Inhibition of TrkB- and TrkC-Signaling Pathways Affects Neurogenesis in the Opossum Developing Neocortex.

We have previously reported that the blockage of TrkB and TrkC signaling in primary culture of opossum neocortical cells affects neurogenesis that involves a range of processes including cell proliferation, differentiation, and survival. Here, we studied whether TrkB and TrkC activity specifically affects various types of progenitor cell populations during neocortex formation in the Monodelphis opossum in vivo. We found that the inhibition of TrkB and TrkC activities affects the same proliferative cellular phenotype, but TrkC causes more pronounced changes in the rate of cell divisions. Additionally, inhibition of TrkB and TrkC does not affect apoptosis in vivo, which was found in cell culture experiments. The lack of TrkB and TrkC receptor activity caused the arrest of newly generated neurons; therefore, they could not penetrate the subplate zone. We suggest that at this time point in development, migration consists of 2 steps. During the initial step, neurons migrate and reach the base of the subplate, whereas during the next step the migration of neurons to their final position is regulated by TrkB or TrkC signaling.

Organisation of the dopamine neuronal subsets within midbrain of the feathertail glider (Acrobates pygmaeus, Acrobatidae, Marsupialia).

The Marsupial feathertail glider has a unique set of morphological, anatomical and behavioural features that make it a promising model for study of primate evolution. Among them it has many locomotor adaptations to arboreal life, such as diagonal gait of movements, gliding, fast climbing and running along branches. These ecological and behavioural specialisations could result in differences in anatomy of the brain systems involved in their integration. It is well acknowledged that dopaminergic neurons are involved in motor control, motivation and cognition. Due to the fact that there are no data on morphological organisation of dopaminergic system in the midbrain of this species, we decided to investigate it using immunohistochemical and quantitative methods. Our study showed that the general distribution and characteristics of the dopaminergic cells within midbrain nuclei of the pygmy acrobat is similar to that in other species, but it lack the substantia nigra compact part - ventral tier and "tail" of the substantia nigra subnuclei. This study provides the first description of the dopaminergic cells and nuclei in the midbrain of the feathertail glider and we hope it will start interest in the neurobiology of this species.

The expression of interleukin-6 and its receptor in various brain regions and their roles in exploratory behavior and stress responses.

We examined the involvement of interleukin-6 (IL-6) and its receptor IL-6Rα on behavior and stress responses in mice. In the open field, both wild-type (WT) and IL-6 deficient mice displayed similar levels of locomotor activity; however, IL-6 deficient mice spent more time in the central part of the arena compared to control WT mice. After behavioral testing, mice were subjected to stress and then sacrificed. The levels of IL-6 and its receptor in their brains were determined. Immunohistochemical labeling of brain sections for IL-6 showed a high level of expression in the subventricular zone of the lateral ventricles and in the border zone of the third and fourth ventricles. Interestingly, 95% of the IL-6-expressing cells had an astrocytic phenotype, and the remaining 5% were microglial cells. A low level of IL-6 expression was observed in the olfactory bulb, hypothalamus, hippocampus, cerebral cortex, cerebellum, midbrain and several brainstem structures. The vast majority of IL-6-expressing cells in these structures had a neuronal phenotype. Stress increased the number of IL-6-immunoreactive astrocytes and microglial cells. The levels of the IL-6Rα receptor were increased in the hypothalamus of stressed mice. Therefore, in this study, we describe for the first time the distribution of IL-6 in various types of brain cells and in previously unreported regions, such as the subventricular zone of the lateral ventricle. Moreover, we provide data on regional distribution and expression within specific cell phenotypes. This highly differential expression of IL-6 indicates its specific roles in the regulation of neuronal and astrocytic functions, in addition to the roles of IL-6 and its receptor IL-6Rα in stress responses.

Effect of light-dark changes on the locomotor activity in open field in adult rats and opossums.

