Euroglia 2007
VIII European Conference on Glial Cells
in Health and Disease
I attended the optional Lecture Course on Glial Cell Biology at the University College of London prior to the start of the meeting. This was an all day lecture series reviewing astrocytes, oligodendrocytes, and microglia. It was an excellent course. My interest with astrocytes started as a technician in Scott Baraban's Lab at UCSF, then progressed with my courses in graduate school and then with my thesis project with Karen Wilcox, but never have I had such a comprehensive review on the all the cells of the glia family!
For due reasons, most courses in the neuroscience curriculums deal with neurons (development, function, disease, etc) and glial cells, though not disregarded, are simply not given their fair coverage. From the perspective of designing a core course in the neurosciences, this of course makes sense. Students need to get a broad overview of the field, and most of the research, historically, has focused on neurons. No doubt they are key players in the nervous system, but our understanding is incomplete without a full appreciation for the roles of glia.
Below, I have summarized the lecture courses and talks I attended during the 5 day conference. I think the reader will see both how organized and comprehensive this conference was, and how much of an impact this meeting had on one graduate student.
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Professor Rhona Mirsky, University College of London (UCL), reviewed schwann cells, the glial cells of the peripheral nervous system (PNS), whose functions include myelination, regulation of the microenvironment, nerve support during development, maintenance, movement, regeneration, and immune interactions with schwann cells. In particular, she focused on the critical role of neuregulin-1 as a signal for both schwann cell survival and myelin maintenance.
Professor Nigel Pringle (UCL) gave a talk titled (humorously) "Astrocytes: in 40 minutes." His talk focused on the problems with glial fibrillary acidic protein (GFAP) as a marker for astrocytes. The presence of this intermediate filament is universally accepted as a marker for astrocytes, but it has become apparent in recent years that astrocytes are not a homogeneous population. We need better markers that can identify the functional heterogeneity of astrocytes. His team is utilizing the Allen Brain Atlas to comb through gene expression profiles of the rodent brain to identify new markers. I was pleased to hear him talk about this subject, for it is something that I have been wondering about for a while now, and that he is using the Allen Brain Atlas. I am hearing references to this wonderful resource more and more.
Professor Richard Reynolds, Imperial College London (ICL), reviewed oligodendrocytes, the myelinating cells of the central nervous system (CNS). In particular he talked about the origins of the oligodendrocyte precursor cells (OPC), the migration of OPC through the spinal cord, and the signaling mechanisms that occur between OPC and axons to initiate myelin formation. This fascinating topic was one of the major themes at the conference. I was amazed at the numbers of posters and presentations that dealt with spinal cord repair, and was also amazed at where the field is right now. The term "stem cells" gets bandied around quite a bit, both politically and scientifically, so it was refreshing to hear (throughout the meeting) about this one particular type of cell that holds promise as one an endogenous stem cell of the brain.
Professor Deanna Taylor (ICL) gave a fascinating review on microglia, the resident mononuclear phagocytes of the brain. These wonderful little guys (15% of the total number of brain cells) constantly monitor and sense the extracellular environment by mobilizing their distal processes. Their filopodia are constantly shortening and elongating until they sense an activation signal to mobilize, then these cells swarm en masse and engulf the invader. I found a link to some of the movies (by Axel Nimmerjahn, who later gave a fascinating symposia at this conference) she showed which can be found here. In particular, check out the high resolution movie S7. In the beginning, you can see the microglia in their surveillance mode, but once the lesion happens, they become motile and move to the lesion site. An interesting point is that although microglia are critical for the brain's immune response and can clear the brain of unwanted invaders and pathogens, excessive microglial activation leads to a pathological environment deleterious to neuronal survival.
Professor Siddhartan Chandran of Cambridge University then gave a rousing review on stem cells, oligodendrocytes, and repair. His talk focused on the major question in stem cell research, which is directed differentiation. How do you get from one cell, the stem cell, to one specific type of cell out of the 216 possibilities (according to Dr. Chandran, there are 216 different cell types in the human body)? As a model, he focused on the OPC as an endogenous stem cell capable of regenerating into astrocytes, neurons, and oligodendrocytes. Like I mentioned, there is a lot of excitement for this little cell.
Professor David Attwell (UCL) then gave a talk on neurotransmitter signaling to astrocytes and oligodendrocytes, and the metabolic coupling of astrocytes and neurons. He gave a review on the somewhat controversial lactate shuttle hypothesis. The idea is that neurons derive their energy from importing lactate that is provided by astrocytes. The model predicts that as neuronal activity increases, and hence more glutamate is released into the synapse, astrocytic glutamate transporters clear the extracellular glutamate, but at the cost of energy depletion in the astrocyte. To counteract the energy demand, astrocytes then dilate the blood vessels to take up glucose. I love this particular topic. It is at the same time, both extremely complicated, in terms of the science, but incredibly beautiful to imagine the neurons and astrocytes in this type of a symbiotic and harmonious partnership.
Finally, Professor Nicola Woodroofe of Sheffield Hallam University gave a review on the properties of cytokines and chemokines in CNS physiology and pathology. The cytokines are small secreted proteins than are responsible for (along with many other functions) the immune response. This was a nice review because it is a topic that I am quite unfamiliar with and realized that it deserves more of my attention.
With the lecture course over, everyone dispersed and headed to Imperial College London for the start of Euroglia. Because the train line was closed, I decided to walk the hour and a half to ICL, and as I was looking over a map to decide directions, I met Marcelo and Anna-Paula, two PhD students from Brazil. They were to become my friends during the conference.
A snapshot of the main lecture auditorium. I counted about 650 people in attendance, and was told that 750 had registered! In the space below, I am going to summarize the talks and posters I attended, but have left out details on purpose.
Tuesday Evening
Opening Plenary Lecture 1
Marie Philbin, Hunter College, New York, USA
Regeneration in the CNS
Dr. Philbin talked about her strategy in developing CNS regenerating drugs for the treatment of spinal cord injury. Because I work along side the anti-convulsant drug discovery program at the University of Utah and we share lab space and lab meetings, drug discovery has been a topic that I have been exposed to a little bit, so this talk was particularly interesting from this perspective. One of the major obstacles in axon regeneration is the inhibitory signal from myelin. By manipulating cAMP, Dr. Philbin is trying to "trick" the axon to become unresponsive to these inhibitory signals in order to promote axon growth, and thus axon regeneration.
Opening Plenary Lecture 2
Patrick Charnay, INSERM, Paris, France
Glial Cell Functions in the PNS
Dr. Charnay discussed his research of the boundary cap cells that exist at the interface of the CNS and PNS. By using conditional knock in animals, he is investigating myelin formation in the PNS.
Wednesday Morning Plenary Lecture
Pierre Magistretti, University of Lausanne, Switzerland
Neuron-glia Metabolic Coupling
Dr. Magistretti introduced to the audience his hypothesis of neuron-glia metabolic coupling. He also discussed at length the metabolic wave that accompanies neuronal activation. As a synaptic event occurs and astrocytes take up glutamate, the response coincides with an increase in intracellular sodium from the coupled transport of glutamate and sodium ions. This signal is then amplified and propagated throughout the astrocytic syncytium as a Na, Ca, glucose, and ATP wave.
He also discussed the different roles of astrocytic and neuronal glucose transport. Though this is an oversimplification, in general, it seems that neurons are utilizing their own glucose transporters (Glut-3) for their basal activity, but as energy demands increase, glucose utilization switches to astrocytes, at which point, the lactate shuttle becomes essential for neuronal energy supply.
In addition, Dr. Magistretti proposed novel questions regarding energy demand. For example, is synaptic plasticity associated with metabolic plasticity? How is metabolic coupling involved in pathogenesis? What is the role of glycogen in the sleep-wake cycle? For each of these questions, there were a series of experiments that clarified his model and definitely piqued my curiosity with regards to these topics.
Morning Symposia
Functional Role of Bergmann Glia-Purkinje Cell Signaling
Axel Nimmerjahn, Stanford University
2-Photon Ca Imaging of Circuit Activity in the Cerebellar Cortex of Live Mice
Ok, this was an amazing talk. As the title suggests, Dr. Nimmerjahn's experimental set up allows for 2-photon calcium imaging of the cerebellum in live, non-anesthetized, behaving mice. One of the major problems in recording from live animals is the possible confounding effects of an anesthetic. He amazed the audience with movies of 2 types of calcium signals in the cerebellum that is correlated with the movement of the animal. One form was a short point like burst, and the other was more in line with what I would think a spreading calcium wave to be like.
