RESUMEN
Foetal onset hydrocephalus is a disease starting early in embryonic life; in many cases it results from a cell junction pathology of neural stem (NSC) and neural progenitor (NPC) cells forming the ventricular zone (VZ) and sub-ventricular zone (SVZ) of the developing brain. This pathology results in disassembling of VZ and loss of NSC/NPC, a phenomenon known as VZ disruption. At the cerebral aqueduct, VZ disruption triggers hydrocephalus while in the telencephalon, it results in abnormal neurogenesis. This may explain why derivative surgery does not cure hydrocephalus. NSC grafting appears as a therapeutic opportunity. The present investigation was designed to find out whether this is a likely possibility. HTx rats develop hereditary hydrocephalus; 30-40% of newborns are hydrocephalic (hyHTx) while their littermates are not (nHTx). NSC/NPC from the VZ/SVZ of nHTx rats were cultured into neurospheres that were then grafted into a lateral ventricle of 1-, 2- or 7-day-old hyHTx. Once in the cerebrospinal fluid, neurospheres disassembled and the freed NSC homed at the areas of VZ disruption. A population of homed cells generated new multiciliated ependyma at the sites where the ependyma was missing due to the inherited pathology. Another population of NSC homed at the disrupted VZ differentiated into ßIII-tubulin+ spherical cells likely corresponding to neuroblasts that progressed into the parenchyma. The final fate of these cells could not be established due to the protocol used to label the grafted cells. The functional outcomes of NSC grafting in hydrocephalus remain open. The present study establishes an experimental paradigm of NSC/NPC therapy of foetal onset hydrocephalus, at the etiologic level that needs to be further explored with more analytical methodologies.
Asunto(s)
Hidrocefalia/terapia , Células-Madre Neurales/trasplante , Animales , Diferenciación Celular , Proliferación Celular , Neurogénesis , RatasRESUMEN
Fetal onset hydrocephalus and abnormal neurogenesis are two inseparable phenomena turned on by a cell junction pathology first affecting neural stem/progenitor cells (NSPCs) and later the multiciliated ependyma. The neurological impairment of children born with hydrocephalus is not reverted by derivative surgery. NSPCs and neurosphere (NE) grafting into the cerebrospinal fluid (CSF) of hydrocephalic fetuses thus appears as a promising therapeutic procedure. There is little information about the cell lineages actually forming the NE as they grow throughout their days in vitro (DIV). Furthermore, there is no information on how good a host the CSF is for grafted NE. Here, we use the HTx rat, a model with hereditary hydrocephalus, with the mutation expressed in about 30% of the litter (hyHTx), while the littermates develop normally (nHTx). The investigation was designed (i) to establish the nature of the cells forming 4 and 6-DIV NE grown from NSPCs collected from PN1/nHTx rats and (ii) to study the effects on these NEs of CSF collected from nHTx and hyHTx. Immunofluorescence analyses showed that 90% of cells forming 4-DIV NEs were non-committed multipotential NSPCs, while in 6-DIV NE, 40% of the NSPCs were already committed into neuronal, glial and ependymal lineages. Six-DIV NE further cultured for 3 weeks in the presence of fetal bovine serum, CSF from nHTx or CSF from hyHTx, differentiated into neurons, astrocytes and ßIV-tubulin+ multiciliated ependymal cells that were joined together by adherent junctions and displayed synchronized cilia beating. This supports the possibility that ependymal cells are born from subpopulations of NSC with their own time table of differentiation. As a whole, the findings indicate that the CSF is a supportive medium to host NE and that NE grafted into the CSF have the potential to produce neurons, glia and ependyma.
