PSA-NCAM immunocytochemistry in the cerebral cortex and other telencephalic areas of the lizard Podarcis hispanica: differential expression during medial cortex neuronal regeneration

The lizard medial cortex, a region homologous to the mammalian dentate gyrus, shows postnatal neurogenesis and the surprising ability to replace its neurons after being lesioned specifically with the neurotoxin 3-acetylpyridine. As the polysialylated form of the neural cell adhesion molecule (PSA-NCAM) is expressed during neuronal migration and differentiation, we have studied its distribution in adult lizards and also during the lesion-regeneration process. In the medial cortex of control animals, many labeled fusiform somata, presumably corresponding to migratory neuroblasts, appeared in the inner plexiform layer. There were also scattered immunoreactive granule neurons in the cell layer. Double immunocytochemistry with 5'-bromodeoxyuridine revealed that some of the PSA-NCAM-expressing cells in the inner plexiform and cell layers were generated recently. PSA-NCAM immunoreactivity was also present in the dorsomedial, dorsal, and lateral cortices, as well as in the dorsal ventricular ridge, the nucleus accumbens, and the nucleus sphericus. Twelve hours after the injection of 3-acetylpyridine, some medial cortex granule neurons appeared degenerated, although some of them still expressed PSA-NCAM. One to 2 days after the injection, most granule neurons appeared degenerated and no PSA-NCAM immunoreactivity was detected in the medial cortex cell layer. Four to 7 days after treatment, abundant labeled fusiform cells populated the inner plexiform layer and some immunoreactive somata were seen in the cell layer. Fifteen to 30 days after the neurotoxin injection, the number of PSA-NCAM expressing granule neurons augmented considerably and the level was still above control levels in lizards that survived 42 days. Our results show for the first time the expression of PSA-NCAM in a reptile brain, where it appears to participate in the migration and differentiation of granule neurons during adult neurogenesis and regeneration.     

A Golgi study of the principal projection neurons of the medial cortex of the lizard Podarcis hispanica

The medial cortex of lizards is a simple three-layered brain region displaying many characteristics that parallel the hippocampal fascia dentata of mammals. Its principal neurons form a morphologically diverse population, partly as a result of the prominent continuous growth of this nervous center. By using the classic Golgi impregnation method, we describe here the morphology of the principal neurons populating the medial cortex of Podarcis hispanica. These were projection neurons giving off descending axons. These axons displayed deep collateral branches provided with prominent axonal boutons, while the main axonal branch reached adjacent cortical areas and the bilateral septum. According to three main classification criteria, dendritic tree pattern, dendritic spine covering, and soma size, we have distinguished eight different types of projection neurons. Five of them, “heavily spiny granular” (monotufted, medium-sized), “heavily spiny bitufted” (large), “spiny bitufted” (medium-sized), “sparsely spiny bitufted” (small), and “superficial multipolar” (small), were found in the cell layer, whereas the three others lay outside this layer and were regarded as ectopic types (“outer plexiform ectopic bitufted,” “inner plexiform ectopic bitufted”, and “inner plexiform monotufted”). Additional secondary criteria, soma position and shape, allowed us to further classify bitufted neurons into three distinct subtypes each: “superficial-round,” “intermediate-fusiform,” and “deep-pyramidal.” Moreover, a variety of small impregnated cells were observed; they probably represented newly generated immature neurons that had not yet completed their development. These cell types were compared with those reported previously in Golgi, immunocytochemical, and electron-microscopy studies, both in the reptilian medial cortex and in the mammalian dentate area. Presumably age-related changes and synaptic relationships of these projection cells in the medial cortex circuitry were analyzed. J. Comp. Neurol. 385:528–564, 1997. © 1997 Wiley-Liss, Inc.     

