Main
Results
Development and organization of the lamprey telencephalon with special reference to the GABAergic system
Pombal MA, Álvarez-Otero A, Pérez-Fernández J, Solveira C,
Megías M.
Frontiers Neuroanatomy 5:20.
Lampreys, together with hagfishes, represent the sister group of gnathostome vertebrates. There is an increasing interest for comparing the forebrain organization observed in lampreys and gnathostomes to shed light on vertebrate brain evolution. Within the prosencephalon, there is now a general agreement on the major subdivisions of the lamprey diencephalon; however, the organization of the telencephalon, and particularly its pallial subdivisions, is still a matter of controversy. In this study, recent progress on the development and organization of the lamprey telencephalon is reviewed, with particular emphasis on the GABA immunoreactive cell populations trying to understand their putative origin. First, we describe some early general cytoarchitectonic events by searching the classical literature as well as our collection of embryonic and prolarval series of hematoxylin-stained sections. Then, we comment on the cell proliferation activity throughout the larval period, followed by a detailed description of the early events on the development of the telencephalic GABAergic system. In this context, lampreys apparently do not possess the same molecularly distinct subdivisions of the gnathostome basal telencephalon because of the absence of a Nkx2.1-expressing domain in the developing subpallium; a fact that has been related to the absence of a medial ganglionic eminence as well as of its derived nucleus in gnathostomes, the pallidum. Therefore, these data raise interesting questions such as whether or not a different mechanism to specify telencephalic GABAergic neurons exists in lampreys or what are their migration pathways. Finally, we summarize the organization of the adult lamprey telencephalon by analyzing the main proposed conceptions, including the available data on the expression pattern of some developmental regulatory genes which are of importance for building its adult shape.
Distal-less-like protein distribution in the larval lamprey forebrain.
Martínez-de-la-Torre M, Pombal MA, Puelles L.
Neuroscience. 2011. 178:270-84.
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. Copyright © 2011 IBRO. Published by Elsevier Ltd. All rights reserved.
New and old thoughts on the segmental organization of the forebrain in lampreys
Pombal MA, Megías M, Bardet SM, Puelles L
Brain Behav Evol. 2009. 74:7-19
Ten years ago, we published the first detailed prosomeric map of the forebrain in lampreys. In the interim, the prosomeric model has been modified and simplified to better explain numerous data on the expression patterns of regulatory genes, as well as data from chemical, hodological and neuroembryological experiments, mostly in amniote vertebrates. In this report we first review the main historical con- cepts of lamprey forebrain organization, relating them to either columnar- or segmental-influenced models and explicit or implicit axial references. Next, our previous prosomeric model of the lamprey forebrain is updated, postulating some new hypotheses on the organization of the secondary prosencephalon. Copyright © 2009 S. Karger AG, Basel
Developmental changes of calretinin immunoreactivity in the lamprey spinal cord.
Viloria A, Rodríguez-Alonso M, Costas V, Pérez-Fernández J, Pombal MA, Megías M.
Brain Res Bull. 2008. 75(2-4):428-32.
We studied the distribution of calretinin immunoreactivity (CR-ir) in the rostral and intermediate levels of the spinal cord of lampreys from embryonic to adult periods. CR-ir was first observed at hatching in motoneurons and primary sensory neurons of the spinal cord, the dorsal cells. During the prolarval period two new cell types showed CR-ir: ganglion cells and interneurons. Motoneurons, dorsal cells, and ganglion cells were strongly positive, whereas interneurons were weakly stained in late prolarvae. The intensity of CR-ir in these four types of cells changed during the larval period. Increase of CR-expression was found in interneurons but a decrease in dorsal cells and in ganglion cells. These changes were more evident in premetamorphic larvae. Postmetamorphic lampreys showed almost no CR-ir in dorsal cells. In adult lampreys, the interneurons showed the highest CR-ir, whereas motoneurons were more weakly stained than in earlier stages of development. Moreover, in adults the dorsal cells and the ganglion cells showed no CR-ir. The present study shows that CR-ir changes during lamprey spinal cord development in different types of neurons, sometimes in opposite ways. This plasticity of CR-expression may indicate different needs from subsets of lamprey spinal cord cells involved in the different locomotor behaviors along its life cycle.