Asunto(s)
Astrocitos/citología , Líquido Cefalorraquídeo/fisiología , Epéndimo/citología , Células Ependimogliales/citología , Hidrocefalia/patología , Células-Madre Neurales/metabolismo , Animales , Diferenciación Celular , Proliferación Celular , Cilios/metabolismo , Modelos Animales de Enfermedad , Humanos , Células Madre Multipotentes/citología , Células Madre Multipotentes/metabolismo , Células-Madre Neurales/citología , Neuronas/citología , RatasRESUMEN
Fetal-onset hydrocephalus is not only a disorder of cerebrospinal fluid (CSF) dynamics, but also a brain disorder. How can we explain the inborn and, so far, irreparable neurological impairment in children born with hydrocephalus? We hypothesize that a cell junction pathology of neural stem cells (NSC) leads to two inseparable phenomena: hydrocephalus and abnormal neurogenesis. All neurons, glial cells, and ependymal cells of the mammalian central nervous system originate from the NSC forming the ventricular zone (VZ) and the neural progenitor cells (NPC) forming the subventricular zone. Several genetic mutations and certain foreign signals all convey into a final common pathway leading to cell junction pathology of NSC and VZ disruption. VZ disruption follows a temporal and spatial pattern; it leads to aqueduct obliteration and hydrocephalus in the cerebral aqueduct, while it results in abnormal neurogenesis in the telencephalon. The disrupted NSC and NPC are released into the CSF and may transform into neurospheres displaying a junctional pathology similar to that of NSC of the disrupted VZ. These cells can then be utilized to investigate molecular alterations underlying the disease and open an avenue into possible NSC therapy.
Asunto(s)
Feto/fisiopatología , Hidrocefalia/patología , Ventrículos Laterales/patología , Células-Madre Neurales/patología , Animales , Humanos , Neuronas/patologíaRESUMEN
The dynamic and molecular composition of the cerebrospinal fluid (CSF) and, consequently, the CSF physiology is much more complex and fascinating than the simplistic view held for decades. Signal molecules either transported from blood to CSF or secreted into the CSF by circumventricular organs and CSF-contacting neurons, use the CSF to reach their targets in the brain, including the pre- and postnatal neurogenic niche. The subcommissural organ (SCO), a highly conserved brain gland present throughout the vertebrate phylum, is one of the sources for signals, as well as the choroid plexus, tanycytes and CSF-contacting neurons. The SCO secretes into the fetal and adult CSF SCO-spondin, transthyretin, and basic fibroblast growth factor. These proteins participate in certain aspects of neurogenesis, such as cell cycle of neural stem cells, neuronal differentiation, and axon pathfinding. Through the CSF, the SCO-secretory proteins may reach virtually any target in the embryonic and adult central nervous system. Since the SCO continues to secrete throughout life span, it seems likely that the neurogenetic property of the SCO compounds would be targeted to the niches where neurogenesis continues in adulthood. This review is aimed to bring into discussion early and new evidence concerning the role(s) of the SCO, and the probable mechanisms by which SCO compounds can readily reach the neurogenic niche of the subventricular zone flowing with the CSF to participate in the regulation of the neurogenic niche. As we unfold the multiples trans-fluid talks between discrete brain domains we will have more tools to influence such talks.
RESUMEN
Disruption/denudation of the ependymal lining has been associated with the pathogenesis of various human CNS disorders, including hydrocephalus, spina bifida aperta, and periventricular heterotopia. It has been traditionally considered that ependymal denudation is a consequence of mechanical forces such as ventricular enlargement. New evidence indicates that ependymal disruption can precede ventricular dilation, but the cellular and molecular mechanisms involved in the onset of ependymal denudation are unknown. Here, we present a novel model to study ependymal cell pathophysiology and demonstrate that selective disruption of N-cadherin-based adherens junctions is sufficient to provoke progressive ependymal denudation. Blocking N-cadherin function using specific peptides that interfere with the histidine-alanine-valine extracellular homophilic interaction domain caused early pathologic changes characterized by disruption of zonula adherens and abnormal intracellular accumulation of N-cadherin. These changes then triggered massive apoptosis of ependymal cells and denudation of brain ventricular walls. Because no typical extrinsic mechanical factors such as elevated pressure or stretching forces are involved in this model, the critical role of N-cadherin-based adherens junctions in ependymal survival/physiology is highlighted. Furthermore, the results suggest that abnormal adherens junctions between ependymal cells should be considered as key components of the pathogenesis of CNS disorders associated with ependymal denudation.
