passionless Droning about autism

Posts Tagged ‘Bilbo

Hello friends –

I have a confession to make.  The fact that a lot of very smart people have ignored or flat out laughed at my arguments bothers me sometimes.  I have applied non-trivial, not to be rebated time and effort to put forth what I considered to be logical views, scientifically defendable and important ideas; and yet people I knew were otherwise rational, and in some cases, very intelligent, just hadn’t seemed to get what I was saying.  Often this was within the context of a discussion argument of vaccination, but my larger concern, that of a non-imaginary, non-trivial increase in children with autism in the past decades, also usually falls on deaf ears.  If “environmental changes” incorporate the chemical milieu of our mother’s wombs, the microbial world our infants are born into, or the ocean of synthetic chemicals we all swim through every day, we have no rational conclusion but that our environment has changed a lot in the past few decades.  Considered within the context of the reality based model where the events of early life can be disproportionally amplified through the lifetime of an organism, clinging to the idea that there has been a stable incidence of autism seems dangerously naïve, at most charitable.

And yet, for the most part, many or most of the people who are alarmed are crackpots.   There were times I questioned myself.  Am I missing something?  Am I chasing phantoms?  Why aren’t any of these other smart people as worried as I am?

A while ago I got a copy of Microglia in the developing brain: A potential target with lifetime effects (Harry et all), a paper that tells me that if nothing else, I have some good company in pondering the potential for disturbances in early life to uniquely affect developmental outcome, in this instance through alterations to the neuroimmune system.  If I am incorrect about the validity of a developmental programming model with lifetime effects, lots of prolific researchers are wrong about the same thing in the same way.  Harry is a very thorough (and terrifying) review of the relevant literature.  Here is the abstract:

Microglia are a heterogenous group of monocyte-derived cells serving multiple roles within the brain, many of which are associated with immune and macrophage like properties. These cells are known to serve a critical role during brain injury and to maintain homeostasis; yet, their defined roles during development have yet to be elucidated. Microglial actions appear to influence events associated with neuronal proliferation and differentiation during development, as well as, contribute to processes associated with the removal of dying neurons or cellular debris and management of synaptic connections. These long-lived cells display changes during injury and with aging that are critical to the maintenance of the neuronal environment over the lifespan of the organism. These processes may be altered by changes in the colonization of the brain or by inflammatory events during development. This review addresses the role of microglia during brain development, both structurally and functionally, as well as the inherent vulnerability of the developing nervous system. A framework is presented considering microglia as a critical nervous system-specific cell that can influence multiple aspects of brain development (e.g., vascularization, synaptogenesis, and myelination) and have a long term impact on the functional vulnerability of the nervous system to a subsequent insult, whether environmental, physical, age-related, or disease-related.

Hell yeah!

The body of Microglia in the developing brain: A potential target with lifetime effects has tons of great stuff.  From the Introduction

The evidence of microglia activation in the developing brain of patients with  neurodevelopmental disorders(e.g., autism) and linkage to human disease processes that have a developmental basis (schizophrenia) have raised questions as to whether developmental  neuroinflammation actively contributes to the disease process. While much of the available data represent associative rather than causative factors, it raises interesting questions regarding the role of these ‘‘immune-type’’ cells during normal brain development and changes that may occur with developmental disorders. Within the area of developmental neurotoxicology, the potential for environmental factors or pharmacological agents to directly alter microglia function presents a new set of questions regarding the impact on brain development.

There is a short section on what is known about the colonization of the brain by microglia, it is a busy, busy environment, and while we are just scratching the surface, microglia seem to be involved in scads of uber-critical operations, many of which pop up in the autism literature.   It is just being confirmed that microglia constitute a distinct developmental path that diverges as an embryo, two papers from 2007 and 2010 are referenced as reasons we now believe microglia are a population of cells that migrate into the CNS before birth and are not replaced from the periphery in adulthood. From there, the beautiful complexity is in full effect; as the microglia develop and populate the brain there are specific spatial and morphological conditions, microglia are first evident at thirteen weeks after conception, and do not reach a stable pattern until after birth.   In fact, it appears that microglia aren’t done finishing their distribution in the CNS until the postnatal period, “With birth, and during the first few postpartum weeks, microglia disseminate throughout all parts of the brain, occupying defined spatial territories without significant overlap (Rezaie and Male, 2003) suggesting a defined area of surveillance for each cell.”

It occurred to me to wonder if there are differences in microglia settlement patterns in males and females in human infants, as has been observed in other models?  Could a spatially or temporally different number of micoglia, or different developmental profiles of microglia based on sex be a participant in the most consistent finding in the autism world, a rigid 4:1 male/female ratio?

Speaking towards the extremely low replacement rates for microglia in adulthood, the authors wonder aloud on the possible effects of perturbations of the process of microglial colonization.

The slow turn-over rate for mature microglia raises an issue related to changes that may occur in this critical neural cell population. While this has not been a primary issue of investigation there is limited data suggesting that microglia maintain a history of previous events. Thus, if this history alters the appropriate functioning of microglia then the effects could be long lasting. Additionally, a simple change in the number of microglia colonizing the brain during development, either too many or too few, could have a significant impact not on only the establishment of the nervous system network but also on critical  cell specific processes later in life.

(Emphasis mine)

Perhaps coincidentally (*cough*), we have abundant evidence of an altered microglial state and population in the autism population; while we do not know that these findings are the result of a disturbance during development, it is an increasingly biologically plausible mechanism, and thus far, I’ve yet to see other mechanisms given much thought, excepting the chance of an ongoing, undetected infection.

There is a brief section concerning the changes found in adult microglial populations in terms of density, form, and gene expression in different areas of the brain, “With further investigation into the heterogeneity of microglia one would assume that a significant number of factors, both cell membrane and secreted, will be found to be differentially expressed across the various subpopulations.”  Nice.

There is a section of the paper on microglial phenotypes, there are a lot of unknowns and the transformation microglia undergo between functional states is even more nebulously understood during  brain development.  “It is now becoming evident that in the developing brain, many of the standards for microglia morphology/activation may require readdressing.”  We haven’t even figured out what they’re doing in the adult brain!

