[Show abstract][Hide abstract] ABSTRACT: Many of the genes involved in brain patterning during development are highly conserved in vertebrates and similarities in their expression patterns help to recognize homologous cell types or brain regions. Among these genes, Pax6 and Pax7 are expressed in regionally restricted patterns in the brain and are essential for its development. In the present immunohistochemical study we analyzed the distribution of Pax6 and Pax7 cells in the brain of six representative species of tetrapods and lungfishes, the closest living relatives of tetrapods, at several developmental stages. The distribution patterns of these transcription factors were largely comparable across species. In all species only Pax6 was expressed in the telencephalon, including the olfactory bulbs, septum, striatum, and amygdaloid complex. In the diencephalon, Pax6 and Pax7 were distinct in the alar and basal parts, mainly in prosomeres 1 and 3. Pax7 specifically labeled cells in the optic tectum (superior colliculus) and Pax6, but not Pax7, cells were found in the tegmentum. Pax6 was found in most granule cells of the cerebellum and Pax7 labeling was detected in cells of the ventricular zone of the rostral alar plate and in migrated cells in the basal plate, including the griseum centrale and the interpeduncular nucleus. Caudally, Pax6 cells formed a column, whereas the ventricular zone of the alar plate expressed Pax7. Since the observed Pax6 and Pax7 expression patterns are largely conserved they can be used to identify subdivisions in the brain across vertebrates that are not clearly discernible with classical techniques.
Frontiers in Neuroanatomy 08/2014; 8(75):1-20. · 4.06 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: Pax6 and Pax7 are transcription factors essential for the development of the CNS. In addition, increasing data, mainly obtained in amniotes, support that they are expressed in subsets of neurons in the adult, likely playing a role in maintaining neuron type identity. In the present study we analyzed the detailed distribution of Pax6 and Pax7 cells in the adult CNS of Xenopus laevis. Immunohistochemistry with antibodies that are required for high-resolution analysis of Pax-expressing cells was conducted. A wide distribution of Pax6 and Pax7 cells throughout the CNS was detected, with distinct patterns that showed only slight overlapping. Only Pax6 was expressed in the telencephalon, including the olfactory bulbs, septum, striatum and amygdaloid complex. In the diencephalon, Pax6 and Pax7 were distinct in the alar and basal parts, respectively, of prosomere 3. Large numbers of Pax6 and Pax7 cells were distributed in the pretectal region (alar plate of prosomere 1) but only Pax6 cells extended into basal plate. Pax7 specifically labeled cells in the optic tectum, including the ventricular zone, and Pax6 cells were the only cells found in the tegmentum. Pax6 was found in most granule cells of the cerebellum and Pax7 expression was found only in the ventricular zone. In the rostral rhombomere 1, Pax7 labeling was detected in cells of the ventricular zone of the alar plate, but numerous migrated cells were located in the basal plate, including the griseum centrale and the interpeduncular nucleus. Pax6 cells also formed a column of scattered neurons in the reticular formation and were found in the octavolateral area. The rhombencephalic ventricular zone of the alar plate expressed Pax7. Dorsal Pax7 cells and ventral Pax6 cells were found along the spinal cord separated from the ventricle, which did not show immunoreactivity. Our results show that the expression of Pax6 and Pax7 is widely maintained in the adult brain of Xenopus, like in urodele amphibians and in contrast to the situation described in amniotes. Therefore, in amphibians these transcription factors seem to be needed to maintain specific entities of subpopulations of neurons in the adult CNS.
Journal of chemical neuroanatomy 04/2014; · 1.75 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: The cholinergic system in the brain has been widely studied in most vertebrate groups, but there is no information available about this neurotransmission system in the brains of holostean fishes, a primitive and poorly understood group of actinopterygian fishes. The present study provides the first detailed information on the distribution of cholinergic cell bodies and fibers in the central nervous system in two holostean species, the Florida gar, Lepisosteus platyrhincus, and the bowfin, Amia calva. Immmunohistochemistry against the enzyme choline acetyltransferase (ChAT) revealed distinct groups of ChAT-immunoreactive (ChAT-ir) cells in the habenula, isthmic nucleus, laterodorsal tegmental nucleus, octavolateral area, reticular formation, cranial nerve motor nuclei and the motor column of the spinal cord, all of which seem to be highly conserved among vertebrates. Some ChAT-ir cells were detected in the basal telencephalon that appear in actinopterygians for the first time in the evolution of this neurotransmission system, whereas the remarkable cholinergic population in the optic tectum is a peculiar characteristic, the presence of which varies throughout evolution, although it is present in all teleosts studied. Abundant cholinergic fibers were found in the pretectal region and optic tectum, where they probably modulate vision, and in the hypothalamus and the interpeduncular neuropil. Some interspecific differences were also observed, such as the presence of ChAT-ir cells in the supraoptoparaventricular band only in Lepisosteus and in in the nucleus subglomerulosus only in Amia. In addition, ChAT-ir fibers in the olfactory bulb were detected only in Amia. Comparison of these results with those from other classes of vertebrates, and a segmental analysis to correlate cell populations, reveal that the pattern of the cholinergic system in holosteans is very close to that in ancestral actinopterygian fishes, as recently described in the bichir (Cladistia), although an important evolutionary novelty in holosteans is the presence of cholinergic cells in the basal telencephalon.