Distribution of adrenomedullin-like immunoreactivity in the brain of the adult sea lamprey.
Pombal MA, López JM, de Arriba Mdel C, González A, Megías M
Brain Res Bull. 2008. 75(2-4):261-5
Adrenomedullin (AM) is a neuropeptide widely distributed in vertebrates. In jawed vertebrates it has been localized in distinct regions of the central nervous system by means of antisera against human AM because the molecule seems to be well conserved across species. In this study, we have analyzed the localization of AM-like immunoreactive (AM-ir) cell bodies and fibers throughout the brain of the adult sea lamprey Petromyzon marinus, by using immunohistochemistry. Several AM-ir cell populations were found in the basal plate of the secondary prosencephalon, being more numerous in the hypothalamus, as well as two in the diencephalon and one in the mesencephalon; in addition two cell populations were found in the rhombencephalic alar plate, one in the isthmic region and other in the nucleus of the solitary tract. Immunolabeled fibers were widespread throughout the lamprey brain, but were more abundant in the basal plate. Of particular interest was the conspicuous innervation of the striatum by AM-ir fibers. In addition, our results indicate that AM-ir cells and fibers are present in the lamprey hypothalamo-neurohypophyseal system, suggesting that AM plays some important role in the control of pituitary gland function.
Epicardial development in lamprey supports an evolutionary origin of the vertebrate epicardium from an ancestral pronephric external glomerulus.
Pombal MA, Carmona R, Megías M, Ruiz A, Pérez-Pomares JM, Muñoz-Chápuli R.
Evol Dev. 2008. 10(2):210-216.
The epicardium is the outer layer of the vertebrate heart. Both the embryonic epicardium and its derived mesenchyme are critical to heart development, contributing to the coronary vasculature and modulating the proliferation of the ventricular myocardium. The embryonic epicardium arises from an extracardiac, originally paired progenitor tissue called the proepicardium, a proliferation of coelomic cells found at the limit between the liver and the sinus venosus. Proepicardial cells attach to and spread over the cardiac surface giving rise to the epicardium. Invertebrate hearts always lack of epicardium, and no hypothesis has been proposed about the origin of this tissue and its proepicardial progenitor in vertebrates. We herein describe the epicardial development in a representative of the most basal living lineage of vertebrates, the agnathan Petromyzon marinus (lamprey). The epicardium in lampreys develops by migration of coelomic cells clustered in a paired structure at the roof of the coelomic cavity, between the pronephros and the gut. Later on, these outgrowths differentiate into the pronephric external glomerulus (PEG), a structure composed of capillary networks, mesangial cells, and podocytes. This observation is consistent with the conclusion that the primordia of the most anterior pair of PEG in agnathans have been retained and transformed into the proepicardium in gnathostomes. Glomerular progenitor cells are highly vasculogenic and probably allowed for the vascularization of a cardiac tube primarily devoid of coronary vessels. This new hypothesis accounts for the striking epicardial expression of Wt1 and Pod1, two transcription factors essential for development of the excretory system.
Afferent connections of the optic tectum in lampreys: an experimental study.
de Arriba MA, Pombal MA
Brain Behav Evol. 2007;69(1):37-68
Tectal afferents were studied in adult lampreys of three species (Ichthyomyzon unicuspis, Lampetra fluviatilis, and Petromyzon marinus) following unilateral BDA injections into the optic tectum (OT). In the secondary prosencephalon, neurons projecting to the OT were observed in the pallium, the subhipoccampal lobe, the striatum, the preoptic area and the hypothalamus. Following tectal injections, backfilled diencephalic cells were found bilaterally in: prethalamic eminence, ventral geniculate nucleus, periventricular prethalamic nucleus, periventricular pretectal nucleus, precommissural nucleus, magnocellular and parvocellular nuclei of the posterior commissure and pretectal nucleus; and ipsilaterally in: nucleus of Bellonci, periventricular thalamic nucleus, nucleus of the tuberculum posterior, and the subpretectal tegmentum, as well as in the pineal organ. At midbrain levels, retrogradely labeled cells were seen in the ipsilateral torus semicircularis, the contralateral OT, and bilaterally in the mesencephalic reticular formation and inside the limits of the retinopetal nuclei. In the hindbrain, tectal projecting cells were also bilaterally labeled in the dorsal and lateral isthmic nuclei, the octavolateral area, the sensory nucleus of the descending trigeminal tract, the dorsal column nucleus and the reticular formation. The rostral spinal cord also exhibited a few labeled cells. These results demonstrate a complex pattern of connections in the lamprey OT, most of which have been reported in other vertebrates. Hence, the lamprey OT receives a large number of nonvisual afferents from all major brain areas, and so is involved in information processing from different somatic sensory modalities. Copyright (c) 2007 S. Karger AG, Basel.
