Margaret Martínez-de-la-Torre

A Segmental Map of Architectonic Subdivisions in the Diencephalon of the Frog Rana perezi: Acetylcholinesterase-Histochemical Observations; pp. 279–289

The work examines frog diencephalic subdivisions from a segmental viewpoint and adds a number of details to the atlas of the bullfrog diencephalon reported by Neary and Northcutt [1983]. Acetylcholinesterase histochemistry was performed on brains of Rana perezi, sectioned either sagittally or in a plane roughly parallel to the optic tract to optimize detection of segmented landmarks. This material provided a very detailed picture of individual neurons, neuropils and some fiber tracts expressing the enzyme in diverse patterns characteristic for each diencephalic region. The main diencephalic areas previously recognized in the bullfrog appeared subdivided into smaller AChE-chemoarchitectonic units. Modified subdivisions are proposed for several entities: preoptic, suprachiasmatic, entopeduncular, ventral thalamic, anterior thalamic, pretectal, hypothalamic and tuberculum posterior regions. A number of cell groups are described for the first time in frogs. This mapping is expected to be useful for the interpretation of immunocytochemical and experimental hodologic results in the diencephalon of frogs and opens new possibilities for comparative analysis.

Morphologic fate of diencephalic prosomeres and their subdivisions revealed by mapping cadherin expression.

The expression of four cadherins (cadherin‐6B, cadherin‐7, R‐cadherin, and N‐cadherin) was mapped in the diencephalon of chicken embryos at 11 days and 15 days of incubation and was compared with Nissl stains and radial glial topology. Results showed that each cadherin is expressed in a restricted manner by a different set of embryonic divisions, brain nuclei, and their subregions. An analysis of the segmental organization based on the prosomeric model indicated that, in the mature diencephalon, each prosomere persists and forms a coherent domain of gray matter extending across the entire transverse dimension of the neural tube, from the ventricular surface to the pial surface. Moreover, the results suggest the presence of a novel set of secondary subdivisions for the dorsal thalamus (dorsal, intermediate, and ventral tiers and anteroventral subregion). They also confirm the presence of secondary subdivisions in the pretectum (commissural, juxtacommissural, and precommissural). At most of the borders between the prosomeres and their secondary subdivisions, changes in radial glial fiber density were observed. The diencephalic brain nuclei that derive from each of the subdivisions were determined. In addition, a number of previously less well‐characterized gray matter regions of the diencephalon were defined in more detail based on the mapping of cadherin expression. The results demonstrate in detail how the divisions of the early embryonic diencephalon persist and transform into mature gray matter architecture during brain morphogenesis, and they support the hypothesis that cadherins play a role in this process by providing a framework of potentially adhesive specificities. J. Comp. Neurol. 421:481–514, 2000.

Conserved pattern of OTP-positive cells in the paraventricular nucleus and other hypothalamic sites of tetrapods.

The paraventricular nucleus complex (Pa) is a component of central neural circuitry that regulates several homeostatic variables. The paraventricular nucleus is composed of magnocellular neurons that project to the posterior pituitary and parvicellular neurons that project to numerous sites in the central nervous system. According to the revised prosomeric model, the paraventricular nucleus is located caudal to the eye stalk along the rostrocaudal dimension of the dorsal hypothalamic alar plate. Caudally, the paraventricular nucleus abuts the prethalamus (prosomere 3), and the entire complex is flanked ventrally and dorsally by Dlx5-expressing domains of the alar plate. The homeodomain transcription factor Orthopedia (Otp) is expressed in several separate hypothalamic sites: the paraventricular nucleus, perimammillary region and arcuate nucleus. In this study, we compared Otp expression in the hypothalamus of mouse (Mus musculus), chick (Gallus gallus), frog (Rana perezi) and axolotol (Ambystoma mexicanum), using immunohistochemical and in situ hybridization techniques. In all cases, Otp-positive cells in the paraventricular nucleus were excluded from Dlx5-expressing adjacent domains. Other positive neuronal populations were observed in the arcuate nucleus and oblique perimammillary band. Expression in the medial amygdala appears to be continuous with the Otp-expressing paraventricular nucleus complex. This area is relatively unevaginated in the amphibian brains, barely evaginated in the chick, and fully evaginated in the mouse. These data led us to conclude that the expression pattern of Otp is topologically highly conserved in tetrapods and is plesiomorphic among chordates.

3 dimensional modelling of early human brain development using optical projection tomography.

