Life and Death for Neurons We lose 20-80% of our neurons during development Most neurons die right around the time that axons are invading the target 1.

Life and Death for Neurons

We lose 20-80% of our neurons during development

Most neurons die right around the time that axons are invading the target

1

Expt: Look at the effect of removing or adding a limb on neuron survival in the spinal cord of the Xenopus frog

Result: Adding a limb increases neuronal survival, while removing one reduces it.

Model: The target is making something in limited quantities that promotes cell survival.

Viktor Hamburger, 1920s

Why do neurons live or die?

2

Experiment : Transplant mouse sarcoma tumors near limb bud.

Result: More neurons in the dorsal root ganglia survive.

What makes neurons survive?

They isolated the factor and called it nerve growth factor.

Nobel 1986: Rita Levi-Montalcini and Stanley Cohen

Identification of nerve growth factor

3

How Nerve Growth Factor was isolated

1. Develop an assay: cultured Dorsal Root Ganglia in a dishAdded tumor extract, neuron outgrowth is dramatically enhanced

2. Identify a rich source of NGF: purified NGF from snake venom

4

NGF is the trophic factor that promotes cell survival

NGF is made by the target neuron

DRG + Sympathetic Neurons

They DIE

They SURVIVE+ NGF

They DIE

venom + NGF Ab

Does NGF really promote cell survival?

5

The Neurotrophic Hypothesis

• The target cells release a factor that promotes cell survival

• This factor is found in limiting quantities

• Cells compete to get enough factor to survive

6

There are many different trophic factors

•Neurotrophins

•NGF TrkA Receptor

•BDNF TrkB Receptor

•NT3 TrkC Receptor

•NT4/5 TrkB Receptor

•TGF-Beta Family

•Interleukin – 6 related cytokins

•FGFs

•SHH

They all also bind the p75 receptor

7

Neurotrophins act through receptor tyrosine kinase signaling cascades

8

Signal transduction for Neurotrophins

9

Other actions of neurotrophins

• Neuronal survival• Nerve growth• Nerve sprouting• Differentiation• Modulation of synaptic transmission• Electrical properties

10

A

Ne uro ns a p p ro a c h ta rg e t

De g e ne ra ting ne uro n

Ta rg e t so urc e o f ne uro tro p hic fa c to r

Lim ite d sup p ly o f ne uro tro p hic fa c to r

B

The Neurotrophic Hypothesis: neurons compete for limiting amounts of a neurotrophin

some neurons survive, other neurons die

11

How do neurons die?

Two kinds of cell death

1) NecrosisDeath by accident, ie trauma to tissue

2) Programmed Cell Death (Apoptosis)Death by designControlled cellular self-destruction

12

Normal Cell

Cell shrinks away from neighbours

Plasma membrane blebbingCytoplasmic and nuclear condensation

Chromatin condenses

Nuclear and cellular fragmentation

Apoptotic Bodies

Phagocytosis

The process of Programmed Cell Death (PCD)

13

PCD Necrosis

Nuclei Chromatin condensation, fragmentation

Irregular chromatin clumping

Cytoplasmic Organelles

Membranes intact Disrupted

vs

Morphological Differences between

Programmed Cell Death and Necrosis

DNA Cleaved into fragments No damage

14

A A

A

A

Morphological Appearance of PCD

Nucleus is fragmented, blebby

15

normal

PCD

Morphological appearance of PCD

DNA is fragmented into 180 bp pieces

16

normal PCD

C. elegans cell death Program

Complete lineage description of all 1090 cells - 131 cells die during development

Mutagen

Increased survival

Decreased survival

Isolate gene

Mutated Gene Phenotype ced3 blocked all 131 embryonic cell deaths ced4 blocked all 131 embryonic cell deaths ced9(g.o.f) absence of cell death, embryonic lethality

What molecules cause Programmed Cell Death?

17

The apoptotic pathway is conserved in worms and humans

18

Most of the molecules involved in PCD are novel

CED3 caspase is a cysteine protease….cleaves proteins

CED4 adaptor activates CED3

CED9 inhibits CED4

EGL-1 inhibits CED9

19

A

Ne uro ns a p p ro a c h ta rg e t

De g e ne ra ting ne uro n

Ta rg e t so urc e o f ne uro tro p hic fa c to r

Lim ite d sup p ly o f ne uro tro p hic fa c to r

B

Do trophic factors inhibit Programmed Cell Death?

20

Neurotrophins increase Bcl-2 expression to inhibit PCD

21

Overexpression (OE) of Bcl-2 gives bigger brains

12% volume increase in Bcl-2 OE brains

Normal

Bcl-2 OE

bcl-2 OE normal

22

Why is PCD used during development?

1. sculpting structures

2. deleting unneeded structures

3. controlling cell numbers

4. eliminating non-functional or harmful cells

23

Diseases Associated with Deregulated Apoptosis

Increased Apoptosis

AIDSNeurodegernative disorders Alzeheimer’s disease, Parkinson’s disease, Amyotrophic lateral sclerosis Retinitis pigmentosaMyelodysplastic syndromes Aplastic anaemiaIschaemic Injury Myocardial infarction, Stroke, Reperfusion injuryToxin-Induced liver disease Alcohol

Cancer Follicular lymphomas carinomas with p53 mutations hormone dependent tumours: breast cancer, prostate cancer, ovarian cancer

Autoimmune Disorders Systemic lupus erythematosus Immune-mediated glomerulonephritus

Viral Infections Herpesvirus, poxvirus, adenovirus

Inhibition of Apoptosis

24

Why does a neuron live or die?

1. Neurotrophins promote cell survival by inhibiting Programmed Cell Death

2. Programmed Cell Death allows death to occur without harming nearby cells

25