Lecture 26; Ischemic Brain injury 3 Flashcards
Describe the stages of ischemic injury;
*** Revisit slide 1
Hypoxia-ischemia
- Primary Injury (Reperfusion) - Latent Phase (~6hrs) Secondary Phase 6-72hrs Tertiary Phase (days-months)
= Brain Injury
Whats the reality of the primary phase injury?
Only very few cells actually die
Bulk cell death in secondary phase due to incapacity to generate ATP
What happens in the secondary phase of injury?
12-72 h after ischemia
Bulk cell death
Seizures
Secondary cell swelling
Secondary energy failure (in the presence of normal oxygen and glucose supply)
Describe what happens to cytochrome oxidase over time;
*** look at the graphs for her three lectures to understand
- Primary phase = Increase in Cyt C becomes oxidised
- Secondary phase (delayed loss) = Drop off of cytox b/c loss of mitochondrial function due to damage and cyt.c is pushed out of the mitochondria (partially responsible for secondary energy failure) as part of apoptotic signalling.
- Sham control demonstrates mitochondrial circadian rhythm with increased activity at night.
Note*** Delayed mitochondrial failure after asphyxia in fetal sheep
What else happens after ischemia?
Delayed seizures and secondary cell swelling after ischemia
During this time there is also secondary cytotoxic edema.
Describe the EEG for ischemia evolving injury;
Ischemia ; Rapid depression of EEG activity as brain shuts down to save energy
Latent phase; remains depressed
Secondary phase; Increased activity, reflecting seizures (high amp, evolving nature) (also secondary cytotoxic edema as measured by impedence)
Fetus age and injury length dependant
What are the signals of ischemic brain injury?
Apoptotic signals Connexin hemichannel opening ATP Glutamate Inflammation
What is ATP release from cells a signal of death/disruption/ a problem?
ATP released from cells is highly toxic and can potentiate inflammation and neighboring cell death
Glutamate- reflects cells undergoing anoxic depolarisaiton
Describe apoptosis in relation to ischemia;
Balance between pro and anti-apoptotic signals
- Increased apoptotic molecules through latent and secondary phase contributing to bulk cell death
- Cyt C released post mitochondrial injury, signals apoptosis and it is elevated for 24hrs
- Caspase 8 (present early) - progressive increase
- Caspase 3 (final executive signal) only triggered (activated) during secondary phase there pro-apoptotic signals triggered early and build up over time to this
= cells probably dont die via apoptosis early on and this means theres a window (latent phase) for intervention
Describe apoptotic morphology;
- Blebbing of the cell membrane
- Cell shrinkage
- Nuclear fragmentation
- Chromatin condensation
- Chromosomal DNA fragmentation
- mRNA decay
- Mitochondrial function preserved during apoptosis
- Energy Dependant process (therefore mitochondria are preserved)
- Programmed cell death
Is it true that primary phase = necrosis, secondary phase = apoptosis?
Its a continuum!!!!!
- Clear apoptosis
- Continuum
- Clear necrosis
A lot of it is mixed morphology.
What is the apoptosis necrosis continuum?
6 things for continuum;
- Partial activation of the caspase cascade
- Intermediate forms of cell degradation
- Incomplete packaging of nuclear and cytoplasmic contents
- Impaired trafficking of mitochondria to axons
- Loss of mitochondrial structural integrity and function
- Loss of cytochrome C activity
Describe the cellular appearance of a necrotic cell;
- Translucent cytoplasm
- disbursed and minimal chromatin condensation
- intact nuclear membrane
Describe the cellular appearance of a continuum cell;
- Incomplete packaging of nuclear and cytoplasmic contents
- Partially condense cytoplasm within intact cell membrane
- Variably swollen and disrupted mitochondria and vacuolated cytoplasm
- Irregularly condensed chromatin
- At least partial segregation of nuclear and cytoplasmic contents
Describe the cellular appearance of a apoptosis cell;
- Intact cell membrane
- Dense cytoplasm
- Highly organised chromatin condensation
- Complete loss of nuclear membrane
What controls the mode of cell death?