Asunto(s)
Uniones Adherentes/metabolismo , Antígenos CD/metabolismo , Apoptosis/fisiología , Encéfalo/citología , Cadherinas/metabolismo , Epéndimo/metabolismo , Uniones Adherentes/efectos de los fármacos , Análisis de Varianza , Animales , Anticuerpos/farmacología , Antígenos CD/química , Antígenos CD/inmunología , Apoptosis/efectos de los fármacos , Cadherinas/química , Cadherinas/inmunología , Bovinos , Relación Dosis-Respuesta a Droga , Impedancia Eléctrica , Fenómenos Electrofisiológicos/efectos de los fármacos , Epéndimo/citología , Epéndimo/ultraestructura , Proteína Ácida Fibrilar de la Glía/metabolismo , Humanos , Etiquetado Corte-Fin in Situ , Microscopía Electrónica de Transmisión , Técnicas de Cultivo de Órganos , Péptido Hidrolasas/inmunología , Fracciones Subcelulares/metabolismo , Fracciones Subcelulares/ultraestructura , Factores de TiempoRESUMEN
Most cells of the developing mammalian brain derive from the ventricular (VZ) and the subventricular (SVZ) zones. The VZ is formed by the multipotent radial glia/neural stem cells (NSCs) while the SVZ harbors the rapidly proliferative neural precursor cells (NPCs). Evidence from human and animal models indicates that the common history of hydrocephalus and brain maldevelopment starts early in embryonic life with disruption of the VZ and SVZ. We propose that a "cell junction pathology" involving adherent and gap junctions is a final common outcome of a wide range of gene mutations resulting in proteins abnormally expressed by the VZ cells undergoing disruption. Disruption of the VZ during fetal development implies the loss of NSCs whereas VZ disruption during the perinatal period implies the loss of ependyma. The process of disruption occurs in specific regions of the ventricular system and at specific stages of brain development. This explains why only certain brain structures have an abnormal development, which in turn results in a specific neurological impairment of the newborn. Disruption of the VZ of the Sylvian aqueduct (SA) leads to aqueductal stenosis and hydrocephalus, while disruption of the VZ of telencephalon impairs neurogenesis. We are currently investigating whether grafting of NSCs/neurospheres from normal rats into the CSF of hydrocephalic mutants helps to diminish/repair the outcomes of VZ disruption.
Asunto(s)
Hidrocefalia/terapia , Uniones Intercelulares/patología , Células-Madre Neurales/patología , Trasplante de Células Madre/métodos , Animales , Diferenciación Celular , Proliferación Celular , Acueducto del Mesencéfalo/patología , Ventrículos Cerebrales/embriología , Ventrículos Cerebrales/patología , Humanos , Hidrocefalia/patología , Células-Madre Neurales/trasplante , Neurogénesis , RatasRESUMEN
Most cells of the developing mammalian brain derive from the ventricular (VZ) and the subventricular (SVZ) zones. The VZ is formed by the multipotent radial glia/neural stem cells (NSCs) while the SVZ harbors the rapidly proliferative neural precursor cells (NPCs). Evidence from human and animal models indicates that the common history of hydrocephalus and brain maldevelopment starts early in embryonic life with disruption of the VZ and SVZ. We propose that a "cell junction pathology" involving adherent and gap junctions is a final common outcome of a wide range of gene mutations resulting in proteins abnormally expressed by the VZ cells undergoing disruption. Disruption of the VZ during fetal development implies the loss of NSCs whereas VZ disruption during the perinatal period implies the loss of ependyma. The process of disruption occurs in specific regions of the ventricular system and at specific stages of brain development. This explains why only certain brain structures have an abnormal development, which in turn results in a specific neurological impairment of the newborn. Disruption of the VZ of the Sylvian aqueduct (SA) leads to aqueductal stenosis and hydrocephalus, while disruption of the VZ of telencephalon impairs neurogenesis. We are currently investigating whether grafting of NSCs/neurospheres from normal rats into the CSF of hydrocephalic mutants helps to diminish/repair the outcomes of VZ disruption.