Parvalbumin-containing neurons in the cerebral cortex of the lizard Podarcis hispanica: Morphology,ultrastructure, and coexistence with GABA,somatostatin, and neuropeptide Y

The morphology, fine structure, and degree of colocalization with GABA, somatostatin, and neuropeptide Y of parvalbumin-containing cells were studied with immunocytochemistry in the cerebral cortex of the lizard Podarcis hispanica. Parvalbumin-containing cells make up a morphologically heterogeneous population of spine-free neurons, displaying the morphological features of nonprincipal cells previously described in Golgi studies. Electron microscopically, parvalbumin-immunoreactive cell bodies are similar in all cortical areas and layers. The perisomatic input is moderate in number, and boutons with either round clear vesicles or flattened vesicles were observed making asymmetric or symmetric synaptic contacts, respectively. Parvalbumin-immunoreactive dendrites are smooth and almost completely covered with synaptic boutons of different types, most of which establish asymmetric contacts. Parvalbumin-immunoreactive boutons are concentrated around cell bodies of principal cells. They are large, containing abundant mitochondria and small pleomorphic vesicles, and establishing symmetric synaptic contacts with somata, proximal dendritic shafts, and axon initial segments of principal cells. Colocalization studies revealed that all the parvalbumin-containing cells are GABA-immunoreactive, representing only a fraction of the GABA-immunopositive cell population, and that parvalbumin- and peptide- (somatostatin and neuropeptide Y) containing cells show a negligible overlap. These results demonstrate that in the cerebral cortex of the lizard Podarcis hispanica, parvalbumin-containing cells represent a subset of nonprincipal GABAergic neurons largely involved in perisomatic inhibition, which are different from the peptide-containing cells, and suggest that they may include both axosomatic and axoaxonic cells. © 1993 Wiley-Liss, Inc.     

CRMP-4 expression in the adult cerebral cortex and other telencephalic areas of the lizard Podarcis hispanica

The control of neuritogenesis is crucial for the development, maturation and regeneration of the nervous system. The collapsin response-mediated protein 4 (CRMP-4) is a member of a family of proteins that are involved in neuronal differentiation and axonal outgrowth. In rodents, this protein is expressed in recently generated neurons such as some granule neurons of the dentate gyrus, as well as in certain differentiated neurons undergoing neurite outgrowth or synaptogenesis during adulthood. Since CRMP-4 protein appears to be highly conserved throughout the evolutionary scale, we have used immunocytochemistry to study its distribution in the lizard cerebral cortex. We have found pronounced CRMP-4 immunolabeling in certain neurons of the medial cortex, the homologous region to the dentate gyrus, but also in the dorsal and lateral cortices. Double labeling with 5'-BrdU indicated that these medial cortex neurons were recently generated. However, it is also possible that many of these cells were not new but undergoing some kind of plasticity implicating neurite outgrowth. Similar CRMP-4-labeled neurons and processes were observed in subcortical regions as the PDVR and the nucleus sphericus. Our results show for the first time the expression of CRMP-4 in a reptile brain, where it appears to be expressed in regions where adult neurogenesis and/or neurite outgrowth occur.     

Distribution of GABAergic interneurons immunoreactive for calretinin, calbindin D28K, and parvalbumin in the cerebral cortex of the lizard Podarcis hispanica.

The types and distribution of cells containing three calcium-binding proteins, calretinin, calbindin D28K, and parvalbumin, have been studied by immunocytochemistry in different areas of the cerebral cortex of lizards. Cross-reactivity of the antisera has been excluded by demonstrating the existence of several cell groups immunoreactive for one but not the other two calcium-binding proteins. In the dorsal and dorsomedial cortices all three proteins coexist in a single subpopulation of gamma-aminobutyric acid (GABA)ergic neurons, the terminals of which form pericellular baskets around cell bodies of bipyramidal neurons. The somata of these neurons are largely restricted to the cellular and inner plexiform layers, but the dendrites usually penetrate all layers, allowing the neurons to sample input from all possible sources. A small number of parvalbumin-containing neurons in the outer plexiform layer do not contain the other two proteins. The medial cortex, which is likely to be homologous to the mammalian dentate gyrus, only contains parvalbumin-immunoreactive neurons. The dendritic trees of these cells appear to avoid the Timm-positive fields receiving input from zinc-rich fiber collaterals, originating from principal cells. The lateral cortex contains calbindin D28K-immunoreactive GABAergic neurons, which lack the other two calcium-binding proteins. These neurons have horizontally running dendrites in the outer plexiform layer, but their axon terminals could not be visualized. The present study uncovered important similarities and differences between the lizard and the mammalian archicortex in the types of neurons containing calcium-binding proteins. As in mammals, different cell types evolved in the lizard to inhibit the perisomatic versus the distal dendritic region of principal cells, the calcium-binding protein-containing neurons being responsible for the former, and neuropeptide-containing neurons for the latter. The results also suggest that further neurochemical diversion of GABAergic interneurons coupled to a functional specialization took place during phylogenetic development from reptiles to mammals.     

