Corrigendum: Downregulation of TrkC Receptors Increases Dendritic Arborization of Purkinje Cells in the Developing Cerebellum of the Opossum, Monodelphis domestica.

[This corrects the article DOI: 10.3389/fnana.2020.00056.].

Roles of the exon junction complex components in the central nervous system: a mini review.

The exon junction complex (EJC) consists of four core proteins: Magoh, RNA-binding motif 8A (Rbm8a, also known as Y14), eukaryotic initiation factor 4A3 (eIF4A3, also known as DDX48), and metastatic lymph node 51 (MLN51, also known as Casc3 or Barentsz), which are involved in the regulation of many processes occurring between gene transcription and protein translation. Its main role is to assemble into spliceosomes at the exon-exon junction of mRNA during splicing. It is, therefore, a range of functions concerning post-splicing events such as mRNA translocation, translation, and nonsense-mediated mRNA decay (NMD). Apart from this, proteins of the EJC control the splicing of specific pre-mRNAs, for example, splicing of the mapk transcript. Recent studies support essential functions of EJC proteins in oocytes and, after fertilization, in all stages of zygote development, as well as the growth of the embryo, including the development of the nervous system. During the development of the central nervous system (CNS), the EJC controls mitosis, regulating both symmetric and asymmetric cell divisions. Reduced levels of EJC components cause microcephaly. In the adult brain, Y14 and eIF4A3 appear to be involved in synaptic plasticity and in learning and memory. In this review, we focus on the involvement of EJC components in brain development and its functioning under normal conditions.

Stress-Dependent Changes in the CacyBP/SIP Interacting Protein S100A6 in the Mouse Brain.

The CacyBP/SIP target S100A6 is widely present in the nervous system, and its up-regulation is associated with certain neurodegenerative diseases. Here, we examined the involvement of S100A6 protein in stress responses in mice. Using Western blotting, we observed a marked change in brainstem structures, whereby stressed mice showed approximately one-third the protein level produced in the control group. A decreased level of S100A6 protein in stressed animals was also detected in the olfactory bulb and the cerebellum and stress-related structures such as the hippocampus and the hypothalamus. Additionally, using immunohistochemistry, high levels of S100A6 expression were observed in astrocytes localized in the border zones of all brain ventricles, tanycytes of the ventro-lateral walls of the hypothalamus, including the arcuate nucleus (ARH) and low levels of this protein were in neurons of the olfactory bulb, the hippocampus, the thalamus, the cerebral cortex, the brainstem and the cerebellum. Although S100A6-expressing cells in all these brain structures did not change their phenotype in response to stress, the intensity of immunofluorescent labeling in all studied structures was lower in stressed mice than in control animals. For example, in the ARH, where extremely strong immunostaining was observed, the number of immunolabeled fibers was decreased by approximately half in the stressed group compared with the controls. Although these results are descriptive and do not give clue about functional role of S100A6 in stress, they indicate that the level of S100A6 decreases in several brain structures in response to chronic mild stress, suggesting that this protein may modify stress responses.

Distribution and function of TrkB receptors in the developing brain of the opossum Monodelphis domestica.

The expression, development pattern, spatiotemporal distribution, and function of TrkB receptors were investigated during the postnatal brain development of the opossum. Full-length TrkB receptor expression was detectable in the newborn opossum, whereas three different short forms that are expressed in the adult brain were almost undetectable in the newborn opossum brain. The highest level of full-length TrkB receptor expression was observed at P35, which corresponds to the time of eye opening. We found that in different brain structures, TrkB receptors were localized in various compartments of cells. The hypothalamus was distinguished by the presence of TrkB receptors not only in cell bodies but also in the neuropil. Double immunofluroscent staining for TrkB and a marker for the identification of the cell phenotype in several brain regions such as the olfactory bulb, hippocampus, thalamus, and cerebellum showed that unlike in eutherians, in the opossum, TrkB receptors were predominantly expressed in neurons. A lack of TrkB receptors in glial cells, particularly astrocytes and oligodendrocytes, provides evidence that TrkB receptors can play a functionally different role in marsupials than in eutherians. The effects of TrkB signaling on the development of cortical progenitor cells were examined in vitro using shRNAs. Blockade of the endogenous TrkB receptor expression induced a decrease in the number of progenitor cells proliferation, whereas the number of apoptotic progenitor cells increased. These changes were statistically significant but relatively small. In contrast, TrkB signaling was strongly involved in regulation of the cortical progenitor cell differentiation process.

