Matthew Zabel, PhD – Brain Blogger http://brainblogger.com Health and Science Blog Covering Brain Topics Wed, 30 May 2018 15:00:03 +0000 en-US hourly 1 https://wordpress.org/?v=4.9.6 Microglia – Part 1, Definitions and Developmental Progression http://brainblogger.com/2015/04/12/microglia-part-1-definitions-and-developmental-progression/ http://brainblogger.com/2015/04/12/microglia-part-1-definitions-and-developmental-progression/#respond Sun, 12 Apr 2015 12:00:45 +0000 http://brainblogger.com/?p=19192 One of the more remarkable advances in neuroscience, perhaps on par with Santiago Ramon y Cajal’s Neuron Doctrine (the theory that distinct neurons are the functional units of the brain), is the discovery of microglia — appropriately by Cajal’s student Pio del Rio Hortega.

Since their discovery, microglia have been the center of controversy in many contexts (i.e. ontogeny, neurodevelopment, synaptic plasticity, disease, etc.). This is not an overstatement. For example, Hortega surmised (correctly), back in the 1920s, that these cells hailed from the mesoderm, rather than the neuroectoderm (like all other brain cells) during an organism’s development.

This prediction, however, was not definitively verified until seven decades later, when it was demonstrated that microglia do not populate the neuroectoderm until embryonic day eight. This insight generated further questions, including: from where do these alien cells specifically originate? This question of microglial ontogeny and other definitions will be the focus of Part I of a five part series on the role of microglia in brain function from development to disease, as potential therapeutic targets and culminating with a fun — possibly unexpected — look at futuristic directions.

First, let us define some important terms. Unfortunately, many contemporary authors continue to use antiquated terminology when discussing microglia and this has become somewhat debilitating to the field (not to mention a pet peeve of mine). In 1908, Elie Metchnikoff won the Nobel Prize for his work on phagocytosis, or “cellular act of eating,” which later lead to his description of the macrophage system. This system is made up of cells collectively called mononuclear phagocytes, resulting in what is now called the mononuclear phagocyte system. It is this classification to which the microglial cell fits. Microglia are, after all, the tissue resident phagocytic cells of the brain.

Now, for the purpose of this article, I would like to define two more classification terms: “circulating” monocytes and “tissue-resident” macrophages. This distinction is important developmentally. It was recently demonstrated that circulating and tissue-resident cells arise from temporally and spatially different stem cell pools. For example, nearly all of our organs contain macrophages specifically designed to function within that organ, mainly performing maintenance duty. These tissue-resident macrophages derive from a developmental organ known as the yolk sac (most anatomists might call it an umbilical vesicle now), which is the sight of primitive hematopoeisis (or blood production). In other words, macrophages designed to function in a specific tissue are made first from a special pool of hematopoeitic stem cells and then leave the yolk sac to colonize their respective organs — such as microglia in the brain, Langerhans cells in the skin, or Kupfer cells in the liver. On the other hand, circulating monocytes are produced in the bone marrow later on in development, have a high turn over rate, and are continually produced over a lifetime. These cells mostly meander through the blood and lymphatic system and usually die after becoming activated — not necessarily so for tissue-resident macrophages.

So, it was shown in 2011 in the journal Science for the first time, that microglial ontogeny begins in the yolk sac. From there, they travel through the newly developed blood stream to reach the brain before the aptly named blood brain barrier closes — forever shunning the entry of any other type of mononuclear phagocyte. Here they spend the rest of their lives actively surveying surrounding brain tissue (among other roles to be discussed in following articles), providing a homeostatic milieu for the brain to function efficiently.

This brings us to our last set of definitions: surveillance, primed, activated and dystrophic microglia. I will not go deeply into each category, but I think it is important to understand that microglia are the most dynamic cells in the brain with their unique ability to transition between these activation states.

I would also like to emphasize that these states are contextual and do not necessarily describe a microglial cell as beneficial or destructive to surrounding brain tissue, but instead are responses to molecular cues necessitating some microglial effector function. For example, a microglial cell can be activated in both developmental synapse pruning (beneficial) and phagocytosis of viable neurons during disease (detrimental). No matter what, microglia are never resting — a term that has also deluded the field — but in constant motion within defined, non-overlapping locales.

You can find a live video I have taken of a retinal microglia in this state here. Instead of “resting” microglia, they are more appropriately termed surveillance microglia, as they extend and retract their processes, feeling around the adjacent tissue, sensing the environment.

