BIOL 2052 - Glia

defining glia

• macroglia - a group of cells - anything that is a glia but is not a microglia

• glial cells do not carry synaptic function but essential for function of NS

• around 90% of the cells in the brain are glia

• without glia the neurons wont work

• of the 90% divided into CNS and PNS

  • PNS - schwann cells

  • CNS - macroglia (85-90%) and microglia (10-15%)

    • macroglia can then further be subdivided into ependymal cells (5%), oligodenrocytes (5%) , astrocytes (80%)

function of glia

MAIN FUNCTIONS

1. provide physical support

2. supply nutrients and oxygen to neurons

3. to insulate one neuron from another and faciliate synaptic communication

4. destroy/remove cell debris

OTHER SPECIFIC FUNCTIONS

• critical development roles

  • glial migration

  • growth/direction of axon/dendrites

• modulate synpatic transmission

• fundemental role in disease/degeneration —> as you get more complex brain function you require more glia and increased size of glia with more glia per neurons

discovery of glia

• Rudolf Virchow - 1956 first discovered glia (glue) didnt think they were functionally relevant

• Deiters then begins to describe them - lack of axons

• Deiters was the first to suggest ectodermic origin - same origin as neurons

• andriezeen then began to classify types of glia

  • ectodermal/fibrous glia in white matter

  • mesoblastic protoplasmic glia in grey matter

• Cajal then proved that these 2 glia both were ectodermically derived but suggested 3 types of glia —> later proposed that there were 4 types of glia:

  • protoplasmic

  • neroglia

  • mesoblastic glia

  • interfasicular glia (oligodendrocytes)

many functions recognised

• plasticity

• electrical insulation

• pathological role

evidence for glial origin

• radial glia have their body in the ventricular surface and body in the ependymal cells

• act as a scaffold for migration of glia

• but these cells persist in adults, they can give rise to astrocytes, microglia, oligodendrocytes

development of the brain

• synaptic pruning and myelination can take up to 20 years (critical for learning)

• Before this neurogenesis also important

• glia become important about halfway through embryonic development when neurons start to connect to each other

  • microglia are the exception - come from an extra neuronal origin and remain in the brain as the BBB closes

O2A progenitor

• stem cells develop into O2A progenitor cells - cells which have already comitted to either development of astrocytes or oligodendrocytes

• O2A progenitor replicates into astrocytes to start but once we have made the astrocytes it becomes an NG2 cell (expressing NG2) which wil form oligodendrocytes

• NG2 cells dormant but can be reactivated if theres a need for astrocytes (E.g: demyelinary disorder

how do NH2 cells make coligodendrocytes

1. immature oligodenrocyte

   • no expressed NG2, expression of O4

2. mature oligodendrocytes

   • express proteins that occur in myelin (OPC and MBP)

   • transcription factors define the transition (oligodendrocyte precursors express high levels fo notch 1 and prox 1)

   • lost notch 1 and increased prox 1 causes progression to next cell stage

schwann cells

• highly related to oligodendrocytes

• similar stepwise development

• neural precursor —> schwann cell precursor —> immature schwann cell which forms either

  • promyelinating schwann cell —> myelinating schwann cell

  • non myelinating schwann cells (provide support)

astrocyte

• neural stem cell —> astrocyte precursor —> astrocyte

• stages of astrocyte lineage poorly defined and lack stage specific markers

• do express certain proteins at certain development points (E.g: GFAP is an intermediate filament required for the mature structure)

• sox 9 is essential for neural stem cell to progress to astrocyte progenitor

• activation of Jak/stat essential for astrocyte precursor to develop to astrocytes

• potential for multiple types of astrocyte

generation of new glia

• radioglia common precursor for glia/neural cells

• cajal had the hypothesis that once the brain was deveoped no new cells formed - turns out to be wrong, new neurons constantly produced in:

  • dentate gyrus

  • ventricular zone

• new neuronal cells important for moving to the olfactory bulb and formation of memories in the hippocampus

• default state for radioglia to generate astrocytes when they dont generate more neuronal cells

• if we can embedd TFs can we repolarise specialised cells to earlier progenitor cells —> this means that the generation of these cells likely to be from the same cell

microglia

• are actually macrophages —> not macrophage like

• one of the immune cells in the brain —> lots of immune cells in the membranes surrounding the brain and within the parenchyma but not much known