Neurons of the medial cortex outer plexiform layer of the lizard Podarcis hispanica: Golgi and immunocytochemical studies

The study of Golgi-impregnated lizard brains has revealed a scarce but heterogeneous neuronal population in the outer plexiform layer of the medial cortex. Some of the neuronal types detected here resemble the neurons of the dentate molecular layer of the mammalian hippocampus. According to their morphology, five intrinsic neuronal types have been clearly identified: short axon aspinous bipolar neuron (type 1, or sarmentous neuron), short axon aspinous juxtasomatic neuron (type 2, or coral neuron), short axon sparsely spinous multipolar neuron (type 3, or stellate neuron), short axon sparsely spinous juxtasomatic multipolar neuron (type 4, or deep stellate neuron, and sparsely spinous juxtasomatic horizontal neuron (type 5, or couchant neuron). Most neuronal types were identified as γ-aminobutyric acid (GABA) and parvalbumin immunoreactive, and are thus probably involved in medial cortex inhibition. Moreover, a small fraction of them displayed ß-endorphin immunoreactivity. The distribution of these neuronal types is not uniform in the laminae of the outer plexiform layer. Type 1 (sarmentous) and type 3 (stellate) neurons overlap the axonal field projection coming from the dorsal cortex and the thalamus, whereas types 4 (deep stellate) and 5 (couchant) neurons overlap ipsi- and contralateral dorsomedial projection fields as well as raphe serotoninergic and opioid immunoreactive axonal plexi. Thus, these neuronal types may be involved in the control of specific inputs to the medial cortex by presumably feed-forward inhibition; nevertheless, feed-back inhibition may also occur regarding type 4 (deep stellate) neurons that extend deep dendrites to the zinc-rich bouton field.     

Laminar distribution and morphology of gamma-aminobutyric acid (GABA)-immunoreactive neurons in the medial and dorsomedial areas of the cerebral cortex of the lizard Podarcis hispanica

The morphology and laminar distribution of immunolabeled neurons in the medial and dorsomedial telencephalic cortices of the lizard Podarcis hispanica were examined in vibratome sections after preembedding gamma-aminobutyric acid (GABA)-immunocytochemistry. In both cortical areas and at all rostrocaudal levels, GABA-immunoreactive neurons were found in all cortical layers, with the largest number (74%) of GABA-positive cells in layer 3. GABA-positive neurons were classified into pyramidlike, vertical-fusiform, multipolar, and horizontal neurons. Cells that could be so classified were counted in each cortical lamina. In the medial cortex, multipolar and horizontal-bipolar cells dominated layer 1. Layer 2 displayed mainly horizontal and pyramidlike cells at its outer margin and pyramidlike cells at its inner margin. In layer 3, horizontal cells were the prevalent group. In the dorsomedial cortex, layer 1 mainly contained small multipolar neurons (35% of layer-1 cells) in its outer third and vertical-fusiform neurons (37% of layer-1 cells) in its inner two thirds. In layer 2, 47% of the few GABA-positive perikarya were pyramidlike. The largest population of neurons in layer 3 was that formed by multipolar cells (45% of layer-3 cells). Ultrastructural examination revealed that GABA-immunoreactive neurons possessed indented euchromatic nuclei with a central nucleolus. Their cytoplasm contained numerous mitochondria and a very well-developed granular endoplasmic reticulum. Their somata were contacted by numerous unstained boutons making asymmetric contacts and by a few symmetric synapses of GABA-positive terminals. Dendrites of GABA-immunoreactive cells were thin, with irregular outlines, and generally aspinous. Like the somata, dendrites were contacted by many unstained asymmetric synapses. Some dendritic profiles also received symmetric contacts from GABA-positive boutons. GABA-positive terminal-like puncta were found throughout the layers, with a maximal concentration in layer 2. Electron microscopy confirmed that nearly all of the puncta represent GABA-positive terminal boutons. Comparison of GABA-immunoreactive cells in Podarcis with those found in the mammalian hippocampus suggests that these cells may be inhibitory neurons, as in the hippocampus of mammals.     