Asunto(s)
Animales , Humanos , Ratas , Hidrocefalia/terapia , Uniones Intercelulares/patología , Células-Madre Neurales/patología , Trasplante de Células Madre/métodos , Diferenciación Celular , Proliferación Celular , Acueducto del Mesencéfalo/patología , Ventrículos Cerebrales/embriología , Ventrículos Cerebrales/patología , Hidrocefalia/patología , Neurogénesis , Células-Madre Neurales/trasplanteRESUMEN
A heterogeneous population of ependymal cells lines the brain ventricles. The evidence about the origin and birth dates of these cell populations is scarce. Furthermore, the possibility that mature ependymal cells are born (ependymogenesis) or self-renewed (ependymal proliferation) postnatally is controversial. The present study was designed to investigate both phenomena in wild-type (wt) and hydrocephalic α-SNAP mutant (hyh) mice at different postnatal stages. In wt mice, proliferating cells in the ventricular zone (VZ) were only found in two distinct regions: the dorsal walls of the third ventricle and Sylvian aqueduct (SA). Most proliferating cells were monociliated and nestin+, likely corresponding to radial glial cells. Postnatal cumulative BrdU-labeling showed that most daughter cells remained in the VZ of both regions and they lost nestin-immunoreactivity. Furthermore, some labeled cells became multiciliated and GLUT-1+, indicating they were ependymal cells born postnatally. Postnatal pulse BrdU-labeling and Ki-67 immunostaining further demonstrated the presence of cycling multiciliated ependymal cells. In hydrocephalic mutants, the dorsal walls of the third ventricle and SA expanded enormously and showed neither ependymal disruption nor ventriculostomies. This phenomenon was sustained by an increased ependymogenesis. Consequently, in addition to the physical and geometrical mechanisms traditionally explaining ventricular enlargement in fetal-onset hydrocephalus, we propose that postnatal ependymogenesis could also play a role. Furthermore, as generation of new ependymal cells during postnatal stages was observed in distinct regions of the ventricular walls, such as the roof of the third ventricle, it may be a key mechanism involved in the development of human type 1 interhemispheric cysts.
Asunto(s)
Encéfalo/patología , Epéndimo/crecimiento & desarrollo , Hidrocefalia/patología , Tercer Ventrículo/fisiopatología , Factores de Edad , Animales , Animales Recién Nacidos , Bromodesoxiuridina/metabolismo , Recuento de Células , Proliferación Celular , Modelos Animales de Enfermedad , Epéndimo/ultraestructura , Regulación del Desarrollo de la Expresión Génica/fisiología , Proteína Ácida Fibrilar de la Glía/metabolismo , Transportador de Glucosa de Tipo 1/metabolismo , Ratones , Ratones Mutantes Neurológicos , Microscopía Electrónica de Rastreo , Antígeno Nuclear de Célula en Proliferación/metabolismo , Tercer Ventrículo/citología , Tubulina (Proteína)/metabolismoRESUMEN
The blood-brain barrier (BBB) is a single uninterrupted barrier that in the brain capillaries is located at the endothelial cells and in the circumventricular organs, such as the choroid plexuses (CP) and median eminence (ME), is displaced to specialized ependymal cells. How do hypothalamic hormones reach the portal circulation without making the BBB leaky? The ME milieu is open to the portal vessels, while it is closed to the cerebrospinal fluid (CSF) and to the arcuate nucleus. The cell body and most of the axons of neurons projecting to the ME are localized in areas protected by the BBB, while the axon terminals are localized in the BBB-free area of the ME. This design implies a complex organization of the intercellular space of the median basal hypothalamus. The privacy of the ME milieu implies that those neurons projecting to this area would not be under the influence of compounds leaking from the portal capillaries, unless receptors for such compounds are located at the axon terminal. Amazingly, the arcuate nucleus also has its private milieu that is closed to all adjacent neural structures and open to the infundibular recess. The absence of multiciliated cells in this recess should result in a slow CSF flow at this level. This whole arrangement should facilitate the arrival of CSF signal to the arcuate nucleus. This review will show how peripheral hormones can reach hypothalamic targets without making the BBB leaky.
