Cytokines and neuronal differentiation

A diverse spectrum of complementary experimental investigations has demonstrated that two major cytokine superfamilies, the transforming growth factor-beta (TGF beta) and the hemopoietins, mediate an extensive range of developmental events in the nervous system that often rivals and frequently exceeds that of the classic neurotrophins. The exponential growth in the identification and physiological analysis of TGF beta subclasses of cytokines that transduce intracellular signals through intrinsic membrane serine/threonine kinase-associated receptor subunits has led to an increased understanding of their complex cellular actions in programming the temporospatial expression and maturation of anatomically distinct neuronal subpopulations. An analysis of the developmental parallels that exist between neural development and hematolymphopoiesis has fostered an expansion in the identification and classification of cellular actions of hematolymphopoietic cytokines that are also active during neural development. During the process of neural maturation, it has also become apparent that an extensive range of cell surface-associated and intracellular signalling molecules that are essential for hematopoietic and immunological development may also represent an important set of effector molecules that are active during neuronal differentiation. These recent advances in cell and molecular biology have allowed us to begin to construct an integrated model of the developmental signalling pathways and diverse cellular processes that are necessary for graded stages of neuronal maturation. These cumulative observations suggest that a dynamic hierarchy of epigenetic and genetic signals is essential for the growth, survival, and maturation of regional neuronal subpopulations that are derived from multipotent progenitor species within the central and peripheral nervous systems.     

A posteriorising factor, retinoic acid, reveals that anteroposterior patterning controls the timing of neuronal differentiation in Xenopus neuroectoderm

During early development of the Xenopus central nervous system (CNS), neuronal differentiation can be detected posteriorly at neural plate stages but is delayed anteriorly until after neural tube closure. A similar delay in neuronal differentiation also occurs in the anterior neural tissue that forms in vitro when isolated ectoderm is treated with the neural inducer noggin. Here we examine the factors that control the timing of neuronal differentiation both in embryos and in neural tissue induced by noggin (noggin caps). We show that the delay in neuronal differentiation that occurs in noggin caps cannot be overcome by inhibiting the activity of the neurogenic gene, X-Delta-1, which normally inhibits neuronal differentiation, suggesting that it represents a novel level of regulation. Conversely, we show that the timing of neuronal differentiation can be changed from late to early after treating noggin caps or embryos with retinoic acid (RA), a putative posteriorising agent. Concommittal with changes in the timing of neuronal differentiation, RA suppresses the expression of anterior neural genes and promotes the expression of posterior neural genes. The level of early neuronal differentiation induced by RA alone is greatly increased by the additional expression of the proneural gene, XASH3. These results indicate that early neuronal differentiation in neuralised ectoderm requires posteriorising signals, as well as signals that promote the activity of proneural genes such as XASH3. In addition, these result suggest that neuronal differentiation is controlled by anteroposterior (A-P) patterning, which exerts a temporal control on the onset of neuronal differentiation.     

Effect of different albuterol dosing regimens in the treatment of acute exacerbation of chronic obstructive pulmonary disease

There is substantial evidence for an activation of the sympathetic nervous system in man as well as in genetic models of hypertension, such as the spontaneously hypertensive rat (SHR), but we are only beginning to understand the central mechanisms that generate changes in sympathetic activity and elevate blood pressure (BP). Significant recent advances have been made in defining the neural pathways involved in BP regulation and in identifying the neurotransmitters these neurones utilise. In this overview, we describe the neural pathways within the medulla oblongata and spinal cord that participate in BP control and examine the role of amino acid neurotransmitters within these pathways. We demonstrate how alterations in these pathways explain the sympathetic activation observed in the SHR and contribute to hypertension in this model. Lastly, we examine the application of modern molecular biological approaches to further our understanding of the neural regulation of the circulation. In these studies, we used the administration of antisense oligonucleotides to interrupt gene expression.     