There is a really cool reference for a study that shows altered microglial function dependent on the age of the organism.

In the adult rodent, ischemia can induce microglia to display either a more ramified and bushy appearance or an amoeboid morphology depending on the level of damage and distance from the infarct site(s). In the immature rodent, ischemia-induced changes in capillary flow or, presumably, altered CNS vascularization can retain the microglia in an amoeboid phenotype for longer and delay the normal ramification process (Masuda et al., 2011).

One way of looking at this would be to say that we should exercise extreme caution in trying to translate our nascent understanding of how mature microglia react when speculating on how immature microglia will act.  To follow up on just how little we know, there is a long discussion about the shortcomings of a the term ‘activated’ microglia with some details on chemical profiles of broadly generalized ‘classically inflammatory, ‘alternatively activated’, ‘anti-inflammatory’, and ‘tissue repair’ phenotypes.

Next up is a dizzyingly list of brain development functions that microglia are known, or suspected to participate in.  Without getting too deep in the weeds, of particular interest to the autism realm, that list includes neurogenesis and differentiation in the cortex [related: Courchesne, me], cell maturation via cytokine generation, axon survival and proliferation [related: Wolff, me],  programmed cell death of Purkinje cells, clearance of ‘early postnatal hippocampul neurons’, and the ‘significant contribution to synaptic stripping or remodeling events’, i.e., pruning (Paolicelli / fractaltine), and even experience dependent microglia / neuron interactions.  Taking all of this (and more) into consideration, the authors conclude “Thus, one can propose that alterations in microglia functioning during synapse formation and maturation of the brain can have significant long-term effects on the final established neural circuitry. “  Ouch.

Next up is a summary of many of the animal studies on microglial participation in brain formation, there is a lot there.  Interestingly (and particularly inconvenient) is the finding that a lot of the functional actions of microglia during development appear to operate after birth.  “Overall, the data suggest that microglial actions may be most critical during postnatal brain maturation rather than during embryonic stages of development.” Doh!

Early life STRESS gets some attention, and for once there is some good news if you look at it the right way.  There is something about a very cool study from Schwarz (et all / Staci Bilbo!) involving drug challenge that peered deep into the underlying mechanisms of an environmental enrichment model; animals given a preferential handling treatment were found by two metrics to have differential microglia response in adulthood with (biologically plausible) observations, increased mRNA levels for IL-10 production, and decreased  DNA methylation; i.e., less restriction on the gene that produces IL-10, and more messenger RNA around to pass off the production orders [totally beautiful!].  There is more including thyroid disruption (though in a way that I found surprising), and the observations of time dependent effects on immue disturbances.  (super inconvenient)

There is so much data that keeps piling on that the authors end up with “Overall, the existing data suggest a critical regulatory role for microglia in brain development that is much expanded from initial considerations of microglia in the context of their standard, immune mediated responses.”

A terrifying concept that I haven’t found time to dedicate a post towards is microglia priming, which gets some attention in Harry.

There is a significant amount of evidence regarding what is often termed ‘‘priming’’ and ‘‘preconditioning’’ events that serve to either exacerbate or provide neuroprotection from a secondary insult, respectively. In these states, the constitutive level of proinflammatory mediators would not be altered; however, upon subsequent challenge, an exaggerated response would be induced. The phenomena of priming represent a phenotypic shift of the cells toward a more sensitized state. . . Exactly how long this primed state will last has not been determined; however, data from microglia suggest that it can extend over an expanded period of time. Preconditioning can also represent changes that would occur not only over the short term but may be long lasting.”

I happen to think that microglia priming is going to be a very important cog in the machinery for this journey when all is said and done; the evidence to support a preconditioning system is strong, and in parallel, the things we see different in autism (and elsewhere) is consistent with a different set of operations of microglia, AND we also have evidence the disturbances that would invoke microglial change are subtle but real risk factors for autism.

What comes next is a type of greatest hits mashup of very cool papers on developmental programming in the CNS.

Galic et al.(2008) examined age related vulnerabilities to LPS in rats to determine critical age periods. Postnatal injection of LPS did not induce permanent changes in microglia or hippocampal levels of IL-1b or TNFa; however, when LPS was given during the critical postnatal periods, PND 7 and 14, an increased sensitivity to drug induced seizures was observed in 8-week-old rats. This was accompanied by elevated cytokine release and enhanced neuronal degeneration within the hippocampus after limbic seizures. This persistent increase in seizure susceptibility occurred only with LPS injection at postnatal day 7 or 14 and not with injections during the first day of life or at PND 20. Similar long-lasting effects were observed for pentylenetetrazol-induced seizures when PND 11 or 16 rat pups were subjected to LPS and hyperthermic seizures (Auvin et al., 2009). These results again highlight this early postnatal period as a ‘‘critical window’’ of development vulnerable to long-lasting modification of microglia function by specific stimuli. Work by Bilbo and co-workers demonstrated LPS-induced deficits in fear conditioning and a water maze task following infection of PND 4 rats with Escherichia coli. In the young adult, an injection of LPS induced an exaggerated IL-1b response and memory deficits in rats neonatally exposed to infection (Bilbo et al., 2005). Consistent with the earlier work by Galic et al. (2008), an age dependency for vulnerability was detected with E. coli-induced infection at PND 30 not showing an increased sensitivity to LPS in later life (Bilbo et al., 2006).

In particular, Galic 2008, or Postnatal Inflammation Increases Seizure Susceptibility in Adult Rats (full paper) was a very formative paper for me; it was elegant in design and showed alarming differences in outcome from a single immune challenge experience, if it occurred during a critical developmental timeframe.  If you haven’t read it, you should.

This paper has a nice way of distilling the complexity of the literature in a readable way.