There have been no reports on how the light-dark changes determine the locomotor activity of animals in the group of high reactivity (HR) and low reactivity (LR). In the present study we have compared selected parameters of the locomotor activity of the HR and the LR groups of the laboratory opossums and Wistar rats during consecutive, light and dark phases in the open field test. Sixty male Wistar adult rats, at an average weight of 350 g each, and 24 adult Monodelphis opossums of both sexes at an average weight of 120 g each were used. The animals' activity for 2 h daily between the hours of 17:30 and 19:30, in line with the natural light-dark cycle were recorded and then analysed using VideoTrack ver.2.0 (Vievpoint France). According to our results, we noted that a change of the experimental conditions from light to dark involves an increase in the locomotor activity in rats and opossums of the HR group, while there is no effect on the activity of the rats and opossums in the LR group. Locomotor activity in the HR rats, both in the light and dark conditions is characterised by a consistent pattern of change - higher activity in the first stage of the recording and a slowdown (habituation) in the second phase of the observation. The locomotor activity of the opossum, during both light and dark conditions, was observed to be at a consistently high level compared to the rats.

Adult neurogenesis in the hedgehog (Erinaceus concolor) and mole (Talpa europaea).

We investigated adult neurogenesis in two species of mammals belonging to the superorder Laurasiatheria, the southern white-breasted hedgehog (order Erinaceomorpha, species Erinaceus concolor) from Armenia and the European mole (order Soricomorpha, species Talpa europaea) from Poland. Neurogenesis in the brain of these species was examined immunohistochemically, using the endogenous markers doublecortin (DCX) and Ki-67, which are highly conserved among species. We found that in both the hedgehog and mole, like in the majority of earlier investigated mammals, neurogenesis continues in the subventricular zone (SVZ) of the lateral ventricles and in the dentate gyrus (DG). In the DG of both species, DCX-expressing cells and Ki-67-labeled cells were present in the subgranular and granular layers. In the mole, a strong bundle of DCX-labeled processes, presumably axons of granule cells, was observed in the center of the hilus. Proliferating cells (expressing Ki-67) were identified in the SVZ of lateral ventricles of both species, but neuronal precursor cells (expressing DCX) were also observed in the olfactory bulb (OB). In both species, the vast majority of cells expressing DCX in the OB were granule cells with radially orientated dendrites, although some periglomerular cells surrounding the glomeruli were also labeled. In addition, this paper is the first to show DCX-labeled fibers in the anterior commissure of the hedgehog and mole. These fibers must be axons of new neurons making interhemispheric connections between the two OB or piriform (olfactory) cortices. DCX-expressing neurons were observed in the striatum and piriform cortex of both hedgehog and mole. We postulate that in both species a fraction of cells newly generated in the SVZ migrates along the rostral migratory stream to the piriform cortex. This pattern of migration resembles that of the 'second-wave neurons' generated during embryonal development of the neocortex rather than the pattern observed during development of the allocortex. In spite of the presence of glial cells alongside DCX-expressing cells, we never found colocalization of DCX protein with a glial marker (vimentin or glial fibrillary acidic protein).

Postnatal treatment with NAN-190 but not with 5-HT1A receptor agonists retards growth of the rat brain.

We investigated the influence of prolonged administration of the 5-HT1A receptor agonists (8-OH-DPAT or buspirone) or its antagonist, NAN-190 to rat pups on development of their cortical barrel field. Pups were injected daily with the drugs starting from the day of birth till either the 5th postnatal day or the 22-25th postnatal day and were perfused one day later. Square areas of their whisker barrel fields were measured on tangential sections of the cortex stained for cytochrome oxidase. Injections of 8-OH-DPAT or buspirone till the 5th postnatal day did not change any of the investigated parameters, while injections of NAN-190 resulted in 15% reduction of the pups' body and brain weight and proportional reduction of the square area of their barrel fields. Groups treated till the 22-25th postnatal day showed similar results. Some of these pups were injected with [C(14)]2-deoxyglucose to investigate the strength of responses of their cortical barrels to stimulation of corresponding vibrissae. The cortical area labeled with 2-deoxyglucose after stimulation of vibrissae of the row C was narrower in the NAN-190 injected rats. This functional deficit was more pronounced than the anatomical one, which resembled the effects of neonatal serotonin depletion (Neuroreport, 1997). Therefore, the results of injecting NAN-190 to the rat pups point to a deficit of trophic developmental influences of serotonin, adding new arguments for the hypothesis of a trophic role of 5-HT1A receptors in the brain development.