Sprouting adult CNS cholinergic axons express NILE and associate with astrocytic surfaces expressing neural cell adhesion molecule

To assess the cellular and molecular substrates for cholinergic axon growth in the adult central nervous system (CNS), we implanted grafts of control and nerve growth factor (NGF)-producing genetically modified fibroblasts within the striatum of rats. Sprouting cholinergic axonal processes that grew into grafts of NGF-producing fibroblasts were fasciculated and followed the surface of astrocytic processes for long distances within the grafts. The close and long distance anatomical relationship between the sprouted axons and the astrocytes supported previous ultrastructural evidence that astrocytes may serve as a cellular substrate for sprouting cholinergic axons in vivo. The sprouted axon processes were associated with the expression of nerve growth factor-inducible large external (NILE) glycoprotein on their surfaces. NILE expression was not seen in control grafts where there was an absence of cholinergic ingrowth. NILE has been demonstrated to play a role in axon fasciculation in a number of other neural systems. The astrocytic processes in both control and NGF-producing fibroblast grafts expressed neural cell adhesion molecule (NCAM), suggesting that NCAM-mediated adhesion may be responsible for the close relationship between the axons and astrocytes within the grafts. NGF-induced heterotypic interactions between neuronal NILE and astroglial NCAM may also be required for adult cholinergic axonal sprouting.     

Lesion-induced plasticity of central neurons: sprouting of single fibres in the rat hippocampus after unilateral entorhinal cortex lesion

In response to a central nervous system trauma surviving neurons reorganize their connections and form new synapses that replace those lost by the lesion. A well established in vivo system for the analysis of this lesion-induced plasticity is the reorganization of the fascia dentata following unilateral entorhinal cortex lesions in rats. After general considerations of neuronal reorganization following a central nervous system trauma, this review focuses on the sprouting of single fibres in the rat hippocampus after entorhinal lesion and the molecular factors which may regulate this process. First, the connectivity of the fascia dentata in control animals is reviewed and previously unknown commissural fibers to the outer molecular layer and entorhinal fibres to the inner molecular layer are characterized. Second, sprouting of commissural and crossed entorhinal fibres after entorhinal cortex lesion is described. Single fibres sprout by forming additional collaterals, axonal extensions, boutons, and tangle-like axon formations. It is pointed out that the sprouting after entorhinal lesion mainly involves unlesioned fibre systems terminating within the layer of fibre degeneration and is therefore layer-specific. Third, molecular changes associated with axonal growth and synapse formation are considered. In this context, the role of adhesion molecules, glial cells, and neurotrophic factors for the sprouting process are discussed. Finally, an involvement of sprouting processes in the formation of neuritic plaques in Alzheimer's disease is reviewed and discussed with regard to the axonal tangle-like formations observed after entorhinal cortex lesion.     

Gonadotropin-releasing hormone neuronal system of the freshwater snails Helisoma trivolvis and Lymnaea stagnalis: possible involvement in reproduction

Peptides of the gonadotropin-releasing hormone (GnRH) family are present in neural and nonneural tissues throughout the chordate phylum. Although GnRH peptides have been implicated in nonreproductive functions, their primary function is to control reproduction by regulating sexual behaviors and inducing gonadotropin hormone release from the pituitary. Evidence suggesting the presence of a similar peptide in the central nervous system (CNS) of the gastropod mollusc Helisoma trivolvis has recently been provided. In the present study, we examined the tissue distribution of the peptide and found that it is likely restricted to the nervous system. The neuronal system containing the endogenous GnRH-like peptide is described further and is shown, in part, to innervate the male reproductive tract. Immunostaining in the closely related snail, Lymnaea stagnalis, showed a conservation in the locations of some immunoreactive neurons. Notably, staining occurred in and adjacent to the lateral lobes of both snails. Because these lobes contain neurons involved in the stimulation of egg laying and GnRH staining occurred in additional areas in the Helisoma CNS that are involved in reproduction, we suggest that the endogenous GnRH-like peptide plays a role in regulating reproduction in freshwater snails.     

