BAX

Within the mitochondrial membrane are members of the bcl-2 family of proteins, some of which are pro-apoptotic and some anti-apoptotic. Antiapoptotic members (in white) prevent homodimerization of pro-apoptotic members, blocking cytochrome c release

NGF withdrawal

Scheme summarizing proposed mechanisms used by Ras to protect SCG neurons from apoptosis. Scheme summarizing proposed mechanisms used by Ras to protect SCG neurons from apoptosis. NGF withdrawal and araC induce neuronal death by different mechanisms that converge at a point upstream of cytochrome c release (38). The Ras/PI3K pathway is both necessary and sufficient to deliver the protective signal against apoptosis induced by NGF deprivation, but none of the downstream effects of this pathway (including Akt) can target any of the components mediating araC-induced release of cytochromec, activation of caspases, or apoptosis execution. Although the Ras/ERK pathway is activated concurrently, it can only protect neurons from araC-induced death. See “Discussion” for further details. Xue L et al. J. Biol. Chem. 2000;275:8817-8824

Oxygen can be modified so that it loses an electron, leaving one unpaired electron—this is a reactive oxygen species. Most common is superoxide or hydroxl radical, both of which can oxidize lipids or proteins and disable their function.

ROS are a huge coponent of apoptotic cell death and a hallmark of neurodegenerative diseases like AD And PD

Affect on transport after H2O2 exposure.

Neurogenesis: History Old Dogma: no new neurons after infancy 1960’s: (Altman) Cell proliferation in olfactory bulb and hippocampus 1980’s: (Nottebohm) Neurogenesis in adult canaries 1990’s: Neurogenesis occurs throughout life span in many species

Adult Neurogenesis Extracellular molecular regulators overlap and cooperate within neurogenic niches in the adult CNS. (a) Neurogenic niches of the dentate gyrus (DG) and forebrain, where neurons born in the subventricular zone (SVZ) migrate through the rostral migratory stream (RMS) to the olfactory bulb (OB). (b) In the dentate gyrus, new neurons are born in the subgranular zone just inside the granule cell layer (GCL). (c) Neurogenesis in the SVZ and neuroblast migration through the RMS to the OB probably results from integration of several cooperating and anatomically overlapping extracellular signaling pathways, involving growth factors, neurotransmitters, guidance factors, metalloproteases and hormones. This is most likely also the case in the dentate gyrus. Additional abbreviations: G, glomeruli of the olfactory bulb; LV, lateral ventricle. Hagg T, TINS 28 (2005)

Adult Hippocampal Neurogenesis (A) The basic tri-synaptic circuit of the hippocampus, showing inputs to and outputs from the DG granule cells. Stellate cells in the entorhinal cortex (EC) provide the major input to the granule cells via the perforant path. Axons of the granule cells, called mossy fibers, synapse on CA3 pyramidal cells, which in turn synapse onto CA1 pyramidal cells, which send their axons out of the hippocampus proper. (B) New granule cells are generated from progenitor cells residing in the subgranular zone (SGZ), at the border between the granule cell layer (GCL) and hilus. As the postmitotic daughter cell differentiates, it moves up slightly into the granule cell layer and develops dendrites that reach through the granule cell layer into the molecular layer and an axon that reaches into the CA3 region. As the new neuron matures, it develops a more mature dendritic morphology. Christie and Cameron, Hippocampus 16 (:2006)

Methods to Study Neural Precursor Cells In Vivo Mitotic Labeling: Thymidine or BrdU Problems: DNA repair, apoptosis Mitotic Staining (IHC): Ki67 and PCNA Variable expression during cell cycle phases (Ki67) and expression in early postmitotic cells (PCNA) Retroviral Labeling: reporter gene expression Advantages: cell identification for electrophysiology, no dilution of marker Disadvantages: tissue damage, infection rates are low and variable, fewer cells and differentiation markers @ long survival times Retroviral labeling: the virus is a modified nonreplicative oncoretrovirus: thus the genes required for replication are deleted and the viral particles will infect cells locally without enabling them to produce additional infectious virus particles Local infusion into the brain and the virus only enters dividing cells via a cell surface receptor Drive expression of GFP

