and Nkx‐2.1 domains and migrated Pax‐6‐positive neurons; the avian pallium expresses Pax‐6, Tbr‐1, and Emx‐1 and also contains a distinct Emx‐1‐negative ventral pallium, formed by the massive domain confusingly called “neostriatum.” These expression patterns extend into the septum and the archistriatum, as they do into the mouse septum and amygdala, suggesting that the concepts of pallium and subpallium can be extended to these areas. The similarity of such molecular profiles in the mouse and chicken pallium and subpallium points to common sets of causal determinants. These may underlie similar histogenetic specification processes and field homologies, including some comparable connectivity patterns. J. Comp. Neurol. 424:409–438, 2000.

Expression patterns of homeobox and other putative regulatory genes in the embryonic mouse forebrain suggest a neuromeric organization

The molecular mechanisms that control regional specification, morphogenesis and differentiation of the embryonic forebrain are not known, although recently several laboratories have isolated homeobox, Wnt and other genes that are candidates for playing roles in these processes. Most of these genes exhibit temporally and spatially restricted patterns of expression within the forebrain. However, analysis of the spatial patterns has been complicated because an understanding of the organization of the embryonic forebrain has been lacking. This article describes a neuromeric model of the forebrain that is consistent with the expression patterns of these genes, and that provides a framework for understanding the morphological relationships within this complex structure.

Forebrain gene expression domains and the evolving prosomeric model

The prosomeric model attributes morphological meaning to gene expression patterns and other data in the forebrain. It divides this territory into the same transverse segments (prosomeres) and longitudinal zones in all vertebrates. The axis and longitudinal zones of this model are widely accepted but controversy subsists about the number of prosomeres and their nature as segments. We describe difficulties encountered in establishing continuity between prosomeric limits postulated in the hypothalamus and intra-telencephalic limits. Such difficulties throw doubt on the intersegmental nature of these limits. We sketch a simplified model, in which the secondary prosencephalon (telencephalon plus hypothalamus) is a complex protosegment not subdivided into prosomeres, which exhibits patterning singularities. By contrast, we continue to postulate that prosomeres p1-p3 (i.e. the pretectum, thalamus and prethalamus) are the caudal forebrain.

Delineation of Multiple Subpallial Progenitor Domains by the Combinatorial Expression of Transcriptional Codes

The mammalian telencephalon is considered the most complex of all biological structures. It comprises a large number of functionally and morphologically distinct types of neurons that coordinately control most aspects of cognition and behavior. The subpallium, for example, not only gives rise to multiple neuronal types that form the basal ganglia and parts of the amygdala and septum but also is the origin of an astonishing diversity of cortical interneurons. Despite our detailed knowledge on the molecular, morphological, and physiological properties of most of these neuronal populations, the mechanisms underlying their generation are still poorly understood. Here, we comprehensively analyzed the expression patterns of several transcription factors in the ventricular zone of the developing subpallium in the mouse to generate a detailed molecular map of the different progenitor domains present in this region. Our study demonstrates that the ventricular zone of the mouse subpallium contains at least 18 domains that are uniquely defined by the combinatorial expression of several transcription factors. Furthermore, the results of microtransplantation experiments in vivo corroborate that anatomically defined regions of the mouse subpallium, such as the medial ganglionic eminence, can be subdivided into functionally distinct domains.

T-Brain-1: A homolog of Brachyury whose expression defines molecularly distinct domains within the cerebral cortex

The mechanisms that regulate regional specification and evolution of the cerebral cortex are obscure. To this end, we have identified and characterized a novel murine and human gene encoding a putative transcription factor related to the Brachyury (T) gene that is expressed only in postmitotic cells. T-brain-1 (Tbr-1) mRNA is largely restricted to the cerebral cortex, where during embryogenesis it distinguishes domains that we propose may give rise to paleocortex, limbic cortex, and neocortex. Tbr-1 and Id-2 expression in the neocortex have discontinuities that define molecularly distinct neocortical areas. Tbr-1 expression is analyzed in the context of the prosomeric model. Topological maps are proposed for the organization of the dorsal telencephalon.