Dynamic expression of the LIM-homeodomain gene Lhx15 through larval brain development of the sea lamprey (Petromyzon marinus).
Osorio J, Megías M, Pombal MA, Retaux S
Gene Expr Patterns 2006. 6(8):873-8
LIM-homeodomain genes encode a family of transcription factors with highly conserved roles in the patterning and regionalisation of the vertebrate brain. The expression of one of those genes, Lhx15, in the embryonic lamprey brain, characterises precise functional subdivisions. In order to analyse the non-embryonic development of the lamprey brain, we chose this gene to perform in situ hybridisations in Petromyzon marinus larvae of different ages. We demonstrate the usefulness of Lhx15 to follow the development and morphogenesis of brain structures and show the dynamical expression of this gene through time. Furthermore, we provide evidence for the evolutionary conservation of the expression of this gene in the spinal cord, notochord and urogenital system.
Distribution of neuropeptide FF-like immunoreactive structures in the lamprey central nervous system and its relation to catecholaminergic neuronal structures.
Pombal MA, Lopez JM, de Arriba MC, Megías M, González A
Peptides 2006. 27(5):1054-72
The neuropeptide FF (NPFF) is an octapeptide of the RFamide-related peptides (FaRPs) that was primarily isolated from the bovine brain. Its distribution in the CNS has been reported in several mammalian species, as well as in some amphibians. Therefore, in order to gain insight in the evolution on the expression pattern of this neuropeptide in vertebrates, we carried out an immunohistochemical study in the sea lamprey, Petromyzon marinus. The distribution of NPFF-like-immunoreactive (NPFF-ir) structures in the lamprey brain is, in general, comparable to that previously described in other vertebrate species. In lamprey, most of the NPFF-ir cells were found in the hypothalamus, particularly in two large populations, the bed nucleus of the tract of the postoptic commissure and the tuberomammillary area. Numerous NPFF-ir cells were also observed in the rostral rhombencephalon, including a population in the dorsal isthmic gray and the reticular formation. Additional labeled neurons were found inside the preoptic region, the parapineal vesicle, the periventricular mesencephalic tegmentum, the descending trigeminal tract, the nucleus of the solitary tract, as well as in the gray matter of the spinal cord. The NPFF-ir fibers were widely distributed in the brain and the spinal cord, being, in general, more concentrated throughout the basal plate. The presence of NPFF-ir fibers in the lamprey neurohypophysis suggests that the involvement of NPFF-like substances in the hypothalamo-hypophyseal system had emerged early during evolution.
Developmental changes of the GABA-immunoreactive fibers in the lamprey spinal cord.
Pombal MA, Ruiz Y, Rodríguez-Alonso M, de Arriba MC, Costas V, Alvarez R, Megías M
Brain Res Bull. 2005. 66:371-375
The changes in distribution and number of GABA immunoreactive (GABA-ir) fibers from postembryonic stages to adulthood in the lamprey spinal cord white matter were studied by using immunocytochemical techniques. From prolarvae to adult spawning animals there was an increase of the number of GABA-ir fibers. Three phases can be distinguished: (a) from prolarvae to middle size larvae (around 50 mm in body length) an increase in the number of GABA-ir fibers per section is observed. Furthermore, an adult-like pattern of GABA-ir fibers distribution is established during this phase. (b) Then, the number of GABA-ir fibers remains stable until metamorphosis, the end of the larval period. (c) Finally, in young postmetamorphic and adult animals the number of GABA-ir fibers is higher than in larvae. These observations, joined to the changes previously reported in the GABA-ir neurons, indicate that at least parts of the GABA inhibitory component of the spinal locomotor network is reorganized during the lamprey life cycle and it may indicate different inhibitory requirements in the locomotor network.