One hypothesis for developmental sensitivity is the heterogeneous roles for inflammatory factors and pro-inflammatory cytokines during development, including their timing-, region and situation-specific neurotrophic properties. Many of the proinflammatory cytokines are lower at birth with a subsequent rapid elevation occurring during the first few weeks of life. In an examination of the developing mouse cortex between PND 5 and 11, mRNA levels for TNFa, IL-1b, and TNFp75 receptor remained relatively constant while a significant increase in mRNA levels of CR3, macrophage-1 antigen (MAC-1), IL-1a, IL-1 receptor 1 (IL- )R1, TNFp55 receptor (TNFp55R), IL-6, and gp130 occurred (Fig. 2). This data suggests that an upregulation of interleukins and cytokine receptors may contribute to enhanced cytokine signaling during normal cortical development.

One hypothesis put forward using a model reliant on postnatal exposure to LPS suggests that these types of exposure may ‘‘reprogram’’ neuroimmune responses such that adult stress results in hyperactivation of the hypothalamic pituitary adrenal (HPA) axis (Mouihate et al., 2010) and corticosterone  changes (Bilbo and Schwarz, 2009).While limited, the available data suggest that events occurring during development, especially postnatal development, have the  potential to cause long term alterations in the phenotype of microglia and that this can be done in a region specific manner.

[extremely inconvenient]

In what could, conceivably, be a coincidence, our available information on the autism brain also shows region specific changes in microglia populations, microglial activation profiles, and oxidative stress.   I do not believe the findings reviewed in Microglia in the developing brain: A potential target with lifetime effects will be meaningless artifacts; the likelihood that our observations of an altered neuroimmune state in autism are not, at least, participatory has become vanishingly small.

Can these findings inform us on the incidence question?  I was lurking on a thread on Respectful Insolence a while ago, and someone gave what I thought was a very succinct way of thinking about the changes that our species has encountered the past few decades; it went something like “we have replaced infection with inflammation”.  That’s a pretty neat way of looking at how things have gotten different for humanity, at least lots of us, and especially those of us in the first world.  We used to get sick and die early; now we live longer, but oftentimes alongside chronic disorders that share a common underlying biological tether point, inflammation.

Any dispassionate analysis of the available data can tell us that we have, indeed, replaced infection with inflammation; we suffer from less death and misery from infection, but more metabolic disorder, more diabetes, more hypertension, more asthma and autoimmune conditions than previous generations.   We have largely replaced good fatty acids with poor ones in our diet.  All of these conditions are characterized by altered immune biomarkers, including an increase in proinflammatory cytokines.   Those are the facts that no one can deny; we have replaced infection with inflammation.

But when we look to the findings of Microglia in the developing brain: A potential target with lifetime effects, it becomes clear that our newfound knowledge of microglial function and crosstalk with the immune system raises some very troubling possibilities.

Lately it has been quite in vogue among a lot of the online posting about autism to at least mention environmental factors which could participate in developmental trajectory leading to autism; that’s a big step, an important and long overdue acknowledgement.  If you pay close attention, you will notice that 99% of these admissions are handcuffed to the word “prenatal”.  This is likely an attempt to deflect precise questions about the robustness of our evaluation of the vaccine schedule, but the big question, the incidence question, still hinges on fulcrum of the genetic versus environmental ratio ; that is a problem for the purveyors of the fairytale because the prenatal environment of our fetuses, the chemical milieu of their development, is qualitatively different compared to generations past.  That chemical soup is their environment; and that environment has unquestionably changed in the past decades as we have replaced infection with inflammation.

Our previous analysis tells us that invoking inflammation outside the brain modifies microglial function inside the wall of the blood brain barrier; good or bad, no honest evaluation of the literature can argue against a lack of effect.  What happens outside the brain affects what happens inside the brain.  If, however, microglia are active participants in brain formation, as a swath of recent research indicates, can this fact give us insight into the incidence question?

Is a state of increased inflammation the pathway between maternal asthma, depression, stress, and obesity being associated with increased risk of autistic offspring?  Have we replaced infection with inflammation plus?

What could be more lethal to the fairytale of a static tale of autism than a positive relationship between a lifestyle characterized by increased inflammation and the chances of having a baby with autism?

Are we totally fucked?

We cannot know the answers unless we have the courage to ask the difficult questions with methods powerful enough to provide good data, and it won’t be easy.  The static rate of autism fairytale is a comforting notion; it expunges responsibility for the coronal mass ejection sized change to our fetuses developing environment, and while hiding behind the utterly frail findings of social soft scientists, we can happily place tin foil hats and accusations of scientific illiteracy on anyone who might be worried that our abilities have outstripped our wisdom.  That is a terrible, cowardly way to approach the incidence question, what we should be doing is exactly the opposite, ridiculing the epidemic sized error bars in prevalence studies and demanding more answers from the hard scientists.  Eventually we will get there and it will be a critical mass of information from studies like Harry that will propel decision makers to abandon the fairytale for a course regulated by dispassionate analysis.

–          pD

Hello friends –

A study with a beautifully terse title, Microglia in the Cerebral Cortex in Autism landed in my inbox the other day.  It adds to the growing literature showing perturbations in neuroimmune system in the autism population, this time by measuring the number of microglia in different parts of the brain.  Here is the abstract:

We immunocytochemically identified microglia in fronto-insular (FI) and visual cortex (VC) in autopsy brains of well-phenotyped subjects with autism and matched  controls, and stereologically quantified the microglial densities. Densities were determined blind to phenotype using an optical fractionator probe. In FI, individuals with autism had significantly more microglia compared to controls (p = 0.02). One such subject had a microglial density in FI within the control range and was also an outlier behaviorally with respect to other subjects with autism. In VC, microglial densities were also significantly greater in individuals with autism versus controls (p = 0.0002). Since we observed increased densities of microglia in two functionally and anatomically disparate cortical areas, we suggest that these immune cells are probably denser throughout cerebral cortex in brains of people with autism.

[Note: You don’t see p-values of .0002 too often!]  This paper is at a high level largely similar to another recent paper, Microglial Activation and Increased Microglial Density Observed in the Dorsolateral Prefrontal Cortex in Autism (discussed on this blog, here).  The authors were clever here, they intentionally used two very anatomically different, and spatially separated parts of the brain to evaluate for microglia population differences, a sort of bonus slice to learn more about the population of microglia in the brain.