Interestingly, there is a very recent paper in PNAS that shows two forms of vesicular release in astrocytes. One, being a very small, defined, kiss-and-run type of event, and the other being a more slow and spreading release event, probably representing a full fusion and exocytosis event. Of course this study was done in cell culture and they had the benefit of imaging in TIRF, so they were able to conclude the size of the objects involved down to the tens of nanometers, but it was cool to see something that kind of resembles that data in the live animal. Might a kiss and run event underlie the small punctate calcium increase while full fusion events represent the spread of a calcium wave? Or, are the calcium signals only representative of the underlying neuronal activity? What signals dictate the two forms of calcium signaling and what is the functional consequence of these different calcium waves? How are these waves related to the metabolic coupling of glia and neurons? Dr. Nimmerjahn is a post-doc at Stanford, so I eagerly talked to him about the imaging group there and was encouraged to contact them for further information.
Christine Rose, University of Dusseldorf, Germany
Imaging Glial Glutamate Uptake in the Cerebellum
Dr. Rose gave a talk on her use of a Na sensitive fluorescent dye to image glutamate transport by Bergmann glial cells. Yet again, I watched with amazement as she demonstrated how she patch clamped a cell while loading it with the fluorescent dye, then eliciting both a current response and a change in fluorescence as the cell detected extracellular glutamate. In this way, she is able to measure the current charge carried by the coupled transport of sodium ions with glutamate transport, and image the spatial spread of glutamate transporter activity in real time. I also interacted with her colleague who had a poster on the use of SR101 (sulforhodamine 101), a commercially available fluorescent dye, to specifically label astrocytes in hippocampal slice to aid in patch clamp recordings. His results suggest that I will be able to use this dye in the lab during my own experiments, and I was very excited to talk with him about his results.
Thomas Bellamy, The Babraham Institute, Cambridge, UK
Plasticity of Neuron to Bergmann Glial Cell Transmission
Dr. Bellamy presented data regarding the plasticity of the synapses that innervate Bergmann glia. Both parallel fiber and climbing fiber synapses are opposed by calcium permeable AMPA receptors on the Bergmann glial cell. He demonstrated that certain inputs and stimuli result in paired pulse facilitation, whereas other conditions resulted in paired pulse depression. Importantly, he showed that these effects are specific to the neuron-glia synapse, as neuron to neuron synapses were unaffected by his experimental setup.
Maria Rubio, University of Connecticut, Storrs, USA
Bidirectional Communication Between Purkinje Cell Synapses and Bergmann Glial Cell Processes
Dr. Rubio presented beautiful immunofluorescent images of the spatial relationships of specific AMPA receptor subtypes and the developmental regulation of these receptors. She also showed fascinating EM images of the translocation of AMPA receptors as the shape of the glial cell processes changes in response to various extracellular and intracellular signals.
Frank Kirchhoff, Max Planck Institute, Gottingen, Germany
Inducible Deletion of AMPA-type Glutamate Receptors in Bergmann Glia
As the title suggests, Dr. Kirchhoff uses a tamoxifen sensitive cre-lox system to conditionally induce the deletion of specific AMPA receptor subtypes. By using these animals, he is able to see the effects of knocking out these genes at both the morphological and behavioral levels.
Actually, his talk on the use of animals with genetic modifications highlights an important point. It was apparent throughout the meeting that sophisticated control of gene expression is yielding very exciting data on cells in the adult animal and it was very cool to see the diversity of experiments that utilized these techniques. Because most of the gene knockouts utilize some form of marker protein, and the knock out is performed in a certain cell types at specific time points, scientists are able to identify at any point which cells were affected by the knock out then track the deletion through time. The other important point is that since these genes can be knocked out with tamoxifen, the knock out is not hampered by any developmental side effects. The result is a visual, functional, and behavioral analysis on the function of a given gene in a given time point in the animal’s lifetime.
Dr. Kirschoff is finding the functional roles of the various AMPA receptor subtypes that are expressed by the Bergmann glia, and their effects on cell morphology and animal behavior.
Afternoon Symposia
The Role of Astrocyte Dysfunction in Epilepsy
Of course, I was very interested to see the speakers at this symposium!
Detlev Boison, DOW Legacy Research, Portland, Oregon, USA
Adenosine, Astrogliosis, and Seizures: A New Perspective of Epileptogenesis
Dr. Boison presented data on adenosine as an endogenous regulator of excitability. With regards to the gliotransmitters involved in modulating synaptic transmission, I found many labs whose focus was adenosine. Adenosine works to keep synaptic activity low, whereas its enzymatic counter part, adenosine kinase (ADK), breaks adenosine down and promotes synaptic activity. Following an acute seizure or brain injury, ADK is rapidly down-regulated whereas adenosine levels rise in the synapse. Thus, it seems that the brain has its own mechanism for neuroprotection and neural quiescence. However, elevated levels of adenosine kinase, or down regulation in adenosine production can lead to pathological hyperexcitability. Therefore, by combining animal models for epilepsy, genetic manipulation, and stem cell transplants, he is trying to use the adenosine system for the therapeutic treatment of epilepsy.
Annamaria Vezzani, Milano, Italy
Astrocytes as a Chronic Source of Inflammation in Epilepsy
Dr. Vezzani presented her work regarding inflammatory signals as possible contributors of the recurrent seizure state. During acute seizures, pro-inflammatory cytokines such as Il-1B are rapidly synthesized by both astrocytes and microglia. Interestingly, while Il-1B levels return to normal levels in microglia, it remains elevated in reactive astrocytes throughout chronic epilepsy. It is also known that reducing Il-1B levels will down regulate seizure activity. Therefore, her work is trying to understand how the immune response, especially this altered state of cytokine production, is contributing to epilepsy.
Devin Binder, UC Irvine, USA
Potential Role of Glial Water Channels in Epilepsy
This was another subject where I have studied little about, but found very interesting. The aquaporin water channels are responsible for water transport in the CNS. Seminal work in acute brain slices demonstrated the effects of changing the extracellular space volume on neuronal excitability. Because the aquaporins control the amount of water inside the cell, they can change its volume and thus change the volume of extracellular space. Dr. Binder uses an aquaporin knock out mouse to investigate what role these water channels have in seizure generation and chronic epilepsy.
Christian Steinhauser, University of Bonn, Germany
Altered Functional Properties of Astrocytes in Human Epileptogenic Hippocampus: Relevance to Seizure Generation
Dr. Steinhauser’s work has been especially important for me. As a student learning the electrophysiology of astrocytes, several papers have been very instrumental in my education. Among these, his papers that utilize the GFAP-GFP transgenic mouse to characterize the GluT and GluR astrocytes of the hippocampus have been crucial in understanding the results of my experiments. For the symposia, Dr. Steinhauser described the astrocyte phenotypes found in resected tissue from patients with Ammon’s horn sclerosis, then talked about a new animal model that recapitulates this very intriguing data. I found the results to be very exciting, and am looking forward to his next papers on this subject.
Evening Plenary Lecture
Helmut Kettenmann
Max Delbrueck Center for Molecular Medicine, Berlin, Germany
Control of Microglial Function by Transmitter Receptors
Dr. Kettenmann discussed his research regarding the ways in which microglia sense signals in the extracellular environment. His group has developed an in situ model to study the response of resting and activated microglia to various transmitters. In his talk, he discussed the effects of GABA, dopamine, noradrenaline, and adenosine. In general, it appears that activation of some of these transmitter receptors encourages microglia to remain in a resting state, supporting the hypothesis that microglia are less prone to be activated in the presence of normal neuronal activity. However, changes in these signals are probably contributing to the switch from resting microglia to the hyper ramified or activated state.
Thursday
Much to my extreme consternation, I discovered on Wednesday that I had printed my poster in the wrong length x width dimensions. Any hope of the poster fitting on the allotted space was erased when I went back to my lodging and unrolled the poster from its tube. I decided that I needed to reprint the poster in the correct dimensions, and that meant working late through the night, then getting to a print shop in the morning. I had traveled all the way to London, and this being the first time I was presenting at this particular meeting, I just could not bare to have a poster that appeared sloppy. Luckily, I found a print shop (Kinko’s!) that was able to print my poster in about 3 hours, and I made it on time to the afternoon symposia.