BackgroundAs development proceeds the human embryo attains an ever more complex three dimensional (3D) structure. Analyzing the gene expression patterns that underlie these changes and interpreting their significance depends on identifying the anatomical structures to which they map and following these patterns in developing 3D structures over time. The difficulty of this task greatly increases as more gene expression patterns are added, particularly in organs with complex 3D structures such as the brain. Optical Projection Tomography (OPT) is a new technology which has been developed for rapidly generating digital 3D models of intact specimens. We have assessed the resolution of unstained neuronal structures within a Carnegie Stage (CS)17 OPT model and tested its use as a framework onto which anatomical structures can be defined and gene expression data mapped.ResultsResolution of the OPT models was assessed by comparison of digital sections with physical sections stained, either with haematoxylin and eosin (H&E) or by immunocytochemistry for GAP43 or PAX6, to identify specific anatomical features. Despite the 3D models being of unstained tissue, peripheral nervous system structures from the trigeminal ganglion (~300 μm by ~150 μm) to the rootlets of cranial nerve XII (~20 μm in diameter) were clearly identifiable, as were structures in the developing neural tube such as the zona limitans intrathalamica (core is ~30 μm thick). Fourteen anatomical domains have been identified and visualised within the CS17 model. Two 3D gene expression domains, known to be defined by Pax6 expression in the mouse, were clearly visible when PAX6 data from 2D sections were mapped to the CS17 model. The feasibility of applying the OPT technology to all stages from CS12 to CS23, which encompasses the major period of organogenesis for the human developing central nervous system, was successfully demonstrated.ConclusionIn the CS17 model considerable detail is visible within the developing nervous system at a minimum resolution of ~20 μm and 3D anatomical and gene expression domains can be defined and visualised successfully. The OPT models and accompanying technologies for manipulating them provide a powerful approach to visualising and analysing gene expression and morphology during early human brain development.

Gene Maps and Related Histogenetic Domains in the Forebrain and Midbrain

Rodent brain gene patterns are readily comparable with counterparts in the human, avian, reptilian, amphibian, teleost, shark and agnathan species. This scenario provides substantial novel evidence for comparative neuroanatomy, both corroborating some earlier conclusions and calling for revision of other concepts. In recent years accumulated observations have shown that gene expression patterns frequently display reproducible boundaries; these tend to be aligned with the axial and dorsoventral dimensions of the neural tube and are topologically invariant during ontogenesis. Many early molecular patterns are remarkably resistant to evolutionary change. Accumulated comparative results over the last decade strongly indicate that there is a common pattern of differentially-specified neural regions among all vertebrates. This chapter studies the molecular approaches to understanding the structural and functional organization of the nervous system which promise new insights into modular brain domains conserved among various species. Fate-map analysis suggests that early differential molecular specification of progenitor regions and subregions in the neural plate and neural tube eventually correlates with specific prospective fates via patterned proliferation, neurogenesis and differentiation.

Development of the serotonergic cells in murine raphe nuclei and their relations with rhombomeric domains

The raphe nuclei represent the origin of central serotonergic projections. The literature distinguishes seven nuclei grouped into rostral and caudal clusters relative to the pons. The boundaries of these nuclei have not been defined precisely enough, particularly with regard to developmental units, notably hindbrain rhombomeres. We hold that a developmental point of view considering rhombomeres may explain observed differences in connectivity and function. There are twelve rhombomeres characterized by particular genetic profiles, and each develops between one and four distinct serotonergic populations. We have studied the distribution of the conventional seven raphe nuclei among these twelve units. To this aim, we correlated 5-HT-immunoreacted neurons with rhombomeric boundary landmarks in sagittal mouse brain sections at different developmental stages. Furthermore, we performed a partial genoarchitectonic analysis of the developing raphe nuclei, mapping all known serotonergic differentiation markers, and compared these results, jointly with others found in the literature, with our map of serotonin-containing populations, in order to examine regional variations in correspondence. Examples of regionally selective gene patterns were identified. As a result, we produced a rhombomeric classification of some 45 serotonergic populations, and suggested a corresponding modified terminology. Only a minor rostral part of the dorsal raphe nucleus lies in the midbrain. Some serotonergic neurons were found in rhombomere 4, contrary to the conventional assumption that it lacks such neurons. We expect that our reclassification of raphe nuclei may be useful for causal analysis of their differential molecular specification, as well as for studies of differential connectivity and function.

Genoarchitectonic profile of developing nuclear groups in the chicken pretectum.

