Rouzanna L. Djavadian

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.

Organic cation/carnitine transporter OCTN3 is present in astrocytes and is up-regulated by peroxisome proliferators-activator receptor agonist.

In the brain beta-oxidation, which takes place in astrocytes, is not a major process of energy supply. Astrocytes synthesize important lipid metabolites, mainly due to the processes taking place in peroxisomes. One of the compounds necessary in the process of mitochondrial beta-oxidation and export of acyl moieties from peroxisomes is l-carnitine. Two Na-dependent plasma membrane carnitine transporters were shown previously to be present in astrocytes: a low affinity amino acid transporter B(0,+) and a high affinity cation/carnitine transporter OCTN2. The expression of OCTN2 is known to increase in peripheral tissues upon the stimulation of peroxisome proliferators-activator receptor alpha (PPARalpha), a nuclear receptor known to up-regulate several enzymes involved in fatty acid metabolism. The present study was focused on another high affinity carnitine transporter-OCTN3, its presence, regulation and activity in astrocytes. Experiments using the techniques of real-time PCR, Western blot and immunocytochemistry analysis demonstrated the expression of octn3 in rat astrocytes and, out of two rat sequences ascribed as similar to mouse OCTN3, XM_001073573 was found in these cells. PPARalpha activator-2-[4-chloro-6-[(2,3-dimethylphenyl)amino]-2-pyrimidinyl]thio]acetic acid (WY-14,643) stimulated by 50% expression of octn3, while, on the contrary to peripheral tissues, it did not change the expression of octn2. This observation was correlated with an increased Na-independent activity of carnitine transport. Analysis by transmission electron microscopy showed an augmented intracellular localization of OCTN3 upon PPARalpha stimulation, mainly in peroxisomes, indicating a physiological role of OCTN3 as peroxisomal membrane transporter. These observations point to an important role of OCTN3 in peroxisomal fatty acid metabolism in astrocytes.

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.

Response to novelty in the laboratory Wistar rat, wild-captive WWCPS rat, and the gray short-tailed opossum (Monodelphis domestica).

Behavior of the laboratory gray short-tailed opossums (Monodelphis domestica), Warsaw Wild Captive Pisula Stryjek rats (WWCPS) and laboratory rats (Wistar) has been registered in the period of familiarization with a new environment and consecutive confrontation with a novel, innocuous object placed in that familiarized environment. In the new environment the sequence of anxiety, investigation, and habituation was shortest in the opossum, longer in the laboratory rat and longest in the WWCPS rat. When placed in it, gray short-tailed opossums investigated the new environment with the shortest delay and most intensity. In reaction to novel objects, opossums and laboratory rats prolonged the time spent in the proximity of the new object, while the WWCPS rat did not show that reaction. Both opossums and laboratory rats increased the number of contacts with the new object, whereas WWCPS rats reduced those contacts. Behavior of all three species and lines grouped in different clusters. Some other quantitative and qualitative differences in behavior of the investigated animals are also described, showing a higher level of anxiety in both lines of rats than in the opossum. Behavioral differences between species and lines of animals used in this study may be attributed to different ecological adaptations of rats and opossums and to the effect of domestication in the laboratory rats. These behavioral differences make comparisons of opossums vs rat, and wild rat vs laboratory rat interesting models for studying the brain mechanisms of anxiety and neotic motivations.

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.

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.

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.

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.

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.

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 Monodelphis opossum 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.