Serotoninergic innervation of nonprincipal cells in the cerebral cortex of the lizard Podarcis hispanica

The mechanism of serotoninergic transmission in the neo- and archicortex of mammals kis complex, including both synaptic and nonsynaptic components, direct actions on principal cells, and indirect effects mediated by GABAergic interneurons. Here we studied the termination pattern and synaptic organization of the serotoninergic afferents in the cerebral cortex of the lizard, Podarcis hispanica, which is considered to correspond in part to the mammalian hippocampal formation, with the aim of unraveling basic, phylogenetically preserved rules in the connectivity of this pathway. We demonstrate that serotoninergic afferents, visualized by immunostaining for serotonin itself, establish multiple synaptic contacts with different subpopulations of nonprincipal cells containing parvalbumin, neuropeptide Y, and opioid peptides. The former two subpopulations contain GABA, whereas the opioid- immunoreactive neurons are most likely GABA-negative cells. Evidence is provided at the electron microscopic level that serotonin-immunoreactive varicosities establish conventional asymmetric synaptic contacts with their nonprincipal targets, but nonsynaptic varicosities also exist. We conclude that, similarly to mammals, a selective synaptic innervation of nonprincipal, possibly inhibitory, neurons is among the mechanisms of serotoninergic modulation of cerebral cortical activity in the lizard. © 1994 Wiley-Liss, Inc.     

Amygdalo-hypothalamic projections in the lizard Podarcis hispanica: A combined anterograde and retrograde tracing study

The cells of origin and terminal fields of the amygdalo-hypothalamic projections in the lizard Podarcis hispanica were determined by using the anterograde and retrograde transport of the tracers, biotinylated dextran amine and horseradish peroxidase. The resulting labeling indicated that there was a small projection to the preoptic hypothalamus, that arose from the vomeronasal amygdaloid nuclei (nucleus sphericus and nucleus of the accessory olfactory tract), and an important projection to the rest of the hypothalamus, that was formed by three components: medial, lateral, and ventral. The medial projection originated mainly in the dorsal amygdaloid division (posterior dorsal ventricular ridge and lateral amygdala) and also in the centromedial amygdaloid division (medial amygdala and bed nucleus of the stria terminalis). It coursed through the stria terminalis and reached mainly the retrochiasmatic area and the ventromedial hypothalamic nucleus. The lateral projection originated in the cortical amygdaloid division (ventral anterior and ventral posterior amygdala). It coursed via the lateral amygdalofugal tract and terminated in the lateral hypothalamic area and the lateral tuberomammillary area. The ventral projection originated in the centromedial amygdaloid division (in the striato-amygdaloid transition area), coursed through the ventral peduncle of the lateral forebrain bundle, and reached the lateral posterior hypothalamic nucleus, continuing caudally to the hindbrain. Such a pattern of the amygdalo-hypothalamic projections has not been described before, and its functional implications in the transfer of multisensory information to the hypothalamus are discussed. The possible homologies with the amygdalo-hypothalamic projections in mammals and other vertebrates are also considered. J. Comp. Neurol. 384:537–555, 1997. © 1997 Wiley-Liss, Inc.     

Distribution of calcitonin gene-related peptide-like immunoreactivity in the brain of the lizard Podarcis hispanica

The present work studies the distribution of calcitonin gene-related peptide-immunoreactive (CGRP-li) neurons and fibers in the brain of a reptile, the lizard Podarcis hispanica. CGRP-li perikarya were not present in the telencephalon. In the thalamus, CGRP-li perikarya were restricted to the posteromedial and posterolateral nuclei. In the hypothalamus, CGRP-li cells were found mainly in the supramammillary and mammillary nuclei. In the midbrain and brainstem, CGRP-li cells appeared in the ventral tegmental area, the parabrachial nucleus, and the motor nuclei of the III-VII, IX, X, and XII cranial nerves. Motoneurons of the ventral horn of the spinal cord were also immunoreactive for CGRP. CGRP-li fibers were seen in the telencephalic hemispheres, where a dense plexus of reactive fibers appeared in the septum and in the lateral striatoamygdaloid transition area. From the latter, CGRP-li fibers entered the posterior dorsal ventricular ridge, the cell layer and deep stratum of the ventral lateral cortex, and various amygdaloid nuclei. Parts of the striatum (nucleus accumbens) and pallidum also displayed CGRP-li innervation. In the diencephalon, CGRP-li innervation was observed in parts of the dorsal thalamus and in the periventricular and medial hypothalamus. The pretectum and deep layers of the optic tectum also showed CGRP-li fibers, and numerous CGRP-li fibers were observed in the midbrain central gray, tegmentum, and pons. Some of the sensory fibers of the trigeminal, vagal, and spinal nerves were also CGRP-li. These results show that the distribution of CGRP-li structures in the reptilian brain is similar to that described for other vertebrates and suggest that the thalamotelencephalic CGRPergic projections appear to be conserved among amniote vertebrates.     

