Biomedical subjects

Aldo Fasolo

Publications and source records attributed to Aldo Fasolo.

16 recordsLinked to original sources

PubMed2007

Hepatocyte growth factor acts as a motogen and guidance signal for gonadotropin hormone-releasing hormone-1 neuronal migration.

Reproduction in mammals is under the control of the hypothalamic neuropeptide gonadotropin hormone-releasing hormone-1 (GnRH-1). GnRH-1-secreting neurons originate during embryonic development in the nasal placode and migrate into the forebrain along olfactory nerves. Gradients of secreted molecules may play a role in this migratory process. In this context, hepatocyte growth factor (HGF) is a potential candidate, because it promotes cell motility in developing brain and has been shown previously to act as a motogen on immortalized GnRH-1 neurons (GN11). In this study, the role of HGF and its receptor Met during development of the GnRH-1 system was examined. GnRH-1 cells express Met during their migration and downregulate its expression once they complete this process. Tissue-type plasminogen activator (tPA), a known HGF activator, is also detected in migratory GnRH-1 neurons. Consistent with in vivo expression, HGF is present in nasal explants, and GnRH-1 neurons express Met. HGF-neutralizing antibody was applied to explants to examine the role of the endogenous growth factor. Migration of GnRH-1 cells and olfactory axon outgrowth were significantly reduced, in line with disruption of a guidance gradient. Exogenous application of HGF to explants increased the distance that GnRH-1 cells migrated, suggesting that HGF also acts as a motogen to GnRH-1 neurons. Functional experiments, performed on organotypic slice cultures, show that creation of an opposing HGF gradient inhibits GnRH-1 neuronal migration. Finally, tPA(-/-):uPA(-/-) (urokinase-type plasminogen activator(-/-)) knock-out mice exhibit strong reduction of the GnRH-1 cell population. Together, these data indicate that HGF signaling via Met receptor influences the development of GnRH-1.

Animals↗

PubMed2006

Differential expression of neuregulins and their receptors in the olfactory bulb layers of the developing mouse.

Neuregulins (NRGs), and their cognate receptors (ErbBs), play essential roles in numerous aspects of neural development and adult synaptic plasticity. The goal of this study was to investigate the developmental expression profiles of these molecules during the olfactory bulb (OB) maturation. The OB is a highly organized structure with cell types and synaptic connections segregated into discrete anatomical layers. We employed a novel approach by combining single-layer microdissection at different development ages, with isoform-specific semi-quantitative RT-PCR and Western blotting to monitor layer-specific developmental profiles of these molecules and alternate splice variants. Layer and age specific regulation was observed for the ErbB4 splice variants JMa/JMb and NRG-1-beta1/beta2 forms. With the exception of the outermost (nerve) layer, ErbB4-JMb and NRG-1-beta1 are expressed throughout the OB and their expressions decrease in the adult age in most layers. In contrast both ErbB4-JMa and NRG-1-beta2 are highly expressed in the granule cell layer in the early postnatal OB. This early postnatal expression correlates with the dramatic change from radial glia to astrocytes and appearance of the bulk of granule cells occurring at this developmental stage.

Age Factors↗

PubMed2006

Neurogenesis in the caudate nucleus of the adult rabbit.

Stem cells with the potential to give rise to new neurons reside in different regions of the adult rodents CNS, but in vivo only the hippocampal dentate gyrus and the subventricular zone-olfactory bulb system are neurogenic under physiological condition. Comparative analyses have shown that vast species differences exist in the way the mammalian brain is organized and in its neurogenic capacity. Accordingly, we have demonstrated recently that, in the adult rabbit brain, striking structural plasticity persists in several cortical and subcortical areas. Here, by using markers for immature and mature neuronal and glial cell types, endogenous and exogenously administered cell-proliferation markers, intraventricular cell tracer injections coupled to confocal analysis, three-dimensional reconstructions, and in vitro tissue cultures, we demonstrate the existence of newly formed neurons in the caudate nucleus of normal, untreated, adult rabbit. Our results suggest that neurogenesis in the caudate nucleus is a phenomenon independent from that occurring in the adjacent subventricular zone, mostly attributable to the activity of clusters of proliferating cells located within the parenchyma of this nucleus. These clusters originate chains of neuroblasts that ultimately differentiate into mature neurons, which represent only a small percentage of the total neuronal precursors. These results indicate that striatum of rabbit represents a favorable environment for genesis rather than survival of newly formed neurons.

Animals↗

PubMed2005

cAMP response element-binding protein regulates differentiation and survival of newborn neurons in the olfactory bulb.