The specific measurement technique in use, staining for specific antibodies, does not give us information regarding the activated/non activated state of the microglia, a determination which must be made with evaluations of morphology, though several other studies have measured this directly, and many more provide indirect evidence of a chronic state of activation of microglia.   Not only did the author s report an increase in population density in the autism group, the number of microglia was also positively correlated between sites; i.e., a patient with more microglia in the visual cortex was also more likely to have more microglia in the fronto-insular.

These findings demonstrate that, at the time of death, there were significantly higher microglial densities in the subjects with autism compared to the control subjects, and that this change in microglial density is widespread throughout the cerebral cortex in autism. The microglial  densities in FI and VC in the same subject were significantly correlated (both measures were available in 10 controls and 8 autistic subjects for a total of 18 subjects) with Pearson’s r2 = 0.4285, p = 0.0024 (Fig. 6). This indicates that the elevation in density is consistent between these areas, and probably throughout the cortex, in both subjects with autism and controls.

Also of interest, in the control group microglia densities tended to decrease with age, but this change was not seen in the autism population.

There is some discussion about a big problem in the autism research world, a very real and meaningful dearth of available tissue samples, this study shared five patients with Morgan, and one from Vargas.  [Note: Sign up to help.  Morbid but necessary.]

The authors went on to ask the exact same question I had, “How and when does the increased density of autistic microglial arrays arise, and how is it maintained?”  Unfortunately, while there aren’t any good answers, I was still a little disappointed with the analysis.  There is a quick rundown of a variety of neuroimmune and peripheral immune findings in autism, and some thoughts on ‘sickness behavior’ with the implicit interconnectedness of the immune state and behaviors, and some discussion on some of the many animal models of maternal immune activation in autism.

In an stroke of amazing serendipity, the authors wonder aloud towards the possibility of a type of distracted worker effect of microglia on neural networks, sort of a bank shot on the autism paradox I struggled with in my previous post when I said,

Are increased neuron number and altered white matter tracts the result of microglia not performing the expected maintenance of the brain?  Are the findings from Courchesne and Wolff the opportunity costs of having a microglia activated during decisive developmental timeframes?

The authors of Microglia in the Cerebral Cortex in Autism state

In contrast, microglia can also phagocytize synapses and whole neurons, thus disrupting neural circuits. For example,when the axons of motor neurons are cut, the microglia strip them of their synapses (Blinzinger and Kreutzberg 1968; Cullheim and Thams 2007; Graeber et al. 1993). Another example of the disruption of circuitry arises from the direct phagocytosis of neurons. Neurons communicate with microglia by emitting fractalkine*, which appears to inhibit their phagocytosis by microglia. Deleting the gene for the microglial fractalkine receptor (Cx3cr1) in a mouse model of Alzheimer’s disease has the effect of preventing the microglial destruction and phagocytosis of layer 3 neurons that was observed in these mice in vivo with 2-photon microscopy (Furhmann* et al. 2010). In particular, Cx3cr1 knockout mice have greater numbers of dendritic spines in CA1 neurons, have decreased frequency sEPSCs and had seizure patterns which indicate that deficient fractalkine signaling* reduces microglia-mediated synaptic  pruning, leading to abnormal brain development, immature connectivity, and a delay in brain circuitry in the hippocampus (Paolicelli* et al. 2011). In summary, the increased density of microglia in people with autism could be protective against other aspects of this condition, and that a possible side-effect of this protective response might involve alterations in neuronal circuitry.

Oh hell yeah.  (* concepts and papers discussed on this blog, here)

Going back to the big dollar question, How and when does the increased density of autistic microglial arrays arise, and how is it maintained?”, the possibility of an ongoing infection was raised as a one option, “The increase of microglial densities in individuals with autism could be a function of neuroprotection in response to harmful microorganisms.”  Vargas had a dedicated section towards a failure to find agents of the peripheral immune system that are consistent with infiltration from the peripheral immune system commonly observed during acute infection, I do not think other papers have looked for that per se, but will cede to someone with better data.  (?)   There was a very weird paper from Italy that pointed to a possible polyomavirus transmission from the father in the autism group, though this study was not referenced in Microglia in the Cerebral Cortex in Autism. [Note:  I showed my wife this paper, and she told me, “Good job with the autism gametes.”  Nice.]  Could a virus cause autism, is a nice discussion on this that includes blog and personal favorites, Fatemi, Patterson, and Persico discussing the possibilities and limitations of the study.  Great stuff!

While I must admit the possibility that the chronically activated microglia in autism are working on purpose, the irony gods mandate that I wonder aloud if certain segments of the autism Some-Jerk-On-The-Internet population will cling to the possibility that autism is caused by a disease in order to disavow a causative role for neuroinflammation?  Those are some tough choices.

There is a discussion on the myriad of ways that microglia could directly participate in autism pathogenesis, starting the discussion off right to the point, “By contrast, there are diseases that arise from intrinsic defects in the microglia themselves which can cause stereotypic behavioral dysfunctions.”  There is a short discussion of Nasu-Hakola disease, something I’d never heard of, which has evidence of an increase in cytokines as a result of genetically driven microglial deficiencies, and shows striking behavioral manifestations.

The possibility of some areas of the brain being more susceptible to alterations than others is there too, “Thus, while changes in microglial density appear to be widespread in brains of autistic individuals, some areas may be more vulnerable than others to its effects.”   Considering this idea alongside the extremely heterogeneous set of symptoms assigned to autism, a curious question to ponder becomes; if neuroinflammation is a participatory process in the behavioral manifestation of autism, could some of the variability in autistic behaviors be explained by spatially specific gradients of microglial activity?  Going further, considering the still largely mysterious migration of microglia into the brain during development, could the temporal origin of microglial activation in autism be a determinant in the eventual behavioral manifestations?  These are tricky questions, and I don’t think that our current methodological capacities are sufficient to start thinking about forming a model for analysis.