Asunto(s)
Núcleo Arqueado del Hipotálamo/anatomía & histología , Barrera Hematoencefálica/fisiología , Líquido Cefalorraquídeo/metabolismo , Hipotálamo/anatomía & histología , Eminencia Media/anatomía & histología , Animales , Núcleo Arqueado del Hipotálamo/metabolismo , Barrera Hematoencefálica/ultraestructura , Hipotálamo/metabolismo , Eminencia Media/metabolismo , Neuronas/citología , Neuronas/fisiología , Hormonas Hipofisarias/metabolismoRESUMEN
The pars tuberalis (PT) is the only pituitary region in close contact with the medial-basal hypothalamus and bathed by cerebrospinal fluid (CSF). Although PT has long been recognized as an endocrine gland, certain aspects of its structure remain obscure. The present investigation has been designed to gain information concerning (1) the cellular organization of PT, (2) the PT/median eminence spatial relationship and (3) the exposure of various cell compartments of PT to CSF. Non-endocrine cells (S100-reactive) appear as the organizer of the PT architecture. The apical poles of these cells line large cistern-like cavities and the processes of these cells establish a close spatial relationship with PT-specific secretory cells, portal capillaries and tanycytes. The cisterns are also endowed with clusters of ciliated cells and with a highly electron-dense and PAS-reactive content. The unique spatial organization of endocrine and non-endocrine cells of the PT supports a functional relationship between both cell populations. PT endocrine cells display a hallmark of PT-specific cells, namely, the paranuclear spot, which is a complex structure involving the Golgi apparatus, a large pool of immature secretory granules and a centriole from which originates a single 9+0 cilium projecting to the intercellular channels. Horseradish peroxidase (HRP) injected into the CSF readily reaches the intercellular channels of PT and the inner channel of the single cilium and is incorporated by the endocytic machinery of the secretory cells. The PT endocrine cells, through their single 9+0 cilium, may act as sensors of the CSF. HRP also reaches the lumen of the cisterns, indicating that this PT compartment is also exposed to CSF. PT endocrine cells establish direct cell-to-cell contacts with hypothalamic beta(1) tanycytes, suggesting a second means of brain-PT communication.
Asunto(s)
Líquido Cefalorraquídeo , Epéndimo/citología , Eminencia Media/citología , Adenohipófisis/citología , Animales , Capilares , Centriolos/ultraestructura , Cilios/ultraestructura , Células Endocrinas/metabolismo , Células Endocrinas/ultraestructura , Endocitosis , Espacio Extracelular , Aparato de Golgi/metabolismo , Aparato de Golgi/ultraestructura , Hipotálamo Medio/citología , Adenohipófisis/metabolismo , Ratas , Proteínas S100/metabolismo , Vesículas Secretoras/ultraestructura , Tercer Ventrículo/citologíaRESUMEN
Increasing evidence suggests that the hypophyseal pars tuberalis (PT) plays a key role in the transduction of light/dark (melatonin) information to the endocrine system. It has been shown that PT-specific cells express melatonin receptors and thyrotropin hormone (TSH) subunits. However, these cells do not resemble thyrotrophs or any other of the pars distalis (PD) cells. There is evidence that PT-specific cells secrete a glycoprotein hormone designated as 'tuberalin'. We have identified a putative tuberalin of 21 kDa (tuberalin II) and have raised antibodies against it. To further investigate whether tuberalin II is a distinct secretory compound of the PT, absorption studies of antituberalin II with TSH or with an extract of the rat PD containing beta-TSH, beta-luteinizing hormone (LH) and the common alpha-subunit of glycoprotein hormones (GSU), were performed. Neither of the absorption tests abolished the immunoreactivity of the PT to antituberalin II, suggesting that tuberalin II is different from TSH or the other PD glycoprotein hormones. Double immunofluorescence analyses using antibodies against tuberalin II, beta-TSH and GSU revealed that in the developing and adult PT there are 3 populations of PT-specific cells expressing tuberalin II and GSU (type 1), beta-TSH and GSU (type 2) and tuberalin II, beta-TSH and GSU (type 3). This further indicates that tuberalin II and beta-TSH correspond to different compounds and that they may be expressed either by different cells types or coexpressed in a 3rd cell type. The distribution and temporal expression of tuberalin II, beta-TSH, beta-LH and GSU were investigated in the developing pituitary gland. At E14.5, tuberalin II and GSU were expressed by cells of the PT primordium but not by the PD and pars intermedia primordia. The onset of expression of beta-TSH, beta-LH and GSU in cells of the PD occurred about 1 day later, further indicating the distinct nature of tuberalin II and supporting the earlier view that the secretion of polypeptides from the fetal rat pituitary gland begins in PT-specific cells.