Thursday Afternoon Symposia
Glial Migration in the Developing and Pathological Brain
The main questions in this symposium were:
1. What are migratory glia?
2. What mediates migration?
3. How are they related to pathology?
4. Does pathology recapitulate development?
This symposium was very interesting because as I have mentioned, many posters and talks dealt with this topic of migrating OPC cells, and as my knowledge in this field is very premature, this symposium allowed for me an in-depth look into this topic.
Peter Cannoll, Columbia University, USA
Glial Progenitor Migration and Proliferation in PDGF-driven Brain Tumors
The tumor cells in brain gliomas are very tough cells to get rid of. Even when a surgeon removes the main tumor, small pockets of cells will proliferate and migrate towards the bloodstream. Gliomas are composed of many different types of cells, but they are abundant with cells that resemble glial progenitors (hence the name). By using a retrovirus marker (which only labels dividing cells), Dr. Cannoll is able to track the migratory patterns of cells out of the proliferative zones, and by manipulating levels of PDGF, he can induce these cells to migrate faster and further, to proliferate at an enhanced rate, inhibit their differentiation, and most importantly, induce tumor formation 100% of the time. By comparing the migratory patterns of normal cells to those that form gliomas, he is uncovering the mechanisms involved in glial migration during development and pathology.
Jacqueline Trotter, University of Mainz, Germany
NG2 and Glial Cell Migration
NG2 is a chondroiton sulfate proteoglycan that is expressed on the surface of OPCs, but is a protein that is down regulated as the OPC differentiates into a mature, myelinating oligodendrocyte. It is known that NG2 is expressed in glioma cells and that the protein is involved in the cytoskeletal modifications required for migration. Dr. Trotter is uncovering the intracellular binding partners for NG2 and presented data that suggests it is in a tripartite complex with AMPA receptors. Knowledge of these binding proteins could allow for a better understanding of what proteins are involved in NG2 expression on the cell membrane, and what role NG2 may have with regards to AMPA receptor expression and cell mobility and differentiation.
Vittorio Gallo, Children’s National Medical Center, Washington D.C., USA
ET-1 Regulates OPC Migration and Differentiation
Endothelins are small, secreted peptides that have caught my attention recently for their ability to modulate gap junction coupling. However, Dr. Gallo presented new data on the role that endothelins might play in other systems. I think that the data he presented has never been published, so I am going to refrain from talking about the details, but he presented some very interesting data regarding how endothelin signaling can modulate OPC migration. His research is important for the question of how we can get our endogenous stem cell population to migrate and myelinate.
Evening Plenary Lecture
Dwight Bergles, Johns Hopkins, USA
Synaptic Communication Between Neurons and Glial Cells in the Mammalian CNS
Historically, synapses have been regarded as cellular components that are exclusive to neurons. However, Dr. Bergles and others have conclusively demonstrated that synapses also exist between neurons and glia, and his lecture presented several lines of work that explored this topic. Combining electrophysiology with light, confocal, and electron microscopy, he demonstrated the properties of these neuron-to-glia synapses on NG2 positive cells in the hippocampus, corpus callosum, and cerebellum. Furthermore, the neuron-glia synapses that are found in these brain regions have unique properties, ie. the functional and anatomical properties of the synapses differ depending on the brain region. For example, in the white matter tracts, it is only a certain subset of the fibers that make synaptic contacts with the NG2 cells, indicating a specific functional role for this pairing.
His research brings up some great questions: what is the functional role that these inputs are playing? How is receptor activation in these cells related to their migratory and myelinating properties? What are the downstream events that occur after receptor activation? Is receptor activation related to differentiation? Are these responses shared among all NG2 cells? I was very impressed with his talk, and during the question period, an audience member even gushed, “I think this is the best talk I have ever seen!”
Friday
Morning Plenary Lecture
Bill Richardson, UCL, London Tracking Progenitor/Stem Cells and their Progeny in the Adult Mouse Brain
Dr. Richardson is combining inducible genetics to both mark a certain cell population, and to ablate that population at a given time during development. He showed the audience the migratory patterns of cells out of the medial and dorsal spinal cord by using these specific genetic markers, and showed where these cells ended up and what became of their phenotype. He is trying to answer the big question of “where do glial cells come from?” Amazingly, he showed that he could ablate every ventral and dorsal OPC population, but OPCs from the diencephalon were still able to migrate and populate areas formerly covered by those populations to overcome the deficit. He also traced the fate of adult progenitor cell populations and showed that they become mature Sox10 positive oligodendrocytes, not GFAP+ astrocytes and occasionally, a NeuN+ neuron.
Morning Symposium
Neurotransmitter Signaling to Oligodendrocytes and Neurological Disease
The classical model of oligodendrocyte (and hence myelin) damage postulates that pathology results from an excessive activation of AMPA receptors expressed on the oligodendrocytes. This symposium was designed to present new data highlighting the importance of NMDA receptors, oxidative stress, and the immune response in mediating oligodendrocyte damage.
Carlos Matute, Universidad del Pais Vasco, Spain
Interactions of Glutamate Signaling and Immune Attack in Damaging Oligodendrocytes
I have been interested in Dr. Matute’s work ever since my preliminary examination that dealt with glutamate receptor mediated glial cell death, and then with our work on the KA1 kainate receptor subunit, so it was a real joy to listen to him speak.
Dr. Matute began his presentation by acknowledging that oligodendrocytes express AMPA, kainate, and NMDA type glutamate receptors, and that excessive activation of these receptors, and hence excessive calcium influx, is deleterious to oligodendrocyte survival. Furthermore, he acknowledged that glutamate receptor antagonists can ameliorate these effects. However, he presented data that shows how activated microglia can release glutamate through reverse transport, and hence, represents a source of glutamate release that is dependent on the immune response and independent from neuronal sources. He also showed how brief activity of oligodendrocyte glutamate receptors primes these cells for complement attack. The complement attack, he showed, was dependent on both kainate receptors and the formation of reactive oxygen species.
Complement attack is part of the innate immune system and involves the formation of (among other proteins) the Membrane Attack Complex (MAC) and is a potent means of clearing the body of pathogens. The MAC complex attacks the plasma membrane of a cell by forming a trans-membrane channel that makes the cell vulnerable to osmotic lysis (it pops a hole in the cell to make it burst).
Ragnihildur Karadottir, UCL, London
Neurotransmitter Signaling to 2 types of NG2 positive Oligodendrocyte Precursor
Dr. Karadottir presented functional data regarding the NMDA receptors expressed on NG2 positive OPCs. By combining cell fill and immunocytochemistry, she showed that the OPCs in the white matter tracts express NMDA receptors with a non-canonical pharmacological profile. Because these cells express the NR3 NMDA receptor subunit, they are not susceptible to the classical Mg block on NMDA receptors. In addition, she presented the currently accepted model of oligodendrocytes, where the AMPA receptors are localized to the soma of the cell, but NMDA receptors are localized to the internal and external folds of the myelin sheath.
She also showed intriguing data that suggests that a subset of NG2+ cells have large TTX sensitive sodium channels that are capable of producing spike activity, and that the synaptic inputs Dr. Bergles showed might only exist on cells with the Na current. Her data suggests that ischemic injury might effect these 2 cell populations differently.
Peter Stys, University of Calgary, Canada
NMDA Receptor Mediated Calcium Accumulation and Oligodendrocyte Damage
Dr. Stys presented amazing data where he used two-photon microscopy to image calcium fluxes in myelin. He showed convincing data that the myelin specific calcium responses were sensitive to NMDA receptor antagonists. His work greatly supports the current idea that NMDA receptors are expressed on the myelin sheath.
Afternoon Symposium
Pannexins: An Alternative Pathway for Cell-Cell Communication
Eliana Scemes, Albert Einstein College of Medicine, New York, USA
Yuri Panchin, Moscow State University, Russia
Rolf Dermietzel, University of Bochum, Germany
Gerhard Dahl, University of Miami, USA
What are pannexins? Do they form gap junctions? Where did they originate during evolution?