Primed microglia describes the state between surveillance and activation. One could think of it as a pre-activation state, where they begin showing signs of morphological changes (i.e. shortening and thickening of their processes) and expression of certain molecules.

From there, microglia become fully activated, taking on a rounded (amoeboid) shape and expressing high levels of molecules associated with inflammation and/or phagocytosis.

Lastly, dystrophic describes a dysfunctional state of microglia, in which their processes begin to fragment or even fuse with other microglia. This is usually associated with chronic activation during disease and not usually part of their functional spectrum. These definitions are currently in a state of flux as the field tries to develop a cohesive theory of microglial function.

When looking at a fluorescent image of a mature, surveying microglia, one will notice how elegantly their processes radiate from the small cell body, forming complex, arborous structures. This is not their perpetual morphologic state. On the contrary, they are born as small, amorphous blobs, reminiscent of peripheral macrophages. In fact, their morphological development is the reverse of their transition through activation states. It is not until they enter the developing brain that they begin to ramify. Once they enter, microglia spread out to evenly tile the entire brain, taking up their positions as noble sentinels.

Research is ongoing to determine what molecular cues developing microglia receive (and from where, but probably the brain) to drive their migration into the brain. A few candidate molecules (e.g. CSF-1, IL-34, TGF-beta; no need to memorize these) have been implicated. It can be imagined that as microglia develop from primitive macrophages in the yolk sac, such molecules would create a signaling gradient that microglia, expressing corresponding receptors, would follow to reach their final desination. Think of leaving a trail of biscuits for your stubborn puppy to follow if you want to get her into the kennel.

Once they enter the brain, what keeps them there and in that complex, ramified morphology? What induces them to clear developmentally apoptotic neurons and prune synapses, both to allow appropriate, efficient wiring of the brain? Again, we think we have some ideas, but the precise molecular mechanisms remain unclear. What is clear, however, is that this area of microglial research is still blossoming and constant strides are being made to understand the development of these frenetic little cells.

This could lead us to some interesting insights that will facilitate therapies for diseases where microglia seem to have a role; a topic we will cover in Part III of this series. Now that we have an idea of microglial ontogeny, we will next discuss what role they play in the healthy developing and adult brain in Part II of Microglia.

References

Alliot F, Godin I, & Pessac B (1999). Microglia derive from progenitors, originating from the yolk sac, and which proliferate in the brain. Brain research. Developmental brain research, 117 (2), 145-52 PMID: 10567732

Ginhoux F, Greter M, Leboeuf M, Nandi S, See P, Gokhan S, Mehler MF, Conway SJ, Ng LG, Stanley ER, Samokhvalov IM, & Merad M (2010). Fate mapping analysis reveals that adult microglia derive from primitive macrophages. Science (New York, N.Y.), 330 (6005), 841-5 PMID: 20966214

Gomez Perdiguero E, Klapproth K, Schulz C, Busch K, Azzoni E, Crozet L, Garner H, Trouillet C, de Bruijn MF, Geissmann F, & Rodewald HR (2015). Tissue-resident macrophages originate from yolk-sac-derived erythro-myeloid progenitors. Nature, 518 (7540), 547-51 PMID: 25470051

Kaufmann SH (2008). Immunology’s foundation: the 100-year anniversary of the Nobel Prize to Paul Ehrlich and Elie Metchnikoff. Nature immunology, 9 (7), 705-12 PMID: 18563076

Rezaie P, & Male D (2002). Mesoglia & microglia–a historical review of the concept of mononuclear phagocytes within the central nervous system. Journal of the history of the neurosciences, 11 (4), 325-74 PMID: 12557654

Image via Johan Swanepoel / Shutterstock.

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Ultrasound to Slow Alzheimer’s Disease? http://brainblogger.com/2015/03/23/ultrasound-to-slow-alzheimers-disease/ http://brainblogger.com/2015/03/23/ultrasound-to-slow-alzheimers-disease/#respond Mon, 23 Mar 2015 15:00:50 +0000 http://brainblogger.com/?p=19098 The media is buzzing after recent publication of an article in the journal Science: Translational Medicine, which suggests that we can treat Alzheimer’s disease non-invasively with ultrasound. As usual though, they may be making more hype than is warranted given a thorough look at the data; or if they would have read the last paragraph, where the authors discuss (some of) the study’s weaknesses.

