Gαz regulates BDNF-induction of axon growth in cortical neurons

The disruption of neurotransmitter and neurotrophic factor signaling in the central nervous system (CNS) is implicated as the root cause of neuropsychiatric disorders, including schizophrenia, epilepsy, chronic pain, and depression. Therefore, identifying the underlying molecular mechanisms by which neurotransmitter and neurotrophic factor signaling regulates neuronal survival or growth may facilitate identification of more effective therapies for these disorders. Previously, our lab found that the heterotrimeric G protein, Gz, mediates crosstalk between G protein-coupled receptors and neurotrophin signaling in the neural cell line PC12. These data, combined with Gαz expression profiles – predominantly in neuronal cells with higher expression levels corresponding to developmental times of target tissue innervation – suggested that Gαz may play an important role in neurotrophin signaling and neuronal development. Here, we provide evidence in cortical neurons, both manipulated ex vivo and those cultured from Gz knockout mice, that Gαz is localized to axonal growth cones and plays a significant role in the development of axons of cortical neurons in the CNS. Our findings indicate that Gαz inhibits BDNF-stimulated axon growth in cortical neurons, establishing an endogenous role for Gαz in regulating neurotrophin signaling in the CNS.     

Comparative analysis of neurotrophin receptors and ligands in vertebrate neurons: tools for evolutionary stability or changes in neural circuits?

To better understand the role of multiple neurotrophin ligands and their receptors in vertebrate brain evolution, we examined the distribution of trk neurotrophin receptors in representatives of several vertebrate classes. Trk receptors are largely expressed in homologous neuronal populations among different species/classes of vertebrates. In many neurons, trkB and trkC receptors are co-expressed. TrkB and trkC receptors are primarily found in neurons with more restricted, specialized dendritic and axonal fields that are thought to be involved in discriminative or 'analytical' functions. The neurotrophin receptor trkA is expressed predominantly in neurons with larger, overlapping dendritic fields with more heterogeneous connections ('integrative' or 'modulatory' systems) such as nociceptive and sympathetic autonomic nervous system, locus coeruleus and cholinergic basal forebrain. Surveys of trk receptor expression and function in the peripheral nervous system of different vertebrate classes reveal trends ranging from dependency on a single neurotrophin to a more complex dependency on increasing numbers of neurotrophins and their receptors, for example, in taste and inner ear innervation. Gene deletion studies in mice provide evidence for a complex regulation of neuronal survival of sensory ganglion cells by different neurotrophins. Although expression of neurotrophins and their receptors is predominantly conserved in most circuits, increasing diversity of neurotrophin ligands and their receptors and a more complex dependency of neurons on neurotrophins might have facilitated the formation of at least some new neuronal entities.     

Neurotransmitter phenotype-specific expression changes in developing sympathetic neurons

During late developmental phases individual sympathetic neurons undergo a switch from noradrenergic to cholinergic neurotransmission. This phenomenon of plasticity depends on target-derived signals in vivo and is triggered by neurotrophic factors in neuronal cultures. To analyze genome-wide expression differences between the two transmitter phenotypes we employed DNA microarrays. RNA expression profiles were obtained from chick paravertebral sympathetic ganglia, treated with neurotrophin 3, glial cell line-derived neurotrophic factor or ciliary neurotrophic factor, all of which stimulate cholinergic differentiation. Results were compared with the effect of nerve growth factor, which functions as a pro-noradrenergic stimulus. The gene set common to all three comparisons defined the noradrenergic and cholinergic synexpression groups. Several functional categories, such as signal transduction, G-protein-coupled signaling, cation transport, neurogenesis and synaptic transmission, were enriched in these groups. Experiments based on the prediction that some of the identified genes play a role in the neurotransmitter switch identified bone morphogenetic protein signaling as an inhibitor of cholinergic differentiation.     