Asunto(s)
Hormonas Glicoproteicas de Subunidad alfa/metabolismo , Glicoproteínas/metabolismo , Hipófisis/crecimiento & desarrollo , Hipófisis/metabolismo , Tirotropina de Subunidad beta/metabolismo , Animales , Recuento de Células , Femenino , Técnica del Anticuerpo Fluorescente , Inmunohistoquímica , Hormona Luteinizante de Subunidad beta/metabolismo , Masculino , Hipófisis/embriología , Ratas , Ratas Sprague-Dawley , Factores de TiempoRESUMEN
Hydrocephalus with hop gait (hyh) is a recessive inheritable disease that arose spontaneously in a mouse strain. A missense mutation in the Napa gene that results in the substitution of a methionine for isoleucine at position 105 (M105I) of alphaSNAP has been detected in these animals. alphaSNAP is a ubiquitous protein that plays a key role in membrane fusion and exocytosis. In this study, we found that male hyh mice with a mild phenotype produced morphologically normal and motile sperm, but had a strongly reduced fertility. When stimulated with progesterone or A23187 (a calcium ionophore), sperm from these animals had a defective acrosome reaction. It has been reported that the M105I mutation affects the expression but not the function of the protein. Consistent with an hypomorphic phenotype, the testes and epididymides of hyh mice had low amounts of the mutated protein. In contrast, sperm had alphaSNAP levels indistinguishable from those found in wild type cells, suggesting that the mutated protein is not fully functional for acrosomal exocytosis. Corroborating this possibility, addition of recombinant wild type alphaSNAP rescued exocytosis in streptolysin O-permeabilized sperm, while the mutant protein was ineffective. Moreover, addition of recombinant alphaSNAP. M105I inhibited acrosomal exocytosis in permeabilized human and wild type mouse sperm. We conclude that the M105I mutation affects the expression and also the function of alphaSNAP, and that a fully functional alphaSNAP is necessary for acrosomal exocytosis, a key event in fertilization.
Asunto(s)
Reacción Acrosómica/fisiología , Ratones Mutantes , Mutación Puntual , Proteínas Solubles de Unión al Factor Sensible a la N-Etilmaleimida/genética , Animales , Epidídimo/metabolismo , Exocitosis/fisiología , Femenino , Fertilidad/fisiología , Fertilización In Vitro , Humanos , Masculino , Ratones , Ratones Endogámicos C57BL , Proteínas Solubles de Unión al Factor Sensible a la N-Etilmaleimida/metabolismo , Espermatozoides/citología , Espermatozoides/metabolismo , Testículo/citología , Testículo/metabolismoRESUMEN
BACKGROUND: The subcommissural organ (SCO) is a highly conserved brain gland present throughout the vertebrate phylum; it secretes glycoproteins into the cerebrospinal fluid (CSF), where they aggregate to form Reissner's fiber (RF). SCO-spondin is the major constituent protein of RF. Evidence exists that the SCO also secretes proteins that remain soluble in the CSF. The aims of the present investigation were: (i) to identify and partially characterize the SCO-secretory compounds present in the SCO gland itself and in the RF of the Sprague-Dawley rat and non-hydrocephalic hyh mouse, and in the CSF of rat; (ii) to make a comparative analysis of the proteins present in these three compartments; (iii) to identify the proteins secreted by the SCO into the CSF at different developmental periods. METHODS: The proteins of the SCO secreted into the CSF were studied (i) by injecting specific antibodies into ventricular CSF in vivo; (ii) by immunoblots of SCO, RF and CSF samples, using specific antibodies against the SCO secretory proteins (AFRU and anti-P15). In addition, the glycosylated nature of SCO-compounds was analysed by concanavalin A and wheat germ agglutinin binding. To analyse RF-glycoproteins, RF was extracted from the central canal of juvenile rats and mice; to investigate the CSF-soluble proteins secreted by the SCO, CSF samples were collected from the cisterna magna of rats at different stages of development (from E18 to PN30). RESULTS: Five glycoproteins were identified in the rat SCO with apparent molecular weights of 630, 450, 390, 320 and 200 kDa. With the exception of the 200-kDa compound, all other compounds present in the rat SCO were also present in the mouse SCO. The 630 and 390 kDa compounds of the rat SCO have affinity for concanavalin A but not for wheat germ agglutinin, suggesting that they correspond to precursor forms. Four of the AFRU-immunoreactive compounds present in the SCO (630, 450, 390, 320 kDa) were absent from the RF and CSF. These may be precursor and/or partially processed forms. Two other compounds (200, 63 kDa) were present in SCO, RF and CSF and may be processed forms. The presence of these proteins in both, RF and CSF suggests a steady-state RF/CSF equilibrium for these compounds. Eight AFRU-immunoreactive bands were consistently found in CSF samples from rats at E18, E20 and PN1. Only four of these compounds were detected in the cisternal CSF of PN30 rats. The 200 kDa compound appears to be a key compound in rats since it was consistently found in all samples of SCO, RF and embryonic and juvenile CSF. CONCLUSION: It is concluded that (i) during the late embryonic life, the rat SCO secretes compounds that remain soluble in the CSF and reach the subarachnoid space; (ii) during postnatal life, there is a reduction in the number and concentration of CSF-soluble proteins secreted by the SCO. The molecular structure and functional significance of these proteins remain to be elucidated. The possibility they are involved in brain development has been discussed.
