Chapter 53: The Generation and Survival of Nerve Cells

An Overall View

• The molecular basis of neuronal generation is similar throughout phylogeny

• Neuronal and glial fates are controlled by local signaling:

-secreted factors direct the differentiation of neural crest cells into neurons and glia

-glial cell differentiation in the central nervous system is also controlled by diffusible factors

• Neuronal fate in the mammalian cortex is influenced by the timing of cell differentiation

An Overall View

• The neurotransmitters phenotype of a neuron is controlled by signals from the neuronal target

• The survival of a neuron is also regulated by signals from the neuronal target:

target cells secrete a variety of neurotrophic factors

elimination of neurotrophic factors and their receptors leads to neuronal death

• Deprivation of neurotrophic factors activates a cell death program in neurons

Notch signaling controls specification of neural cell identity in Drosophila-

Lateral Inhibition

A small cluster of cells in proneural region of ectoderm in Drosophila acquire the potential to give rise to neuronal precursors.

Neuronal fate is decided by a process of signaling between adjacent cells involving two neurogenic genes delta and notch. Delta functions as a ligand and notch is its receptor.

A, Notch signaling is initially similar in pro-neural region of ectoderm that express achaete-scute class basic helix-loop-helix genes.

B, one cell begins to express higher level of delta, thereby activating notch in neighboring cells.

C, rapid amplification by feedback pathway involving suppressor of hairless and enhancer of split.

Notch-mediated lateral inhibition

• Amplification of an initial bias to create different cell types.

• A difference between two initially equivalent cells may arise randomly. Alternatively, interacting cells may have an intrinsic bias or an extrinsic bias.

• For instance, cells that have received different proteins in an asymmetric cell division will be intrinsically biased.

• Those that have received different signals will be extrinsically biased.

A Drosophila sensory bristle comprises four types of cells, hair cell, socket cell, sheath cell, and a sensory neuron.

The development of these distinct cells depends on the asymmetric cell divisions of a sensory organ precursor (SOP) cell.

Numb inhibits notch signaling and controls neural fate in Drosophila.

A typical Drosophila sensory bristle comprises four cells-the hair cell, socket cell, sheath cell, and a sensory neuron, all of which are derived from sensory organ precursor (SOP) cells.

Asymmetric distribution of numb (a ubiquitin ligase associated with the down-regulation of Notch) protein within SOP cells allow numb to be inherited by only one daughter cells after cell division.

The cell that does not receive numb expresses a high level of notch activity, giving rise to bristle and socket cells. The cells without notch give rise to neurons.

Wild-type No notch function No numb function

socket

bristle

sheath

neuron

Within the SOP cell, numb protein is localized to one side only. When the SOP divides, numb is inherited by only one of the daughter cells, cell IIB that gives rise to the sensory neuron and sheath cell.

Within the IIA or IIB cells, again only one daughter cell can inherit numb protein.

Subcellular localization of numb during mitosis of Drosophila neuroblasts.

Neuroblasts in Drosophila arise by segregating basally from the overlying epithelial cell layer. The neuroblast then divides asymmetrically along its apical-basal axis to generate a larger apical cell retaining neuroblast characteristics and a smaller basal ganglion mother cell. During this division numb and several other proteins become segregated to the ganglion mother cells.

Insc: inscuteable, a cytoskeletal protein transiently localizedto apical side of the neuroblasts.Miranda act as an adaptor protein to localize prospero, numb, andstaufen to the basal cortical membrane.

Three domains of primary neuron differentiation in the neural plate of Xenopus laevis develop as parallel stripes: lateral stripe become sensory neurons; intermediate stripes become interneurons; medial stripes become motor neurons (next slide). Before onset of neuronal differentiation, delta is expressed only in these three stripes, notch is expressed in both stripes and inter-stripe region.

Manipulations of notch signaling result in altered numbers of neurons generated within these stripes.

notch inhibitionnotch activation

inhibition of neuronaldifferentiation

increase of neuronnumbers

neurogenin or neuroD

additional neurondifferentiation

Control of Neuronal and Glial Fate in PNS by Local Signaling

Neural crest is a migratory cell population that emerges from the dorsal neural tube. Neural crest cell can give rise to many distinct cell types including neurons and Schwann cells.

Different environments encountered by neural crest cells during migration, and the distinct signals produced by these environment, appear to have a critical role in controlling their fate.