Earlier results on molecularly coded progenitor domains in the chicken pretectum revealed an anteroposterior subdivision of the pretectum in precommissural (PcP), juxtacommissural (JcP), and commissural (CoP) histogenetic areas, each specified differentially (Ferran et al. [2007] J Comp Neurol 505:379–403). Here we examined the nuclei derived from these areas with regard to characteristic gene expression patterns and gradual histogenesis (eventually, migration patterns). We sought a genoarchitectonic schema of the avian pretectum within the prosomeric model of the vertebrate forebrain (Puelles and Rubenstein [2003] Trends Neurosci 26:469–476; Puelles et al. [2007] San Diego: Academic Press). Transcription‐factor gene markers were used to selectively map derivatives of the three pretectal histogenetic domains: Pax7 and Pax6 (CoP); FoxP1 and Six3 (JcP); and FoxP2, Ebf1, and Bhlhb4 (PcP). The combination of this genoarchitectonic information with additional data on Lim1, Tal2, and Nbea mRNA expression and other chemoarchitectonic results allowed unambiguous characterization of some 30 pretectal nuclei. Apart from grouping them as derivatives of the three early anteroposterior domains, we also assigned them to postulated dorsoventral subdomains (Ferran et al. [2007]). Several previously unknown neuronal populations were detected, thus expanding the list of pretectal structures, and we corrected some apparently confused concepts in the earlier literature. The composite gene expression map represents a substantial advance in anatomical and embryological knowledge of the avian pretectum. Many nuclear primordia can be recognized long before the mature differentiated state of the pretectum is achieved. This study provides fundamental notions for ultimate scientific study of the specification and regionalization processes building up this brain area, both in birds and other vertebrates. J. Comp. Neurol. 517:405–451, 2009.

Early pretectal gene expression pattern shows a conserved anteroposterior tripartition in mouse and chicken

A changing network of gene activity settles the molecular basis of regionalization in the nervous system. As a consequence, analysis of combined gene expressions patterns represents a powerful initial approach to decode the complex process that drives neurohistogenesis and generates distinct morphological features. We have started to do a comparative screening of molecular regionalization in the mouse and chicken pretectal region at selected developmental stages. The pretectal region is composed of alar and roof plate derivatives of prosomere 1. This is a poorly understood region, best characterized in avian embryos and adults because nuclear cytoarchitectonic delimitation is clearer in these animals. During the early regionalization process the main pretectal boundaries and histogenetic/progenitor domains are established. We explore here Pax3, Pax6 and Six3 mRNA expression (and PAX3 immunoreactivity) in both chicken and mice, with the aim to compare their respective patterns. Our focus is centered on stages HH22-HH24 in chicken and embryonic days E11.5-E12.5 in mice. We found that, in both vertebrates, the same three main anteroposterior subdivisions are distinguished by these markers. They were defined as precommissural, juxtacommissural and commissural pretectal domains. These preliminary data represent an initial scaffold to explore more detailed pretectal regionalization processes and provide an important new key to approach unresolved pretectal homologies between vertebrates.

Chapter 8 – Hypothalamus

Publisher Summary Neuroanatomical concepts of the hypothalamus, the forebrain territory that controls homeostasis and drives in vertebrates, are over 100 years old. Neuroanatomists of the 19th century generally considered the hypothalamus to consist of the tuberal or infundibular area, a median prominence at the brain ventral surface at the location of the pituitary gland, framed in front and at the sides by the optic chiasm and tracts. The neural plate fate maps of amniotes and anamniotes are strictly comparable from a topological point of view. The relative positions of the prospective brain areas can be extrapolated from one species to another. The prospective longitudinal or axial dimension of the brain is best represented by the neural/non-neural border of the neural plate, which will transform into the longitudinal roof plate of the closed neural tube. This developmental analysis suggests that all adult median forebrain derivatives found between the anterior commissure and the mamillary area are equally rostralmost loci in the brain. At the open neural plate, the rostral forebrain primordium contains the prospective hypothalamus as a large area centered upon the alar, basal, and floor portions of the terminal wall, its caudal part covering as well a rostral part of the lateral wall, whereas the prospective telencephalon forms a thin band at the dorsal periphery of the alar portion of these domains, including the corresponding roof area. A handful of genes, coding either for transcription factors or secreted protein morphogens, participate in early patterning and regional identity specification along the AP and DV dimensions of the whole forebrain, including the hypothalamus. The complex networked interplay of many molecular signals that control brain development represents a dynamic ontogenetic system that tends to reach different equilibrium states in different parts of the neuroepithelial wall.

Distal-less-like protein distribution in the larval lamprey forebrain.

A polyclonal antibody against the Drosophila distal-less (DLL) protein, cross-reactive with cognate vertebrate proteins, was employed to map DLL-like expression in the midlarval lamprey forebrain. This work aimed to characterize in detail the separate diencephalic and telencephalic DLL expression domains, in order to test our previous modified definition of the lamprey prethalamus [Pombal and Puelles (1999) J Comp Neurol 414:391-422], adapt our earlier schema of prosomeric subdivisions in the lamprey forebrain to more recent versions of this model [Pombal et al. (2009) Brain Behav Evol 74:7-19] and reexamine the pallio-subpallial regionalization of the lamprey telencephalon. We observed a large-scale conservation of the topologic distribution of the DLL protein, in consonance with patterns of Dlx expression present in other vertebrates studied. Moreover, evidence was obtained of substantial numbers of DLL-positive neurons in the olfactory bulb and the cerebral hemispheres, in a pattern consistent with possible tangential migration out of the subpallium into the overlying pallium, as occurs in mammals, birds, frogs and teleost fishes.