bHLH transcription factors and mammalian neuronal differentiation

The basic helix-loop-helix (bHLH) factor Mash1 is expressed in the developing nervous system. Null mutation of Mash1 results in loss of olfactory and autonomic neurons and delays differentiation of retinal neurons, indicating that Mash1 promotes neuronal differentiation. Other bHLH genes, Math/NeuroD/Neurogenin, all expressed in the developing nervous system, have also been suggested to promote neuronal differentiation. In contrast, another bHLH factor, HES1, which is expressed by neural precursor cells but not by neurons, represses Mash1 expression and antagonizes Mash1 activity in a dominant negative manner. Forced expression of HES1 in precursor cells blocks neuronal differentiation in the brain and retina, indicating that HES1 is a negative regulator of neuronal differentiation. Conversely, null mutation of HES1 up-regulates Mash1 expression, accelerates neuronal differentiation, and causes severe defects of the brain and eyes. Thus, HES1 regulates brain and eye morphogenesis by inhibiting premature neuronal differentiation, and the down-regulation of HES1 expression at the right time is required for normal development of the nervous system. Interestingly, HES1 can repress its own expression by binding to its promoter, suggesting that negative autoregulation may contribute to down-regulation of HES1 expression during neural development. Recent studies indicate that HES1 expression is also controlled by RBP-J, a mammalian homologue of Suppressor of Hairless [Su(H)], and Notch, a key membrane protein that may regulate lateral specification through RBP-J during neural development. Thus, the Notch-->RBP-J-->HES1-Mash1 pathway may play a critical role in neuronal differentiation.     

Expression of the human antigen SPAG2 in the testis and localization to the outer dense fibers in spermatozoa

There is increasing evidence to suggest that opioid peptides may have widespread effects as regulators of growth. To evaluate the hypothesis that endogenous opioids control cellular proliferation during neural development, we have used in situ hybridization to examine opioid peptide and receptor mRNA expression in neuroepithelial zones of fetal rat brain and spinal cord. Our data show that proenkephalin mRNA is widely expressed in forebrain germinal zones and choroid plexus during the second half of gestation. In contrast, prodynorphin mRNA expression is restricted to the periventricular region of the ventral spinal cord. Little mu or delta receptor mRNA expression was detected in any regions of neuronal proliferation prior to birth. However, kappa receptor mRNA is widely expressed in hindbrain germinal zones during the 3rd week of gestation. Our present findings support the hypothesis that endogenous opioids may regulate proliferation of both neuronal and non-neuronal cells during central nervous system development. Given the segregated expression of proenkephalin mRNA in forebrain neuroepithelium and kappa receptor mRNA within hindbrain, different opioid mechanisms may regulate cell division in rostral and caudal brain regions.     

P19 cells differentiate into glutamatergic and glutamate-responsive neurons in vitro

The neurotransmitter L-glutamate has been associated with a number of developmental events within the central nervous system including synaptogenesis and the refinement of topographically ordered neural maps. As a model for studying such events at the molecular level, we have examined the expression of glutamate and glutamate receptors in neurons that develop from P19 cells in response to retinoids. We report here that many P19-derived neurons do contain glutamate in secretory vesicles and that this glutamate appears to function as a neurotransmitter. The neurotransmitter GABA is also present in these cultures and both glutamate and GABA appeared to co-localize in some neuronal processes. Both neurotransmitters were released from the neurons in response to membrane depolarization. These neurons also express various glutamate receptor subunits including GluR1, GluR4 and NMDAR1 as detected by immunological methods. Using whole-cell patch-clamping, we have recorded spontaneous postsynaptic potentials which increase in both amplitude and frequency with time in culture and which are sensitive to the glutamate antagonist kynurenic acid Thus, P19-derived neurons mature in culture and form electrically active neural networks involving glutamate and glutamate receptors.     