Septal complex of the telencephalon of the lizard Podarcis hispanica. II. afferent connections

The afferent connections to the septal complex were studied in the lizard Podarcis hispanica (Lacertidae) by means of a combination of retrograde and anterograde tracing. The results of these experiments allow us to classify the septal nuclei into three main divisions. The central septal division (anterior, lateral, dorsolateral, ventrolateral, and medial septal nuclei plus the nucleus of the posterior pallial commissure) receives a massive, topographically organized, cortical projection (medial, dorsal, and ventral areas) and widespread afferents from the tuberomammillary hypothalamus and the basal telencephalon. Moreover, it receives discrete projections from the dorsomedial anterior thalamus, the ventral tegmentum, the midbrain raphe, and the locus coeruleus. The ventromedial septal division (ventromedial septal nucleus) receives a massive projection from the anterior hypothalamus, dense serotonergic innervation, and a faint amygdalohypothalamic projection, but it is devoid of direct cortical input. The midline septal division (nucleus septalis impar and dorsal septal nucleus) receives a nontopographic cortical projection (dorsomedial and dorsal cortices) and afferents from the preoptic hypothalamus, the dorsomedial anterior thalamus, the midbrain central gray, and the reptilian A8 nucleus/substantia nigra. Our results indicate that the cortex provides a physiologically complex, massive input to the septum that terminates over the whole dendritic tree of septal cells. In contrast, most of the ascending afferents make axosomatic contacts by means of pericellular nests. The chemical nature of the main septal afferents and the comparative implications of the available hodological data on the organization of the septal complex of tetrapod vertebrates are discussed. J. Comp. Neurol. 383:489-511, 1997. © 1997 Wiley-Liss, Inc.     

Size variation and cell proliferation in chemosensory brain areas of a lizard (Podarcis hispanica): effects of sex and season

Many lizards rely on chemoreception for crucial aspects of their biology, including exploration, prey and predator detection, and intraspecific communication. Here we investigate sex and seasonal variation in size and proliferative activity in chemosensory areas of the lizard brain. We captured adult Iberian wall lizards ( Podarcis hispanica ) of either sex in the breeding (April) and non-breeding (November) season, injected them with 5-bromo-2'-deoxyuridine (BrdU) and killed them 3 weeks later. We removed the brains, measured the length of the olfactory bulbs, and counted BrdU-labelled cells in the main and accessory olfactory bulbs (MOB, AOB), lateral cortex (LC) and nucleus sphericus (NS). Our results show that, relative to body size, males have larger MOBs and AOBs than females; however, relative to brain size, males have larger AOBs, but not larger MOBs than females. Additionally, males produce more new cells than females in the olfactory bulbs, LC and NS. We failed to detect significant seasonal changes or sex × season interaction in size or proliferative activity in these areas. Sex differences in the addition of newly generated cells – mainly neurons – may be partly responsible for the size differences in chemosensory brain areas. The presence of sexual dimorphism in AOB is expected given the available behavioural evidence, which suggests that males of P. hispanica are more responsive than females to socially relevant chemical stimuli. This is the first demonstration of sexual dimorphism in size and proliferative activity in chemosensory areas of a non-mammalian species.     