• immune census of the brain: many cell types clustered within regions of the brain but many sub populations within these populations

• macrophage subpopulation:

  • choroid plexus macrophage

  • perivascular macrophage

  • meningeal macrophage

• even just residing in the brain gives some kind of similarity to any kind of macrophages

• we can predict what functions these macrophages might have

basic characteristics of microglia

• highly ramified → tile the brain parenchyma in a mosaic like distribution (no overlap)

• projections move, cell body doesnt

• difference between white and grey matter microglia:

  • white: highly polarised

  • grey: more ramified

• different densities between regions

• equipped with immune sensors to return the brain to homeostatic state

systemic sensing microglia

• can phagocytose synaptic elements, dead cells etc

• postulated that microglia can prune synapses which need to be removed and can modulate synaptic activity

• good at sensing inflammation

• communicate with the rest of the immune system

discovery of microglia

• discovered with nissal staining

• first named based off their less round nucleus (stabenzellen)

• observed with a shape change in animals with rabies

• showed that perivascular microglia derived from bone marrow

development of microglia

• erythromyeloid progenitors derived from yolk sac (not part of the embryo but has stem cells) give rise to all macrophage population

• colonise the brain (whilst the BBB is open)

• after closure we cant get cell migration to the brain

• foetal liver main heamopoetic organ giving rise to cells

• they eminate from the liver, colonise the other organs and develop macrophage populations

• in the brain microglia are sustained by cell renewal, whilst in other organs macrophages can eb replaced by bone marrow stem cells

• in cases of damage to BBB some macrophages might enter the brain and differentiate into microglia like cells

microglia lineage

1. EMP (erythromyeloid progenitor cells) colonise the brain

2. differentiate into premacrophage (common to other macrophage cell types)

3. premacrophage (A1CD45+CX3CR1 is low, so is F4/80) —> premicroglial cell (A2CD45+, CX3CR1 high, F4/80 high)

   • this commits the cells to microglia

   • conversion of A1-A2 is controlled by the upregulation of TF PU.1 (genetic factors which do not depend on the environment)

   • conversion of A2 —> microglia driven by the environment (surrounded by IL34, TGFbeta which is not found anywhere else)

   • only when cells start to develop into A2 do they express proteins characteristic of macrophages

transcription and functional diversity

• microglia not equal —> diff environment drives diff phenotype (degree of heterogeneity)

• genes and TFs which are upregulated in cerebellum that are not upregulated in the striatum and the cortex

• degree of similarity in the hippocampus and cerebellar cortex but still distinct

• cells provide info about energy metabolism, immune response etc of tissue

• distance of dissimilarity with other macrophage is huge but equally all macrophages in the brain are not similar

• populations of microglia associated with injury, development, aging, postnatal development etc.

• there is heterogeneity of macrophages linked to environmental/developmental factors

• microglial density is not affected by aging —> must either be very long lived or proliferating - turns out to be cycling proven by:

  • can dose cells with Brdu and doesnt affect cell cycling

  • allows us to know which cells are replicating at a certain time

  • if we check at different periods we can compare rate of apoptosis compared to rate of growth

  • microglia proliferate at a surprisingly high rate - BIM dependent apoptosis balances microglial proliferation and apoptosis allowing constant turnover (6.6 times in mouse lifetime)

role of astrocytes

• neurogenesis in the adult brain

• neuronal guidance in developmental rols of radioglia

• regulation of synaptogenesis and synaptic maturation

• structural —> responsible for defining and connecting domains that include neurons, synaptic vessels, blood vessels

• communicate through gap juctions

• make up the BBB

tripartate synapse

• majority of synapses have a third astrocyte component (60%)

• pre and post synaptic neuron sealed by astrocyte neurotransmitter in contact with astrocyte

• 80% of large perforated synapses are enwrapped by astrocytes

• in the cerebellum, innervation of the purkinje cells with bergmann cells (astrocyes of the cerebellum) each enwrapping 2000-6000 synaptic contacts

• possibility to integrate and modulate activity as recieve inputs from hundreds of cells

evidence for tripartate synapses

• astrocytes are excitable, can produce a transient change in IC Ca2+ conc through release from ER stores

• lots of astrocytes respond shortly after the stimulation of an axon

• astrocytes communicate bidirectionally:

  • able to detect neurotransmitters and other signal released from the neurons at the synapse and can release their own neurotransmitters or gliotransmitters that are capable of exciting neurons