Neonatal depletion of serotonin increases the numbers of callosally projecting neurons in cat visual areas 17 and 18.

We investigated the influence of neonatal depletion of serotonin on the developmental reduction of callosal connections in cat visual cortex. Neonatal kittens were injected with 5,7-dihydroxytryptamine. At the age of 3 months, Fast Blue was injected into visual areas of one hemisphere in these and control cats and retrogradely labeled perikarya were mapped in the opposite hemisphere. In both groups callosally projecting neurons were found in a 3-5 mm wide belt centered on the transient zone of areas 17 and 18. However, numbers of labeled neurons were twice higher in the serotonin-depleted cats. We postulate that normally serotonin intensifies the process of axon pruning by augmenting developmental plasticity, therefore its depletion reduced the plasticity and more axons targeting callosal zones were stabilized, even though ectopic projections were still eliminated.

Reorganization of the corticotectal projections introduced by neonatal monocular enucleation in the Monodelphis opossum and the influence of serotoninergic depletion.

The influence of neonatal serotoninergic lesion (performed with s.c. injection of 5,7-dihydroxytryptamine) on the plasticity of the developing corticotectal projection was studied in the gray short-tailed opossum (Monodelphis domestica). As a first step, the placement and density of neurons projecting from the visual cortical areas to the superior colliculus was established in the adult opossum. Injections of retrogradely transported fluorescent dyes into the superior colliculus of intact three-month-old animals labeled neurons of cortical layer V. In this species, there are three visual areas: the striate area and two secondary areas, the laterally placed peristriate area and the medial visual area. The population of the labeled neurons was denser in peristriate and medial visual areas than in the striate area. Secondly, the influence of neonatal monocular enucleation on the extent of this projection was investigated, alone or in combination with a serotoninergic lesion. Injection of dyes into the superior colliculi of three-month-old animals that were unilaterally enucleated on the second postnatal day also labeled neurons of cortical layer V. However, the density of the cortical neurons projecting to the superior colliculus contralateral to the remaining eye was much lower. This reduction was most profound in the striate visual area. No significant modifications of this projection were found on the side ipsilateral to the remaining eye. In another group of opossums, unilateral enucleation on the second postnatal day was combined with serotoninergic lesion. Brains of some of the treated pups were immunostained for serotonin on the fifth postnatal day. At this age, 70-80% of serotoninergic axons in the brain were missing. However, in about three weeks these axons had regrown, and their density in the neocortex was approximately the same as in the control animals. We conclude that severe reduction of the serotoninergic innervation during the early postnatal period did not influence the plastic changes induced in the corticotectal projection by unilateral enucleation.

Spontaneous behavior of the gray short-tailed opossum (Monodelphis domestica) in the elevated plus-maze: comparison with Long-Evans rats.

We observed the spontaneous behavior of a laboratory marsupial--the gray short-tailed opossum (Monodelphis domestica)--in the elevated plus-maze (EPM) during six consecutive sessions and compared it with the behavior of Long-Evans rats. During the first exposure to the maze both species spent most of the time in the enclosed arms but opossums showed much higher frequency of entries into the open arms and stayed there longer. On the third and subsequent days opossums reduced their entries into the open arms and spent more time on the central square, where unlike rats they frequently groomed their lower belly and hind legs. During the last sessions they started spending more time in the enclosed arms. It is concluded that probably opossums, like rats show a stable anxiety evoked by open space. However, in the rat anxiety prevails over motivation to explore a new environment, while in the opossum it is initially at equilibrium with curiosity which habituates slower than in the rat. Results are discussed in the context of different ecology of the gray opossum that actively searches and hunts quickly moving insects. Thigmotaxic behavior, while strong in both species, dominates spontaneous behavior of the rat, but not opossum.

Localization of the 5-HT1A receptors in the brain of opossum Monodelphis domestica.