The role of neurotrophins in axonal growth, guidance, and regeneration

Neurotrophins are proteins that regulate neuronal survival, axonal growth, synaptic plasticity and neurotransmission. They are members of the neurotrophic factors family and include factors such as the nerve growth factor (NGF), the brain derived neurotrophic factor (BDNF), the neurotrophin-3 (NT-3), and the neurotrophin-4/5 (NT-4/5). These molecules bind to two types of receptors: i) tyrosine kinase receptors (TrkA, TrkB, TrkC) and ii) a common neurotrophin receptor (p75NTR). The two receptor types can either suppress or enhance each other's actions. Neurotrophins have a multifunctional role both in the central and peripheral nervous system. They have been suggested as axonal guidance molecules during the growth and regeneration of nerves. It has also been proven that they stimulate axonal growth by mediating the polymerization and accumulation of F-actin in growth cones and axon shafts. Neurotrophins, as other neurotrophic factors, have been shown that they reduce neuronal injury by exposure to excitotoxins, glucose deprivation, or ischemia. Furthermore, the nerve regeneration promoting effect of these growth factors is well documented for many different models of central or peripheral nervous system injury. Several studies have shown that exogenous administration of these factors has protective properties for injured neurons and stimulates axonal regeneration. Based on these properties, these molecules may be used as therapeutic agents for treating degenerative diseases and traumatic injuries of both the central and peripheral nervous system.     

Dissecting the diverse actions of pro- and mature neurotrophins

The neurotrophins mediate diverse actions in the developing peripheral and central nervous systems. They are initially synthesized as precursor forms, or proneurotrophins, that are cleaved to release C-terminal mature forms that bind to Trk receptor tyrosine kinases to enhance synaptic plasticity and neuronal survival. Recent studies suggest that proneurotrophins are not inactive precursors, but signaling proteins that can activate the p75 receptor to mediate diverse responses. Proneurotrophins can activate a heteromeric receptor complex of p75 and sortilin to initiate cell death, or bind to p75 in hippocampal neurons to enhance long term depression. Thus, neurotrophin actions are regulated by the form of the neurotrophin (pro- or mature) secreted by cells, by extracellular proteolytic cleavage of proneurotrophins to generate mature forms, and by the expression of neurotrophin receptors Trk, or p75 and sortilin, that are selectively activated by mature or proneurotrophins, respectively. Here, recent studies are reviewed that reveal that pro- and mature neurotrophins have distinct and sometimes opposing actions in regulating cell death and survival in development and in pathophysiologic states, in regulating neurotrophin secretion, and in modulating synaptic plasticity.     

Satellite glial cells in sympathetic and parasympathetic ganglia: In search of function

Glial cells are established as essential for many functions of the central nervous system, and this seems to hold also for glial cells in the peripheral nervous system. The main type of glial cells in most types of peripheral ganglia - sensory, sympathetic, and parasympathetic - is satellite glial cells (SGCs). These cells usually form envelopes around single neurons, which create a distinct functional unit consisting of a neuron and its attending SGCs. This review presents the knowledge on the morphology of SGCs in sympathetic and parasympathetic ganglia, and the (limited) available information on their physiology and pharmacology. It appears that SGCs carry receptors for ATP and can thus respond to the release of this neurotransmitter by the neurons. There is evidence that SGCs have an uptake mechanism for GABA, and possibly other neurotransmitters, which enables them to control the neuronal microenvironment. Damage to post- or preganglionic nerve fibers influences both the ganglionic neurons and the SGCs. One major consequence of postganglionic nerve section is the detachment of preganglionic nerve terminals, resulting in decline of synaptic transmission. It appears that, at least in sympathetic ganglia, SGCs participate in the detachment process, and possibly in the subsequent recovery of the synaptic connections. Unlike sensory neurons, neurons in autonomic ganglia receive synaptic inputs, and SGCs are in very close contact with synaptic boutons. This places the SGCs in a position to influence synaptic transmission and information processing in autonomic ganglia, but this topic requires much further work.     