One concept I was surprised to not receive attention was a developmental programming model, where animal studies tell us that if something happens during critical developmental timeframes, the effect can propagate into adulthood.    In fact, one study, Enduring consequences of early-life infection on glial and neural cell genesis within cognitive regions of the brain (Bland et all)  exposed four day old animals to e-coli, which found, among other things, “significantly more microglia in the adult DG of early-infected rats”, something seemingly of considerable salience to the current findings, especially considering the known risk factors of early infections as autism risk factors.  In Bland, no external agent other than an infection during early life was necessary; this is the essence of the developmental programming model, even after the infection was long since cleared, patterns of physiology were imprinted, the animals recovered from e-coli but were changed from the experience.  This my biggest issue with the possibility of an as of yet undefined, and continued evidence free pathogen or process that is causing the immune abnormalities we see in autism, it mandates we ignore existing biologically plausible models that fit well within known risk factors for autism.  Why?

Another area this paper was curiously silent on is the data regarding differences in males and females in the timeframes of microglial migration into the brain, something I’d like to learn much more about soon.  As an example, Sex differences in microglial colonization of the developing rat brain [yet another by blog favorite, Staci Bilbo] reported “the number and morphology of microglia throughout development is dependent upon the sex and age of the individual, as well as the brain region of interest” among other findings broadly consistent with a beautiful complexity.  This is interesting fodder for a discussion concerning possibly the most persistent finding in autism, a very high male to female ratio that has a series of possible explanations [somewhat discussed on this blog, here].

So we know more, but still have only increased our knowledge incrementally.  It is increasingly likely that an increased number of microglia in many areas of the brain is characteristic of autism, but the whys, hows, whens, wheres, and whoms still hold many mysteries.  The more things change, the more they stay the same.

–          pD

Hello friends –

I’ve had a couple of interesting papers land in my pubmed feed the past few weeks that seem to be tangentially touching on something that has been at the back of my mind for a long time; namely, the repeated findings of a state of an ongoing immune response in the CNS of the autism population, coupled with a behavioral state that is either static, or in many cases, showing gradual improvement over time.  [Discussions of ongoing immune response in the brain in autism, here, here, or here].  This is exactly the opposite of what I expected.  Most of the conditions I had generally associated with a state of neuroinflammation, i.e., Alzheimer’s or Parkinson’s show a behavioral profile opposite to autism over time, i.e., a deterioration of skills and cognitive abilities.   The diagnosis for these conditions is never a straight line or a gradual curve upwards, but a dispassionately reliable trajectory of a downward spiral.

This is something that has been really bugging me a lot as a riddle, I’ve mentioned it here in comments, and other places on the Internet.  While outright signs of neuroinflammation are clearly associated with conditions you would rather not have, as opposed to have, we must admit that the available evidence tells us that  we cannot just wave our hands, say ‘neuroinflammation!’, and know much more than the broad strokes.  [Note: In my early days of my AutismNet life, my view was somewhat less nuanced.]  I think that part of what was bothering me is the result of an oversimplified model in my mind’s eye, but I’d formed that model on top of a set of measurements that had empirical precision but underpowered understandings, alongside a more fundamental lack of knowledge.

We know a little more now.

The first paper that really got me thinking along these lines was Synaptic pruning by microglia is necessary for normal brain development, (discussed on this blog, here), which provided evidence of microglial involvement in the ‘pruning’ of synapses, an important step in brain development thought to streamline neural communication by optimizing neuron structure.  This was the first paper I’d read that hinted at microglia participation in ‘normal’ brain function; it was only very recently that microglia were considered to have any role in non pathological states.  Another paper, Microglial Cx3cr1 knockout prevents neuron loss in a mouse model of Alzheimer’s disease, also implicated microglia in synaptic pruning.

Then I got myself a copy of The role of microglia at synapses in the healthy CNS: novel insights from recent imaging studies. It is a review of several recent studies on the non-excited life of microglia.

In the healthy brain, quiescent microglia continuously remodel their shape by extending and retracting highly motile processes. Despite a seemingly random sampling of their environment, microglial processes specifically interact with subsets of synaptic structures, as shown by recent imaging studies leading to proposed reciprocal interactions between microglia and synapses under non-pathological conditions. These studies revealed that various modalities of microglial dynamic behavior including their interactions with synaptic elements are regulated by manipulations of neurotransmission, neuronal activity and sensory experience. Conversely, these observations implied an unexpected role for quiescent microglia in the elimination of synaptic structures by specialized mechanisms that include the phagocytosis of axon terminals and dendritic spines. In light of these recent discoveries, microglia are now emerging as important effectors of neuronal circuit reorganization.

This review by Tremblay was published in 2012, evidence of the nascent nature of our available data on microglial involvement in the normal brain environment; Tremblay states that part of the reason this type of finding is so recent is the relative difficulty of measuring microglia in non excited states.  They were the electrons of brain measurements; our previous attempts to measure them were capable of causing them to change morphology.

The roles of ‘resting’ or immunologically quiescent microglia have remained relatively unknown (also see Tremblay et al., 2011). This is largely due to the difficulties of studying microglia in their non-activated state. Microglia respond promptly to any changes occurring in their environment, and therefore experimental ex vivo and in vitro preparations inevitably result in transformation of their normally prevailing behavior.

Nice.

Anyway, some new whizbang technologies (i.e., in vivo two-photon laser scanning microscopy)[?] are allowing researchers to peer into the ho-hum everyday activities of ‘non activated’ microglia, and what they are finding is that the term ‘activated microglia’ might be a bit of a misnomer, microglia have been participating in brain function all along, it is just that our filters were insignificantly powered to detect some of their actions until very recently.   Several studies have shown that so called ‘resting’ microglia are constantly evaluating their environment with protusions that seemed to operate rather quickly in relationship to other types of neurons.

This unexpected behavior suggested that resting or surveillant microglia may continuously survey the brain parenchyma as part of their immune function, which would justify the substantial expenditure of energy required to continuously maintain microglial dynamics in the normal brain, without excluding the possibility of an additional, distinct contribution to normal brain physiology

Several papers are reviewed that utilized a couple of highly technical methods, including double roll your own transgenic mouse models to visualize the interactions of microglia in a non excited state and synapses.  Specific areas of the brain were measured in different studies, microglia were observed transiently engaging with neurons and seemed to target some dendrites for removal.  The authors speculate that this could be a mechanism by which neuronal network maintenance, plasticity, could be affected.