Morphological Fate of Rhombomeres in Quail/Chick Chimeras: A Segmental Analysis of Hindbrain Nuclei

Quail rhombomeres two to six (r2‐r6) were individually grafted homotopically into the hindbrain of chick embryos at 2 days of incubation. Nine to 10 days after the operation the chimeric embryos were fixed and processed for parallel cytoarchitectural and immunocytochemical study (with an anti‐quail antibody) in order to map the anatomical fate of the grafted tissue. Emphasis was placed on conventionally identified and distinct neuronal populations composing the sensory and motor longitudinal columns. Grafted rhombomeres consistently developed as complete transverse slices of the chimeric hindbrain. Interrhombomeric cell migration was either sparse or restricted to specific nuclei. The cranial nerve motor nuclei showed rhombomeric origins consistent with the patterns described in early embryos. Unexpectedly, alar r2 was found to form the auricular part of the cerebellum. As regards the cochlear nuclei, we found that nucleus angularis derives from r3 to r6, nucleus laminaris from r5 to r6, nucleus magnocellularis from r6 to r7 and nucleus olivaris superior from r5. The nuclei of the lateral lemniscus originated between r1 and r3. We also delimited the respective rhombomeric subdivisions of the sensory vestibular and trigeminal columns, both of which extend from r1 caudalwards throughout the hindbrain. There were consistently some interrhombomeric neuronal migrations inside the vestibular column, some motor nuclei and the reticular formation, involving only one rhombomere length. The pontine nuclei, which extended from r1 to r7, showed neuronal migrations that crossed several rhombomeres. On the whole, these results represent the first anatomical analysis of the mature avian hindbrain in terms of rhombomere‐derived domains.

DLX‐1, DLX‐2, and DLX‐5 expression define distinct stages of basal forebrain differentiation

The homeobox genes in the Dlx family are required for differentiation of basal forebrain neurons and craniofacial morphogenesis. Herein, we studied the expression of Dlx‐1, Dlx‐2, and Dlx‐5 RNA and protein in the mouse forebrain from embryonic day 10.5 (E10.5) to E12.5. We provide evidence that Dlx‐2 is expressed before Dlx‐1, which is expressed before Dlx‐5. We also demonstrate that these genes are expressed in the same cells, which may explain why single mutants of the Dlx genes have mild phenotypes. The DLX proteins are localized primarily to the nucleus, although DLX‐5 also can be found in the cytoplasm. During development, the fraction of Dlx‐positive cells increases in the ventricular zone. Analysis of the distribution of DLX‐1 and DLX‐2 in M‐phase cells suggests that these proteins are distributed symmetrically to daughter cells during mitosis. We propose that DLX‐negative cells in the ventricular zone are specified progressively to become DLX‐2‐expressing cells during neurogenesis; as these cells differentiate, they go on to express DLX‐1, DLX‐5, and DLX‐6. This process appears to be largely the same in all regions of the forebrain that express the Dlx genes. In the basal telencephalon, these DLX‐positive cells differentiate into projection neurons of the striatum and pallidum as well as interneurons, some of which migrate to the cerebral cortex and the olfactory bulb. J. Comp. Neurol. 414:217–237, 1999.

The mouse Dlx-2 (Tes-1) gene is expressed in spatially restricted domains of the forebrain, face and limbs in midgestation mouse embryos.

The pattern of RNA expression of the murine Dlx-2 (Tes-1) homeobox gene is described in embryos ranging in age from E8.5 through E11.5. Dlx-2 is a vertebrate homologue of the Drosophila Distal-less (Dll) gene. Dll expression in the Drosophila embryo is principally limited to the primordia of the brain, head and limbs. Dlx-2 is also expressed principally in the primordia of the forebrain, head and limbs. Within these regions it is expressed in spatially restricted domains. These include two discontinuous regions of the forebrain (basal telencephalon and ventral diencephalon), the branchial arches, facial ectoderm, cranial ganglia and limb ectoderm. Several mouse and human disorders have phenotypes which potentially are the result of mutations in the Dlx genes.