Brain Behavior and Evolution 03/2013; · 2.89 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: The patterns of expression of a set of conserved developmental regulatory transcription factors and neuronal markers were analyzed in the alar hypothalamus of Xenopus laevis throughout development. Combined immunohistochemical and in situ hybridization techniques were used for the identification of subdivisions and their boundaries. The alar hypothalamus was located rostral to the diencephalon in the secondary prosencephalon and represents the rostral continuation of the alar territories of the diencephalon and brainstem, according to the prosomeric model. It is composed of the supraoptoparaventricular (dorsal) and the suprachiasmatic (ventral) regions, and limits dorsally with the preoptic region, caudally with the prethalamic eminence and the prethalamus, and ventrally with the basal hypothalamus. The supraoptoparaventricular area is defined by the orthopedia (Otp) expression and is subdivided into rostral and caudal portions, on the basis of the Nkx2.2 expression only in the rostral portion. This region is the source of many neuroendocrine cells, primarily located in the rostral subdivision. The suprachiasmatic region is characterized by Dll4/Isl1 expression, and was also subdivided into rostral and caudal portions, based on the expression of Nkx2.1/Nkx2.2 and Lhx1/7 exclusively in the rostral portion. Both alar regions are mainly connected with subpallial areas strongly implicated in the limbic system and show robust intrahypothalamic connections. Caudally, both regions project to brainstem centers and spinal cord. All these data support that in terms of topology, molecular specification, and connectivity the subdivisions of the anuran alar hypothalamus possess many features shared with their counterparts in amniotes, likely controlling similar reflexes, responses, and behaviors.
The Journal of Comparative Neurology 03/2013; 521(4):Spc1. · 3.66 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: The LIM-homeodomain transcription factor Islet1 (Isl1) has been widely used as a marker of neuronal differentiation in the developing visual system of different classes of vertebrates, including mammals, birds, reptiles, and fish. In the present study, we analyzed the spatial and temporal distribution of Isl1-immunoreactive cells during Xenopus laevis retinal development and its relation to the formation of the retinal layers, and in combination with different markers of cell differentiation. The earliest Isl1 expression appeared at St29-30 in the cell nuclei of sparse differentiating neuroblasts located in the vitreal surface of the undifferentiated retina. At St35-36, abundant Isl1-positive cells accumulated at the vitreal surface of the neuroepithelium. As development proceeded and through the postmetamorphic juveniles, Isl1 expression was identified in subpopulations of ganglion cells and in subsets of amacrine, bipolar, and horizontal cells. These data together suggest a possible role for Isl1 in the early differentiation and maintenance of different retinal cell types, and Isl1 can serve as a specific molecular marker for the study of retinal cell specification in X. laevis.
The Scientific World Journal 01/2013; 2013:740420. · 1.73 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: Pax7 is a member of the highly conserved Pax gene family that is expressed in restricted zones of the central nervous system (CNS) during development, being involved in early brain regionalization and the maintenance of the regional identity. Using sensitive immunohistochemical techniques we have analyzed the spatiotemporal pattern of Pax7 expression in the brain of the anuran amphibian Xenopus laevis, during development. Pax7 expression was first detected in early embryos in the basal plate of prosomere 3, roof and alar plates of prosomere 1 and mesencephalon, and the alar plate of rhombomere 1. As development proceeded, Pax7 cells were observed in the hypothalamus close to the catecholaminergic population of the mammillary region. In the diencephalon, Pax7 was intensely expressed in a portion of the basal plate of prosomere 3, in the roof plate and in scattered cells of the thalamus in prosomere 2, throughout the roof of prosomere 1, and in the commissural and juxtacommissural domains of the pretectum. In the mesencephalon, Pax7 cells were localized in the optic tectum and, to a lesser extent, in the torus semicircularis. The rostral portion of the alar part of rhombomere 1, including the ventricular layer of the cerebellum, expressed Pax7 and, gradually, some of these dorsal cells were observed to populate ventrally the interpeduncular nucleus and the isthmus (rhombomere 0). Additionally, Pax7 positive cells were found in the ventricular zone of the ventral part of the alar plate along the rhombencephalon and the spinal cord. The findings show that the strongly conserved features of Pax7 expression through development shared by amniote vertebrates are also present in the anamniote amphibians as a common characteristic of the brain organization of tetrapods.