In the mature healthy CNS, neuronal networks are continuously remodeled through the formation, modification and elimination of synaptic structures (see Fortin et al. (2011) for molecular mechanisms of structural plasticity) in relation with behavioral and sensory experience.

And

To determine a possible role of surveillant microglia in the structural remodeling of synaptic structures under normal physiological conditions, Tremblay et al. (2010b) also examined the size changes of spines and terminals before, during and after microglial contacts. Spines contacted by microglial processes during imaging (30–120 min sessions) were found to be smaller initially than those which remained non-contacted. Spines, but not terminals, also underwent transient increases in size during microglial contact, with smaller spines showing the most pronounced changes. Surprisingly, chronic imaging over 2 days further revealed a statistically significant difference in the elimination rate of microglia-contacted spines: spines contacted by microglia were more frequently eliminated than non-contacted spines (24 versus 7%; P  0.05), and in all cases, only the small spines were seen to disappear. These observations suggest that despite an apparently random sampling of the parenchyma, microglial processes specifically target a subset of small, structurally dynamic and transient dendritic spines.

There is also some description of studies that seemed to indicate that the microglial/synapse interactions could be modified through environmental stimulus, two experiments were described involving sensory deprivation and consequent changes in microglia activity.  Other experiments described changes in microglial surveillance as a result of induced changes in neuronal excitability by chemical agonists or antagonists of glutamate receptors.  [Perhaps this is the basis of the curious findings in Neuroprotective function for ramified microglia in hippocampal excitotoxicity?]

In their concluding statements, Tremblay provides a good description of just how little we know, and in a style that I love, poses open questions for the newer rounds of literature to address.

Since the recent studies have barely scratched the surface (of the brain in this case), the modalities of microglial interactions with excitatory and inhibitory synapses throughout the CNS, much as their functional significance and particular cellular and molecular mechanisms still remain undetermined. For example, in which contexts do quiescent microglia directly phagocytose axon terminals and dendritic spines, use other mechanisms such as proteolytic remodeling of the extracellular space, or refrain from intervening?  How do surveillant microglia recognize and respond to the various molecular signals in their environment, including dynamic changes in neurotransmission and neuronal activity at individual synapses? How do these immune cells cooperate with other glial cells, as well as peripheral myeloid cells, in maintaining or shaping neuronal architecture and activity? And, as in the case of microglial memory of past immune challenges (see Bilbo et al., 2012), do surveillant microglia somehow remember their previous behavioral states, the flux of information processing in the brain, or the structural changes of synaptic elements in recent and not so recent windows of intervention?

The last sentence there, I think, is especially salient considered within a context of developmental programming.

So what we’ve learned is that decades after the discovery of microglia cells as the immune regulators in the CNS, they appear to also be participating in more fundamental maintenance of the neural structure of our brains; there is increasing evidence of direct relationships in synaptic and axonal removal as well as roles in neurotransmission and the regulation of excitability.   Is more on the horizon?

But what about autism and our apparent autism paradox of a static or improving behavioral state alongside conditions of immune activation within the CNS?

Well, I have also been thinking about two brain scanning studies that have come out not too long ago, Neuron Number in Children With Autism (Courchesne et all) , which found increased numbers of neurons in the autism cohort, and Differences in White Matter Fiber Tract Development Present From 6 to 24 Months in Infants With Autism (Wolff et all) which found that the autism group showed denser bundled of white matter, so called wiring, between different parts of the brain.  In both of these studies mention is made of the fact that it was possible that their findings, increased cell numbers could be the result of inappropriate removal of excess neurons during development.

Apoptotic mechanisms during the third trimester and early postnatal life normally remove subplate neurons, which comprise about half the neurons produced in the second trimester. A failure of that key early developmental process could also create a pathological excess of cortical neurons.

and

For example, differences in structural organization prior to a period of experience-dependent development related to social cognition (52–54) may decrease neural plasticity through limitations on environmental input, preventing typical neural specialization (52). These alterations could have a ripple effect through decreasing environmental responsiveness and escalating invariance*, thus canalizing a specific neural trajectory that results in the behavioral phenotype that defines ASDs. In typical development, the selective refinement of neural connections through axonal pruning (55) along with constructive processes such as myelination (56) combine to yield efficient signal transmission among brain regions. One or both of these mechanisms may underlie the widespread differences in white matter fiber pathways observed in the current study. 

* 😦

So, we have growing evidence of microglial participation of neural maintenance alongside growing evidence of impaired maintenance in the autism cohort.

Can our autism paradox be explained by microglia converging in the center of these related lines of thought?  Is the answer to our riddle that the ongoing immune response in the brain is not sufficiently powered, or targeted, to cause increasing loss of abilities, but instead, was enough to keep critical, once in a lifetime chances for brain organization from occurring?  Are increased neuron number and altered white matter tracts the result of microglia not performing the expected maintenance of the brain?  Are the findings from Courchesne and Wolff the opportunity costs of having a microglia activated during decisive developmental timeframes?

That is a pretty neat idea to consider.

Even without the Courchesne and Wolff, the findings that specifically mention impaired network maintenance as possible culprits, the findings of active participation of ‘non-active’ microglia in brain optimization and normal processes is a very problematic finding for another autism canard, the idea that findings of neuroinflammation may not be pathological.  The intellectually honest observer will admit that the crux of this defense lay in vaccine count trial testimony presented by John Hopkin’s researchers after their seminal neuroinflammation paper was published.  Unfortunately, the vigor with which this testimony is trotted out online does not match the frequency with which such ideas actually percolate into the literature.

But with the data from Tremblay, Paolicelli, and others, such an idea becomes even more difficult to defend, we must now speculate on a mechanism by which either microglia could be in an excited state and continue to perform streamlining of the neural structure, or insist that it is possible that microglia were not excited during development, and something else happened to interfere with neuron numbers, and then, subsequently the microglia became chronically activated.