• all astrocytes communicate via gap junctions - can illicit a response in astrocyte not in contact with the original syanpase - must be a non neuronal way of communicating

• works via glutamate signalling

glutamate signalling

• astricytes clear glutamine from the synaptic cleft

• glutamine is a target for kainate receptors, metabolic receptors, glutamate receptors and NMDA receptors

• the ability of astrocytes taking up the Glu prevents ot from interacting with neurons and modulates activity

• also allows astrocytes to synchronise neurons in the next cycle of activation (synchranous depolarisation)

other giotransmitters

• ATP targets P2X receptors, P2Y receptors (present on astrocyte and neuronal membrane

• ATP produced in astrocyte which can signal to neighbouring astrocyte and either cause it to produce more ATP, Ca2+ etc or it can signal to neuron and tell it to release more or less gluatamate

• can also modulate AMPA receptors on post synaptic membrane

• ATP release mechanism not well understood - known to be connected to Ca2+ waves, likely related to SNARE proteins

distal regulation: example

• astrocytes of teh hippocampal startum oriens form tripartate synapses with axonal projections from the alveolus

• alveolus projections can be either glutameric or cholinergic synapses with the stratum oriens but teh astrocytes of this region respond with changes in Ca2+ conc (only to cholinergic activation of alveolus projections)

• not just a passive response, actively responding

• astrocytes produce more glutamate at cholinergic synapse due to cholinergic signalling —> keep neurons signalling regardless of neurons activity

integration and modulation of activity

• the hippocampus striatum oriens astrocytes which respond to synaptic activity from the glutameric neurons originating in the chaffer collateral and cholinergic neurons originating in the alvus

• they produce changes in Ca2+ conc that are non linear with synaptic input

• additionally, the same stimuli are capable of producing either a potentiates Ca2+ response at low frequencies of stimulation or a depressed Ca2+ conc at high frequencies of stimulation

the blood brain barrier

• barrier between the intercerebral blood vessels and the brain parenchyma

• endothelial cells closed by tight junctions

• every peice of capillary wrapped by astrocyte end foot

• 2 components: bood and parenchymal compartment

• present everywhere throughout the brain except circumventricular organs, neurophyphoris, pineal gland, subfornical organ and neuroendocrine signalling (need to be able to check blood content fast)

• every solute must pass through endotheial cells: selectively permeable to essential nutreints to enter and metabolites to have

• specific trasporters at the endothelial cells include:

  • amino acid transporter

  • energy dependent ABC transporters which excrete xenobiotics (impermeable to drugs, antibiotics etc)

  • GLUT1 glucose transporters

  • ion exchangers

• transporters at the astrocyte endfeet include:

  • glucose transporters, uptake and distribute to neurons

  • K+ chnnels

  • water channels

astrocyte and spinal chord injury

• damage to the BBB causes damage to the environment of the brain which the astrictyes react to:

• spinal chord injury can causse separation of pre and post synaptic neuron as well as blood components in spinal chord which activates glial cells

• astrocytes from glial scar to form a barrier to protect the environment - prevent flow of damaging molecules into the tissue (proliferate and join end feet)

• protection from secondary injury

• glial scar is hard to remove and remains for a long time - doesnt allow neurons to grow through it

• causes complete separation of ascending and descending signals

• once all of the dead waste/debris cleared up it forms a cystic cavity surrounded by astroctyes

• physical and molecular barriers

• astroctyes also secrete growth promoting molecules in attemot to regenerate

• growth cone retreats if it comes in contact with astrocytes

• inability of cells to cross the barrier not caused by astrocyte but intrinsic failiure of neurons to grow through glial scar

  • PI3K key pathway for growth cone, hen PTEN activated it inhibits this

  • when PTEN inhibited the axon is able to grow through glial scar

myelination

1. oligodendrocytes

   • all myelination in CNS is done by oligodendrocytes

   • multiple axons myelinated by OG cell (avg 10/cell)

2. schwann cell

   • myelinate in PNS

   • schwann cells myelinate in a 1:1 ratio

3. myelination is dependent on axon diameter

   • lamellae refers to the number of layers of glial cell membrane wrappiing around the axon

   • radial growth of axons and myelin sheath are interdependent

   • ratio of axons to myelin lamellae is called the g ratio and it is constant in the CNS and the PNS (1:10)