RESUMEN
The present investigation was designed to investigate the fate of the large pool of neurohypophyseal hormones that is never released into the blood. Normal Sprague-Dawley and taiep mutant rats were investigated under normal water balance, after dehydration and after dehydration-rehydration. Lectin histochemistry and light- and electron-microscopic immunocytochemistry using antibodies against vasopressin, oxytocin, and neurophysins used at low (1:1,000) and high (1:15,000) dilutions allowed to distinguish (1) recently packed immature granules, as those located in the perikaryon; (2) mature; and (3) aged granules. The distribution of these granules within the different domains of the neurosecretory axons located in the neural lobe, namely, undilated segments, swellings, terminals, and Herring bodies, and the response of these compartments to dehydration and dehydration-rehydration allowed to roughly follow the routing of the granules through such axonal domains. It is suggested that granules may move backward and forward between the terminals and the swellings. At variance, aged granules located in Herring body are retained in this compartment and would finally become degraded. Herring bodies displayed distinct lectin binding and immunocytochemical properties, allowing to distinguish them from axonal swellings. After a dehydration-rehydration cycle, immunocytochemistry and electron microscopy revealed that Herring bodies were no longer present in the neural lobe and that several terminals had degenerated. It is concluded that (1) the neurophysin axons may undergo remodeling under appropriate stimuli and (2) Herring bodies are a specialized and plastic domain of the magnocellular neurosecretory neuron involved in the disposal of aged neurosecretory granules. No differences were detected at the neural lobe level between normal and mutant rats subjected to the same experimental conditions.
Asunto(s)
Envejecimiento/metabolismo , Axones/fisiología , Neurofisinas/metabolismo , Neurohipófisis/ultraestructura , Animales , Axones/química , Axones/ultraestructura , Neurosecreción , Ratas , Ratas Sprague-DawleyRESUMEN
Tanycytes are bipolar cells bridging the cerebrospinal fluid (CSF) to the portal capillaries and may link the CSF to neuroendocrine events. During the perinatal period a subpopulation of radial glial cells differentiates into tanycytes, a cell lineage sharing some properties with astrocytes and the radial glia, but displaying unique and distinct morphological, molecular, and functional characteristics. Four populations of tanycytes, alpha(1,2) and beta(1,2), can be distinguished. These subtypes express differentially important functional molecules, such as glucose and glutamate transporters; a series of receptors for neuropeptide and peripheral hormones; secretory molecules such as transforming growth factors, prostaglandin E(2), and the specific protein P85; and proteins of the endocytic pathways. This results in functional differences between the four subtypes of tanycytes. Thus, alpha(1,2) tanycytes do not have barrier properties, whereas beta(1,2) tanycytes do. Different types of tanycytes use different mechanisms to internalize and transport cargo molecules; compounds internalized via a clathrin-dependent endocytosis would only enter tanycytes from the CSF. There are also differences in the neuron-tanycyte relationships; beta(1,2) tanycytes are innervated by peptidergic and aminergic neurons, but alpha(1,2) tanycytes are not. Important aspects of the neuron-beta(1) tanycyte relationships have been elucidated. Tanycytes can participate in the release of gonadotropin-releasing hormone (GnRH) to the portal blood by expressing estrogen receptors, absorbing molecules from the CSF, and providing signal(s) to the GnRH neurons. Removal of tanycytes prevents the pulse of GnRH release into the portal blood, the peak of luteinizing hormone, and ovulation. The discovery in tanycytes of new functional molecules is opening a new field of research. Thus, thyroxine deiodinase type II, an enzyme generating triiodothyronine (T(3)) from thyroxine, appears to be exclusively expressed by tanycytes, suggesting that these cells are the main source of brain T(3). Glucose transporter-2 (GLUT-2), a low-affinity transporter of glucose and fructose, and ATP-sensitive K(+) channels are expressed by tanycytes, suggesting that they may sense CSF glucose concentrations.