The transcription factor cAMP response element-binding protein (CREB) is involved in multiple aspects of neuronal development and plasticity. Here, we demonstrate that CREB regulates specific phases of adult neurogenesis in the subventricular zone/olfactory bulb (SVZ/OB) system. Combining immunohistochemistry with bromodeoxyuridine treatments, cell tracer injections, cell transplants, and quantitative analyses, we show that although CREB is expressed by the SVZ neuroblasts throughout the neurogenic process, its phosphorylation is transient and parallels neuronal differentiation, increasing during the late phase of tangential migration and decreasing after dendrite elongation and spine formation. In vitro, inhibition of CREB function impairs morphological differentiation of SVZ-derived neuroblasts. Transgenic mice lacking CREB, in a null CREM genetic background, show reduced survival of newborn neurons in the OB. This finding is further supported by peripheral afferent denervation experiments resulting in downregulation of CREB phosphorylation in neuroblasts, the survival of which appears heavily impaired. Together, these findings provide evidence that CREB regulates differentiation and survival of newborn neurons in the OB.

Animals↗

PubMed2005

Chain formation and glial tube assembly in the shift from neonatal to adult subventricular zone of the rodent forebrain.

The subventricular zone (SVZ) is regarded as an embryonic germinal layer persisting at the end of cerebral cortex neurogenesis and capable of generating neuronal precursors throughout life. The two distinct compartments of the adult rodent forebrain SVZ, astrocytic glial tubes and chains of migrating cells, are not distinguishable in the embryonic and early postnatal counterpart. In this study we analyzed the SVZ of mice and rats around birth and throughout different postnatal stages, describing molecular and morphological changes which lead to the typical structural arrangement of adult SVZ. In both species studied, most changes occurred during the first month of life, the transition being slightly delayed in mice, in spite of their earlier development. Important modifications affected the glial cells, eventually leading to glial tube assembly. These changes involved an overall reorganization of glial processes and their mutual relationships, as well as gliogenesis occurring within the SVZ which gives rise to glial cell subpopulations. The neuroblast cell population remained qualitatively quite homogeneous throughout all the stages investigated, changes being restricted to the relationships among cells and consequent formation of chains at about the third postnatal week. Electron microscopy showed that chain formation is not directly linked to glial tube assembly, generally preceding the occurrence of complete glial ensheathment. Moreover, chain and glial tube formation is asymmetric in the medial/lateral aspect of the SVZ, being inversely related. The attainment of an adult SVZ compartmentalization, on the other hand, seems linked to the pattern of expression of adhesion and extracellular matrix molecules.

Animals↗

PubMed2004

Stathmin expression modulates migratory properties of GN-11 neurons in vitro.

Expression of stathmin, a microtubule-associated cytoplasmic protein, prominently localized in neuroproliferative zones and neuronal migration pathways in brain, was investigated in the GnRH neuroendocrine system in vivo and the function was analyzed using an in vitro approach. Here we present novel data demonstrating that GnRH migrating neurons in nasal regions and basal forebrain areas of mouse embryos express stathmin protein. In addition, this expression pattern is dependent on location, as GnRH neurons reaching the hypothalamus are stathmin negative. Immortalized GN-11 cells, which retain many characteristics of migrating GnRH neurons, strongly express stathmin mRNA and protein. The role of stathmin in GnRH migratory properties was evaluated using GN-11 cell line. We up-regulated [stathmin-transfected clones (STMN)+] and down-regulated (STMN-) the expression of stathmin in GN-11 cells, and we investigated changes in cell morphology and motility in vitro. Cells overexpressing stathmin assume a spindle-shaped morphology and their proliferation, as well as their motility, is higher with respect to parental cells. Furthermore, they do not aggregate and express low levels of cadherins compared with control cells. STMN- GN-11 cells are endowed with multipolar processes, and they show a decreased motility and express high levels of cadherin protein. Our findings suggest that stathmin plays a permissive role in GnRH cell motility, possibly via modulation of cadherins expression.

Animals↗

PubMed2004

ErbB4 expression in neural progenitor cells (ST14A) is necessary to mediate neuregulin-1beta1-induced migration.