Expression of TrkC receptors in the developing brain of the Monodelphis opossum and its effect on the development of cortical cells.

In this study, we investigated the distribution, localization and several various functions of TrkC receptors during development of the Monodelphisopossum brain. Western blotting analysis showed that two different forms of the TrkC receptor, the full-length receptor and one of its truncated forms, are abundantly expressed in the opossum brain. The expression of TrkC receptors was barely detected in the brain of newborn opossums. At postnatal day (P) 3, the expression of full-length TrkC remained at low levels, while moderate expression of the TrkC truncated form was detected. The expression levels of both forms of this protein gradually increased throughout development, peaking at P35. We found that in different neocortical areas located both at the rostral and caudal regions of the cortex, up to 98% of BrdU-labeled cells forming cortical layers (II-VI) had prominently expressed TrkC. To assess which developmental processes of cortical cells are regulated by TrkC receptors, three different shRNAs were constructed. The shRNAs were individually tested in transfected cortical progenitor cells grown on culture plates for 2 days. The effects of the shRNA-TrkC constructs were similar: blockade of TrkC receptors decreased the number of Ki67-positive and apoptotic cells, and it did not change the number of TUJ-positive neurons in vitro. Thus, the lack of TrkC receptors in cultured progenitor cells provided insight on the potential role of these receptors in the regulation of proliferation and cell survival but not in the differentiation of cortical cells.

Lipopolysaccharide injected to pregnant mice affects behavior of their offspring in adulthood.

We studied consequences of maternal immune response on the course of pregnancy and the behavior of adult offspring. Mice in late gestation (day 16-17) were injected with lipopolysaccharide (LPS). Treatment of pregnant mice with high doses of LPS resulted in fetal resorption or stillbirths. Pregnant mice treated with low doses (100 or 300 micrograms/kg) of LPS gave birth to normal numbers of pups. However, behavior of the offspring was altered. Adult offspring of dams injected at a dose of 300 micrograms/kg of LPS traveled longer distances in the open field and spent more time in the central part of the arena, than mice in the control group. Female mice of this group spent more time in open arms of the elevated plus maze, in comparison to female control mice. Results of the Morris water maze test showed impairment of spatial learning and memory in male offspring born to LPS-injected dams. Furthermore, in the nest building test adult mice born from LPS challenged pregnancies constructed worse quality nests, which points to the presence of hippocampal dysfunction. These findings indicate that maternal bacterial infections during pregnancy may alter offspring behavior in adult life.

Neurotrophins and their receptors in early development of the mammalian nervous system.

Neurotrophins belonging to the class of growth factors and including nerve growth factor (NGF), brain-derived neurotrophic factor (BDNF), neurotrophin-3 (NT-3) and neurotrophin-4/5 (NT-4/5) are widely recognized as essential factors in the developing central nervous system (CNS). Neurotrophins are synthesized as precursor forms (proneurotrophins). Mature forms of neurotrophins exert their effect by binding to specific tyrosine kinases receptors (TrkA, TrkB and TrkC) as well as via the p75 receptor, a member of the tumor necrosis factor receptor superfamily while proneurotrophins interact with the receptor p75 or co-receptor complex of p75 and sortilin, that is a Vps10p domain-containing transmembrane protein. Expression of neurotrophins corresponds with the onset of neurogenesis in developing mammalian species. BDNF is low in early embryonic stages of development, while NT-3 highly expresses in the developing CNS. Expression of neurotrophins receptors mainly overlaps at early development. Data concerning early distribution of neurotrophins and their receptors in the nervous system and results in mice with targeted disruptions of neurotrophin or receptor genes show that neurotrophins and their receptors play distinct roles in control and regulation of the most crucial developmental processes such as proliferation, migration, differentiation, survival, apoptosis and synaptic plasticity.