Frontiers in Neuroanatomy 01/2013; 7:48. · 4.06 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: The sequence of appearance of calretinin and calbindin-D28k immunoreactive (CRir and CBir, respectively) cells and fibers has been studied in the brain of the urodele amphibian Pleurodeles waltl. Embryonic, larval and juvenile stages were studied. The early expression and the dynamics of the distribution of CBir and CRir structures have been used as markers for developmental aspects of distinct neuronal populations, highlighting the accurate extent of many regions in the developing brain, not observed on the basis of cytoarchitecture alone. CR and, to a lesser extent, CB are expressed early in the central nervous system and show a progressively increasing expression from the embryonic stages throughout the larval life and, in general, the labeled structures in the developing brain retain their ability to express these proteins in the adult brain. The onset of CRir cells primarily served to follow the development of the olfactory bulbs, subpallium, thalamus, alar hypothalamus, mesencephalic tegmentum, and distinct cell populations in the rhombencephalic reticular formation. CBir cells highlighted the development of, among others, the pallidum, hypothalamus, dorsal habenula, midbrain tegmentum, cerebellum, and central gray of the rostral rhombencephalon. However, it was the relative and mostly segregated distribution of both proteins in distinct cell populations which evidenced the developing regionalization of the brain. The results have shown the usefulness in neuroanatomy of the analysis during development of the onset of CBir and CRir structures, but the comparison with previous data has shown extensive variability across vertebrate classes. Therefore, one should be cautious when comparing possible homologue structures across species only on the basis of the expression of these proteins, due to the variation of the content of calcium-binding proteins observed in well-established homologous regions in the brain of different vertebrates.
Brain Structure and Function 07/2012; · 7.84 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: Fluorescence microscope image of double immunhistochemistry for the detection of tyrosine hydroxylase (green) and the transcription factor Nkx2.1 (red) in a transverse section through the hypothalamus of the turtle, Pseudemys scripta. The catecholaminergic neurons in the suprachiasmatic nucleus are revealed by its content in tyrosine hydroxylase, whereas Nkx2.1 labels the ventricular and subventricular zones of this hypothalamic region. The Journal of Comparative Neurology, Volume 520, Number 3, pages 453-478.
The Journal of Comparative Neurology 02/2012; 520(3):Spc1. · 3.66 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: The patterns of distribution of a set of conserved brain developmental regulatory transcription factors and neuronal markers were analyzed in the hypothalamus of the juvenile turtle, Pseudemys scripta. Combined immunohistochemical techniques were used for the identification of the main boundaries and subdivisions in the optic, paraventricular, tuberal, and mammillary hypothalamic regions. The combination of Tbr1 and Pax6 with Nkx2.1 allowed identification of the boundary between the telencephalic preoptic area, rich in Nkx2.1 expression, and the prethalamic eminence, rich in Tbr1 expression. In addition, at this level Nkx2.2 expression defined the boundary between the telencephalon and the hypothalamus. The dorsalmost hypothalamic domain was the supraoptoparaventricular region that was defined by the expression of Otp/Pax6 and the lack of Nkx2.1/Isl1. It is subdivided into rostral, rich in Otp and Nkx2.2, and caudal, only Otp-positive, portions. Ventrally, the suprachiasmatic area was identified by its catecholaminergic groups and the lack of Otp, and could be further divided into a rostral portion, rich in Nkx2.1 and Nkx2.2, and a caudal portion, rich in Isl1 and devoid of Nkx2.1 expression. The expressions of Nkx2.1 and Isl1 defined the tuberal hypothalamus, whereas only the rostral portion expressed Otp. Its caudal boundary was evident by the lack of Isl1 in the adjacent mammillary area, which expressed Nkx2.1 and Otp. All these results provide an important set of data on the interpretation of the hypothalamic organization in a reptile, and hence make a useful contribution to the understanding of hypothalamic evolution.
The Journal of Comparative Neurology 09/2011; 520(3):453-78. · 3.66 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: Lungfishes (dipnoans) are currently considered the closest living relatives of tetrapods. The organization of the cholinergic systems in the brain has been carefully analyzed in most vertebrate groups, and major shared characteristics have been described, although traits particular to each vertebrate class have also been found. In the present study, we provide the first detailed information on the distribution of cholinergic cell bodies and fibers in the central nervous system in two representative species of lungfishes, the African lungfish (Protopterus dolloi) and the Australian lungfish (Neoceratodus forsteri), as revealed by immunohistochemistry against the enzyme choline acetyltransferase (ChAT). Distinct groups of ChAT immunoreactive (ChAT-ir) cells were observed in the basal telencephalon, habenula, isthmic nucleus, laterodorsal tegmental nucleus, cranial nerve motor nuclei, and the motor column of the spinal cord, and these groups seem to be highly conserved among vertebrates. In lungfishes, the presence of a cholinergic cell group in the thalamus and the absence of ChAT-ir cells in the tectum are variable traits, unique to this group and appearing several times during evolution. Other characters were observed exclusively in Neoceratodus, such as the presence of cholinergic cells in the suprachiasmatic nucleus, the pretectal region and the superior raphe nucleus. Cholinergic fibers were found in the medial pallium, basal telencephalon, thalamus and prethalamus, optic tectum and interpeduncular nucleus. Comparison of these results with those from other classes of vertebrates, including a segmental analysis to correlate cell populations, reveals that the cholinergic systems in lungfishes largely resemble those of amphibians and other tetrapods.