Mash1 regulates neurogenesis in the ventral telencephalon

Previous studies have shown that mice mutant for the gene Mash1 display severe neuronal losses in the olfactory epithelium and ganglia of the autonomic nervous system, demonstrating a role for Mash1 in development of neuronal lineages in the peripheral nervous system. Here, we have begun to analyse Mash1 function in the central nervous system, focusing our studies on the ventral telencephalon where it is expressed at high levels during neurogenesis. Mash1 mutant mice present a severe loss of progenitors, particularly of neuronal precursors in the subventricular zone of the medial ganglionic eminence. Discrete neuronal populations of the basal ganglia and cerebral cortex are subsequently missing. An analysis of candidate effectors of Mash1 function revealed that the Notch ligands Dll1 and Dll3, and the target of Notch signaling Hes5, fail to be expressed in Mash1 mutant ventral telencephalon. In the lateral ganglionic eminence, loss of Notch signaling activity correlates with premature expression of a number of subventricular zone markers by ventricular zone cells. Therefore, Mash1 is an important regulator of neurogenesis in the ventral telencephalon, where it is required both to specify neuronal precursors and to control the timing of their production.     

Neural stem and progenitor cells: a strategy for gene therapy and brain repair

The damaged adult mammalian brain is incapable of significant structural self-repair. Although varying degrees of recovery from injury are possible, this is largely because of synaptic and functional plasticity rather than the frank regeneration of neural tissues. The lack of structural plasticity of the adult brain is partly because of its inability to generate new neurons, a limitation that has severely hindered the development of therapies for neurological injury or degeneration. However, a variety of experimental studies, as well as moderately successful clinical engraftment of fetal tissue into the adult parkinsonian brain, suggests that cell replacement is evolving as a valuable treatment modality. Neural stem cells, which are the self-renewing precursors of neurons and glia, have been isolated from both the embryonic and adult mammalian central nervous system. In the adult human brain, both neuronal and oligodendroglial precursors have been identified, and methods for their harvest and enrichment have been established. Neural precursors have several characteristics that make them ideal vectors for brain repair. They may be clonally expanded in tissue culture, providing a renewable supply of material for transplantation. Moreover, progenitors are ideal for genetic manipulation and may be engineered to express exogenous genes for neurotransmitters, neurotrophic factors, and metabolic enzymes. Thus, the persistence of neuronal precursors in the adult mammalian brain may permit us to design novel and effective strategies for central nervous system repair, by which we may yet challenge the irreparability of the structurally damaged adult nervous system.     

The correction of immunoendocrine disorders in patients with hyperplastic processes in the endometrium

The nervous systems of invertebrates and vertebrates consist of neuronal networks of varying complexity, and the elucidation of the organization of these networks is essential if we are to understand neural function. Up until the mid-19th Century gross dissection was the primary tool available to scientists to study the nervous system. The development of neurohistological techniques, electrical stimulation, and observation of neural function in humans and animals following injury added rapidly to our understanding of the nervous system during the following century. Over the last 3 decades investigators seeking to unravel the complexities of neural circuits have made use of analytical methods based upon the biological properties of neurons, including orthograde and retrograde axonal transport of tracer substances, the expression of particular genes and gene products that can be assessed with immunocytochemical or in situ methods, and the imaging of the utilization of oxygen or glucose by active populations of neurons. Advances in neuroscience have led to an enormous expansion in our knowledge of normal neural functioning and how that function is altered by injury or disease. Modern studies of neuronal organization have been at the center of our increased understanding of how the brain works.     

Lyme disease (Lyme borreliosis)

Unlike the peripheral nervous system (PNS), the mammalian central nervous system (CNS) clearly lacks the robust regenerative characteristics and capacity of the former. Despite this fact, two unique regions of the adult mammalian CNS possess such regenerative potential and are capable of active regeneration following injury or structural compromise. These unique areas are the olfactory system and the neurohypophyseal system of the endocrine hypothalamus. Furthermore, it has been clearly demonstrated that primordial neuroblasts regarded as stem cells emerge from the subependymal parenchyma of the walls and floor of the third cerebral ventricle, migrate to the ventricular surface and undergo compensatory synaptogenesis within one week following hypophysectomy. In situ hybridization studies have unequivocally demonstrated that the up-regulation of nitric oxide synthase (NOS) is essential for neural (axonal) regeneration and neuronal (stem cell) migration to occur. Moreover, neuronal migration is reliably inhibited following the administration of the NO antagonist, nitroarginine. The current investigation serves to confirm a remarkable degree of plasticity and regeneration in the adult mammalian neurohypophyseal system coupled with the emergence of primordial neuroblasts that undergo apparent differentiation, migration and compensatory synaptogenesis in response to the up-regulation of NO that occurs following the trauma of hypophysectomy. Evidence from the current investigation appears to confirm that specialized glia of the neurohypophyseal system, the so-called pituicyte, proliferate following hypophysectomy and may serve as a growth matrix or structural template that may target and direct regenerating Supraoptic (SON) and Paraventricular (PVN) axons toward endothelial primordia in the regenerating neural stem and lobe.     