Gap junctional communication is thought to be universal among all multi cellular organisms (metazoa). Connexins were identified over 20 years ago as the vertebrate gap junction, but attempts to clone these proteins in invertebrates have failed. However, a new protein family was identified as the gap junctions in invertebrates and they were called innexins (invertebrate analog of connexins). Surprisingly, proteins with sequence homology to innexins were found in vertebrates, and the entire protein family is now called pannexins.
In the CNS, two pannexin genes, panx1 and panx2, are expressed by most, if not all, neural cells. The big debate in the field, it appears, is whether these pannexins are able to form gap junctions in vertebrates, since they form gap junctions in invertebrates and in vitro systems. The alternative hypothesis is whether these pannexins form gap junction hemichannels. If they are responsible for forming hemichannels, are they responsible for the release of ATP?
It seems that panx1 is expressed on postsynaptic sites of principal cells and interneurons of the cortex and hippocampus, and co-localizes with PSD-95. It might be a novel component of the postsynaptic protein complex. In other work, panx1 hemichannels were shown to open with changes in intracellular calcium levels, and that the channels were permeable to ATP. It seemed from the symposium that active research is uncovering the properties of these elusive channels, but the verdict is still out regarding their functions and localiztion.
Evening Plenary Lecture
Ron McKay, NIH, USA
Neural Stem Cells and Gliogenesis
Dr. Mckay is interested in the big question of how we get from an embryonic stem cell to a differentiated, functional cell in the CNS. He showed an in vitro cell tracing system wherein he followed the migration pattern of a newly derived pluripotent embryonic stem cell line. He took a picture every 10 minutes and followed the cell for 2 weeks, then stained the brain slice to determine what type of cells the ES cells had differentiated into. He found that not all ES cells migrated to the same locations, and that some cells had bi-potent or tri-potent potential, that is, a single cell was giving rise to neurons, astrocytes, and oligodendrocytes. Other cells migrated to a location and just differentiated into a single cell type. He demonstrated that both the lineage of the cell, and the signal processing that occur en route are important. In other work, he showed the potential for enhancing the endogenous repair system for the treatment of Parkinson’s disease.
Saturday Morning
Plenary Lecture
Linda Watkins, University of Colorado, Boulder, USA
Glial Regulation of Pain and Addiction
Pathological pain is a debilitating condition where normal stimuli such as warmth, coolness, and light pressure are misperceived as pain by the sensory system. Current drugs are ineffective in controlling this pain state. Dr. Watkins believes this is because the drugs target neurons. Her work is showing that glial activation is playing a role in modulating pain states, and believes that drugs would be more efficacious if they were targeting activated glia.
Pain can arise as a function of macrophage activation. As the macrophage engulfs invading pathogens, they produce pro-inflammatory cytokines, which in turn activate glial cells in the spinal cord, who then release their own pro-inflammatory cytokines. This glial activation can modulate the activity of dorsal horn neurons, and in concert with neuronal activation, produce the sensory condition known as pain.
Interestingly, her work is showing that the upregulation of glial pro-inflammatory cytokines is correlated with pathological pain states. Blocking the actions of the pro-inflammatory cytokine reduces pain, and reverses pathological pain.
In my opinion, her most interesting work was the roles activated glia are playing in modulating the analgesic efficacy of opioids. She showed how the production of the proinflammatory cytokines by astrocytes is masking the effects of opioids and even showed how she and her colleagues at Avigen Pharmaceutics are controlling the activated glial state to improve the efficacy of opioids, and amazingly, to reduce dependency.
Saturday Symposium
Diverse Regulatory Roles for Gliotransmitters at he Tripartite Synapse
Aside from the astrocytes and epilepsy symposium, this symposium, held on the last day of the conference, was the other symposium I really wanted to see. Due to my thesis work and curiosity, I have become really interested in the glial gliotransmitter release story, and I have been following the work of all the presenters in this symposium for some time. With regards to epilepsy and modulation of activity at the synapse, I feel that astrocytes, and their release of such diverse molecules as glutamate, D-serine, and ATP, must be considered in the normal and pathological function of the synapse.
Andrea Volterra, University of Lausanne, Switzerland
Glutamate Exocytosis from Astrocytes Controls Synaptic Strength
Dr. Volterra’s papers have been very instrumental in my understanding of the glial contribution to synaptic transmission. In 2004, his group demonstrated that astrocytes contain a vesicular compartment that undergoes calcium dependent exocytosis, the first observation of its kind. Recently, his work has focused on how astrocytic gliotransmitter release can modulate presynaptic transmitter release.
For the talk, he focused on synaptic strengthening. He showed data that was obtained from paired patch clamp recordings in the hippocampus of a dentate granule cell and an outer molecular layer astrocyte. He showed how the release of gliotransmitters can modulate the strength at this synapse, and preliminary work is suggesting that the mechanism may be post synaptic AMPA receptor insertion.
He found that only 1/3 of the cells they recorded from were paired. In other words, only about a third of the time could they see astrocytic modulation at the synapse. I am intrigued by this number, because it is line with other experiments. For example, the original Bezzi 2004 paper describing astrocytic vesicles also shows that only 32% of GFAP positive cells are VGLUT positive, suggesting that only a subset of GFAP positive astrocytes contain the necessary machinery to fill vesicles with glutamate. Also, the slow inward current data from Fellin, 2004, also shows that un-caging calcium in astrocytes results in a detectable slow inward current in pyramidal cells only 38% of the time. Might these suggest heterogeneity in the astrocyte population, such that in any given space, only a third are capable of releasing gliotransmitters? If this were the case, is this population static, or could it change with development, experience, learning, etc.? Or might this be an artifact of recording in acute slices?
Stephan Oliet, Inserm, France
Glial D-serine Controls NMDA Receptor Dependent Plasticity in the Rat Hypothalamus
Dr. Oliet talked about another story that I have been very interested in, the glial release of the endogenous NMDA receptor co-agonist D-serine. The general idea is that astrocytes sense presynaptic glutamate release, and respond by releasing D-serine in a calcium and vesicle dependent manner. Using the super optic nucleus of the hypothalamus as a model, Dr. Oliet demonstrated the dynamic nature of glial modulation of a synapse. During lactation, astrocytes retreat their processes such that the synaptic endings are less ensheathed. This results in less D-serine at the synaptic cleft and changes the nature of the synaptic communication that is occurring. The same stimuli, or neuronal activity, can then have very different effects on the synaptic potentiation or depression occurring at the synapse.
Jaideep Bains, University of Calgary, Canada
Glial-derived ATP Promotes AMPA Receptor Insertion and Increases the Strength of Glutamatergic Synapses
As his title suggests, Dr. Bains talked about the release of ATP from astrocytes and how ATP is controlling synaptic strength by promoting the insertion of AMPA receptors at the post synaptic synapse. He also uses the hypothalamus as his model system and showed with various experiments the importance of ATP release on AMPA insertion. It was, again, very intriguing to learn about this diverse molecule, and the mechanism of its release, and its downstream effects.
Philip Haydon, University of Pennsylvania, USA
Astrocytic Purinergic Signaling Coordinates Synaptic Networks
And finally, the last symposium speaker for the entire conference, and it was also the speaker I was most interested to listen to. I have followed Dr. Haydon’s seemingly continuous flow of papers out of the Silvio O. Conte Center for Integration at the Tripartite Synapse, and my own thought process with regards to epilepsy and hyper excitability have definitely been influenced by these papers.
Although I was hoping that he would talk about epilepsy, he instead focused on the adenosine signaling story, but I was happy to learn. He talked about the dn-SNARE inducible knock in mouse. This animal allows the inducible and conditional poisoning of the astrocytic SNARE protein responsible for vesicle fusion. Therefore, with one fell swoop, he can eliminate all contributions from the astrocytic vesicular release of gliotransmitters.
With this model, he showed intriguing data on how astrocytes are controlling the sleep-wake cycle! Animals that are sleep derived will sleep longer on successive nights, if they are allowed to. That is, if we are sleep deprived for several nights in a row, the first night that we can, we usually try to sleep as long as possible. Amazingly, dn-SNARE mice do not show this sleep recovery. How this happens is an intriguing story wrapped in adenosine, sleep wave cycles, and oscillations. I will have to wait until his papers come out for me to fully digest the huge amount of data that he presented.
Euroglia 2009: PARIS, FRANCE
I must be there! Its hard to describe the feeling after a conference like this.
All I know is that I want to be at the next Euroglia meeting.