The study authors, from Queensland Brain Institute in Australia, used repeated scanning ultrasound in conjunction with intravenous injection of microbubbles (a gas core wrapped in a lipid shell) to safely induce blood brain barrier opening in a mouse model of Alzheimer’s disease (AD). The current understanding of how this method works is to vibrate the bubbles using ultrasound, causing them to expand and contract. This action displaces the vessel wall and transiently releases the tight junctions between blood vessel cells (endothelial cells) that keep a tight lock on the barrier.

Interestingly, they found that in doing so, they could reduce the number of the amyloid-beta plaques that are a characteristic (and defining) pathology of the disease. In the Amyloid Cascade Hypothesis, a dogmatic theory that has dominated the field for several decades, the aggregation of the amyloid-beta protein (a metabolite of the Amyloid Precursor Protein) into plaques results in neuronal death. This hypothesis has been continually revised and now blames the pre-aggregated, lower molecular weight amyloid oligomers for the ensuing destruction.

We will first begin by looking at the animal model in use. The APP23 mouse model of AD is widely used, but like any model has its weaknesses. The problem with many of the AD mouse models is that they do not necessarily recapitulate the most common human form of AD: sporadic. These models rely on genetic manipulation of the amyloid precursor protein to generate more amyloid-beta than physiologic levels seen in humans with the disease — basically, way too much. Right off the bat, they are biasing their results.

But let’s say, hypothetically, that the APP23 model is sufficient to answer questions about the causes of AD. The experimental groups consist of a wild type (normal) mouse, APP23 mouse without ultrasound (also called the “sham” group) and APP23 mouse with ultrasound (the treated group). Both of the APP23 groups received the microbubble injection. The authors show visually and biochemically that they can reduce levels of many forms of amyloid-beta with microbubble-coupled ultrasound compared to the sham group at no ancillary risk to the animal.

When they look at the behavioral manifestations of this, however, the effects are not as robust. In fact, it looks as if the sham group may get worse, which is difficult to explain. This is evident when the APP23 mouse (without ultrasound or microbubbles) is compared to the sham group. The treated group does show a slight restoration in the behavioral outcome, but considering the variable overlap with the sham group, it is difficult to interpret this as clinically significant.

Now, how is this working? Enter, microglia — maybe. As the innate immune cells of the brain, they have previously been implicated in the removal of amyloid-beta from the brain (including in my own work). The authors do show, rather convincingly, that microglia are eating up a lot of amyloid-beta in the treated group; almost twice as much as microglia in the sham group. That is a lot of amyloid for microglia, as they have occasionally been accused of inefficiency in this context.

The authors try to tie this in with the opening of the blood brain barrier by suggesting that albumin (the most common fluid-phase blood protein) leaks into the brain and somehow helps microglia gobble up higher levels of amyloid in the treated group. Not only is this ambiguously demonstrated in a cell culture model (in vitro) without any mechanistic evidence, but it sounds more like magic. Thankfully the authors concede that additional work should be completed on this front.

In looking for other mechanisms in which this reduction in amyloid could be occurring, the authors investigated microglial activation state, in which the only altered characteristics were those suggestive of phagocytosis (i.e. microglial cell morphology was more rounded and they expressed higher levels the phagocytic marker CD68). This does lend credence to their amyloid phagocytosis hypothesis, and they suggest this is somehow a direct result from the ultrasound stimulation. This seems a bit specious, so let us look at this scenario a different way.

The results of this study are predicated on opening of the blood brain barrier and the influx of albumin, which magically helps microglia consume more amyloid. Why the authors went with this explanation of amyloid-beta reduction and not the low hanging fruit of infiltrating peripheral monocytes, is beyond me.

It has been shown in many studies that peripheral monocytes (which become macrophages when they enter into a tissue) are much more efficient at engulfing amyloid-beta than endogenous brain microglia. Unfortunately, unless molecularly scrutinized, the differences between microglia and monocytes are nuanced, and the methods used in this study fail to differentiate them. The lack of distinction between the two cell types is a hot topic in neurodegenerative diseases at the moment and it surprises me that the reviewers and editors of the journal overlooked this obvious weakness in the study.

To be fair, the ability to open up the blood brain barrier non-invasively is a huge strength of this study and despite the lack of evidence for how it works to reduce levels of amyloid-beta, there are other important potential uses for this technique. One such example would be to enhance delivery of drugs to the central nervous system, a feat by which the neuroscience, neurology and neurosurgery communities are currently baffled. Once an effective drug is developed, ultrasound could be the means for targeted delivery to the brains of Alzheimer’s patients.