This paper describes the distribution of 5-HT1A receptors in the brain of opossum Monodelphis domestica. They were visualized by immunohistological staining with an antibody against the amino acid sequence (170-186) of this receptor that was previously successfully used in the rat and monkey. As in Eutherians, high levels of immunostaining were present in the septum, hippocampus, raphe nuclei and some other brain stem nuclei. Neocortex, several thalamic nuclei and hypothalamus showed moderate density of the labeled structures. Moderate levels of 5-HT1A receptors were also observed in the caudate nucleus and putamen, unlike in the rat, in which labeling in these nuclei was almost absent. Another difference with the rat was observed in the neocortex: in the opossum immunostaining was absent in the layer 4 of many cortical areas. In general, distribution and density of this important receptor in the opossum is very similar to that described in the rat and monkey and therefore it follows a general mammalian pattern.

Neonatal serotonin depletion modifies development but not plasticity in rat barrel cortex.

Effects of serotonin depletion (induced by neonatal injection of 5,7-dihydroxytryptamine) upon dimensions of cortical barrels and their metabolic activation, and upon effects of neonatal vibrissectomy sparing row C, were examined in 1-month-old rats. Dimensions of row C barrels, and of [14C]2-deoxyglucose (2-DG) labelling in the cortex obtained after stimulation of the row C vibrissae, were measured. Serotonin depletion did not change dimensions of barrels, but reduced the extent of 2-DG labelling of cortical representation of the row C whiskers by 30%. Vibrissectomy sparing this row resulted in an expansion of the row C barrels and of 2-DG labelling in the barrel cortex that were similar in both control and serotonin-depleted rats.

Areas PMLS and 21a of cat visual cortex: two functionally distinct areas.

We have compared the receptive field properties of neurons recorded from visuotopically corresponding regions of area 21a and the posteromedial lateral suprasylvian area (PMLS) of cat visual cortex. In both areas, the great majority of neurons were orientation-selective and binocular, and their responses to moving contours were modulated by simultaneous in-phase or anti-phase motion of large textured background stimuli ('visual noise'). However, despite the great hodological similarity between the two areas, PMLS neurons had on average significantly higher peak discharge rates, exhibited substantially greater direction selectivity indices, and preferred substantially higher stimulus velocities than area 21a neurons. Furthermore, the majority of binocular neurons in the PMLS area and in area 21a were dominated respectively by contralateral and ipsilateral eyes. Finally, while 46% of PMLS neurons were excited by movement of visual noise per se, only 25% of area 21a neurons could be excited by such stimuli. We argue that the PMLS area, like its presumed primate homologue the middle-temporal (MT) area, is mainly involved in motion analysis. By contrast, area 21a appears to be involved in pattern analysis rather than motion analysis. It is likely that phylogenetically area 21a derives from the PMLS area.

Evolutionary ancient roles of serotonin: long-lasting regulation of activity and development.

Biogenic monoamines (catecholamines, indoleamines and histamine) are evolutionary old and important modulators of long-lasting changes in the functional state of cells. They are found in many protozoans and in almost all metazoans. Monoamines preserve their evolutionary old functions (first of all being intracellular signals and later hormones and growth factors) even in those animals in which they acquired the function of neurotransmitter. The older functions of serotonin, an important member of the family of indoleamines, are reviewed here. Described are: presence of serotonin in organisms at various phylogenetic levels; its role in embryonal, foetal and postnatal development, especially in the development of the central nervous system. It is concluded that in none of these functions serotonin is the only factor, but it is an ubiquitous and important modulator of a vast array of processes and functions taking part in development and plasticity.

Areas PMLS and 21a of cat visual cortex are not only functionally but also hodologically distinct.