Cloning of human ENC-1 and evaluation of its expression and regulation in nervous system tumors

We recently identified and characterized a novel murine gene, ENC-1, that is expressed primarily in the nervous system and encodes an actin-binding protein. To gain insight into a potential role for ENC-1 gene in the processes of cell differentiation and malignant transformation in the human nervous system, we first cloned and characterized the human homologue of ENC-1. The human ENC-1 gene appeared to be highly expressed in adult brain and spinal cord, and in a number of cell lines derived from nervous system tumors we detected low steady-state levels of ENC-1 mRNA. We used a neuroblastoma differentiation model, the retinoic acid-induced neuronal differentiation of SMS-KCNR cells, to study the regulation of the ENC-1 gene during neural crest cell differentiation. We found that the expression of ENC-1 increased dramatically in the differentiated SMS-KCNR cells as compared to control undifferentiated cells. These results suggest that ENC-1 expression plays a role during differentiation of neural crest cells and may be down regulated in neuroblastoma tumors.     

A genetic analysis of the stoned locus and its interaction with dunce, shibire and Suppressor of stoned variants of Drosophila melanogaster

The genetic complementation patterns of both behavioral and lethal alleles at the stoned locus have been characterized. Mosaic analysis of a stoned lethal allele suggests that stoned functions either in the nervous system or in both the nervous system and musculature, but is not required for gross neural development. The behavioral alleles stnts and stnC, appear to be defective in a diametrically opposite sense, show interallelic complementation, and indicate distinct roles for the stoned gene product in the visual system and in motor coordination. A number of other neurological mutations have been investigated for their possible interaction with the viable stoned alleles. Mutations at two loci, dunce and shibire, act synergistically with the stnts mutations to cause lethality, but fail to interact with stnC. A third variant (Suppressor of stoned) has been identified which can suppress the debilitation associated with the stnts mutations. These data, together with a previously identified interaction between the stnts and tan mutants, indicate a central role for the stoned gene product in neuronal function, and suggests that the stoned gene product interacts, either directly or indirectly, with the neural cAMP second messenger system, with the synaptic membrane recycling pathway via dynamin, and with biogenic amine metabolism.     

Neuronal circuitry of the lower urinary tract; central and peripheral neuronal control of the micturition cycle

A new presentation technique is introduce to describe the neuronal circuitry involved in the control of the uropo?tic system and its control mechanisms during the micturition cycle. This method is based on the preparation of flow charts and is applied to the discussion of four qualitative models which are derived from the literature. Opinions concerning the reflex arcs and supraspinal connections said to be involved in micturition and continence are different and sometimes contradictory. Little is known about supraspinal (inter)connections and their function in micturition control is still fragmentary. The control mechanisms which terminate voiding are not totally clear. Moreover, the role of the pelvic floor musculature in the control of the lower urinary tract is probably underestimated. The flow charts presented in this paper contribute to the future design of a single complete qualitative model representing the general central and peripheral nervous connections and control mechanisms. Such a model would provide an approach for future research in neuromodulation and neurostimulation of the uropo?tic system and a reduced version could be used for quantitative modelling, e.g. in neural network simulations.     