References

Hardy J (2006). Alzheimer’s disease: the amyloid cascade hypothesis: an update and reappraisal. Journal of Alzheimer’s disease : JAD, 9 (3 Suppl), 151-3 PMID: 16914853

Leinenga G, & Götz J (2015). Scanning ultrasound removes amyloid-? and restores memory in an Alzheimer’s disease mouse model. Science translational medicine, 7 (278) PMID: 25761889

Town T, Laouar Y, Pittenger C, Mori T, Szekely CA, Tan J, Duman RS, & Flavell RA (2008). Blocking TGF-beta-Smad2/3 innate immune signaling mitigates Alzheimer-like pathology. Nature medicine, 14 (6), 681-7 PMID: 18516051

Zabel M, Schrag M, Crofton A, Tung S, Beaufond P, Van Ornam J, Dininni A, Vinters HV, Coppola G, & Kirsch WM (2013). A shift in microglial ?-amyloid binding in Alzheimer’s disease is associated with cerebral amyloid angiopathy. Brain pathology (Zurich, Switzerland), 23 (4), 390-401 PMID: 23134465

Image via Akbudak Rimma / Shutterstock.

Shutterstock ID: 65181187

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What Have Your Glia Done For You Today? http://brainblogger.com/2015/03/16/what-have-your-glia-done-for-you-today/ http://brainblogger.com/2015/03/16/what-have-your-glia-done-for-you-today/#respond Mon, 16 Mar 2015 11:00:43 +0000 http://brainblogger.com/?p=19006 How should we approach solving the most fundamental questions in neuroscience? Some would say that our current endeavors into mapping the complement of neuronal connections (the connectome) within the human brain will bring us the most fruitful gains; and they could be right. This is the main focus of several (expensive) big data initiatives such as the BRAIN Initiative, the Human Connectome Project and the Human Brain Project.

However, a major shortcoming of these projects is the lack of goals slated to mapping the other half of the brain; a complement of cells known as glia, of which there are three predominant populations. These cells make up fifty percent of the total brain volume and are integral to normal brain function. A complete understanding of the brain’s “support cells”, as they are also called, along with mapping the neuronal wiring, will be paramount for a global theory of how the brain works to bring about consciousness and our plethora of behaviors.

Microglia

Neuroglia consist mainly of astrocytes, oligodendrocytes and microglia, the last of which do not actually originate in the brain. Instead, microglia (my current field of research), originate in the fetal yolk sac (a structure involved in primitive hematopoesis, or blood formation) where many other primitive and tissue resident macrophages are born. Contrary to original theories, microglia migrate into the brain early in embryonic life and reside there as a stable, self-sustaining population of cells, which are never replaced by bone marrow-derived cells from the periphery.

In essence, unlike other glia, our microglia grow old with us. As a pool of tissue-specific macrophages, microglia constitute the immune cells of the central nervous system. They are capable of mounting an immune response against foreign invaders (e.g. blood cells or proteins and bacteria) as well as clear neuronal debris after injury. As such, they have, somewhat insultingly, been dubbed the “garbage men of the brain.”

It has become clear that this is short-sighted. For example, microglia mediate the elimination of supernumerary synapses from the developing brain through a mechanism known as phagocytosis. Essentially, they gobble up the unnecessary synapses that the brain generates during development, a process without which many developmental and behavioral abnormalities can occur. Understanding this process is thus essential for determining how the brain’s connections are wired.

Extending their services into postnatal life, microglia also maintain the status quo within the brain. For example, when microglia are completely depleted from the central nervous system, adult mice show deficits in learning. In other words, your microglia can make you smarter through learning-induced synaptic remodeling; they act to balance removal of unused or inappropriate synapses allowing neuronal energy expenditure to be allocated to forming efficient synapses.

What important programs do these little “electricians” employ to bring about an appropriately wired brain? How can we manipulate them to prevent or curb neurological and mental illnesses? These are important questions that the neuroscientific community is currently asking, but must be put into the wider framework of mapping the human brain.

Astrocytes

Astrocytes (or astroglia) perform the most diverse set of functions of the glial cells, including sharing with microglia the role of synaptic maintenance. How could they not? A single astrocyte can contact up to one million synapses. At the microstructure level this is known as the “tri-partate synapse”.