I attended the optional Lecture Course on Glial Cell Biology at the University College of London prior to the start of the meeting. This was an all day lecture series reviewing astrocytes, oligodendrocytes, and microglia. It was an excellent course. My interest with astrocytes started as a technician in Scott Baraban's Lab at UCSF, then progressed with my courses in graduate school and then with my thesis project with Karen Wilcox, but never have I had such a comprehensive review on the all the cells of the glia family!
For due reasons, most courses in the neuroscience curriculums deal with neurons (development, function, disease, etc) and glial cells, though not disregarded, are simply not given their fair coverage. From the perspective of designing a core course in the neurosciences, this of course makes sense. Students need to get a broad overview of the field, and most of the research, historically, has focused on neurons. No doubt they are key players in the nervous system, but our understanding is incomplete without a full appreciation for the roles of glia.
Below, I have summarized the lecture courses and talks I attended during the 5 day conference. I think the reader will see both how organized and comprehensive this conference was, and how much of an impact this meeting had on one graduate student.
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Professor Rhona Mirsky, University College of London (UCL), reviewed schwann cells, the glial cells of the peripheral nervous system (PNS), whose functions include myelination, regulation of the microenvironment, nerve support during development, maintenance, movement, regeneration, and immune interactions with schwann cells. In particular, she focused on the critical role of neuregulin-1 as a signal for both schwann cell survival and myelin maintenance.
Professor Nigel Pringle (UCL) gave a talk titled (humorously) "Astrocytes: in 40 minutes." His talk focused on the problems with glial fibrillary acidic protein (GFAP) as a marker for astrocytes. The presence of this intermediate filament is universally accepted as a marker for astrocytes, but it has become apparent in recent years that astrocytes are not a homogeneous population. We need better markers that can identify the functional heterogeneity of astrocytes. His team is utilizing the Allen Brain Atlas to comb through gene expression profiles of the rodent brain to identify new markers. I was pleased to hear him talk about this subject, for it is something that I have been wondering about for a while now, and that he is using the Allen Brain Atlas. I am hearing references to this wonderful resource more and more.
Professor Richard Reynolds, Imperial College London (ICL), reviewed oligodendrocytes, the myelinating cells of the central nervous system (CNS). In particular he talked about the origins of the oligodendrocyte precursor cells (OPC), the migration of OPC through the spinal cord, and the signaling mechanisms that occur between OPC and axons to initiate myelin formation. This fascinating topic was one of the major themes at the conference. I was amazed at the numbers of posters and presentations that dealt with spinal cord repair, and was also amazed at where the field is right now. The term "stem cells" gets bandied around quite a bit, both politically and scientifically, so it was refreshing to hear (throughout the meeting) about this one particular type of cell that holds promise as one an endogenous stem cell of the brain.
Professor Deanna Taylor (ICL) gave a fascinating review on microglia, the resident mononuclear phagocytes of the brain. These wonderful little guys (15% of the total number of brain cells) constantly monitor and sense the extracellular environment by mobilizing their distal processes. Their filopodia are constantly shortening and elongating until they sense an activation signal to mobilize, then these cells swarm en masse and engulf the invader. I found a link to some of the movies (by Axel Nimmerjahn, who later gave a fascinating symposia at this conference) she showed which can be found here. In particular, check out the high resolution movie S7. In the beginning, you can see the microglia in their surveillance mode, but once the lesion happens, they become motile and move to the lesion site. An interesting point is that although microglia are critical for the brain's immune response and can clear the brain of unwanted invaders and pathogens, excessive microglial activation leads to a pathological environment deleterious to neuronal survival.
Professor Siddhartan Chandran of Cambridge University then gave a rousing review on stem cells, oligodendrocytes, and repair. His talk focused on the major question in stem cell research, which is directed differentiation. How do you get from one cell, the stem cell, to one specific type of cell out of the 216 possibilities (according to Dr. Chandran, there are 216 different cell types in the human body)? As a model, he focused on the OPC as an endogenous stem cell capable of regenerating into astrocytes, neurons, and oligodendrocytes. Like I mentioned, there is a lot of excitement for this little cell.
Professor David Attwell (UCL) then gave a talk on neurotransmitter signaling to astrocytes and oligodendrocytes, and the metabolic coupling of astrocytes and neurons. He gave a review on the somewhat controversial lactate shuttle hypothesis. The idea is that neurons derive their energy from importing lactate that is provided by astrocytes. The model predicts that as neuronal activity increases, and hence more glutamate is released into the synapse, astrocytic glutamate transporters clear the extracellular glutamate, but at the cost of energy depletion in the astrocyte. To counteract the energy demand, astrocytes then dilate the blood vessels to take up glucose. I love this particular topic. It is at the same time, both extremely complicated, in terms of the science, but incredibly beautiful to imagine the neurons and astrocytes in this type of a symbiotic and harmonious partnership.
Finally, Professor Nicola Woodroofe of Sheffield Hallam University gave a review on the properties of cytokines and chemokines in CNS physiology and pathology. The cytokines are small secreted proteins than are responsible for (along with many other functions) the immune response. This was a nice review because it is a topic that I am quite unfamiliar with and realized that it deserves more of my attention.
With the lecture course over, everyone dispersed and headed to Imperial College London for the start of Euroglia. Because the train line was closed, I decided to walk the hour and a half to ICL, and as I was looking over a map to decide directions, I met Marcelo and Anna-Paula, two PhD students from Brazil. They were to become my friends during the conference.
A snapshot of the main lecture auditorium. I counted about 650 people in attendance, and was told that 750 had registered! In the space below, I am going to summarize the talks and posters I attended, but have left out details on purpose.
Tuesday Evening
Opening Plenary Lecture 1
Marie Philbin, Hunter College, New York, USA
Regeneration in the CNS
Dr. Philbin talked about her strategy in developing CNS regenerating drugs for the treatment of spinal cord injury. Because I work along side the anti-convulsant drug discovery program at the University of Utah and we share lab space and lab meetings, drug discovery has been a topic that I have been exposed to a little bit, so this talk was particularly interesting from this perspective. One of the major obstacles in axon regeneration is the inhibitory signal from myelin. By manipulating cAMP, Dr. Philbin is trying to "trick" the axon to become unresponsive to these inhibitory signals in order to promote axon growth, and thus axon regeneration.
Opening Plenary Lecture 2
Patrick Charnay, INSERM, Paris, France
Glial Cell Functions in the PNS
Dr. Charnay discussed his research of the boundary cap cells that exist at the interface of the CNS and PNS. By using conditional knock in animals, he is investigating myelin formation in the PNS.
Wednesday Morning Plenary Lecture
Pierre Magistretti, University of Lausanne, Switzerland
Neuron-glia Metabolic Coupling
Dr. Magistretti introduced to the audience his hypothesis of neuron-glia metabolic coupling. He also discussed at length the metabolic wave that accompanies neuronal activation. As a synaptic event occurs and astrocytes take up glutamate, the response coincides with an increase in intracellular sodium from the coupled transport of glutamate and sodium ions. This signal is then amplified and propagated throughout the astrocytic syncytium as a Na, Ca, glucose, and ATP wave.
He also discussed the different roles of astrocytic and neuronal glucose transport. Though this is an oversimplification, in general, it seems that neurons are utilizing their own glucose transporters (Glut-3) for their basal activity, but as energy demands increase, glucose utilization switches to astrocytes, at which point, the lactate shuttle becomes essential for neuronal energy supply.
In addition, Dr. Magistretti proposed novel questions regarding energy demand. For example, is synaptic plasticity associated with metabolic plasticity? How is metabolic coupling involved in pathogenesis? What is the role of glycogen in the sleep-wake cycle? For each of these questions, there were a series of experiments that clarified his model and definitely piqued my curiosity with regards to these topics.
Morning Symposia
Functional Role of Bergmann Glia-Purkinje Cell Signaling
Axel Nimmerjahn, Stanford University
2-Photon Ca Imaging of Circuit Activity in the Cerebellar Cortex of Live Mice
Ok, this was an amazing talk. As the title suggests, Dr. Nimmerjahn's experimental set up allows for 2-photon calcium imaging of the cerebellum in live, non-anesthetized, behaving mice. One of the major problems in recording from live animals is the possible confounding effects of an anesthetic. He amazed the audience with movies of 2 types of calcium signals in the cerebellum that is correlated with the movement of the animal. One form was a short point like burst, and the other was more in line with what I would think a spreading calcium wave to be like.