The development of somatostatin immunoreactive neurons in cat visual cortical areas

The development of somatostatin immunoreactive (SOM-ir) neurons in cat striate and extrastriate cortex was studied to determine whether temporal changes in the morphology, distribution and density of SOM-ir neurons during development would provide clues to the emergence of specific cortical areas. The visual cortical areas examined included areas 17–19 and 7, posteromedial lateral suprasylvian, posterolateral lateral suprasylvian cortex and splenial visual area. We observed that the pattern of SOM-ir neurons in the cortical plate reflects the maturation of the cortical plate. At 1 week of age, SOM-ir neurons were only found in layers V and VI of the developing cortex; by 2 weeks of age, SOM-ir neurons were found in layer IV; and by 3 weeks of age, SOM-ir neurons were located in all layers of the cortex except layer I. SOM-ir neurons in the subplate were much more numerous under lateral cortical areas than under medial areas. This difference decreased over the first 2 postnatal weeks and by the 14th day after birth (P14), the distribution and numbers of SOM-ir neurons in the subplate/white matter had reached the adult pattern. The timing of exuberant SOM expression in the subplate suggests a function in the formation of visual corticocortical connections which begin to develop during the first postnatal week in the kitten.     

Efferent connections of the lateral cortex of the lizard gekko gecko: Evidence for separate origins of medial and lateral pathways from the lateral cortex to the hypothalamus

The lateral cortex of the lizard Gekko gecko is composed of three parts: a dorsal and ventral part located rostrally and a posterior part located caudally. In order to obtain detailed information about the efferent connections of these lateral cortex subdivisions, iontophoretic injections of the anterograde tracers Phaseolus vulgaris leucoagglutinin and biotinylated dextran were made in the various parts. The main projection from the dorsal part terminates in the caudal part of the medial cortex. Other cortical projections were noted to the ipsi- and contralateral lateral cortex, the large-celled part of the medial cortex, and the dorsal cortex. Additional fibers were found bilaterally in the anterior olfactory nucleus and the external amygdaloid nucleus. The ventral part of the lateral cortex projects mainly to the ipsilateral, posterior part of the dorsal ventricular ridge and the external amygdaloid nucleus. Minor contralateral projections to these nuclei were also found. Other projections were observed to travel to the caudal part of the medial cortex, to the nucleus sphericus, and bilaterally to the lateral cortex and the anterior olfactory nucleus. The posterior part of the lateral cortex has similar efferent connections as the dorsal part and should be regarded as the caudal continuation of the dorsal part. Because previous studies have shown that the medial cortex and the amygdaloid complex project to different hypothalamic areas, we conclude that the dorsal and ventral parts of the lateral cortex transmit olfactory information to separate hypothalamic areas that are probably involved with different types of behavior. © 1995 Wiley-Liss, Inc.     

A sulphide-osmium procedure for detection of metal-containing synaptic boutons in the lizard cerebral cortex

Two procedures of the Timm sulphide-silver method for the ultrastructural location of heavy metals and a sulphide-osmium procedure proposed here were applied to studying the Timm-reacting axonal boutons of the lizard cerebral cortex. It was shown that metal-containing boutons which would be labelled when using the Timm method, could be labelled by the sulphide-osmium procedure as well, i.e. perfusion and immersion in a glutaraldehyde fixative saturated with H₂S vapour and then osmium postfixation. When the sulphide-osmium procedure was used, Timm-reacting boutons displayed an electron-dense loading in some of their synaptic vesicles. The histochemical significance of this labelling is suggested; metal contents (possibly zinc) in the synaptic vesicles are first transformed into metal sulphides that afterwards interact with OsO₄ and render an electron-dense labelling. Moreover, chain-like electron-dense aggregates, which could not be detected in the Timm method, have been shown in endoplasmic reticulum cisternae using the sulphide-osmium procedure.The importance of both the Timm method and the sulphide-osmium procedure in ultrastructural studies in the lizard cerebral cortex could be paramount, as they define specifically one of the projection systems in the reptilian cortex. The sulphide-osmium procedure could be more useful than the Timm method since it is a simple procedure that avoids physical development, allows a better ultrastructural quality and a more precise location of the labelling.     

