Berkeley Feb 2004

Outer membrane

Inter-membrane space

Inner membrane

Matrix

Cristae

ATP

ADP + Pi

H+H+

ATP hydrolysis

++--

ATP

ADP + Pi

H+ H+

Energy (redox energy, photons etc)

The proton circuit: Continuous generation

of ATP

Electrons enter the respiratory chain at a redox potential

of–300mV

Ubiquinone pool at 0mV

Cytochrome c at +250mV H2O

at +800mV

Complex I

Complex IIComplex III

Complex IV

ATP synthase

Proton extrusion generates large membrane potential (~150mV) and small pH gradient pH (~0.5pH units)

+150mV, -0.5pH

Total driving force for protons = protonmotive force p = pH

Electrical circuit

Vo

ltag

e 1

.5V

Electron current

Mitochondrion

Vo

ltag

e 0

.2V

Proton current

Mitochondrial membrane

+

_

+

_

Mitochondria work like an electrical circuit.

•The 'battery' is the respiratory chain

Mitochondrion

Vo

ltag

e 0

.2V

Proton current

Mitochondrial membrane

+

_

I III IV

The respiratory chain has 3 proton pumps in parallel with respect to the proton circuit

The proton circuit is governed by Ohm’s law:

Current = voltage/resistance

roughly:Respiration rate = constant x membrane potential / resistance to proton re-entry*

*through ATP synthase or leakage across membrane

m

Matrix Ca 2+

ATP generation

Reactiveoxygenspecies

NADP+

REDUCTION

Glutathionereduction

Respiratory chain

Mitochondria and ‘stress’ i.e. under which circumstances might theMitochondria’s ability to maintain a high ATP/ADP ratio and a reducing

environment fail?

A. Oxygen limitation (stroke, heart attack, near drowning)

synaptic cleft 1M

cytoplasm 10mM

synaptic vesicles 100mM

'Classic EM images from the lab of John Heuser (Washington University)'

GLUTAMATE COMPARTMENTATION


GLUTAMATE EXCITOTOXICITY

1. (BIOENERGETIC DEFICIT RESULTING FROM OXYGEN

DEPROIVATION CAUSES ATP COLLAPSE, FAILURE OF

PLASMA MEMBRANE SODIUM PUMPS AND

MASSIVE GLUTAMATE RELEASE)



2. POST-SYNAPTIC NMDA RECEPTORS

PATHOLOGICALLY ACTIVATED



3. MASSIVE Ca 2+ ENTRY AND ACCUMULATION BY

MITOCHONDRIA



4. MITOCHONDRIAL Ca 2+ LOADING CAN INITIATE DELAYED CELL DEATH


environment fail?

B. Respiratory chain restriction

I

II

III IVQ

Rotenone

Malonate

Some specific mitochondrial inhibitors

Langston JW, Ballard P, Tetrud JW, Irwin I (1983) Chronic Parkinsonism in humans due to a product of meperidine-analog synthesis. Science 219: 979-980.

Four persons developed marked parkinsonism after using an illicit drug intravenously. Analysis of the substance injected by two of these patients revealed primarily 1-methyl-4-phenyl-1,2,5,6-tetrahydropyridine (MPTP) with trace amounts of 1-methyl-4-phenyl-4-propionoxy-piperidine (MPP+). it is proposed that this chemical selectively damages cells in the substantia nigra

Nicklas WJ, Vyas I, Heikkila RE (1985) Inhibition of NADH-linked oxidation in brain mitochondria by 1-methyl-4-phenyl-pyridine, a metabolite of the neurotoxin, 1-methyl-4-phenyl-1,2,5,6-tetrahydropyridine. Life Sci 36: 2503-2508. (MPP+), a major metabolite of the neurotoxin, (MPTP) inhibited the oxidation of NADH-linked substrates by brain mitochondrial preparations. Compromise of mitochondrial oxidative capacity by MPP+ could be an important factor in mechanisms underlying the toxicity of MPTP

Betarbet R, Sherer TB, MacKenzie G, Garcia-Osuna M, Panov AV, Greenamyre JT (2000) Chronic systemic pesticide exposure reproduces features of Parkinson's disease. Nat Neurosci 3: 1301-1306.

chronic, systemic inhibition of complex I by the lipophilic pesticide, rotenone, causes highly selective nigrostriatal dopaminergic degeneration. These results indicate that chronic exposure to a common pesticide can reproduce the anatomical, neurochemical, behavioral and neuropathological features of PD.