Interestingly, there is a very recent paper in PNAS that shows two forms of vesicular release in astrocytes. One, being a very small, defined, kiss-and-run type of event, and the other being a more slow and spreading release event, probably representing a full fusion and exocytosis event. Of course this study was done in cell culture and they had the benefit of imaging in TIRF, so they were able to conclude the size of the objects involved down to the tens of nanometers, but it was cool to see something that kind of resembles that data in the live animal. Might a kiss and run event underlie the small punctate calcium increase while full fusion events represent the spread of a calcium wave? Or, are the calcium signals only representative of the underlying neuronal activity? What signals dictate the two forms of calcium signaling and what is the functional consequence of these different calcium waves? How are these waves related to the metabolic coupling of glia and neurons? Dr. Nimmerjahn is a post-doc at Stanford, so I eagerly talked to him about the imaging group there and was encouraged to contact them for further information.
Christine Rose, University of Dusseldorf, Germany
Imaging Glial Glutamate Uptake in the Cerebellum
Dr. Rose gave a talk on her use of a Na sensitive fluorescent dye to image glutamate transport by Bergmann glial cells. Yet again, I watched with amazement as she demonstrated how she patch clamped a cell while loading it with the fluorescent dye, then eliciting both a current response and a change in fluorescence as the cell detected extracellular glutamate. In this way, she is able to measure the current charge carried by the coupled transport of sodium ions with glutamate transport, and image the spatial spread of glutamate transporter activity in real time. I also interacted with her colleague who had a poster on the use of SR101 (sulforhodamine 101), a commercially available fluorescent dye, to specifically label astrocytes in hippocampal slice to aid in patch clamp recordings. His results suggest that I will be able to use this dye in the lab during my own experiments, and I was very excited to talk with him about his results.
Thomas Bellamy, The Babraham Institute, Cambridge, UK
Plasticity of Neuron to Bergmann Glial Cell Transmission
Dr. Bellamy presented data regarding the plasticity of the synapses that innervate Bergmann glia. Both parallel fiber and climbing fiber synapses are opposed by calcium permeable AMPA receptors on the Bergmann glial cell. He demonstrated that certain inputs and stimuli result in paired pulse facilitation, whereas other conditions resulted in paired pulse depression. Importantly, he showed that these effects are specific to the neuron-glia synapse, as neuron to neuron synapses were unaffected by his experimental setup.
Maria Rubio, University of Connecticut, Storrs, USA
Bidirectional Communication Between Purkinje Cell Synapses and Bergmann Glial Cell Processes
Dr. Rubio presented beautiful immunofluorescent images of the spatial relationships of specific AMPA receptor subtypes and the developmental regulation of these receptors. She also showed fascinating EM images of the translocation of AMPA receptors as the shape of the glial cell processes changes in response to various extracellular and intracellular signals.
Frank Kirchhoff, Max Planck Institute, Gottingen, Germany
Inducible Deletion of AMPA-type Glutamate Receptors in Bergmann Glia
As the title suggests, Dr. Kirchhoff uses a tamoxifen sensitive cre-lox system to conditionally induce the deletion of specific AMPA receptor subtypes. By using these animals, he is able to see the effects of knocking out these genes at both the morphological and behavioral levels.
Actually, his talk on the use of animals with genetic modifications highlights an important point. It was apparent throughout the meeting that sophisticated control of gene expression is yielding very exciting data on cells in the adult animal and it was very cool to see the diversity of experiments that utilized these techniques. Because most of the gene knockouts utilize some form of marker protein, and the knock out is performed in a certain cell types at specific time points, scientists are able to identify at any point which cells were affected by the knock out then track the deletion through time. The other important point is that since these genes can be knocked out with tamoxifen, the knock out is not hampered by any developmental side effects. The result is a visual, functional, and behavioral analysis on the function of a given gene in a given time point in the animal’s lifetime.
Dr. Kirschoff is finding the functional roles of the various AMPA receptor subtypes that are expressed by the Bergmann glia, and their effects on cell morphology and animal behavior.
Afternoon Symposia
The Role of Astrocyte Dysfunction in Epilepsy
Of course, I was very interested to see the speakers at this symposium!
Detlev Boison, DOW Legacy Research, Portland, Oregon, USA
Adenosine, Astrogliosis, and Seizures: A New Perspective of Epileptogenesis
Dr. Boison presented data on adenosine as an endogenous regulator of excitability. With regards to the gliotransmitters involved in modulating synaptic transmission, I found many labs whose focus was adenosine. Adenosine works to keep synaptic activity low, whereas its enzymatic counter part, adenosine kinase (ADK), breaks adenosine down and promotes synaptic activity. Following an acute seizure or brain injury, ADK is rapidly down-regulated whereas adenosine levels rise in the synapse. Thus, it seems that the brain has its own mechanism for neuroprotection and neural quiescence. However, elevated levels of adenosine kinase, or down regulation in adenosine production can lead to pathological hyperexcitability. Therefore, by combining animal models for epilepsy, genetic manipulation, and stem cell transplants, he is trying to use the adenosine system for the therapeutic treatment of epilepsy.
Annamaria Vezzani, Milano, Italy
Astrocytes as a Chronic Source of Inflammation in Epilepsy
Dr. Vezzani presented her work regarding inflammatory signals as possible contributors of the recurrent seizure state. During acute seizures, pro-inflammatory cytokines such as Il-1B are rapidly synthesized by both astrocytes and microglia. Interestingly, while Il-1B levels return to normal levels in microglia, it remains elevated in reactive astrocytes throughout chronic epilepsy. It is also known that reducing Il-1B levels will down regulate seizure activity. Therefore, her work is trying to understand how the immune response, especially this altered state of cytokine production, is contributing to epilepsy.
Devin Binder, UC Irvine, USA
Potential Role of Glial Water Channels in Epilepsy
This was another subject where I have studied little about, but found very interesting. The aquaporin water channels are responsible for water transport in the CNS. Seminal work in acute brain slices demonstrated the effects of changing the extracellular space volume on neuronal excitability. Because the aquaporins control the amount of water inside the cell, they can change its volume and thus change the volume of extracellular space. Dr. Binder uses an aquaporin knock out mouse to investigate what role these water channels have in seizure generation and chronic epilepsy.
Christian Steinhauser, University of Bonn, Germany
Altered Functional Properties of Astrocytes in Human Epileptogenic Hippocampus: Relevance to Seizure Generation
Dr. Steinhauser’s work has been especially important for me. As a student learning the electrophysiology of astrocytes, several papers have been very instrumental in my education. Among these, his papers that utilize the GFAP-GFP transgenic mouse to characterize the GluT and GluR astrocytes of the hippocampus have been crucial in understanding the results of my experiments. For the symposia, Dr. Steinhauser described the astrocyte phenotypes found in resected tissue from patients with Ammon’s horn sclerosis, then talked about a new animal model that recapitulates this very intriguing data. I found the results to be very exciting, and am looking forward to his next papers on this subject.
Evening Plenary Lecture
Helmut Kettenmann
Max Delbrueck Center for Molecular Medicine, Berlin, Germany
Control of Microglial Function by Transmitter Receptors
Dr. Kettenmann discussed his research regarding the ways in which microglia sense signals in the extracellular environment. His group has developed an in situ model to study the response of resting and activated microglia to various transmitters. In his talk, he discussed the effects of GABA, dopamine, noradrenaline, and adenosine. In general, it appears that activation of some of these transmitter receptors encourages microglia to remain in a resting state, supporting the hypothesis that microglia are less prone to be activated in the presence of normal neuronal activity. However, changes in these signals are probably contributing to the switch from resting microglia to the hyper ramified or activated state.
Thursday
Much to my extreme consternation, I discovered on Wednesday that I had printed my poster in the wrong length x width dimensions. Any hope of the poster fitting on the allotted space was erased when I went back to my lodging and unrolled the poster from its tube. I decided that I needed to reprint the poster in the correct dimensions, and that meant working late through the night, then getting to a print shop in the morning. I had traveled all the way to London, and this being the first time I was presenting at this particular meeting, I just could not bare to have a poster that appeared sloppy. Luckily, I found a print shop (Kinko’s!) that was able to print my poster in about 3 hours, and I made it on time to the afternoon symposia.