Superficial migratory pathway just beneath the ectoderm: pigment cells of the skin.

Intermediate pathway via somite: sensory ganglia.

Further medial pathway: sympathetic ganglia and the cells of adrenal medulla.

BMP and mash 1 induce differentiation of premigratory neural crest cells into autonomic ganglia.

Glial growth factor (GGF), a family member of neuregulin gene, is expressed on surface of autonomic neurons at early differentiation stage. GGF may direct glial differentiation and inhibit neurogenesis of nearby neural crest cells.

Glial cell differentiation in CNS is also controlled by diffusible factors.

PDGF secreted by astrocytes maintains the proliferation of oligodendrocyte progenitor cells. However, postnatally these progenitor cells begin to lose sensitivity to PDGF, despite the presence of PDGF in local environment, and differentiate into oligodendrocytes.

Later, astrocytes begin to secret CNTF that helps to promoter the differentiation of progenitor cells into astrocytes.

astrocyte oligodendrocyte

progenitor

Timing of cell differentiation influences neuronal fate in mammalian cortex

The neurons of the cerebral cortex are generated in the ventricular zone, an epithelial layer of progenitor cells lining the lateral ventricles. Once left cell cycle, the immature neurons migrate out of the ventricular zone to form the cortical plate, later becoming the gray matter of the cerebral cortex.

To reach to cortical plate the neurons migrate on radial glial cells, a specialized class of glial cells that retain contact with both the ventrical and pial surfaces.

Cortical cells obey an inside-first, outside-last program of neurogenesis. Neurons born within VZ at early stages migrate to the deepest layers of cortical plate. Neurons generated at later stages migrate past the earlier-generated neurons to form the more superficial layers of cortex.

Control of Subtype Identity of Newly Generated Cortical Neurons

Experiment 1: transplantation of older progenitor cells into ventricular zone of younger host.Results: older progenitor cells cannot acquire the fate of younger neurons.

Experiment 2: transplantation of younger progenitor cells into older animals.Results: if transplanted early in cell cycle, the cells adopt host fate; if transplanted late in final cell cycle, cells maintain original identity.

As development proceeds, not only do the signals that direct neurons to specific laminae change, the competence of neurons responding to those signals also changes.

The plane of division of progenitor cells in the VZ of cerebral cortex influences their fate: vertical cleavage of progenitor cells generates two similar daughters that retain apical connections. Horizontal cleavage produces asymmetric division. Apical daughter remains contact with apical surface and the basal daughter loses its apical contact, migrating

away from VZ and later becoming a postmitotic neuron.

Control of Neurotransmitter Phenotype by Neuronal Target

Most sympathetic neurons use norepinephrine as primary transmitter. However, sympathetic neurons innervate the exocrine sweat glands in the foot pads use acetylcholine instead.

Transplantation of foot pad sweat gland from a newborn rat into the skin normally innervated by noradrenergic sympathetic neurons results in cholinergic transmitter phenotype in these neurons.

Sympathetic neurons switch gradually from noradrenergic to cholinergic properties, passing through a stage where the neurons release both norepinephrine and acetylcholine.

LIF and CNTF switch sympathetic neurons from noradrenergic to cholinergic neuro-transmitter phenotype.

Differentiation of chromaffin cells from neural crest precursors are triggered by migration of these cells into adrenal gland with glucocorticoids.

NGF induces precursors to differentiate into sympathetic neurons with norepinephrine. small translucent vesicles

large dense-core granules

Neurotrophic Factors

Trophic, from the Greek meaning nourish, refers to the interaction between nerve cells that take days, months, or the life time of the animals. In contrast the conventional synaptic effects of neuronal signal transmission are usually measured in milliseconds or seconds.

Neurotrophic factors, including the class of neurotrophin, support the survival, growth, and differentiation of many neuronal populations in both central and peripheral nervous system.

Size or activity of muscle target controls motor neuron survival

A. Removal of a developing limb results in a marked decrease in the number of motor neurons in a chicken embryo.

B. Increasing the size of the limb target reduces the extent of naturally occurring neuronal death during development.

C. Blocking muscle activity prevents the developmental death of motor neurons. Direct stimulation of the muscle enhances the death of motor neurons. Activity of the target cells could inhibit the production of neurotrophic factor.

Neurotrophic Factor HypothesisNeurotrophic Factor Hypothesis was first form

ulated by Viktor Hamburger, Rita Levi-Montalcini and their colleagues in 1940s.