Prepubertal estrogen exposure modifies neurotrophin receptor expression in celiac neurons and alters ovarian innervation

Estradiol is a key hormone in the regulation of reproductive processes acting both on peripheral organs and sympathetic neurons associated to reproductive function. However, many of its regulatory effects on the development and function on the sympathetic neurons have not been completely clarified. Sympathetic neurons located in the celiac ganglion projects to visceral, vascular and glandular targets, and contribute to ovarian innervation, being the main source of sympathetic fibers. In the present study, we analyze the effects of elevated levels of exogenous estrogen during the prepubertal period in post-ganglionic sympathetic neurons. Estrogen exposure induced a significant increase in sympathetic celiac neuronal size and modified the expression of neurotrophin receptor p75. This change affected mainly small and medium size neurons. The effect of estrogens on innervation of celiac target organs was heterogeneous, inducing a significant increase in catecholaminergic innervation of the ovary, but not of the pyloric muscular layers. These findings further support the role of estrogen as a modulator of neuronal plasticity and suggest that estrogen could modify some features involved in the relation between sympathetic immature peripheral neurons and their target organs throughout a neurotrophin-dependent mechanism.     

Energetics and oxidative stress in synaptic plasticity and neurodegenerative disorders

As in other cells, neurons use adenosine triphosphate (ATP) as an energy source to drive biochemical processes involved in various cell functions, and produce reactive oxygen species (ROS) as “by products” of oxidative phosphorylation. However, the electrical excitability and structural and synaptic complexity of neurons present unusual demands upon cellular systems that produce or respond to ATP and ROS. Mitochondria in axons and presynaptic terminals provide sources of ATP to drive the ion pumps that are concentrated in these structures to rapidly restore ion gradients following depolarization and neurotransmitter release. Mitochondria may also play important roles in the regulation of synaptic function because of their ability to regulate calcium levels and ROS production. ROS generated in response to synaptic activity are now known to contribute to the regulation of long-term structural and functional changes in neurons, and the best-known example is the nitric oxide radical. The high-energy demands of synapses, together with their high levels of ROS production, place them at risk during conditions of increased stress, which occur in aging, neurodegenerative disorders such as Alzheimer’s and Parkinson’s diseases, and after acute traumatic and ischemic insults. Energy depletion and/or increased oxidative damage to various synaptic proteins can result in a local dysregulation of calcium homeostasis and synaptic degeneration. Accordingly, recent studies have shown that dietary and pharmacological manipulations that improve energy efficiency and reduce oxyradical production can prevent synaptic degeneration and neuronal death in experimental models of neurodegenerative disorders. A better understanding of the molecular control of subcellular energy production and utilization, and of the functional relationships between energy metabolism, ion homeostasis, and cytoskeletal and vesicular dynamics, will provide novel insight into mechanisms of neuronal plasticity and disease.     

Inhibition of glial maturation by bone morphogenetic protein 2 in a neural crest-derived cell line

Bone morphogenetic proteins (BMPs) regulate developmental decisions in many neural and nonneural lineages. BMPs influence both CNS neuronal and glial development and promote neuronal differentiation in neural crest derivatives. We investigated the actions of BMP2 on glial differentiation in the peripheral nervous system using NCM1 cells, a neural crest-derived cell line with the properties of peripheral glial precursor cells. BMP2 prevented the acquisition of a mature Schwann cell-like morphology, blocking the expression of mature genes and maintaining expression of several early glial markers. We provide evidence that BMP2 activates the GFAP promoter and define signaling pathways underlying this regulation. Our results demonstrate a novel role for BMPs as inhibitors of glial differentiation in the peripheral nervous system and suggest that BMPs may regulate the developmental timing of glial maturation.     