In several cats, paired visuotopically matched injections of retrogradely transported fluorescent dyes, diamidino yellow (DY) and fast blue (FB), were made into two visuotopically organized, functionally distinct extrastriate cortical areas, the posteromedial lateral suprasylvian area (PMLS area) and area 21a respectively. After an appropriate survival time, the numbers of thalamic, claustral and cortical cells which were single-labelled with each dye as well as the numbers of cells in these structures labelled with both dyes (double-labelled cells) were assessed. The clear majorities of thalamic cells projecting to PMLS area (DY labelled cells) and to area 21a (FB labelled cells) were located in the ipsilateral lateral posterior-pulvinar complex with smaller proportions located in the laminae C and the medial intralaminar nucleus of the ipsilateral dorsal lateral geniculate nucleus and several nuclei of the rostral intralaminar thalamic group. Despite the fact that DY labelled (PMLS-projecting) and FB labelled (area 21 a-projecting) cells in all thalamic nuclei were well intermingled, only 1-5% of retrogradely labelled thalamic cells projected to both areas (cells double-labelled with both dyes). Small proportions of retrogradely labelled cells were located in the ipsilateral and to a lesser extent the contralateral dorsocaudal claustra. The proportions of claustral neurons retrogradely labelled with both dyes varied from 4 to 9%. Over half of the cortical neurons labelled retrogradely from area 21a or PMLS area were located in the supragranular layers of the ipsilateral area 17, with smaller proportions located in the supragranular layers of the ipsilateral areas 18 and 19 and even smaller proportions located in mainly but not exclusively, the infragranular layers of the ipsilateral areas 21b and 20a. Again despite strong spatial intermingling of neurons labelled with DY and these labelled with FB, the proportions of associational cortical neurons double-labelled with both dyes were small (2 to 5.5%). Finally, small proportions of neurons retrogradely labelled with DY or FB were located, mainly but not exclusively, in the supragranular layers of the contralateral areas 17, 18, 19 and 21a. Again, the proportions of the double-labelled neurons in the contralateral cortices were small (1-4.5%). Thus, the present study indicates that despite the fact that the diencephalic and telencephalic inputs to the visuotopically corresponding parts of area 21a and PMLS area originate from the same nuclei, areas and layers, the two areas receive their afferents from the largely separate populations of neurons.

Prenatal monocular enucleation induces a selective loss of low-spatial-frequency cortical responses to the remaining eye.

During early development, interactions between the two eyes are critical in the formation of eye-specific domains within the lateral geniculate nucleus and the visual cortex. When monocular enucleation is done early in prenatal life, it induces remarkable anatomical and functional reorganizations of the visual pathways. Behavioral data have shown a loss in sensitivity to low-spatial-frequency gratings in cats. To correlate the behavioral observations with a possible change in the analysis of contrast at the level of primary visual areas we recorded visual evoked potentials at the 17/18 border in two cats enucleated prenatally (gestational age at enucleation, 39-42 days), three neonatal, two control animals, and one animal with a surgical removal of Y-ganglion fibers. Our results show a strong attenuation in the amplitude of response at all contrast values for gratings of low spatial frequency in prenatally enucleated cats, whereas neonatally enucleated and control animals present responses of comparable amplitude. We conclude that the behavioral results reflect the reduced sensitivity for low frequencies of visual cortical neurons. In addition, we define a critical period for the development of the contrast-sensitivity function that seems to be limited to the prenatal gestation period. We suggest that the prenatal interruption of binocular interactions leads to a functional elimination of the Y-ganglion system.

Extent of bilateral collateralization among pontomesencephalic tegmental afferents to dorsal lateral geniculate nuclei of pigmented and albino rats.

In adult pigmented and albino rats, small amounts of different fluorescent dyes (Fast Blue and Fluoro-Gold) were pressure-injected into the dorsal lateral geniculate nuclei, each nucleus (right or left) being injected with one dye only. After postinjection survival of three days, the distribution of neurons retrogradely labelled by each dye was analysed. Consistent with previous studies, in each strain each dye labelled a large number of neurons in the several ipsilateral visuotopically or retinotopically organized structures--visual cortices, retino-recipient layers of the superior colliculi and the pretectal nuclei. A substantial number of retrogradely labelled neurons was also found in the contralateral parabigeminal nucleus. A few retrogradely labelled neurons were found in the ipsilateral and (to a lesser extent) contralateral dorsolateral divisions of the periaqueductal gray matter, as well as in the ipsilateral parabigeminal nucleus and the caudal part of the lateral hypothalamus. However, in all the above structures there was a paucity of cells retrogradely labelled with both dyes (double-labelled cells). By contrast, in each strain, several "modulatory" nuclei (containing cholinergic and aminergic cells) of the pontomesencephalic tegmentum--dorsal raphe, pedunculopontine tegmental nucleus, parabrachial nucleus, laterodorsal tegmental nucleus and locus coeruleus--contained significant numbers of cells projecting to both ipsilateral and contralateral dorsal lateral geniculate nuclei. In each nucleus, ipsilaterally and contralaterally projecting cells constituted, respectively, about 65-70% and about 30-35% of retrogradely labelled cells. About 25% of the contralaterally projecting cells (i.e. about 5-10% of all retrogradely labelled tegmental neurons) were double-labelled with both dyes. Double-labelled cells were intermingled with single-labelled cells projecting ipsilaterally or contralaterally. The proportions of the ipsilaterally, contralaterally and bilaterally projecting neurons in the modulatory components of the pontomesencephalic tegmentum were virtually identical in pigmented and albino strains. It appears that in both strains the visuotopically organized structures convey to the dorsal lateral geniculate nuclei information related mainly to the contralateral visual field. The projections from these structures might play an important role in regulating transmission of visual information in the retinotopically distinct parts of each dorsal lateral geniculate nucleus. By contrast, the projections from the modulatory nuclei of the pontomesencephalic tegmentum are likely to contribute to the functional synchronization of both dorsal lateral geniculate nuclei during the sleep-wakefulness cycle and saccadic eye movements.