It is at this juncture of the pre- and post-synaptic clefts (where the electrical signal of one neuron is coupled via a chemical signal – a neurotransmitter – to the next neuron) and the astroglial process, that astrocytes do most of their work. Here, they can regulate the rate and strength of synaptic transmission across neurons by releasing or sequestering neurotransmitters and modifying extracellular ion concentrations. Not only do they “listen” and “talk” to neurons, but they also provide metabolic support of endothelial cells, which line the brain vasculature, making up the blood brain barrier. At this site, they are also well positioned to regulate the amount of blood flow to a local brain region.

Recently, astrocytes have made public headlines with the discovery of their involvement in bulk clearance of brain metabolic waste during sleep, the importance of which is not lost on us after staying up too late the night before an exam.

Oligodendrocytes

Oligodendrocytes (oligos for short), the central nervous system equivalent of the Schwann cell in the peripheral nervous system, wraps a fatty sheath around the axons of neurons. This sheath known as myelin provides insulation to the axon and allows for much faster propagation of the electrical signal and thus faster communication and transmission of information. Think of it as the insulation around electrical wires on your favorite appliance.

Oligos seem to only function in this capacity, but we do not actually know that – a rough search on PubMed shows about 25 times more papers are published on neurons than oligodendrocytes. For example, a study published this week in the journal Science found that oligos are much more diverse than previously known. The study authors found that this cell type alone could be sub-divided into six classes based on their molecular signatures. We do not yet know the function of each or how each type may play a role in brain cytoarchitecture to facilitate faster, more efficient neuronal signaling, but it should become a priority.

Each of the glial functions listed above (and we have left out other types of glia, such as Bergmann glia, NG2 glia and ependymal cells, but we know even less about them) are essential to global brain plasticity and learning. Because of their intimate relationship with neurons, glial cells hold a central position in influencing cognitive processes. Author of The Other Brain, R. Douglas Fields suggested in a Nature Comment:

“[That] these include processes requiring the integration of information from spatially distinct parts of the brain, such as learning or the experiencing of emotions, which take place over hours, days and weeks, not in milliseconds or seconds.”

In other words, because glia work on the slower chemical level, not via electrical signals, they may facilitate paradigms such as learning, which do not happen instantaneously like say, visual perception. I could not agree more. But at this time, we can only speculate as to how glia might impart their “magical elixir” on neurons, facilitating the workings of the most complex biological entity in the known universe.

Over a century after the start of glial research, we still lack fundamental knowledge of glial numbers, diversity, distribution throughout the brain and function in the context of neurons and between themselves. The “neurotechnologies” that arise from the neurocentric initiatives should additionally be allocated to studying these questions.

If our minds, which emerge from the tangled wet mass of neurons, make us who we are, then paraphrasing my mentor’s mantra sums it up best: What have your glia done for you today?

References

Araque A, Parpura V, Sanzgiri RP, & Haydon PG (1999). Tripartite synapses: glia, the unacknowledged partner. Trends in neurosciences, 22 (5), 208-15 PMID: 10322493

Kettenmann, H. & Ransom, B. R. (eds) Neuroglia 3rd edn (Oxford Univ. Press, 2013).

Paolicelli RC, Bisht K, & Tremblay MÈ (2014). Fractalkine regulation of microglial physiology and consequences on the brain and behavior. Frontiers in cellular neuroscience, 8 PMID: 24860431

Parkhurst CN, Yang G, Ninan I, Savas JN, Yates JR 3rd, Lafaille JJ, Hempstead BL, Littman DR, & Gan WB (2013). Microglia promote learning-dependent synapse formation through brain-derived neurotrophic factor. Cell, 155 (7), 1596-609 PMID: 24360280

Xie L, Kang H, Xu Q, Chen MJ, Liao Y, Thiyagarajan M, O’Donnell J, Christensen DJ, Nicholson C, Iliff JJ, Takano T, Deane R, & Nedergaard M (2013). Sleep drives metabolite clearance from the adult brain. Science (New York, N.Y.), 342 (6156), 373-7 PMID: 24136970

Zeisel A, Muñoz-Manchado AB, Codeluppi S, Lönnerberg P, La Manno G, Juréus A, Marques S, Munguba H, He L, Betsholtz C, Rolny C, Castelo-Branco G, Hjerling-Leffler J, & Linnarsson S (2015). Brain structure. Cell types in the mouse cortex and hippocampus revealed by single-cell RNA-seq. Science (New York, N.Y.), 347 (6226), 1138-42 PMID: 25700174

Image via Jose Luis Calvo / Shutterstock.

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