Reduction of the number of new cells reaching olfactory bulbs impairs olfactory perception in the adult opossum.

In adult mammals cells generated in the subventricular zone (SVZ) migrate to olfactory bulbs (OB). Functional significance of this continuous neurogenesis is not clear. We injected opossums (Monodelphis domestica) for seven consecutive days with a 5HT(1A) agonist (8-OH-DPAT or buspirone) or its antagonist WAY100635. One hour after each of these injections bromodeoxyuridine (BrdU) a marker of dividing cells was also injected. Two months later, when newly generated neurons settled in the OB and matured the ability of these opossums to detect hidden food by olfactory cues was tested. Afterwards, numbers of BrdU-labeled cell nuclei in their OB were counted and a phenotype of labeled cells established. In all groups investigated the majority of new cells differentiated into neurons (55-76%) and a lower proportion into astroglia (6-12%). Numbers of BrdU-labeled cells differed depending on the applied treatment: both agonists of the 5HT(1A) receptor increased these numbers, while its antagonist decreased them. The increased number of new OB interneurons did not change the time required for finding all three food items and therefore did not improve the opossums' performance in this test of the olfactory perception. However, opossums that had the reduced number of new generated OB cells searched longer for each food item and in consequence took three times longer to find all three crickets, than did opossums from other groups. In conclusion, lower numbers of new neurons in the opossums OB correlated with their worse behavioral performance in a test based on olfactory perception.

The partial 5-HT1A receptor agonist buspirone enhances neurogenesis in the opossum (Monodelphis domestica).

We demonstrate for the first time that neurogenesis in the adult Monodelphis opossum has a typical mammalian pattern and occurs only in the dentate gyrus (DG) and subventricular zone (SVZ) of the lateral ventricles. In these two brain regions neurogenesis is present throughout the lifespan, although its rate is reduced by half in the old age. Treatment with buspirone, a partial 5-HT1A receptor agonist which is used in human clinic as an anxiolytic agent, boosts proliferation in the SVZ and DG in both adult and aged opossums. The neuronal phenotype dominates among newly generated cells in both non-treated and buspirone-treated opossums. We suggest that if functional importance of adult neurogenesis is in improving olfactory discrimination and generation of hippocampus-dependent memory, both spatial and emotional, then administration of drugs increasing the rate of neurogenesis via activation of 5-HT1A receptors may be a valuable aid in combating problems of the advanced age.

Thalamic nuclei in the opossum Monodelphis domestica.

We investigated nuclear divisions of the thalamus in the gray short-tailed opossum (Monodelphis domestica) to gain detailed information for further developmental and comparative studies. Nissl and myelin staining, histochemistry for acetylcholinesterase and immunohistochemistry for calretinin and parvalbumin were performed on parallel series of sections. Many features of the Monodelphis opossum thalamus resemble those in Didelphis and small eutherians showing no particular sensory specializations, particularly in small murid rodents. However, several features of thalamic organization in Monodelphis were distinct from those in rodents. In the opossum the anterior and midline nuclear groups are more clearly separated from adjacent structures than in eutherians. The dorsal lateral geniculate nucleus (LGNd) starts more rostrally and occupies a large part of the lateral wall of the thalamus. As in other marsupials, two cytoarchitectonically different parts, alpha and beta are discernible in the LGNd of the opossum. Each of them may be subdivided into two additional bands in acetylcholinesterase staining, while in murid rodents the LGNd consists of a homogeneous mass of cells. Therefore, differentiation of the LGNd of the Monodelphis opossum is more advanced than in murid rodents. The medial geniculate body consists of three nuclei (medial, dorsal and ventral) that are cytoarchitectonically distinct and stain differentially for parvalbumin. The relatively large size of the MG and LGNd points to specialization of the visual and auditory systems in the Monodelphis opossum. In contrast to rodents, the lateral dorsal and lateral posterior nuclei in the opossum are poorly differentiated cytoarchitectonically.