Target cells promote the development and functional maturation of neurons derived from a sympathetic precursor cell line

The role of target interactions in the development and functional maturation of peripheral neurons was investigated using an immortalized sympathetic precursor cell line. bMAH cells underwent neuronal differentiation in response to neurotrophic factors, but maintained an immature neuronal phenotype characterized by small cell bodies and continued cell division. Co-culture with cardiac myocytes, a target of sympathetic innervation, promoted the appearance of large-diameter postmitotic bMAH neurons. Analysis of bMAH maturation in the presence and absence of co-cultured myocytes indicated that myocyte-derived factors promoted the survival of maturing bMAH neurons prior to their acquisition of nerve growth factor dependence. Myocyte interactions also promoted the functional maturation of bMAH neurons, leading to an increase in the localization of synaptic vesicle proteins into neuritic varicosities and the acquisition of sympathetic-like intrinsic electrical properties. Like primary sympathetic neurons, mature bMAH neurons formed functional connections to cardiac myocytes as measured by evoked postsynaptic responses in connected myocytes. The effects of myocyte co-culture on developing bMAH neurons could be mimicked by myocyte conditioned medium, indicating that cardiac myocytes produce soluble factors that promote the appearance of mature neurons. These experiments indicate that targets of innervation play a role in directing the development and final maturation of peripheral neurons.     

Microglial control of neuronal death and synaptic properties

Microglia have long been characterized by their immune function in the nervous system and are still mainly considered in a beneficial versus detrimental dialectic. However a review of literature enables to shed novel lights on microglial function under physiological conditions. It is now relevant to position these cells as full time partners of neuronal function and more specifically of synaptogenesis and developmental apoptosis. Indeed, microglia can actively control neuronal death. It has actually been shown in retina that microglial nerve growth factor (NGF) is necessary for the developmental apoptosis to occur. Similarly, in cerebellum, microglia induces developmental Purkinje cells death through respiratory burst. Furthermore, in spinal cord, microglial TNFalpha commits motoneurons to a neurotrophic dependent developmental apoptosis. Microglia can also control synaptogenesis. This is suggested by the fact that a mutation in KARAP/DAP12, a key protein of microglial activation impacts synaptic functions in hippocampus, and synapses protein content. In addition it has been now demonstrated that microglial brain-derived neurotrophin factor (BDNF) directly regulates synaptic properties in spinal cord. In conclusion, microglia can control neuronal function under physiological conditions and it is known that neuronal activity reciprocally controls microglial activation. We will discuss the importance of this cross-talk which allows microglia to orchestrate the balance between synaptogenesis and neuronal death occurring during development or injuries.     

Coexpression of cholinergic and noradrenergic phenotypes in human and nonhuman autonomic nervous system

It has long been known that the sympathetic innervation of the sweat glands is cholinergic in most mammalian species and that, during development, rodent sympathetic cholinergic sweat gland innervation transiently expresses noradrenergic traits. We show here that some noradrenergic traits persist in cholinergic sympathetic innervation of the sweat glands in rodents but that lack of expression of the vesicular monoamine transporter renders these cells functionally nonnoradrenergic. Adult human sweat gland innervation, however, is not only cholinergic but coexpresses all of the proteins required for full noradrenergic function as well, including tyrosine hydroxylase, aromatic amino acid decarboxylase, dopamine beta-hydroxylase, and the vesicular monoamine transporter VMAT2. Thus, cholinergic/noradrenergic cotransmission is apparently a unique feature of the primate autonomic sympathetic nervous system. Furthermore, sympathetic neurons innervating specifically the cutaneous arteriovenous anastomoses (Hoyer-Grosser organs) in humans also possess a full cholinergic/noradrenergic cophenotype. Cholinergic/noradrenergic coexpression is absent from other portions of the human sympathetic nervous system but is extended in the parasympathetic nervous system to intrinsic neurons innervating the heart. These observations suggest a mode of autonomic regulation, based on corelease of norepinephrine and acetylcholine at parasympathocardiac, sudomotor, and selected vasomotor neuroeffector junctions, that is unique to the primate peripheral nervous system.     