Thursday Afternoon Symposia
Glial Migration in the Developing and Pathological Brain
The main questions in this symposium were:
1. What are migratory glia?
2. What mediates migration?
3. How are they related to pathology?
4. Does pathology recapitulate development?
This symposium was very interesting because as I have mentioned, many posters and talks dealt with this topic of migrating OPC cells, and as my knowledge in this field is very premature, this symposium allowed for me an in-depth look into this topic.
Peter Cannoll, Columbia University, USA
Glial Progenitor Migration and Proliferation in PDGF-driven Brain Tumors
The tumor cells in brain gliomas are very tough cells to get rid of. Even when a surgeon removes the main tumor, small pockets of cells will proliferate and migrate towards the bloodstream. Gliomas are composed of many different types of cells, but they are abundant with cells that resemble glial progenitors (hence the name). By using a retrovirus marker (which only labels dividing cells), Dr. Cannoll is able to track the migratory patterns of cells out of the proliferative zones, and by manipulating levels of PDGF, he can induce these cells to migrate faster and further, to proliferate at an enhanced rate, inhibit their differentiation, and most importantly, induce tumor formation 100% of the time. By comparing the migratory patterns of normal cells to those that form gliomas, he is uncovering the mechanisms involved in glial migration during development and pathology.
Jacqueline Trotter, University of Mainz, Germany
NG2 and Glial Cell Migration
NG2 is a chondroiton sulfate proteoglycan that is expressed on the surface of OPCs, but is a protein that is down regulated as the OPC differentiates into a mature, myelinating oligodendrocyte. It is known that NG2 is expressed in glioma cells and that the protein is involved in the cytoskeletal modifications required for migration. Dr. Trotter is uncovering the intracellular binding partners for NG2 and presented data that suggests it is in a tripartite complex with AMPA receptors. Knowledge of these binding proteins could allow for a better understanding of what proteins are involved in NG2 expression on the cell membrane, and what role NG2 may have with regards to AMPA receptor expression and cell mobility and differentiation.
Vittorio Gallo, Children’s National Medical Center, Washington D.C., USA
ET-1 Regulates OPC Migration and Differentiation
Endothelins are small, secreted peptides that have caught my attention recently for their ability to modulate gap junction coupling. However, Dr. Gallo presented new data on the role that endothelins might play in other systems. I think that the data he presented has never been published, so I am going to refrain from talking about the details, but he presented some very interesting data regarding how endothelin signaling can modulate OPC migration. His research is important for the question of how we can get our endogenous stem cell population to migrate and myelinate.
Evening Plenary Lecture
Dwight Bergles, Johns Hopkins, USA
Synaptic Communication Between Neurons and Glial Cells in the Mammalian CNS
Historically, synapses have been regarded as cellular components that are exclusive to neurons. However, Dr. Bergles and others have conclusively demonstrated that synapses also exist between neurons and glia, and his lecture presented several lines of work that explored this topic. Combining electrophysiology with light, confocal, and electron microscopy, he demonstrated the properties of these neuron-to-glia synapses on NG2 positive cells in the hippocampus, corpus callosum, and cerebellum. Furthermore, the neuron-glia synapses that are found in these brain regions have unique properties, ie. the functional and anatomical properties of the synapses differ depending on the brain region. For example, in the white matter tracts, it is only a certain subset of the fibers that make synaptic contacts with the NG2 cells, indicating a specific functional role for this pairing.
His research brings up some great questions: what is the functional role that these inputs are playing? How is receptor activation in these cells related to their migratory and myelinating properties? What are the downstream events that occur after receptor activation? Is receptor activation related to differentiation? Are these responses shared among all NG2 cells? I was very impressed with his talk, and during the question period, an audience member even gushed, “I think this is the best talk I have ever seen!”
Friday
Morning Plenary Lecture
Bill Richardson, UCL, London Tracking Progenitor/Stem Cells and their Progeny in the Adult Mouse Brain
Dr. Richardson is combining inducible genetics to both mark a certain cell population, and to ablate that population at a given time during development. He showed the audience the migratory patterns of cells out of the medial and dorsal spinal cord by using these specific genetic markers, and showed where these cells ended up and what became of their phenotype. He is trying to answer the big question of “where do glial cells come from?” Amazingly, he showed that he could ablate every ventral and dorsal OPC population, but OPCs from the diencephalon were still able to migrate and populate areas formerly covered by those populations to overcome the deficit. He also traced the fate of adult progenitor cell populations and showed that they become mature Sox10 positive oligodendrocytes, not GFAP+ astrocytes and occasionally, a NeuN+ neuron.
Morning Symposium
Neurotransmitter Signaling to Oligodendrocytes and Neurological Disease
The classical model of oligodendrocyte (and hence myelin) damage postulates that pathology results from an excessive activation of AMPA receptors expressed on the oligodendrocytes. This symposium was designed to present new data highlighting the importance of NMDA receptors, oxidative stress, and the immune response in mediating oligodendrocyte damage.
Carlos Matute, Universidad del Pais Vasco, Spain
Interactions of Glutamate Signaling and Immune Attack in Damaging Oligodendrocytes
I have been interested in Dr. Matute’s work ever since my preliminary examination that dealt with glutamate receptor mediated glial cell death, and then with our work on the KA1 kainate receptor subunit, so it was a real joy to listen to him speak.
Dr. Matute began his presentation by acknowledging that oligodendrocytes express AMPA, kainate, and NMDA type glutamate receptors, and that excessive activation of these receptors, and hence excessive calcium influx, is deleterious to oligodendrocyte survival. Furthermore, he acknowledged that glutamate receptor antagonists can ameliorate these effects. However, he presented data that shows how activated microglia can release glutamate through reverse transport, and hence, represents a source of glutamate release that is dependent on the immune response and independent from neuronal sources. He also showed how brief activity of oligodendrocyte glutamate receptors primes these cells for complement attack. The complement attack, he showed, was dependent on both kainate receptors and the formation of reactive oxygen species.
Complement attack is part of the innate immune system and involves the formation of (among other proteins) the Membrane Attack Complex (MAC) and is a potent means of clearing the body of pathogens. The MAC complex attacks the plasma membrane of a cell by forming a trans-membrane channel that makes the cell vulnerable to osmotic lysis (it pops a hole in the cell to make it burst).
Ragnihildur Karadottir, UCL, London
Neurotransmitter Signaling to 2 types of NG2 positive Oligodendrocyte Precursor
Dr. Karadottir presented functional data regarding the NMDA receptors expressed on NG2 positive OPCs. By combining cell fill and immunocytochemistry, she showed that the OPCs in the white matter tracts express NMDA receptors with a non-canonical pharmacological profile. Because these cells express the NR3 NMDA receptor subunit, they are not susceptible to the classical Mg block on NMDA receptors. In addition, she presented the currently accepted model of oligodendrocytes, where the AMPA receptors are localized to the soma of the cell, but NMDA receptors are localized to the internal and external folds of the myelin sheath.
She also showed intriguing data that suggests that a subset of NG2+ cells have large TTX sensitive sodium channels that are capable of producing spike activity, and that the synaptic inputs Dr. Bergles showed might only exist on cells with the Na current. Her data suggests that ischemic injury might effect these 2 cell populations differently.
Peter Stys, University of Calgary, Canada
NMDA Receptor Mediated Calcium Accumulation and Oligodendrocyte Damage
Dr. Stys presented amazing data where he used two-photon microscopy to image calcium fluxes in myelin. He showed convincing data that the myelin specific calcium responses were sensitive to NMDA receptor antagonists. His work greatly supports the current idea that NMDA receptors are expressed on the myelin sheath.
Afternoon Symposium
Pannexins: An Alternative Pathway for Cell-Cell Communication
Eliana Scemes, Albert Einstein College of Medicine, New York, USA
Yuri Panchin, Moscow State University, Russia
Rolf Dermietzel, University of Bochum, Germany
Gerhard Dahl, University of Miami, USA
What are pannexins? Do they form gap junctions? Where did they originate during evolution?
Gap junctional communication is thought to be universal among all multi cellular organisms (metazoa). Connexins were identified over 20 years ago as the vertebrate gap junction, but attempts to clone these proteins in invertebrates have failed. However, a new protein family was identified as the gap junctions in invertebrates and they were called innexins (invertebrate analog of connexins). Surprisingly, proteins with sequence homology to innexins were found in vertebrates, and the entire protein family is now called pannexins.