Generation recruitment and death of brain cells throughout the life cycle of Sorex shrews (Lipotyphla).

Young shrews of the genus Sorex that are born in early summer reduce their body size before wintering, including a reduction of brain weight of 10-30%. In the spring they mature sexually, double their body weight and regain about half of the loss in brain weight. To investigate the mechanisms of brain weight oscillations we studied the rate of cell death and generation in the brain during the whole life cycle of the common shrew (Sorex araneus) and pygmy shrew (S. minutus). After weaning, shrews generate new brain cells in only two mammalian neurogenic zones and approximately 80% of these develop into neurones. The increase of the shrew brain weight in the spring did not depend on recruitment of new cells. Moreover, adult Sorex shrews did not generate new cells in the dentate gyri. Injections of 5-HT1A receptor agonists in the adult shrews induced neurogenesis in their dentate gyri, showing the presence of dormant progenitor cells. Generation of new neurones in the subventricular zone of the lateral ventricles and their recruitment to olfactory bulbs continued throughout life. TUNEL labelling showed that the rate of cell death in all brain structures, including the proliferation zones and olfactory bulb, was very low throughout life. We conclude that neither cell death nor recruitment significantly contributes to seasonal oscillations and the net loss of brain weight in the Sorex shrews. With the exception of dentate gyrus and olfactory bulb, cellular populations of brain structures are stable throughout the life cycle of these shrews.

The visual system in subterranean African mole-rats (Rodentia, Bathyergidae): retina, subcortical visual nuclei and primary visual cortex.

We have studied the visual system of subterranean mole-rats of the rodent family Bathyergidae, for which light and vision seem of little importance. The eye diameter varies between 3.5mm in Bathyergus suillus and 1.3mm in Heterocephalus glaber. The small superficial eyes have features typical of sighted animals (clear optics, well-developed pupil and well-organized retina) and appear suited for proper image formation. The retinae are rod-dominated but possess rather high cone proportions of about 10%. The total number of retinal ganglion cells and optic nerve fibres ranges between 6000 in Bathyergus suillus and 2100 in Heliophobius argenteocinereus. Visual acuity (estimated from counts of peak ganglion cell density and axial length of the eye) is low, ranging between 0.3 and 0.5 cycles/degree. The retina projects to all the visual structures described in surface-dwelling sighted rodents. The suprachiasmatic nucleus is large and receives bilateral retinal input. All other visual nuclei are reduced in size and receive almost exclusively contralateral retinal projections of varying magnitude. The primary visual cortex is small and, in comparison to other rodents, displaced laterally. In conclusion, the African mole-rats possess relatively well-developed functional visual subsystems involved in photoperiodicity, form and brightness discrimination. In contrast, visual subsystems involved in coordination of visuomotor reflexes are severely reduced. This pattern suggests the retention of basic visual capabilities. Residual vision may enable subterranean mammals to localize breaches in the burrows that let in light thus providing a cue to enable mole-rats to reseal such entry points and to prevent entry of predators.

Acoustic startle response in the opossum Monodelphis domestica in comparison with the Wistar albino rat.

The acoustic startle response (ASR) was studied in 20 gray short-tailed opossums (Monodelphis domestica) and its characteristics were compared with those obtained from responses of 20 Wistar albino rats. The animals were exposed to 10 ms, 110 dB white noise acoustic pulses in the Coulbourn apparatus. Amplitude of ASR was normalized to the weight of animals and then analyzed. The results show that probability of a freezing response is much higher in the opossums that generally respond with lower startle amplitudes in comparison to rats. These differences may reflect different emotional characteristics of the two species, different reactions to fear in opossum and/or different ecological specializations of the two species.