Long-spined polymorphic neurons of the medial cortex of lizards: a Golgi, Timm, and electron-microscopic study

The morphology, ultrastructure, and principal synaptic input of long-spined neurons located in the inner plexiform layer of the medial cortex in three related species of lizards is described. Golgi impregnations have been used to define the external morphology of these neurons and their axonal trajectories. Their most striking characteristic is the presence of very long spines or "microdendrites" especially abundant on the distal dendritic segments. Axons have ascendent trajectories, pass through the cell layer, and ramify in the outer plexiform layer. Combined Golgi-electron microscopy as well as standard electron microscopy permitted the definition of the ultrastructure of these neurons. Timm and sulfide-osmium methods permitted the detection and characterization of their principal synaptic input (i.e., zinc-containing boutons). Gamma aminobutyric acid (GABA)-immunostained sections in one of the species studied allowed the identification of GABA-immunoreactive somata which had the same morphology and ultrastructure as long-spined neurons; these GABA-immunoreactive somata and their processes were found in the same location as long-spined neurons. This suggests that at least some long-spined polymorphic neurons are GABA-ergic and presumably inhibitory. Finally, the neurobiological significance of these long-spined neurons is discussed and briefly compared with that of similar neurons of the hilus of the fascia dentata of the rat.     

Modifications in cortical ependyma of the lizard, Podarcis hispanica, during postnatal development

The ependymal surface, cell number, cell size and cell density were measured in adult and neonatal exemplars of the lizard, Podarcis hispanica. The ependymal monostratified surface increases with age, while that of sulcal areas decreases and the ependymocyte number remains constant. The mitotic activity found in the neonatal sulcal areas could explain the postnatal increase of neuronal number described for the cerebral cortex of the studied species.     

Distribution of neuropeptide Y (NPY) in the cerebral cortex of the lizards Psammodromus algirus and podarcis hispanica: co-localization of NPY, somatostatin, and GABA.

The aim of the present study was to analyze the distribution and characteristics of NPY immunoreactive structures in the cerebral cortex of lizards and to investigate the degree of co-existence of this neuropeptide with somatostatin and GABA. The immunoperoxidase method was applied to vibratome sections as well as to semithin sections. NPY neurons are multipolar or fusiform and were unevenly distributed throughout the brain cortex. Within the medial, dorsomedial and dorsal cortices, most NPY perikarya were located in the plexiform layers, especially in the deep one. This suggests that these cells could be regarded as interneurons. In the lateral cortex, NPY neurons were found throughout all layers. The dorsomedial cortex displayed the highest NPY cell density. Here, neuronal perikarya projected many immunoreactive processes toward two distinct zones: the deep plexiform layer of the medial cortex and the superpositio medialis. The NPY neurons of the dorsomedial cortex differed from the other NPY cortical immunoreactive cells in that the latter displayed very few immunoreactive processes. A high degree of co-existence among NPY, somatostatin, and GABA (approx. 80%) was found. This co-existence rate is very similar to that reported in mammals and suggests that co-localization is a phylogenetically ancient phenomenon.     

Efferent connections of the dorsal cortex of the lizard Gekko gecko studied with Phaseolus vulgaris-leucoagglutinin

The efferent connections from the dorsal cortex of the lizard Gekko gecko have been studied with the anterograde tracer Phaseolus vulgaris-leucoagglutinin. It appeared that the dorsal cortex is not a homogeneous structure as far as the efferent connections are concerned. All parts of the dorsal cortex project to the septum. All parts except the most medial project to the dorsal ventricular ridge, amygdala, nucleus periventricularis hypothalami, area lateralis hypothalami, and the anterior olfactory nucleus. The most medial part, in addition to the septal projections, is connected with the medial cortex and the contralateral medial and dorsal cortices. From the rostral part additional projections could be traced to the nucleus dorsolateralis hypothalami, nucleus ventromedialis thalami, nucleus dorsolateralis thalami, striatum, pallial thickening, medial cortex, nucleus olfactorius anterior, and the main and accessory olfactory bulbs. From the caudal part additional projections exist to the nucleus dorsomedialis thalami, nucleus accumbens, and the contralateral dorsal cortex. A system of intrinsic connections exists that can be subdivided into four subsystems, each of which subserves the interconnections within four subdivisions of the cortex: 1) the superficial medial part, 2) the deep medial part, 3) the caudal lateral and caudal intermediate parts, and 4) the rostral lateral and rostral intermediate parts. Connections between these four areas are scarce. From the present results the conclusion is drawn that the dorsal cortex of the lizard Gekko gecko has many hodological aspects in common with the ventral subiculum of mammals. The present results do not support the hypothesis that the dorsal cortex is the reptilian equivalent of the mammalian neocortex.