Projection of visuotopically organized afferents to the dorsal thalamus in the opossum, Monodelphis domestica.

Retrogradely transported dyes, Fluorogold and Fast Blue were injected into both sides of the dorsal thalamus in the Monodelphis opossum. Projection of the presumed primary visual cortical area, superior colliculus and parabigeminal nucleus to the dorsal lateral geniculate nucleus and the lateral posterior--lateral intermedius nuclear complex were described. They show close similarities to the homologous projections in the North American Opossum, insectivores and some rodents. In comparison with rat, cortico-thalamic and tecto-thalamic projections in the Monodelphis are less numerous. The peculiarity of cytoarchtitectonics of cortical layer 6 is described and discussed.

Parabigeminal, pretectal and hypothalamic afferents to rat's dorsal lateral geniculate nucleus. Comparison between albino and pigmented strains.

In adult pigmented and albino rats different fluorescent dyes were injected into the dorsal lateral geniculate nuclei of opposite sides. Differences between the strains occur mainly in parabigemino-geniculate and pretecto-geniculate projections. Both the major contralateral and the minor ipsilateral parabigemino-geniculate projections in albinos were clearly smaller then those in pigmented rats. In pigmented rats but not in albinos the parabigemino-geniculate projections originated mainly from the region where the vertical meridian is represented and contained a small number of neurones projecting bilaterally. In each strain, a small number of retrogradely labelled neurones was found in the ipsilateral and contralateral lateral hypothalami.

Properties of the visually driven neurons of cat's hippocampal regions CA 1 and CA 3.

Neurons in areas CA 1 and 3 of cat's dorsal hippocampus were studied. Fifteen percent of the investigated cells were influenced by visual stimuli. Eighty five such neurons were investigated. The organization of their receptive fields was tested with stationary and moving visual stimuli. Twenty eight percent of neurons had small receptive fields (10-20 deg square). Forty one neurons responded to stationary flashing spots. They were ON-OFF, ON and OFF types with phasic (66%) and tonic (34%) characteristics. Seventy five responded to dark and bright stimuli moving across their receptive fields. Twenty five neurons were direction-sensitive and 21 responded better to the dark moving stimuli than to the bright ones. No significant differences in the response properties of neurons in the CA 1 and CA 3 fields were observed.

The inhibitory components in the responses of the lateral suprasylvian area neurons to moving stimuli in cats.

The inhibitory components in the neuronal responses of the cat's lateral suprasylvian area (LSA) to moving bright and dark stimuli were investigated. The LSA neurons could be divided into two groups. Neurons of the first group (33%) do not reveal spatial displacement of the inhibitory zones and show displacement of the discharge centers in the receptive field only for one polarity of contrast of moving stimuli, either brighter or darker than the background. The second group (67%) contained the neurons which showed a spatial displacement of the inhibitory components and discharge centers in the receptive field for either polarity of contrasts of the moving stimuli. Tested with stationary flashing stimuli, the majority of neurons in both groups had overlapping ON-OFF discharge regions within their receptive fields. The results obtained with moving stimuli of different speeds and with the masking method suggest the rebound origin of the inhibitory responses in LSA neurons.