Apparent scarcity of glial fibrillary acidic protein expression in the brain of the pygmy shrew Sorex minutus as revealed by immunocytochemistry.

We examined astroglial cells in the brain of the pygmy shrew Sorex minutus (Insectivora). For that purpose we labeled glial fibrillary acidic protein (GFAP) immunohistochemically in brain sections with a polyclonal antibody. Antigen retrieval experiments were performed to counteract formaldehyde fixation masking of GFAP epitopes. Our results showed remarkable paucity of GFAP-immunoreactive cells and fibers in the cerebral cortex and nuclei, as well as in the majority of the diencephalic and mesencephalic structures. In the forebrain, significant numbers of GFAP-containing astrocytes were found only in the ependyma and subventricular zones, superficial part of layer I of the cerebral cortex, and the majority of white matter structures. In the diencephalon, habenular nuclei were rich in GFAP-immunopositive astrocytes and labeled radial fibers were extended between median eminence and the third ventricle. A considerably higher density of labeled astrocytes was detected in the caudal brainstem and cerebellum. In contrast, in the mouse brain, immunoreactive astrocytes were present in large quantities in various structures. Staining of sections of the shrew brain against glutamine synthetase revealed abundance of immunofluorescent astrocytes in many areas, especially in the shrew cerebral cortex. It seems probable that in the shrew brain only a limited fraction of astroglia expresses GFAP, while other astroglial cells can be detected with different markers. It is possible that the rodent type of astroglial GFAP expression might not be common to insectivores and probably to some other mammalian orders.

Behavior of the gray short-tailed opossum (Monodelphis domestica) in the open field and in response to a new object, in comparison with the rat.

We compared the behavior of the gray short-tailed opossums (Monodelphis domestica) and Long-Evans rats during repeated exposures to the open-field (OF) test. Animals were videotaped for 10 min on four consecutive days. A new object was placed in the center of the field on the third day and it was present there again on the fourth day. The rate of locomotor activity in the opossum was always higher than that in the rat. On the first exposure to the open field, both species showed strong thigmotaxy. On the second day, opossums shifted a significant part of their activity to the internal and central parts of the field, while thigmotaxy dominated in the rats' behavior till the end of the experiment. The frequency and time of exploration of a new object placed on the central square was higher in the opossums than in rats. They also showed higher frequency of rearings and lower defecation scores, while the time of grooming was similar to the rats'. These results, that are consistent with those of our earlier experiments in the elevated plus maze (EPM), show that in response to novelty Monodelphis opossums change their behavior from defensive to exploratory faster than rats and then explore it more intensely. These differences may be either a result of different ecologies or evolution of the two species.

Life-long stability of neurons: a century of research on neurogenesis, neuronal death and neuron quantification in adult CNS.

In this chapter we provide an extensive review of 100 years of research on the stability of neurons in the mammalian brain, with special emphasis on humans. Although Cajal formulated the Neuronal Doctrine, he was wrong in his beliefs that adult neurogenesis did not occur and adult neurons are dying throughout life. These two beliefs became accepted "common knowledge" and have shaped much of neuroscience research and provided much of the basis for clinical treatment of age-related brain diseases. In this review, we consider adult neurogenesis from a historical and evolutionary perspective. It is concluded, that while adult neurogenesis is a factor in the dynamics of the dentate gyrus and olfactory bulb, it is probably not a major factor during the life-span in most brain areas. Likewise, the acceptance of neuronal death as an explanation for normal age-related senility is challenged with evidence collected over the last fifty years. Much of the problem in changing this common belief of dying neurons was the inadequacies of neuronal counting methods. In this review we discuss in detail implications of recent improvements in neuronal quantification. We conclude: First, age-related neuronal atrophy is the major factor in functional deterioration of existing neurons and could be slowed down, or even reversed by various pharmacological interventions. Second, in most cases neuronal degeneration during aging is a pathology that in principle may be avoided. Third, loss of myelin and of the white matter is more frequent and important than the limited neuronal death in normal aging.