Methods to Study Neural Precursor Cells In Vivo Electrophysiology and Retrograde Axonal Labeling Function and integration Neuronal identity Phenotypic Identification & Progression of Differentiation Double and triple staining methods Stem Cell markers (nestin/GFAP) Glia versus neurons Early(DCX, PSA-NCAM; Tuj1 and mature (NeuN) neurons Neurotransmitter phenotype

Neurogenesis in the Adult Human Brain? NeuN BrdU BrdU positive cells have a rounded apprearance and are found in the GCL BrdU cells found adjacent to ependymal lining in SVZ of human caudate Cells with elongated nuclei resembling migrating cells are in SVZ BrdU

Adult Neurogenesis Positive Regulation (Proliferation/Survival) Environmental Enrichment Exercise Memory Formation Dietary restriction Trophic Factors (shh, IGF-1, bFGF, HB-EGF, VEGF, BDNF) 5-HT (Antidepressants) Hormones (Estrogen, Thyroid) Pathological Conditions (Stroke, Seizure, Disease Models)

Adult Neurogenesis Negative Regulation (Proliferation/Survival) Aging Inflammation Depression Stress Glucocorticoids Neurotransmitters: Glumatate, ACh, DA Chemotherapy Methamphetamine Opiates

Evidence for Neurogenesis in the Human Brain Huntington’s Disease: PCNA/GFAP/Tuj1 Increased cell birth in subependymal zone More pronounced with increasing severity Alzheimer’s Disease PSA-NCAM, DCX, TUC-4 Elevated numbers of immature neurons in subgranular zone, granule cell layer, CA1 Stroke Cell cycle (Ki67, PCNA) and lineage restricted makers (Tuj1, DCX,PSA-NCAM, TUC-4) Increased expression in ischemic tissue surrounding infarcts (perivascular areas) There is an idea that when confronted with injury, neurons may undergo a type of plasticity that allows them to re-adopt a less differentiated state. This may (likely) involve epigenetic reprogramming. What advantages might there be to undifferentiated state?

Neuronal Phenotype of New Born Cells Following Stroke in Human CNS DCX/GFAP TuJ1 DCX/Ki67 TuJ1/Ki67 Fig. 2. Neuronal character of cells in the ischemic penumbra of human cerebral cortex after stroke. (a) The early neuronal marker DCX (green) is expressed in the cytoplasm of numerous cells in the ischemic penumbra, where its expression does not overlap with that of the astroglial marker GFAP (red). (Scale bar, 150 µm.) (b) The neuronal marker III-tubulin (red) is highly expressed in the ischemic penumbra (center of panel), is less highly expressed in adjacent normal cortex (top of panel), and is absent in the ischemic core (bottom of panel). (Scale bar, 200 µm.) (c) DCX (green) is expressed in the cytoplasm of a cell with Ki-67-positive (red) nucleus. (Scale bar, 5 µm.) (d) III-tubulin (red) is expressed in the cytoplasm of a cell with Ki-67-positive (green) nucleus. (Scale bar, 7 µm.) Jin, Kunlin et al. (2006) Proc. Natl. Acad. Sci. USA 103, 13198-13202

Caveats of Human Neurogenesis Studies Sources of cells are unclear Integration and Function have not been addressed Few cells are generated and in limited sites A potential endogenous source for repair? Therapeutic targets: environment and drugs?

Remaining Questions Regarding Adult Neurogenesis… • How do different extracellular signaling cascades cooperate to affect signaling within the neural precursors? Is there a link between neurotransmitters and growth factors? Which molecules induce specific neuronal phenotypes?

Remaining Questions Regarding Adult Neurogenesis… • Which molecules regulate axonal and dendritic growth, and functional integration of the new neurons? Are the molecular regulators in humans the same as in rodents, and how do neurological disorders affect these?