Activation of the receptor tyrosine kinase ErbB4 leads to various cellular responses such as proliferation, survival, differentiation, and chemotaxis. Two pairs of naturally occurring ErbB4 isoforms differing in their juxtamembrane (JMa/JMb) and C termini (cyt1/cyt2) have been described. To examine the role of ErbB4 in neuron migration, we cloned and stably transfected each of the four ErbB4 isoforms in ST14A cells (a neural progenitor cell line derived from the striatum of embryonic day 14 rats) endogenously expressing the other members of the ErbB family: ErbB1, ErbB2, and ErbB3. Using immunoprecipitation assays, we showed that the neuregulin-1beta1 (NRG1beta1) stimulus induced ErbB4 tyrosine phosphorylation and phosphatidylinositol 3-kinase (PI3K) recruitment and activation (as demonstrated by Akt phosphorylation) either directly (ErbB4 cyt1 isoform) or indirectly (ErbB4 cyt2 isoform). We examined the ability of the four ErbB4 isoforms to induce chemotaxis and cell proliferation in response to NRG1beta1 stimulation. Using migration assays, we observed that only ErbB4-expressing cells stimulated with NRG1beta1 showed a significant increase in migration, whereas the growth rate remained unchanged. Additional assays showed that inhibition of PI3K (but not of phospholipase Cgamma) dramatically reduced migratory activity. Our data show that ErbB4 signaling via PI3K activation plays a fundamental role in controlling NRG1beta1-induced migration.

Animals↗

PubMed2004

Subventricular zone-derived neuronal progenitors migrate into the subcortical forebrain of postnatal mice.

The presence of a germinal layer and the capacity to generate neurons, once thought restricted to the embryonic brain, persists in the forebrain of both postnatal and adult mammals. The two regions in which this phenomenon has been extensively demonstrated are the hippocampal dentate gyrus and the lateral ventricle subventricular zone (SVZ). SVZ-derived cells migrate along the rostral migratory stream into the olfactory bulb, where they differentiate into local interneurons. In this study, using tracer injections into the SVZ at different postnatal ages, we investigated the occurrence of secondary migratory pathways in the mouse subcortical forebrain. During the course of the first week postnatal, in addition to the well-characterized rostral migratory stream, SVZ-derived progenitors migrate in a ventral migratory mass across the nucleus accumbens into the basal forebrain and along a ventrocaudal migratory stream originating at the elbow between the vertical and horizontal limbs of the rostral migratory stream. These cells give rise to granule neurons in the Islands of Calleja and olfactory tubercle pyramidal layer, respectively. In adult, a very small number of cells continue to migrate along the ventrocaudal migratory stream, whereas no migration was observed across the nucleus accumbens. These data demonstrate that in early postnatal and, to a minor extent in adult mice, SVZ-derived cells contribute new neurons to the subcortical forebrain.

Aging↗

PubMed2004

Cholecystokinin modulates migration of gonadotropin-releasing hormone-1 neurons.

Expression of the brain-gut peptide cholecystokinin (CCK) in the developing olfactory-gonadotropin-releasing hormone-1 (GnRH-1) neuroendocrine systems was characterized, and the function of CCK in these systems was analyzed both in vivo and in vitro. We present novel data demonstrating that CCK transcript and protein are expressed in sensory cells in the developing olfactory epithelium and vomeronasal organ, with both ligand and receptors (CCK-1R and CCK-2R) found on olfactory axons throughout prenatal development. In addition, migrating GnRH-1 neurons in nasal regions express CCK-1R but not CCK-2R receptors. The role of CCK in olfactory-GnRH-1 system development was evaluated using nasal explants, after assessing that the in vivo expression of both CCK and CCK receptors was mimicked in this in vitro model. Exogenous application of CCK (10(-7) m) reduced both olfactory axon outgrowth and migration of GnRH-1 cells. This inhibition was mediated by CCK-1R receptors. Moreover, CCK-1R but not CCK-2R antagonism caused a shift in the location of GnRH-1 neurons, increasing the distance that the cells migrated. GnRH-1 neuronal migration in mice carrying a genetic deletion of either CCK-1R or CCK-2R receptor genes was also analyzed. At embryonic day 14.5, the total number of GnRH-1 cells was identical in wild-type and mutant mice; however, the number of GnRH-1 neurons within forebrain was significantly greater in CCK-1R-/- embryos, consistent with an accelerated migratory process. These results indicate that CCK provides an inhibitory influence on GnRH-1 neuronal migration, contributing to the appropriate entrance of these neuroendocrine cells into the brain, and thus represent the first report of a developmental role for CCK.

Animals↗

PubMed2004

Effects of progesterone derivatives, dihydroprogesterone and tetrahydroprogesterone, on the subependymal layer of the adult rat.