Brain Structure and Function 08/2011; 217(2):549-76. · 7.84 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: Major common features have been reported for the organization of the basal telencephalon in amniotes, and most characteristics were thought to be acquired in the transition from anamniotes to amniotes. However, gene expression, neurochemical, and hodological data obtained for the basal ganglia and septal and amygdaloid complexes in amphibians (anamniotic tetrapods) have strengthened the idea of a conserved organization in tetrapods. A poorly characterized region in the forebrain of amniotes has been the bed nucleus of the stria terminalis (BST), but numerous recent investigations have characterized it as a member of the extended amygdala. Our study analyzes the main features of the BST in anuran amphibians to establish putative homologies with amniotes. Gene expression patterns during development identified the anuran BST as a subpallial, nonstriatal territory. The BST shows Nkx2.1 and Lhx7 expression and contains an Islet1-positive cell subpopulation derived from the lateral ganglionic eminence. Immunohistochemistry for diverse peptides and neurotransmitters revealed that the distinct chemoarchitecture of the BST is strongly conserved among tetrapods. In vitro tracing techniques with dextran amines revealed important connections between the BST and the central and medial amygdala, septal territories, medial pallium, preoptic area, lateral hypothalamus, thalamus, and prethalamus. The BST receives dopaminergic projections from the ventral tegmental area and is connected with the laterodorsal tegmental nucleus and the rostral raphe in the brainstem. All these data suggest that the anuran BST shares many features with its counterpart in amniotes and belongs to a basal continuum, likely controlling similar reflexes, reponses, and behaviors in tetrapods.
The Journal of Comparative Neurology 06/2011; 520(2):330-63. · 3.66 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: Networked gene activities control the evolutionarily conserved histogenetic organization of the central nervous system of vertebrates. Genoarchitectonic studies contribute to the analysis of each morphogenetic field by identifying distinct progenitor domains and corresponding derivatives whose pattern of gene expression shows a unique combinatory code. Previous studies in the pretectal region (caudal diencephalon) have defined three anteroposterior genoarchitectonic domains that are conserved in birds and mammals. Here, we have studied the embryonic pretectal genoarchitecture in the amphibian Xenopus laevis, in order to determine whether it is possible to define a comparable anteroposterior tripartition of the amphibian pretectal area. The expression patterns of 14 genes mapped from early embryonic stages to metamorphic climax allowed us to define the boundaries of the pretectum, the expected precommissural, juxtacommissural, and commissural anteroposterior domains, and some dorsoventral subdivisions. Taken together, our data provide evidence for a conserved pattern of pretectal domains and subdomains, shared by amniotes and amphibian anamniotes (tetrapods), understandable as part of a general Bauplan in vertebrates.
The Journal of Comparative Neurology 04/2011; 519(6):1024-50. · 3.66 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: The analysis of the distribution of the calbindin-D28k and calretinin immunoreactive (CBir and CRir) systems recently described in the brain of anuran and urodele amphibians was very useful for the interpretation of many otherwise indistinct brain regions and cell masses. In the present study we have followed a similar approach to investigate the distribution of CBir and CRir cell bodies and fibers in the brain of Dermophis mexicanus, a member of the much neglected third amphibian order of gymnophionans. The pattern of distribution obtained showed particular characteristics in Dermophis, such as the existence of abundant CRir elements in the olfactory bulbs and CBir and CRir cell populations in pallial areas. The distinct distribution of the two proteins allowed the tentative identification of currently described subregions, mainly in the amygdaloid complex and hypothalamic areas. The analysis of the diencephalon and brainstem distribution framed in the neuromeric model highlighted common traits with other amphibians but also specific features. Therefore, the immunohistochemical detection of calcium-binding proteins has served to discern cell populations and has helped to demonstrate neuronal heterogeneity. However, it should be pointed out that a straightforward comparison based only on the presence of these proteins should not be made due to the great variability observed in well-established homologous regions in the brain of different vertebrates, as evidenced within the class Amphibia.
Brain Behavior and Evolution 01/2011; 77(4):231-69. · 2.89 Impact Factor