Expression of Dbx1, Neurogenin 2, Semaphorin 5A, Cadherin 8, and Emx1 Distinguish Ventral and Lateral Pallial Histogenetic Divisions in the Developing Mouse Claustroamygdaloid Complex

We studied the lateral and ventral pallial divisions of the claustroamygdaloid complex by means of analysis of expression patterns of the developmental regulatory genes Tbr1, Dbx1, Neurogenin 2, Emx1, Cadherin 8, and Semaphorin 5A in mouse developing telencephalon, from embryonic day 12.5 until birth. Our results indicate that these genes help to distinguish distinct lateral and ventral pallial histogenetic divisions in the embryonic telencephalon. Tbr1 is broadly expressed in both lateral and ventral pallial histogenetic divisions (the lateroventral migratory stream plus the mantle) during early and intermediate embryonic development; its signal becomes weak in parts of the mantle during late embryonic development. Dbx1 is strongly and specifically expressed in progenitor cells (ventricular zone) of the ventral pallium during early embryonic development, but there is no signal of this gene in the rest of the pallium nor the subpallium. Neurogenin 2 and Semaphorin 5A are both expressed in a ventral subdivision of the lateroventral migratory stream (called by us the ventral migratory stream). Further, specific nuclei of the claustral complex and pallial amygdala show strong expression of Neurogenin 2 and/or Semaphorin 5A, including the ventromedial claustrum and endopiriform nuclei, the lateral and basomedial amygdalar nuclei, the anterior and posteromedial cortical amygdalar areas, plus the amygdalo‐hippocampal area. We interpret these nuclei or areas of the claustroamygdaloid complex as possible derivatives of the ventral pallium. In contrast, during embryonic development the dorsolateral claustrum, the basolateral amygdalar nucleus, and the posterolateral cortical amygdalar area do not express or show weak expression of Neurogenin 2 or Semaphorin 5A, but express selectively and strongly Cadherin 8 plus Emx1, and may be derivatives of the lateral pallium. The lateral pallial and ventral pallial divisions of the claustroamygdaloid complex appear to have some different sets of connections, although this requires further investigation. J. Comp. Neurol. 474:504–523, 2004.

Postembryonic neural proliferation in the zebrafish forebrain and its relationship to prosomeric domains

 Large gaps of knowledge exist regarding postembryonic brain morphogenesis of the zebrafish Danio rerio (Cyprinidae, Teleostei). The zebrafish represents – together with the frog (Xenopus), chick and mouse – one of four major models for the genetic study of early brain development. Here, we used normal silver-stained Bodian material and immunohistochemical material stained with a monoclonal antibody against the proliferating cell nuclear antigen (PCNA, cyclin) to study the morphogenetic appearance and location of proliferation zones of the zebrafish brain between day 1 and day 10, focussing on the forebrain at day 5 postfertilization. Our results directly demonstrate that the dorsal telencephalic proliferation zone (i.e. the pallium) extends – consistent with the process of eversion – some distance laterally on top of the telencephalon. The subpallial telencephalic proliferation consists of dorsal and ventral zones. The preoptic region also includes dorsal and ventral proliferation zones. In the diencephalon proper, separate proliferation zones are present in the habenula, and in the periventricular cell masses of the dorsal thalamus, the ventral thalamus, and the pretectum. More ventrocaudally, the latter three massive proliferation zones appear to be replaced each by thinner, but distinct proliferation zones. Two of them represent ventrocaudal continuations of the dorsal and ventral thalamus and lie in the region referred to as the posterior tubercular area in adult teleostean neuroanatomy. The third lies in the region of the nucleus of the medial longitudinal fascicle. In addition, several hypothalamic proliferation zones are present. The data for the diencephalon are largely in agreement with the neuromeric model of brain organization of Puelles and Rubenstein (1993), which is mostly based on amniote data. Generally, the understanding of the prosomeric origin of teleostean prosencephalic cell masses may be regarded as pivotal for their comparative interpretation.