4. interdependence of glia axons

   • the loss of the axon results in degeneration of oligodendrocyte and differentiation in schwann cells

   • conversely, aons degenerate in absence of support cells (OG and schwann cells)

schwann cells

NON MYELINATING SCHWANN CELLS

• non myelinating schwann cells usually surround bundles providing trophic support

• express L1 and NCAM which are not found on myelinating schwann cells

PERISYNAPTIC SCHWANN CELLS

• 3rd type of schwann cells are perisynaptic schwann cells (found in the NMJ)

• ensheath the synaptic terminal

• respond to synaptic activity with Ca2+ waves

• able to modulate synaptic activity by regulating EC ion levels and inducing post synaptic Ach receptor aggregation

olfactory bulb ensheathing cells

• myelinating cells that are not schwann cells or OG cells

• found in olfactory bulbs

• interesting type of glia - interface the PNS and CNS at the cribiform plate

• myelinating cells moving towards olfactory plate

• have roles similar to microglia —> phagocytose etc but also secrete neurotrophic factors which allow axons to cross glial scar

• secrete glial markers similar to astrocytes (GFAP, s100, p75)) but also radial glial markers such as nestin and vimentin

• due to their mixed role and allows cells to cross glial scar they have been proposed as a therapy to spinal injury

myelin

• fatty insulated layer that facilitates saltatory conduction

• myelin sheath wrapped arund axons to form concentric layers of lamella

• every membrane has a different composition

• myelin leaves gaps - nodes of Ranvier which are specialised axonal areas where action potentials can propogate

• myelin sheath between the nodes of ranvier = inter nodes

• specific molecules which allow for interaction of membrane and OGs

• close to the node = paranode (closest to NOR) and juxtaparanode (closest to the internode)

• moleular interactions at the paranode and juxtaparanode determine the clustering of K+ and Na+ channels that are key for saltatory conduction

  • E.g: contactin 2 and Caspr 2 are only expressed in juxtaparanode annd allow K+ channels to not invade node of ranvier

  • at the paranode: contactin 1 and Caspr which interacts with NF155 on the membrane of myelin cells

  • molecular patterning determines function of the compartments

myelin: lipids

• extracellular face and cytoplasmic face alternating

• composed 70% of lipids, majority are cholesterol, some glycolipids, some phospholipids in a ratio (4:3:2)

• its rich in glycosphingolipids, mainly GalC which is used as a marker

• composed of gangliosides (complex lipids present in grey matter of the brain) which differs in the CNS from the PNS

  • in the PNS LM1 and GM3, in the CNS GM4

myelin: protein

• the other 30% of myelin is composed of protein, mostly shared between CNS and PNS

  • in the CNS main ones are MBP, PLP which fuse EC and cytoplasmic faces (also present in PNS myelin but the functions unclear)

  • in PNS main protein is P0, mediating fusion of lamellae, but PMP22 and Cx32 are also important for keeping the membrane connected

  • MAG is present in both CNS and PNS —> important for connection of pile layers with axons, bind to specific gangliosides on the surface of the axon

myelination - process

PHASE 1: Axonal contact

• only if axon grows thicker than 0.7um in the PNS or 0.2um in the CNS

• loss of NCAM from axonal surface triggers myelination, similarly L1 expressed at premyelination, tagging the axons to be myelinated

• partner molecules in myelinating cells completely resolved

• contact with axons triggers differentiation of OPCs into oligodendrocytes, starting to express myelin products (GalC, CNP, MBP etc)

PHASE 2: axon ensheathment and establishment of internode segment

• extension of initiator process that spirals along the axon (using MAG and PLP to stick)

• myelination of multiple axons follwed by remodelling phase when non ensheathment processes are lost

• initial clustering of Na+ channels around nodes of Ranvier (happens in multiple axons at one time)

PHASE 3/4: remodelling and maturation

• subsequentl wraps of myelin are produced which fuse to eachother dependent on PLP and MBP

• maturation of nodes of Ranvier (synchronised expression of molecular pairs of axons and myelin)

• some of the inital connections may be lost (remodelling)

• when the final no of axons myelinated all other processes of OG lost

multiple sclerosis

• immune system develops and autoimmune attack of the CNS forming plaques of lesions

• generation of autoantibodies against myelin comonents

• effects mostly the CNS and spinal chord

• can allow Abs into the brain

• most common type is relapse and remittance MS where the phases of demyelination are followed by remittance