This is unlikely, and unlikelier still when we consider that anyone proposing such a model must do so with enough robustness to overcome a biologically plausible pathway supported by a variety of studies.  And that is only if there was anything underneath the vapor!  Make no mistake, if you ever press someone to actually defend, with literature, the mechanisms by which a state of chronic neuroinflammation might be beneficial in autism, or even the result of something else that also causes autism, no further elucidation of that mechanism is ever forthcoming.  There isn’t anything there.

At some point, it becomes incumbent of people wishing to make an argument that they propose a biologically plausible mechanism if they wish to continue to be taken seriously.  If they cannot, if the literature cannot be probed to make such a case with more empirical support than it might be, the notion so add odds with available evidence should be summarily discarded, unless and until a transcendent set of findings is presented.  There should always be room for more findings in our worldview, but precious limited space for faith in the face of contradictory findings.

–          pD

Hello friends –

Lately I’ve found myself reading papers and knowing and owning several of the references; tragically I can’t tell if I’m reading the right research and am onto something, or I am chasing phantoms and my web of pubmed alerts and reading interests are funneling my reference list into a narrowing echo chamber of sorts.   With that warning in mind, we can proceed to poking around several papers, only some of which mention autism per se.  Along the way, we will see evidence supporting the possibility of a biologically plausible mechanism of developmental programming of the neuroimmune environment, a sequence of events that may lead to impaired synaptic pruning in (some cases of?) autism.

By now, everyone has seen/read/heard about one form or another of the ‘a massive asteroid is going to destroy the world’ story.  One of the common survival strategies from an asteroid strike involves altering the path of the asteroid so that it misses the Earth.  The thoughtful analysis of this problem allows for the physics based reality of the problem, moving an asteroid out of an extinction based trajectory involves just a little work when the asteroid is ten thousand gazillion miles away, but a lot more work when it is only a gazillion miles away.  Upon careful evaluation living organisms display similar behavior, relatively minor disturbances in early life can alter the developmental trajectory, while that same disturbance later in life is unable to materially affect the organism beyond a transient effect.   The accumulated evidence that early life experiences can shape the adult outcome is nearly impossible to dispute with any remaining intellectual honesty, the question is instead, is how large is the effect in autism?

This analogy adequately symbolizes one of the more beautiful and terrifying concepts I’ve come across researching autism, that of ‘developmental programming’, which I blogged some about here, but essentially is the idea that there are critical timeframes during which environmental impacts can have long term persistent effects on a wide range of outcomes.  The most robustly replicated findings involve changes to metabolic profiles in response to abnormal prenatal nutritional environments, but there is also evidence of various other effects, including neurological, and reputable speculation, that autism, may in fact, be in part, a disorder of developmental programming.

Secondarily, there has long been speculation of problems in the removal of ‘excess’ synapses, i.e., ‘synaptic pruning’ in the autism population.   This culling of synapses begins in fetal life continuing throughout adolescence and the repeated observations of increased head circumference during infancy as a risk factor for autism has resulted in the idea that altered synaptic pruning maybe involved in autism.

In the last month or so several rather serendipitously themed papers have been published with tantalizing clues about some of the finer grained mechanisms of synaptic pruning, the possibility of impaired synaptic pruning in the autism population, and a known risk factor for autism that models a developmental programming event sequence that may tie them together.

First off, we have Synaptic pruning by microglia is necessary for normal brain development, (Paolicelli et all) with a very straightforward title, that has this dynamite in the abstract: (snipped for length)

These findings link microglia surveillance to synaptic maturation and suggest that deficits in microglia function may contribute to synaptic abnormalities seen in some neurodevelopmental disorders.

This paper is short, but pretty cool, and very nice from a new territory perspective.  It also speaks directly towards one of the increasingly hilarious obfuscations you will sometimes see raised in online discussions about immunological findings in autism, namely, that we can’t know if the state of chronic inflammation in the CNS observed in autism is harmful or beneficial.   [hint: It might not be causative, but it isn’t beneficial.]

Here’s is a snippet from the Introduction:

Time-lapse imaging has shown that microglia processes are highly motile even in the uninjured brain and that they make frequent, but transient contact with synapses. This and other observations have led to the hypothesis that microglia monitor synaptic function and are involved in synapse maturation or elimination.  Moreover, neurons during this period up-regulate the expression of the chemokine fractalkine, Cx3cl1, whose receptor in the central nervous system is exclusively expressed by microglia and is essential for microglia migration. If, in fact, microglia are involved in scavenging synapses, then this activity is likely to be particularly important during synaptic maturation when synaptic turnover is highest.

Nice.  A time dependent participation by microglia in the critical process of optimization of neuron numbers, a process we are still very much groping our way in the dark towards untangling.  The researchers focused in on a particular molecular target, a chemical messenger of the immune system, fractalkine, and found that without fractalkine, the process of synaptic turnover was impaired.

A couple of tests were performed, first immunohistochemistry (i.e., exceedingly clever manipulation of antibodies to determine the presence or absence of proteins in very specific locations) which demonstrated that microglia were, in fact, ‘engulfing synaptic material’ in animals during periods of synaptic maturation.

Secondly, so called ‘knock out mice’ (i.e., genetically engineered mice constructed without the ability to make a specific protein, in this case, fractalkine) were used evaluate for changes in synaptic form and function based on a lack of fractalkine.  Changes in dendritic spine density were observed in the knock out mice group, with much higher densities in a very specific type of neuron during the second and third postnatal week of life.  The authors indicate this is a key timeframe in synaptic pruning, and state their findings are “suggesting a transient deficient synaptic pruning in Cx3cr1 knockout mice “.  The effect of not having fractalkine on spine density was time dependent as shown below.

Several other measurements were taken, including synaptic firing frequencies, which also implicated an increased surface area for synapses on dendritic spines, consistent with impaired pruning.  Time dependent effects on synaptic efficiency and seizure susceptibility were also found, which the led the authors to conclude that the findings were “consistent with a delay in brain circuit development at the whole animal level.”