Indirect evidence suggests that in the subependymal layer (SEL) steroid hormones could be partially involved in the modulation of neurogenesis, but little or nothing is known about a direct effect of these molecules on this cellular system. The possible effect of progesterone (P) and/or its neuroactive metabolites, dihydroprogesterone (DHP) and tetrahydroprogesterone (THP), on the two cellular components of the SEL (i.e., proliferating/migrating neuroblasts and protoplasmic astrocytes) has been analyzed in adult male rat. P, DHP, and THP were administered by intraventricular injections and after 2 days the SEL was analyzed by immunohistochemistry by using anti-glial fibrillary acidic protein (GFAP) and anti-vimentin antibodies, to label the glial compartment, anti-polysialylated form of the neural cell adhesion molecule (PSA-NCAM), anti-Stathmin, and anti-beta III Tubulin antibodies to label the migrating neuroblasts. Furthermore, the newly formed cells were identified by using intraventricular injections of 5-bromo-2'-deoxyuridine (BrdU) detected immunohistochemically. Our results demonstrate that DHP and THP treatments drastically decrease the number of BrdU-labeled cells within the SEL. THP, DHP, and to a lesser extent P, administrations also induce molecular and structural modifications of the SEL glial compartment. On the whole, the present results indicate that neuroactive derivatives of P (i.e., DHP and THP) exert direct effects on adult neurogenesis, strongly affecting both neuroblasts and astrocytes of the SEL.

20-alpha-Dihydroprogesterone↗

PubMed2004

Bioactive recombinant neuregulin-1, -2, and -3 expressed in Escherichia coli.

The neuregulins (NRGs) are a family of signaling proteins that are ligands for receptor tyrosine kinase of the ErbB family (namely ErbB3 and ErbB4). To date, four different neuregulin genes have been identified (neuregulin1-4). While NRG1 isoforms have been extensively studied, little is yet known about the other genes of the family. We report the expression of recombinant NRG1beta1, NRG2alpha, NRG2beta, and NRG3 as recombinant fusion proteins in Escherichia coli. The cDNA encoding for the EGF-like domain of each protein was cloned from the mouse olfactory bulb and inserted into the pET-19b vector allowing for bacterial expression of the protein fused to an N-terminal His tag. The recombinant NRGs expressed in the inclusion bodies were solubilized under denaturing conditions, purified by affinity chromatography, and refolded via dialysis in the presence of reducing agents. Purified recombinant NRGs were active as they bound to their receptors and induced their phosphorylation. In particular, and in agreement with data on the native proteins, all the molecules were able to bind and activate ErbB4 while only the rNRG1 and the two rNRG2 (but not rNRG3) bound ErbB3.

Amino Acid Sequence↗

PubMed2003

PACAP and NGF regulate common and distinct traits of the sympathoadrenal lineage: effects on electrical properties, gene markers and transcription factors in differentiating PC12 cells.

To determine the possible role of pituitary adenylate cyclase-activating polypeptide (PACAP) in the development of the sympathoadrenal cell lineage, we have examined the effects of this neurotrophic peptide, in comparison to nerve growth factor (NGF), on the morphology, electrophysiological properties, expression of neuronal and neuroendocrine marker genes, and activity of transcription factors during differentiation of sympathoadrenal-derived cells, using the rat pheochromocytoma PC12 cell model. Both PACAP and NGF elicited rapid neurite outgrowth, which was accompanied by induction of cell excitability and the development of both sodium and calcium currents. Concurrently, PACAP and NGF increased the expression of a marker of synaptic vesicles. By contrast, PACAP, but not NGF, regulated the expression of different constituents of neuroendocrine large dense core vesicles in PC12 cells. Furthermore, PACAP and NGF differentially regulated the expression of mammalian achaete-scute homologue and paired homeobox 2b genes, transcription factors instrumental for sympathoadrenal development. To compare downstream effectors activated by PACAP and NGF, we studied the effects of these factors on the binding activity of consensus 12-O-tetradecanoylphorbol-13-acetate- and cAMP-responsive elements to nuclear extracts of differentiating PC12 cells. We found that both PACAP and NGF markedly increase the binding activity of these cis-regulatory sequences and that PACAP preferentially recruits activator protein-1-like transcription factors to these elements. Taken together, these results show that PACAP and NGF exert common as well as different effects on neuronal and neuroendocrine traits in differentiating PC12 cells, strongly suggesting that these two trophic factors could play complementary roles in the development of the sympathoadrenal cell lineage.

Adrenal Glands↗

PubMed2003

Neurogenesis in the subependymal layer of the adult rat: a role for neuroactive derivatives of progesterone.