Choline acetyltransferase-immunoreactive neurons in the retina of adult and developing lampreys
Pombal MA, Abalo XM, Rodicio MC, Anadón R, González A.
Brain Res. 2004. 993:154-163
The presence of choline acetyltransferase-immunoreactive (ChATir) amacrine cells is reported for the first time in the retinas of three species of lamprey (Lampetra fluviatilis, Ichthyomyzon unicuspis, and Petromyzon marinus). In the three species, the ChATir cells were mainly distributed in the inner plexiform layer (IPL), which in lampreys extends from the inner nuclear layer (INL) to the inner limiting membrane. These cells had a bipolar, triangular or stellate appearance, and gave rise to processes coursing in the inner plexiform layer. In transforming lampreys, ChATir processes formed two asymmetrical inner and outer subplexuses in the inner plexiform layer, which is reminiscent of the distribution of processes of ChATir cells in the On and Off sublaminae reported in jawed vertebrates. The larval retina lacked ChAT immunoreactivity, and ChATir cells and processes appeared at early metamorphosis throughout the retina, exhibiting in late transforming stages an organization similar to that of adults. This first report of ChATir cells in the lamprey retina indicates that the appearance of cholinergic circuits in the retina of vertebrates occurred before the divergence of the agnathan and gnathostome lines.
Development of GABA-immunoreactive cells in the spinal cord of the sea lamprey, P. marinus
Ruiz Y, Pombal MA, Megías M.
J Comp Neurol 2004. 470:151-163
The lamprey spinal cord increases in length and size during all its life cycle and thus it is expected that new cells will be generated. This implies that the locomotor circuits must be continuously remodeled. Key elements in the cellular network controlling locomotor behavior are inhibitory cells. Here, we studied the g-aminobutyric acid immunoreactive (GABA-ir) cells in the lamprey spinal cord during postembryonic development. Three major populations of GABA-ir cells were identified according to their distribution: those located in the gray matter, those contacting the cerebrospinal liquid (LC cells), and those located in the white matter. The results show: a) the number of GABA-ir cells per segment increase from prolarvae (<10 mm) to adulthood; b) the lower number of GABA-ir cells in 100 m of spinal cord is 66 7, found in premetamorphic larvae, and the highest is 107 6, found in postmetamorphic animals; c) the gray matter and LC GABA-ir cells show different variations in number depending on the developmental period. Thus, in the 10 mm larvae the gray matter GABA-ir cells are much abundant than LC cells, whereas in the young postmetamorphics the contrary occurs. Most of the GABA-ir cells located in the white matter were classified as edge cells. They increase in number from the beginning of the prolarval period, where there are not white matter positive cells, to the middle larval period where there are 9 4 GABA-ir edge cells per segment. This value was unaltered in later periods, where GABA-ir edge cells represent 20-30 % of the total number of edge cells per segment. The increase in number of GABA-ir cells in these populations during specific point of the lamprey life cycle may indicate different inhibitory requirements of the locomotor circuit at different developmental periods.
Calbindin and calretinin immunoreactivities identify different types of neurons in the adult lamprey spinal cord.
Megías M, Alvarez-Otero R, Pombal MA.
J Comp Neurol 2003. 455(1):72-85
The central pattern generator for locomotion in vertebrates is composed of different spinal neuronal populations that generate locomotor movement. In the lamprey spinal cord, several classes of interneurons have been identified based on morphologic and physiological criteria and integrated in the spinal cord circuits implicated in the generation of locomotion. However, the lack of histochemical markers for most of the interneurons makes it difficult to study whole populations along the spinal cord. We have investigated the immunoreactivity with antibodies raised against calbindin and calretinin. Several types of neurons could be classified: (1) strongly immunoreactive neurons located dorsomedially, (2) moderately immunoreactive neurons located laterally, (3) small weakly immunoreactive neurons, d) ventromedial neurons, (4) liquor contacting cells, and (5) motoneurons. The ventromedial group of calbindin-immunoreactive neurons also is immunoreactive for serotonin and, therefore, represents the ventromedial group of dopamine/serotonin spinal neurons. Some of the lateral calbindin-immunoreactive neurons may be CC-type cells (cells with caudal-crossed axons), because they are retrogradely labeled by tracer injections into the contralateral spinal cord. Other well-characterized cell types, such as sensory dorsal cells, lateral interneurons, descending propriospinal edge cells, and spinobulbar giant interneurons are negative for both calbindin and calretinin. Therefore, calbindin and calretinin are useful markers for the study of cell populations that may be integrated in locomotor circuits. Copyright 2002 Wiley-Liss, Inc.