In the CNS, two pannexin genes, panx1 and panx2, are expressed by most, if not all, neural cells. The big debate in the field, it appears, is whether these pannexins are able to form gap junctions in vertebrates, since they form gap junctions in invertebrates and in vitro systems. The alternative hypothesis is whether these pannexins form gap junction hemichannels. If they are responsible for forming hemichannels, are they responsible for the release of ATP?
It seems that panx1 is expressed on postsynaptic sites of principal cells and interneurons of the cortex and hippocampus, and co-localizes with PSD-95. It might be a novel component of the postsynaptic protein complex. In other work, panx1 hemichannels were shown to open with changes in intracellular calcium levels, and that the channels were permeable to ATP. It seemed from the symposium that active research is uncovering the properties of these elusive channels, but the verdict is still out regarding their functions and localiztion.
Evening Plenary Lecture
Ron McKay, NIH, USA
Neural Stem Cells and Gliogenesis
Dr. Mckay is interested in the big question of how we get from an embryonic stem cell to a differentiated, functional cell in the CNS. He showed an in vitro cell tracing system wherein he followed the migration pattern of a newly derived pluripotent embryonic stem cell line. He took a picture every 10 minutes and followed the cell for 2 weeks, then stained the brain slice to determine what type of cells the ES cells had differentiated into. He found that not all ES cells migrated to the same locations, and that some cells had bi-potent or tri-potent potential, that is, a single cell was giving rise to neurons, astrocytes, and oligodendrocytes. Other cells migrated to a location and just differentiated into a single cell type. He demonstrated that both the lineage of the cell, and the signal processing that occur en route are important. In other work, he showed the potential for enhancing the endogenous repair system for the treatment of Parkinson’s disease.
Saturday Morning
Plenary Lecture
Linda Watkins, University of Colorado, Boulder, USA
Glial Regulation of Pain and Addiction
Pathological pain is a debilitating condition where normal stimuli such as warmth, coolness, and light pressure are misperceived as pain by the sensory system. Current drugs are ineffective in controlling this pain state. Dr. Watkins believes this is because the drugs target neurons. Her work is showing that glial activation is playing a role in modulating pain states, and believes that drugs would be more efficacious if they were targeting activated glia.
Pain can arise as a function of macrophage activation. As the macrophage engulfs invading pathogens, they produce pro-inflammatory cytokines, which in turn activate glial cells in the spinal cord, who then release their own pro-inflammatory cytokines. This glial activation can modulate the activity of dorsal horn neurons, and in concert with neuronal activation, produce the sensory condition known as pain.
Interestingly, her work is showing that the upregulation of glial pro-inflammatory cytokines is correlated with pathological pain states. Blocking the actions of the pro-inflammatory cytokine reduces pain, and reverses pathological pain.
In my opinion, her most interesting work was the roles activated glia are playing in modulating the analgesic efficacy of opioids. She showed how the production of the proinflammatory cytokines by astrocytes is masking the effects of opioids and even showed how she and her colleagues at Avigen Pharmaceutics are controlling the activated glial state to improve the efficacy of opioids, and amazingly, to reduce dependency.
Saturday Symposium
Diverse Regulatory Roles for Gliotransmitters at he Tripartite Synapse
Aside from the astrocytes and epilepsy symposium, this symposium, held on the last day of the conference, was the other symposium I really wanted to see. Due to my thesis work and curiosity, I have become really interested in the glial gliotransmitter release story, and I have been following the work of all the presenters in this symposium for some time. With regards to epilepsy and modulation of activity at the synapse, I feel that astrocytes, and their release of such diverse molecules as glutamate, D-serine, and ATP, must be considered in the normal and pathological function of the synapse.
Andrea Volterra, University of Lausanne, Switzerland
Glutamate Exocytosis from Astrocytes Controls Synaptic Strength
Dr. Volterra’s papers have been very instrumental in my understanding of the glial contribution to synaptic transmission. In 2004, his group demonstrated that astrocytes contain a vesicular compartment that undergoes calcium dependent exocytosis, the first observation of its kind. Recently, his work has focused on how astrocytic gliotransmitter release can modulate presynaptic transmitter release.
For the talk, he focused on synaptic strengthening. He showed data that was obtained from paired patch clamp recordings in the hippocampus of a dentate granule cell and an outer molecular layer astrocyte. He showed how the release of gliotransmitters can modulate the strength at this synapse, and preliminary work is suggesting that the mechanism may be post synaptic AMPA receptor insertion.
He found that only 1/3 of the cells they recorded from were paired. In other words, only about a third of the time could they see astrocytic modulation at the synapse. I am intrigued by this number, because it is line with other experiments. For example, the original Bezzi 2004 paper describing astrocytic vesicles also shows that only 32% of GFAP positive cells are VGLUT positive, suggesting that only a subset of GFAP positive astrocytes contain the necessary machinery to fill vesicles with glutamate. Also, the slow inward current data from Fellin, 2004, also shows that un-caging calcium in astrocytes results in a detectable slow inward current in pyramidal cells only 38% of the time. Might these suggest heterogeneity in the astrocyte population, such that in any given space, only a third are capable of releasing gliotransmitters? If this were the case, is this population static, or could it change with development, experience, learning, etc.? Or might this be an artifact of recording in acute slices?
Stephan Oliet, Inserm, France
Glial D-serine Controls NMDA Receptor Dependent Plasticity in the Rat Hypothalamus
Dr. Oliet talked about another story that I have been very interested in, the glial release of the endogenous NMDA receptor co-agonist D-serine. The general idea is that astrocytes sense presynaptic glutamate release, and respond by releasing D-serine in a calcium and vesicle dependent manner. Using the super optic nucleus of the hypothalamus as a model, Dr. Oliet demonstrated the dynamic nature of glial modulation of a synapse. During lactation, astrocytes retreat their processes such that the synaptic endings are less ensheathed. This results in less D-serine at the synaptic cleft and changes the nature of the synaptic communication that is occurring. The same stimuli, or neuronal activity, can then have very different effects on the synaptic potentiation or depression occurring at the synapse.
Jaideep Bains, University of Calgary, Canada
Glial-derived ATP Promotes AMPA Receptor Insertion and Increases the Strength of Glutamatergic Synapses
As his title suggests, Dr. Bains talked about the release of ATP from astrocytes and how ATP is controlling synaptic strength by promoting the insertion of AMPA receptors at the post synaptic synapse. He also uses the hypothalamus as his model system and showed with various experiments the importance of ATP release on AMPA insertion. It was, again, very intriguing to learn about this diverse molecule, and the mechanism of its release, and its downstream effects.
Philip Haydon, University of Pennsylvania, USA
Astrocytic Purinergic Signaling Coordinates Synaptic Networks
And finally, the last symposium speaker for the entire conference, and it was also the speaker I was most interested to listen to. I have followed Dr. Haydon’s seemingly continuous flow of papers out of the Silvio O. Conte Center for Integration at the Tripartite Synapse, and my own thought process with regards to epilepsy and hyper excitability have definitely been influenced by these papers.
Although I was hoping that he would talk about epilepsy, he instead focused on the adenosine signaling story, but I was happy to learn. He talked about the dn-SNARE inducible knock in mouse. This animal allows the inducible and conditional poisoning of the astrocytic SNARE protein responsible for vesicle fusion. Therefore, with one fell swoop, he can eliminate all contributions from the astrocytic vesicular release of gliotransmitters.
With this model, he showed intriguing data on how astrocytes are controlling the sleep-wake cycle! Animals that are sleep derived will sleep longer on successive nights, if they are allowed to. That is, if we are sleep deprived for several nights in a row, the first night that we can, we usually try to sleep as long as possible. Amazingly, dn-SNARE mice do not show this sleep recovery. How this happens is an intriguing story wrapped in adenosine, sleep wave cycles, and oscillations. I will have to wait until his papers come out for me to fully digest the huge amount of data that he presented.
Euroglia 2009: PARIS, FRANCE
I must be there! Its hard to describe the feeling after a conference like this.
All I know is that I want to be at the next Euroglia meeting.
Posted: Thu - September 6, 2007 at 11:37 AM
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Published On: Sep 14, 2007 04:34 PM
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Published On: Sep 14, 2007 04:34 PM