Beal MF, Brouillet E, Jenkins B, Henshaw R, Rosen B, Hyman BT (1993) Age-dependent striatal excitotoxic lesions produced by the endogenous mitochondrial inhibitor malonate. J Neurochem 61: 1147-1150.

Abstract: Intrastriatal injection of malonate, a reversible inhibitor of succinate dehydrogenase (SDH), produced age dependent striatal lesions… The results strengthen the possibility that a subtle impairment of energy metabolism may play a role in the pathogenesis of Huntington's disease.


environment fail?

C. Ca2+ overload of mitochondria and activation of the permeability transition

extr

a-m

itoch

ondr

ial f

ree

C

a c

onc

ent

ratio

n (

M)

2+

mitochondria

Ca2+

Ca2+

Ca2+

'set-point'

Ca 2+

Permeability transition

Permeability transition leads to matrix swelling, unfolding of inner membrane, bursting of outer membrane and release of cytochrome

c


environment fail?

D. Mitochondria and pro-apoptotic stress

PlasmaMembrane

OuterMitochondrial

Membrane

InnerMitochondrial

Membrane

PutativeBAx/Bid channel

CQ

9-05

Deathreceptor

Procaspase-8 Caspase-8

Bid t-Bid

Bax

Bax*

C

Apaf-1+

Procaspase-9

Caspase-9

Activation of downstreameffector caspases

e.g. caspase-3

Bcl-2(-)(+)

Ca 2+

Reactive Oxygen Species

Cell Death

Apoptoticsignal

Cytochrome c

Caspase activation

Cell Death

Apoptosis Necrosis


environment fail?

E. Mitochondria and oxidative stress

superoxide

superoxide

Reactive oxygen species (ROS): superoxide anion produced by complexes I and III (increased at high membrane potential)

O2 + e- = O2.-

cf:

O2 + 4e- + 4H+ = 2H2O

O2.-

H2O2

Superoxide dismutases; SOD1, CuZn, cytoplasmic

SOD2, Mn, mitochondrial matrix

SOD

Glutathione peroxidase

H2O

GSH GSSG

NADPH NADP+

NADH NAD+

Glutathione reductase

transhydrogenase

NO

ONOO-

Mitochondria and ‘stress’ i.e. under which circumstances might the mitochondria’s ability to maintain a high ATP/ADP ratio and a

reducing environment fail?

F. Defects in the mitochondrial genome

cyt b

ND5

16s rRNA

12s rRNA

ND6

LHON 1448

4C

LHON 14459A

O (origin)H

ND4

ND4LND3

CO

XIIIATPase 6

ATP

ase 8

CO

XII

COXI

ND2

ND1MELAS 3243G

LHON 11778A

NA

RP 8993G

/C

ME

RR

F 8344G

LHON 3460A

common d

eletio

n

O (origin)L

9-06

Heavy

chain

Light

chain

Table 9.1 some mitochondrial mutations

location

Pearson's syndrome, Kearns-Sayresyndrome, chronic progressiveexternal ophthalmoplegia (CPEO)

‘Common deletion’ of4977 base pairsbetween A8 and ND5.

All mt protein synthesisabolished due to lack of tRNAs

MELAS (mitochondrialencephalomyopathy, lactic acidosisand stroke-like episodes).

tRNAleu(UUR)

All mt protein synthesisabolished due to lack of tRNAs

MERRF (myoclonus, epilepsy,with ragged-red fibers)

tRNAlys All mt protein synthesisabolished due to lack of tRNAs

Leber’s hereditary opticneuropathy (LHON),

6 point mutations inComplex I ND genes

loss of Complex I activity

NARP (neuropathy, ataxia andretinitis pigmentosa).

A6 Inhibition of ATP synthase

Schapira AHV (1998) Mitochondrial dysfunction in neurodegenerative disorders. Biochim Biophys Acta Bio-Energetics 1366: 225-233

Mutations of mitochondrial DNA (mtDNA) are associated with a wide spectrum of disorders encompassing the myopathies, encephalopathies and cardiomyopathies, in addition to organ specific presentations such as diabetes mellitus and deafness. Parkinson's disease , Huntington's disease , Friedreich's ataxia. In any event, mitochondria present an important target for future strategies for 'neuroprotection' to prevent or retard neurodegeneration.

•Paperback: 288 pages •Publisher: Academic Press; ISBN: 0125181213; •$49.95

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