Severity of the insult
The maturity of the cell type injured
The glutamate receptor subtypes stimulated
The degree of cellular calcium overload
Mitochondrial dysfunction
Depletion of cellular energy supply
What leads to mitochondrial dysfunction?
Injured by reactive oxygen species
High intracellular Ca2+
Loss of membrane potential
Leads to loss of species through Mitochondrial permeability transition pore (MPTP)
- outer membrane rupture
- H loss (intermembrane space)
- Cyt c exits and activated procaspase 9
What happens in the MPTP in secondary energy failure?
Protons and other molecules move across the outer mitochondrial membrane
Disrupt the electrochemical gradient
Disrupts the electron transport chain
Stops ATP production
Mitochondrial become permeable to molecules <1.5kDaCa2+ release damage neighbouring mitochondria
Mitochondrial swelling
Rupture of the outer mitochondrial membrane
Release of cytochrome C
Triggers apoptosis
What happens during brian injury that allows inflammation?
Breakdown of the blood-brain barrier
Macrophage infiltration
Microglial activation
Cytokine release
During injuryu what happens when microglia become activated?
Normally microglia are inhibited by neurons as surveillance cells.
But during they can become activated and form M1, M1/M2 and M2 phenotypes
Describe the different microglia phenotypes;
M1 = Profinflam M2 = Anti-inflam
Phenotypic continuum of course
M1-M2 classificationM1 (classical): pro-inf lammatory microglia
M2a (alternative)
M2b (type II)
M2C (deactivated)
M2 microglia: involvement in Type II responses, immunoregulation and tissue remodelling
What is astrocyte function?
Maintain homeostasis
Glutamate uptake and recycling
H+ uptakeK+ uptake and redistribution
Tripartite synapse
Astrocytic syncytium coupled by gap junctions
Important role in healthy tissue and after injury
What is the astrocyte response to injury?
Activation
Proliferation
Hypertrophy
May form glial scars after stroke and spinal cord injury
Exacerbate inflammatory response
What are connexin hemichannels?
1/2 gap junctions
- These can open under normal conditions allowing molecules to move freely from the cyotplasm i.e water, ca, glutamate
What happens to the connexin hemichannels in stroke?
- Open up esp. in stroke ischemic inflammation
- Cytokines open these
- Abnormal cell function
What did they find in studies that blocked the hemi channels?
Blocking hemichannelsfrom 90 min after ischemia improved recovery of brain activity, reduced seizures, reduced cell death
•Continuing blockade for 25 hour was better than one hour
What did the follow up study of the hemichannels show?
In a follow-up study, I showed Connexin hemichannelscontribute to spread of injury in first three hours after ischemia
Release glutamate
Release ATP
Cell swelling
Seizure activity
What is the role of ATP in signalling?
Acts as a signalling molecule
P2X ionotropicand P2Y metabatropicreceptors
Stimulate microglial activation and inflammation
Summerise the latent phase;
Restoration of oxidative metabolism
Resolution of cell swelling
Resolution of extracellular glutamate concentration
Activation of cell death pathways (Bax, Caspase-9)
Connexin hemichannel opening
ATP releaseInf lammation -cytokines
Blood brain barrier breakdown
Summerise the secondary phase;
Seizures
Cell swelling
Cell death via apoptosis, necrosis and the apoptotic necrotic continuum
Secondary energy failure
Mitochondrial permeability transition pore
Mitochondrial collapse Inflammation -microglia and astrocytes
Does injury end after the secondary phase?
Long-term inf lammation
Ongoing cell death
Loss of trophic support
Impaired connectivity
What to study?
What is focal/global brain ischemia?
What is the pattern of injury evolution after focal brain ischemia?
How does the injury spread in the penumbra?
Describe the phases of injury in global brain ischemia
Mechanisms of injury during the primary, latent and secondary phase/ what happens
learn esp about hemi channels as her area