Drawing depicting the different subpopulations of neurons immunoreactive for CB and CR found in the lamprey spinal cord. Dorsal view.
SDL: strongly immunoreactive neurons located dorsomedially; ML: moderately immunoreactive neurons located laterally; Sm: small weakly immunoreactive neurons; CC: liquour contacting cells.
Immunocytochemical localization of calretinin in the olfactory system of the adult lamprey, Lampetra fluviatilis.
Pombal MA, de Arriba MC, Sampedro C, Alvarez R, Megias M.
Brain Res Bull 2002. 57(3-4):281-2833
The distribution of calretinin immunoreactive (CR-ir) structures in the adult lamprey (Lampetra fluviatilis) olfactory system was studied by using immunocytochemical techniques. In the olfactory epithelium, a subpopulation of olfactory receptor cells was CR-ir. In the olfactory bulbs, three different cell populations were observed. Large CR-ir cells (mitral cells) were located medially to the olfactory glomeruli and occasionally between them. In the inner cellular layer, two types of CR-ir perikarya were found: numerous small CR-ir cells (granule cells) and some medium-sized CR-ir cells (putative displaced periglomerular cells). In addition, different intensities of CR-ir fibers were present in particular rootlets of the olfactory nerves, as well as in particular glomeruli. The presence of CR-ir cells and fibers in all layers of the lamprey olfactory bulbs supports the idea that this protein is present in pathways underlying the processing of sensory information throughout evolution.
Distribution of choline acetyltransferase-immunoreactive structures in the lamprey brain.
Pombal MA, Marín O, González A.
J Comp Neurol 2001. 431(1):105-26
The distribution of cholinergic neurons and fibers was studied immunohistochemically in the brain of two species of lampreys (Petromyzon marinus and Lampetra fluviatilis), by using an antiserum against choline acetyltransferase (ChAT). The results obtained in both species were similar, but there appeared some interspecies differences. In the forebrain, cholinergic cells were present in the striatum, preoptic region, paraventricular nucleus, pineal and parapineal organs, habenula, and pretectum. The cranial nerve motoneurons (III, IV, V, VI, VII, IX, and X), the first and second spino-occipital nerves (so), and the ventral horn of the spinal cord showed a strong ChAT immunoreactivity. Additional cholinergic neurons were observed: the mesencephalic M5 nucleus of Schober, two different cell populations in the isthmic region, the efferent component of the eighth nerve, putative preganglionic parasympathetic cells, cells in the solitary tract nucleus, and the rhombencephalic reticular formation. Cholinergic fibers were widely distributed in the brain. Comparison with previous studies in other vertebrates suggests that major cholinergic pathways, like tectal innervation from the isthmic region, are also present in lampreys. Of particular interest was the prominent projection to the neurohypophysis from cholinergic neurons in the preoptic region and paraventricular nucleus. Present data were analyzed within the segmental paradigm, as was previously done in other vertebrates. Our results reveal that the organization of many cholinergic systems in the lamprey as, for example, in the striatal, preoptic, and isthmic regions, comprises features of the anamniote brain that remain common to all living amniotes studied so far, thus being conservative to a surprisingly high degree. Therefore, the distribution of ChAT-immunoreactive structures in the lamprey brain is, in general, comparable to that previously described in other vertebrate species. Copyright 2001 Wiley-Liss, Inc.
Other publications:
GABA-immunoreactive internuclear neurons in the ocular motor system of lampreys.
Meléndez-Ferro M, Pérez-Costas E, González MJ, Pombal MA, Anadón R, Rodicio MC.
Brain Res 2000. 855(1):150-7