Rapid estrogen signaling in the brain: implications for the fine-tuning of neuronal circuitry

Rapid actions of estrogens were first described >40 years ago. However, the importance of rapid estrogen-mediated actions in the CNS is only now becoming apparent. Several lines of evidence demonstrate that rapid estrogen-mediated signaling elicits potent effects on molecular and cellular events, resulting in the "fine-tuning" of neuronal circuitry. At an ultrastructural level, the details of estrogen receptor localization and how these are regulated by the circulating hormone and age are now becoming evident. Furthermore, the mechanisms that allow membrane-associated estrogen receptors to couple with intracellular signaling pathways are also now being revealed. Elucidation of complex actions of rapid estrogen-mediated signaling on synaptic proteins, connectivity, and synaptic function in pyramidal neurons has demonstrated that this neurosteroid engages specific mechanisms in different areas of the brain. The regulation of synaptic properties most likely underlies the fine-tuning of neuronal circuitry. This in turn may influence how learned behaviors are encoded by different circuitry in male and female subjects. Importantly, as estrogens have been suggested as potential treatments of a number of disorders of the CNS, advancements in our understanding of rapid estrogen signaling in the brain will serve to aid in the development of potential novel estrogen-based treatments.     

Mechanisms underlying synaptic vulnerability and degeneration in neurodegenerative disease

Recent developments in our understanding of events underlying neurodegeneration across the central and peripheral nervous systems have highlighted the critical role that synapses play in the initiation and progression of neuronal loss. With the development of increasingly accurate and versatile animal models of neurodegenerative disease it has become apparent that disruption of synaptic form and function occurs comparatively early, preceding the onset of degenerative changes in the neuronal cell body. Yet, despite our increasing awareness of the importance of synapses in neurodegeneration, the mechanisms governing the particular susceptibility of distal neuronal processes are only now becoming clear. In this review we bring together recent developments in our understanding of cellular and molecular mechanisms regulating synaptic vulnerability. We have placed a particular focus on three major areas of research that have gained significant interest over the last few years: (i) the contribution of synaptic mitochondria to neurodegeneration; (ii) the contribution of pathways that modulate synaptic function; and (iii) regulation of synaptic degeneration by local posttranslational modifications such as ubiquitination. We suggest that targeting these organelles and pathways may be a productive way to develop synaptoprotective strategies applicable to a range of neurodegenerative conditions.     

Relationship between differentiation and terminal mitosis: chick sensory and ciliary neurons differentiate after terminal mitosis of precursor cells, whereas sympathetic neurons continue to divide after differentiation

A population of undifferentiated cells has been characterized during the early development of nodose and ciliary ganglia. This population is defined by the absence of surface markers specific for neurons (tetanus toxin receptor, Q211 antigen) and for glial cells (O4 antigen). These undifferentiated cell populations were isolated from the ganglia and were shown to contain neuronal precursor cells that were able to differentiate in vitro into neurons, as characterized by morphology and surface antigens. Undifferentiated cells were detected during the period of neuronal birth, indicating that dividing neuronal precursor cells do not express neuron-specific surface markers. This was directly shown by 3H-thymidine-labeling studies using nodose ganglia, ciliary ganglia, and dorsal root ganglia. In sympathetic ganglia, however, no undifferentiated neuronal precursor cells were detectable at developmental stages when sympathetic neurons are born. 3H-Thymidine injected during that stage at E7 was incorporated into cells expressing the neuronal markers tetanus toxin receptor and Q211 antigen. Quantitative fluorimetric determination of the DNA content of dissociated sympathetic ganglion cells demonstrated the presence of a population of Q211-positive sympathetic ganglion cells in the G2 phase of the cell cycle. E7 sympathetic ganglion cells expressing neuronal surface markers were also shown to be able to divide in vitro. We have concluded that the relationship between terminal mitosis and the onset of differentiation differs between ganglia of the chick peripheral nervous system: Sympathetic ganglion cells continue to divide after the acquisition of neuronal properties, whereas neuronal precursor cells from other autonomic and sensory ganglia start to differentiate after a terminal mitosis.