The survival of developing neurons: a review of afferent control

Developmental cell death is a major event of neurogenesis, and emphasis has systematically been placed on the roles of either the peripheral targets or central postsynaptic neurons in the control of neuronal survival. In this article, the main types of experimental design used to test the control of neuronal death by the afferent supply are compared with analogous data indicating neurotrophic support by the targets. It is argued that targets and afferents may have equivalent roles and interact in the control of neuron numbers during development of the vertebrate nervous system. Possible mechanisms of anterograde trophic control include contact-mediated cell interactions, activity-dependent processes mediated by neurotransmitters or neuromodulators, modulation of the levels of cytoplasmic free calcium and the involvement of neurotrophic factors.     

Cholinergic strategies for Alzheimer's disease

Alzheimer's disease is a devastating degenerative disorder of the central nervous system that results in gradual deterioration of cognitive function and severe alteration of personality. Degeneration of neurons in the nucleus basalis Meynert, the origin of the major cholinergic projections to the neocortex, occurs early in the course of the disease, and is correlated with the cognitive decline. This link between cholinergic dysfunction in the basal-cortical system and cognitive deficits has focused scientific efforts on developing tools to elucidate the neurobiological role of the cholinergic system in cognition and to develop therapeutic interventions in the disorder. An important step in understanding the mechanisms underlying cognitive dysfunction has been the development of in vivo rodent models that mimic some of the features of Alzheimer's disease. Acute excitotoxic or immunotoxic lesions of the nucleus basalis in rodents have revealed a role of the basal-cortical system in attention, learning and memory. More recent advances in developing mouse gene technology offer newer models to systematically examine the underlying neuropathological cascade leading to dysfunctions in mnemonic processing. Using in vivo rodent models, several cholinergic enhancement strategies have been tested and proven to be effective in alleviating lesion-induced cognitive deficits, including neuropharmacological approaches (acetylcholinesterase inhibitors), neurotrophic factor administration (nerve growth factor), and transplantation of cholinergic-enriched fetal grafts. Successful results have also been obtained using ex vivo gene transfer to deliver nerve growth factor or acetylcholine to compromised regions of the basal-cortical system. Gene therapy may be of particular interest for clinical applications, because this approach provides a method for topographically restricted and selective delivery of therapeutic genes and their products to afflicted areas of the brain. Advanced techniques in molecular biology (e.g., exogenous regulatable gene transfer) and newly developed tools of modern neuroscience (e.g., neural precursor cells) will be important contributions for deciphering the biological bases of neuronal degeneration and for refining therapeutic strategies for Alzheimer's disease.     

Glial cells in the enteric nervous system contain glial fibrillary acidic protein

The complex nervous networks found throughout the mammalian gut--the enteric nervous system--are histologically, ultrastructurally, and, to some extent, functionally--similar to the central nervous system. The glial cells of the small enteric ganglia are generally classified as Schwann or satellite cells, since they are found in the peripheral nervous system, possess nuclei which ultrastructurally resemble those of Schwann cells and are derived from the neural crest. However, it has been argued that these cells resemble astrocytes of the central nervous system with respect to gross and fine structure, and their relationship with the enteric neurones and their processes. In immunohistochemical studies of these cells, both in frozen sections of gut wall and in tissue culture preparations of the enteric plexuses, we found evidence that the enteric glial cells are rich in glial fibrillary acidic protein (GFAP), a protein associated with the 100 A glial intermediate filaments, and hitherto believed to be specific to astrocytes of the central nervous system only.     

Gene therapy of central nervous system tumors

Recently, remarkable advances have been achieved in molecular and genetic researches of different kinds of general diseases, as well as in basic and clinical studies using gene therapy for central nervous system diseases. For brain tumors, clinical trials have been already started in more than 10 clinical protocols and more than 100 patients with malignant brain tumors. Nevertheless, there are still major issues that remain to be resolved for achieving better clinical results, such as delivery system of genetic material, regulatory methods of the intracellular expression of the transgene, antitumor efficacy, and tumor selectivity. In this paper, molecular genetic studies and the current state of gene therapy for neurological diseases, especially brain tumors, are described, and the future direction of this fascinating approach is discussed.