• commonly involves white matter

• damages oligodendrocytes, causes demyelination

• there are attempts to remyelinate but they are not complete

PATHOPHYSIOLOGY

• key cause is BBB breakdown; causes entry of T cells

• antibodies enter and recognise myelin

• followed by chronic inflammation —. recruit immune cells of CNS (astrocytes, microglia)

• contributes to the cycle of damage leading to inflammation

progression of MS

PHASE 1: early disease

• antibodies are in circulation but brain is protected by BBB

• can lead to immune cell accumulation in perivascular space (cytokine production which leads to leakiness of BBB)

• antibodies can now attack white matter of the CNS

• this brings in T cells which are able to permeate BBB and lead to destruction of myelin and damage to axon

• once the myelin is degraded, reslts in degradation products in spinal fluid

• when it reaches new ganglia it causes another cycle of reactivation

PHASE 2: chronic phase

• immune cells and cytokines engage miroglia in cycle

  • E.g: chemokines, CCL2, GMCSF which activate reactive O species

• activation of glia also leads to activation of oligodendrocyte precursor cells

• damage to the myelin results in hanges such as Na+ channel rearrangement which affects neuron fuction

• in long term the axon damage can result in astrocytes forming a glial scar

• remyelination is often partially successful - able to tell via MRI (but will also never return to the original state of myelination)

microglial surveillance

• mosaic like distribution with “territory” to protect

• active protection

• protects from

  • extravasion of unwanted molecules

  • cells that need to be removed

  • damage to the BBB

• microglia extend their processes to limit and react to damage

• much more rapidly than other cells

microglial diversity

• show morphological diversity - observed via stain

• structural axonal cues can dictate this

• regional density differences:

  • hippocampus CA1 region is more dense, thalamus density is lower

  • grey matter/white matter density is different

• have different turnover rates

  • slow in the cortex, fast in dentate gyrus

  • fast turnover means slower replacement potential

• able to drive transcriptional differences in microglia samples by either cutting them post morten and sequencing or just sequencing

  • some core markers - these always present in microglia

  • also environmental dependent transcriptional molecules which can eb turned on/off depending on the environment

  • full maturation required environmental factors

  • gene expression profile in diseased microglia not neccessarily driven by microglia but by interaction with alzheimers brain

basic functions

1. ability of microglia to remove synaptic elements

2. ability to phagocytose

are microglia responsible for removal of synapses?

• dorsolateral geniculate nucleus has territories for each eye (majority contralateral, some ipsilateral)

• add stain to each side of teh eyes retinal ganglion cells and you can tell where the retinal ganglion cells project to in the dorsolateral geniculate nucleus

• at some point we must get rid of some synapses to rewire circuitry so we retain some contrilateral projections

• we find some of the tracer in microglia —> phagocytose some of the synapse

• area which ends up being contralateral area the stain in microglia from ipsilateral fibres

• remove synapses which must be removed

• not known whether microglia are responsible - suggested that complement receptor 3 drives phagocytosis but microglia responsible for the degradation

clearance of apoptytic cells

• form phagocytic pouches, found to contain nuclei

• metabolically demanding

• occurs in dentate gyrs - contains cells with neurogenic potential

• huge no of radioglia produce amplifying neuroprogenitor cells (excess of cells)

  • these need to be removed by microglia

  • huge no of cells cleared

immune roles of microglia

• equipped with molcules that allow them to react to environment

• M1/M2 activation, M1=inflammation, M2= antiinflammatory response

• no longer tak about M1/m2 response

• population of microglia in steady state which can develop into a range of phenotypes which can change between eachother

• gradient responses dependent on metabolic states, proteins etc (likely dictated by genetic background)

alzheimers

• innate immunity is a key driver for alheimers

• cognition deteriorates when infected

• many genes in associated with alheimers are associated with immunity

• homeostatic microglia progress to dff phenotype in a 2 step process:

  • STEP 1: convert into disease associated miroglia

  • STEP 2: uses trem 2 to convert to diseased population of microglia which look nothing like normal microglia —> limits the growth of amyloid pathology

• in disease, huge increase in microglia (not specific to alzheimers)

• accumulation of amyloid beta, previously thought that this drove neurodegeneration

• however, we can intervene and prevent degradation by altering microgia

• microglia transducers of disease —> at some point they become involved ad amyloid beta drives the diseased phenotype