Target cells of developing neurons produce a limited amount of an essential nutrient, or trophic factor, that is taken up by the nerve terminals.

Neurotrophic factors bind specific receptors and are internalized and transported retrogradely to the neuronal cell somata, where they promote neuronal survival. Neurons without access to adequate amounts of these factors die by apoptosis.

Neurotrophic Factor Hypothesis

retrograde transport

neutrophic factor

target cellsneurons

apoptotic neurons

surviving neurons

During the development of nervous system, neuronal populations undergo a process of naturally occurring cell death at a time when their axons are innervating target areas. This mechanism ensure a balance between the size of an innervating population and the size of its target territory, thereby regulating the number of neurons and neuronal connection in developing CNS.

What might be the purpose of such a mechanism?

Identification of the First Neurotrophic Factor, NGF

NGF was first discovered by Levi-Montalcini in 1951.NGF consists of three subunits, , , and . The biol

ogically active component of NGF contains two identical 118-amino acid subunits, with a molecular weight of 13,259 in each subunit.

Mature -NGF is synthesized from prepro- NGF (27 kDa) and exists within the CNS as a homodimer.

While the monomer also exhibits growth promoting activity, it is the dimeric form that is physiologically important.

Five neurotrophins have been isolated and divided into three classes.

Nerve growth factor (NGF) trkANeurotrophin-6 trkABrain-derived neurotrophic factor

(BDNF) trkBNeurotrophin-4/5 (NT-4/5) trkBNeurotrophin-3 (NT-3) trkC

trk: tyrosine kinase receptor homodimer

Similar low affinity binding to pan-neurotrophin receptor p75NTR

The entire family of neurotrophins exhibits a distant relationship to transforming growth factor (TGF) family.

Ciliary neurotrophic factor (CNTF) and leukemia inhibitory factor (LIF) are structurally related to several cytokines including interleukin-6 and granulocyte-colony-stimulating factor.

heterodimercomplex

Targetgenes

Both CTNF and LIF go through the signaling pathways whereby dimerized receptor complexes activate associated cytoplasmic Janus kinase (JAK) protein tyrosine kinases, which phosphorylate many substrates including signal transducer and activator of transcription (STAT) transcription factors.

Glia-derived neurotrophic factor (GDNF) is another distant relative of TGF- and shares some structural features with neurotrophins.

Distinct neuronal subtypes depend on different neurotrophic factors

Apoptosis versus Necrosis

Apoptosis (also known as programmed cell death) is defined as a process of active cell death, characterized by cell and nuclear shrinkage, chromatin condensation and cleavage of DNA into oligonucleosomal-sized fragments.

Necrosis is characterized by cellular swelling and lysis, causing an inflammatory response and random DNA fragmentation.

Deprivation of neurotrophic factors activates apoptosis in neurons: hint

Sympathetic neurons in cultures are maintained in continuous presence of NGF and die otherwise. Surprisingly, inhibiting synthesis of protein (by cycloheximide) or RNA (by actinomycin D) at the time of NGF removal prevents cell death, indicating cell death induced by NGF withdrawal requires de novo RNA/protein synthesis. Thus, neurotrophins suppress an endogenous default death program.

Genetic studies on nematode worm C. elegans have identified cell death genes (ced) that either block or promote apoptosis.

bcl-2 family

ced-4 family

caspase family Ced-3

Ced-4

Ced-9

Apaf-1 (apoptosis activating factor-1)

Caspases 1-14

Caspases are generated in an inactive precursor form. Cleavage of the caspase precursor results in the removal of a prodomain and the subsequent assembly of proteolytically active protein.

Apaf-1 is thought to interact with the caspase precursor via a caspase recruitment domain or death effector domain. Bcl-2 related proteins appear to inhibit caspase activation by binding directly to Apaf-1, thereby preventing its ability to trigger pro-caspase cleavage.

Neurotrophic factor deprivation triggers caspase activation and apoptosis. NGF/trk signal activates bcl-2, which in turn inhibits Apaf-1 activity to block caspase cleavage. Removal of NGF permits cleavage of pro-caspases and results in cell death.

Activation of a specific caspase often leads to proteolytic cleavage of other caspases, thus initiating a cascade of caspase activation that leads eventually to the proteolysis of non-caspase proteins essential for cell viability.