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.

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.

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.

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.

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.

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.

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.

The fine structure of the receptive fields of visually driven neurons in the cat's lateral suprasylvian area.

The fine structure of the receptive fields of the lateral suprasylvian area neurons was investigated in the pretrigeminal cat preparation. A majority of the receptive fields consisted of subregions with different qualitative characteristics according to their response to moving visual stimuli. There was an asymmetry in the spatial distribution of inhibitory mechanisms over the receptive field. The steady illumination of the receptive field usually enhanced the inhibitory processes, whereas darkness, decreased the effectivity of inhibitory influences on the neuron. Some receptive fields of neurons reacted vigorously to the motion of borders of visual stimuli. It is suggested that the differences in response patterns to moving stimuli depend in part on the heterogenous fine structure of their receptive fields.

Dynamic and static properties of neurons in the lateral suprasylvian area of the cat.

The response properties and receptive field organization of 106 LSA neurons were investigated in pretrigeminal preparations using moving dark and bright stimuli with particular reference to the dynamic and static structure of receptive fields. About 61percent neurons revealed equal sensitivity to the motions of dark and bright stimuli. A majority of them had an ON-OFF static structure of receptive fields. Nearly 24 percent of cells had higher sensitivity to the motion of dark stimuli in comparison with the bright ones. Their receptive fields constituted the OFF-uniform spatial distribution when tested by stationary flashing lights. No clear-cut correlations were found between the static and dynamic properties of LSA neurons. It was shown that the same region of the receptive field is responsible for the reactions to dark or bright moving stimuli. A group of dark-sensitive neurons (24 percent of the total) was described, some examples of which had no stationary receptive fields at all.

Responses of neurons in cat's lateral suprasylvian area to moving light and dark stimuli.

About 55% of the cells in the cat lateral suprasylvian area were truly direction-sensitive to moving light and dark spots. For these cells the preferred direction of stimulus movement was the same for the light and dark spots. The most significant effects of the reversal of stimulus contrast was a change in the strength of the responses. A population of "dark-sensitive" neutrons was found that responded vigorously to the movement of dark spots but only very weakly to the movement of light spots. These neurons were only weakly sensitive to stationary flashing lights and responded with On/Off discharges over the whole of their receptive fields.

Retinotopic organization of the lateral suprasylvian area of the cat.

The retinotopic organization of the lateral suprasylvian area (L.S.A.) was investigated by microelectrode recording from single neurons. 560 penetrations were made on the medial and lateral banks of the middle suprasylvian sulcus (M.S.S.). The receptive field positions, of single neurons were mapped in relation to the retinal landmarks. A striking variability of the visuotopic organization from one animal to another was observed. The most persistent organization was the representation of the lower contralateral quadrant of the visual field in the rostra1 and middle parts of the L.S.A. and a slight shift to the upper contralateral quadrant of the visual field in the posterior regions of the L.S.A.