Asunto(s)
Hipotálamo Medio/citología , Hipotálamo Medio/fisiología , Sistemas Neurosecretores/fisiología , Animales , Astrocitos/citología , Astrocitos/metabolismo , Astrocitos/fisiología , Barrera Hematoencefálica/citología , Barrera Hematoencefálica/fisiología , Encéfalo/citología , Encéfalo/fisiología , Líquido Cefalorraquídeo/fisiología , Glándulas Endocrinas/citología , Glándulas Endocrinas/fisiología , Endocitosis/fisiología , Epéndimo/química , Epéndimo/citología , Femenino , Hormona Liberadora de Gonadotropina/sangre , Hormona Liberadora de Gonadotropina/líquido cefalorraquídeo , Hipotálamo Medio/metabolismo , Masculino , Neuroglía/citología , Neuroglía/metabolismo , Neuroglía/fisiología , Neuronas/fisiología , Sistemas Neurosecretores/citología , Ratas , Células Madre/citología , Células Madre/fisiologíaRESUMEN
The subcommissural organ (SCO) is a brain gland located in the roof of the third ventricle that releases glycoproteins into the cerebrospinal fluid, where they form a structure known as Reissner's fiber (RF). On the basis of SCO-spondin sequence (the major RF glycoprotein) and experimental findings, the SCO has been implicated in central nervous system development; however, its function(s) after birth remain unclear. There is evidence suggesting that SCO activity in adult animals may be regulated by serotonin (5HT). The use of an anti-5HT serum showed that the bovine SCO is heterogeneously innervated with most part being poorly innervated, whereas the rat SCO is richly innervated throughout. Antibodies against serotonin receptor subtype 2A rendered a strong immunoreaction at the ventricular cell pole of the bovine SCO cells and revealed the expected polypeptides in blots of fresh and organ-cultured bovine SCO. Analyses of organ-cultured bovine SCO treated with 5HT revealed a twofold decrease of both SCO-spondin mRNA level and immunoreactive RF glycoproteins, whereas no effect on release of RF glycoproteins into the culture medium was detected. Rats subjected to pharmacological depletion of 5HT exhibited an SCO-spondin mRNA level twofold higher than untreated rats. These results indicate that 5HT down-regulates SCO-spondin biosynthesis but apparently not its release, and suggest that 5HT may exert the effect on the SCO via the cerebrospinal fluid.
Asunto(s)
Moléculas de Adhesión Celular Neuronal/genética , Moléculas de Adhesión Celular Neuronal/metabolismo , Regulación hacia Abajo , Regulación de la Expresión Génica , Serotonina/metabolismo , Órgano Subcomisural/metabolismo , Transcripción Genética , Secuencia de Aminoácidos , Animales , Secuencia de Bases , Bovinos , Líquido Cefalorraquídeo/química , Líquido Cefalorraquídeo/metabolismo , Masculino , Datos de Secuencia Molecular , Técnicas de Cultivo de Órganos , Ratas , Ratas Sprague-Dawley , Receptor de Serotonina 5-HT2A/metabolismo , Alineación de Secuencia , Órgano Subcomisural/citologíaRESUMEN
SCO-spondin is a large-molecular mass glycoprotein, secreted by the subcommissural organ (SCO), which has been implicated in neuronal development during ontogeny of the central nervous system. The expression of SCO-spondin is not restricted to the SCO but it also occurs in the floor plate, a key structure participating in neuronal differentiation and patterning of the neural tube. It has been postulated that SCO-spondin detected in the floor plate is released into the lumen of the neural tube, but this new route of secretion of floor plate cells needs to be further substantiated. For this purpose, we standardized long-term organ culture of bovine floor plate and performed morphological, immunological, biochemical, and gene expression analyses. The study of floor plate explants and their conditioned media allowed us to demonstrate that: (1) organ-cultured floor plate cells are actively secretory for up to 25 days; (2) SCO-spondin gene is actively transcribed and translated by the cultured floor plate cells; (3) SCO-spondin is released into the culture medium via the apical cell pole; and (4) upon release, SCO-spondin does not aggregate in the conditioned medium but remains soluble. Furthermore, in the cultured floor plate cells, SCO-spondin may be secreted through a route bypassing the Golgi apparatus.