The subependymal layer (SEL) of the adult mammalian brain provides a continuous supply of newborn cells that migrate to the olfactory bulb (OB) where they differentiate into interneurons. These newly generated cells migrate tangentially to the OB within a dense meshwork of astrocytic cells, organized to form tangentially oriented channels (glial tubes). The central nervous system is able to synthesize a variety of steroids. Among these, we analyzed the effects of progesterone (P) and its neuroactive metabolites dihydroprogesterone (DHP) and tetrahydroprogesterone (THP), administered by intraventricular injection, on the SEL of the adult rat. We found that THP and DHP, but not their precursor P, modify glial tubes organization and decrease immunoreactivity for glial associated proteins in SEL astrocytes. Moreover P metabolites reduce the proliferative activity within the SEL.

20-alpha-Dihydroprogesterone↗

PubMed2002

BMP mRNA and protein expression in the developing mouse olfactory system.

The bone morphogenetic proteins (BMPs) play fundamental roles during the organization of the central nervous system. The presence of these proteins has also been demonstrated in regions of the adult brain that are characterized by neural plasticity. In this study, we examined the expression of BMP4, 6, and 7 mRNAs and proteins in the murine olfactory system. The olfactory system is a useful model for studying cell proliferation and neural differentiation because both of these processes persist throughout life in the olfactory epithelium (OE) and olfactory bulb (OB). Our results demonstrate a differential expression of BMP4, 6, and 7 in the embryonic, postnatal, and adult olfactory system. In particular, BMP4 and BMP7 showed similar immunostaining patterns, being expressed in the olfactory region from the earliest stages studied (embryonic day 15.5) to adulthood. During development BMPs were expressed in the OE, olfactory bulb nerve layer, glomerular layer (GL), mitral cell layer (MCL), and subventricular zone. During the first postnatal week of life, BMP4 and 7 immunoreactivity (-ir) was particularly evident in the GL, MCL, and in the subependymal layer (SEL), which originates postnatally from the subventricular zone. In adults, BMP4 and 7 immunostaining was present in the GL and SEL. Within the SEL, BMP4 and 7 proteins were expressed primarily in association with the astrocytic glial compartment. BMP6-ir was always found in mature olfactory receptor neurons and their axonal projections to the OB. In summary, these data support the hypothesis that BMPs play a role in the morphogenesis of the olfactory system during development and in its plasticity during adulthood.

Animals↗

PubMed2002

Cell migration in the rostral migratory stream.

Adult neurogenesis in the olfactory bulb of rodents is provided by cells which migrate tangentially from their site of genesis into the forebrain subependymal layer (SEL). This migration involves 'chains' of neuroblasts sliding into a meshwork of astrocytic cells and processes (glial tubes). The analysis of this process in postnatal rodents and in adult rabbits reveals different types of relationships occurring both among the migrating cells and between these cells and the glial structures of the SEL.

Animals↗

PubMed1992

Bulbectomy Enhances Neurogenesis and Cell Turnover of Primary Olfactory Neurons But Does Not Abolish Carnosine Expression.

Primary olfactory neurons located in the olfactory neuroepithelium project to the ipsilateral olfactory bulb and undergo a continuous process of neurogenesis and differentiation. We describe, in the adult rat, the kinetics of proliferation, differentiation and survival of primary olfactory neurons either in the presence or absence of their target, the olfactory bulb. The experimental design included unilateral bulbectomy, coupled with a single bromodeoxyuridine pulse 35 days after surgery. The rate of proliferation and survival of olfactory neurons was then examined by immunohistochemistry for bromodeoxyuridine, and the differentiation status by in situ hybridization for calmodulin messenger RNA in immature and mature olfactory neurons and immunohistochemistry for the dipeptide carnosine in mature olfactory neurons. We show that primary olfactory neurons can synthesize carnosine in the absence of the olfactory bulb. However, the number of carnosine-immunopositive neurons in the absence of their target is dramatically reduced to less than one-fourth, whereas the number of olfactory neurons expressing calmodulin messenger RNA is only slightly reduced. The numeric reduction of carnosine-positive neurons in the target-deprived neuroepithelium is correlated with a dramatic reduction in the survival rate of olfactory neurons, since newly generated olfactory neurons are completely lost 35 days after the bromodeoxyuridine pulse. In contrast, in the normal olfactory neuroepithelium almost one-third of newly generated olfactory neurons survive 35 days after the bromodeoxyuridine pulse. On the whole, these data indicate that most of the primary olfactory neurons have a short lifespan but that once they have connected with the olfactory bulb they may persist longer, and suggest that throughout adulthood olfactory neurons are overproduced, differentiate independently from their target, and then undergo a process of target-induced neuronal selection.

Journal Article↗