For additional evidence of fractalkine participation in synaptic maintenance, we can look to the opposite direction, where researchers evaluating neuron loss in an Alzheimers model reported “Knockout of the microglial chemokine receptor Cx3cr1, which is critical in neuron-microglia communication, prevented neuron loss”.  Taken together, the conclusion that fractalkine processing is involved with neuron maintenance is highly likely, and correspondingly, highly unlikely to be a set of spurious findings.

There’s a couple paragraphs on potential mechanisms by which fractalkine could be interacting with microglia to achieve this effect, with the authors claiming that their data and other data generally supports a model wherein microglia were not effectively recruited to appropriate locations in the brain due to a lack of fractalkine, or, a ‘transient reduction in microglia surveillance.’

The conclusion is a good layman level wrap up that speaks toward the Interconnectedness of the brain and the immune system:

In conclusion, we show that microglia engulf and eliminate synapses during development. In mice lacking Cx3cr1, a chemokine receptor expressed by microglia in the brain, microglia numbers were transiently reduced in the developing brain and synaptic pruning was delayed. Deficient synaptic pruning resulted in an excess of dendritic spines and immature synapses and was associated with a persistence of electrophysiological and pharmacological hallmarks of immature brain circuitry. Genetic variation in Cx3cr1 along with environmental pathogens that impact microglia function may contribute to susceptibility to developmental disorders associated with altered synapse number. Understanding  microglia-mediated synaptic pruning is likely to lead to a better understanding of synaptic homeostasis and an appreciation of interactions between the brain and immune system

That’s all pretty cool, but there was precious little discussion of autism, except in the general sense of a ‘developmental disorder associated with altered synapse number’.   [But the references do speak to autism, the first reference provided, Dendritic Spines in Fragile X Mice displays a significant relationship to autism, and it describes how another flavor of knock out mice, this time designed to mimic Fragile-X, exhibit a ‘developmental delay in the downregulation of spine turnover and in the transition from immature to mature spine subtypes.’  Go figure!]

The other reason Paolicelli is of particular interest to the autism discussion is one of the major players in this study, the microglia (i.e., the resident immune cells of the CNS), have been found to be ‘chronically activated’ in the autism brain by direct  measurement in two studies (here, and here, [and by me, here]), and tons of other studies have shown indirect evidence of an ongoing state of immunological alertness in the autism brain.

Considering this is a brand new paper, I do not believe that there are any studies illuminating the results of a state of chronic activation of microglia on the process of synaptic pruning per se.  I will, however, go on the record that such an effect is very, very likely, and the logical leap is microscopically small that there will be some detrimental impact to such a state.  The inverse argument, a scenario wherein there could be a state of chronic microglial activation that does not interfere with microglia participation in the synaptic pruning requires logical acrobatics worthy of Cirque Du Soleil.  I am open to evidence, however.

So, from Paolicelli, we know that a ‘transient reduction in microglial surveillance’ induced by a reduction in the ability to production fractalkine can result in a condition ‘consistent with a delay in brain circuit development at the whole animal level’.

Next up, we have a paper that was all over the JerkNet in the days and weeks following its release, Neuron number and size in prefrontal cortex of children with autism.  This is a cool study, and likely a very important paper, but I must say that a lot of the online commentary exhibits an irrational exuberance towards one part of the findings.   Here is part of the abstract.

Children with autism had 67% more neurons in the PFC (mean, 1.94 billion; 95% CI, 1.57-2.31) compared with control children (1.16 billion; 95% CI, 0.90-1.42; P = .002), including 79% more in DL-PFC (1.57 billion; 95% CI, 1.20-1.94 in autism cases vs 0.88 billion; 95% CI, 0.66-1.10 in controls; P = .003) and 29% more in M-PFC (0.36 billion; 95% CI, 0.33-0.40 in autism cases vs 0.28 billion; 95% CI, 0.23-0.34 in controls; P = .009). Brain weight in the autistic cases differed from normative mean weight for age by a mean of 17.6% (95% CI, 10.2%-25.0%; P = .001), while brains in controls differed by a mean of 0.2% (95% CI, -8.7% to 9.1%; P = .96). Plots of counts by weight showed autistic children had both greater total prefrontal neuron counts and brain weight for age than control children.  [PFC == prefrontal cortex]

Essentially the authors used a variety of mechanisms to measure neuron number in a specific area of the brain, the prefrontal cortex, and found large variations (increases) in the autism group.   The prefrontal cortex is thought to be involved in ‘planning complex coginitive behaviors’, and ‘moderating correct social behavior’, among others, so this was a smart place to look.

The implicit hype on the internet is that this firmly indicates a ‘prenatal cause’ to autism, but if you read the paper, read what Courchense has said, and read recent literature, you know that the simplicity of this as a singular prenatal cause of autism is long broad strokes, and short on appreciation of the subtlety that textures reality.

A link @ LBRB sent me to the team at The Thinking Person’s Guide To Autism, who had a very nice transcription of a talk given by Courchesne at IMFAR 2011.  Here is a snipet that started my wheels turning.

What we see in autism is either an excess proliferation, producing an overabundance of neuron numbers, or the excess might be due to a reduced ability to undergo naturally occurring cell death. Or it could be both. We don’t know which and our data don’t speak to that, although our data do suggest that it’s probably both.

Finally, our evidence shows that across time, there’s a prolonged period of apoptosis, removal and remodeling of circuits. In order to get back to where neuron numbers are supposed to be, it takes a very long time for the autistic brain. In the normal developing brain, this takes just a few months. In autism, it’s a couple of decades.

[Note how well this fits within the model described by Paolicelli, i.e., “consistent with a delay in brain circuit development at the whole animal level”.  ]

I would highly recommend anyone who has read this far to go read the entire post @ TPGTA sometime.

As far as synaptic pruning goes, here is the associated segment of the paper:

Apoptotic mechanisms during the third trimester and early postnatal life normally remove subplate neurons, which comprise about half the neurons produced in the second trimester. A failure of that key early developmental process could also create a pathological excess of cortical neurons. A failure of subplate apoptosis might additionally indicate abnormal development of the subplate itself. The subplate plays a critical role in the maturation of layer 4 inhibitory functioning as well as in the early stages of thalamocortical and corticocortical connectivity development.inhibitory functioning and defects of functional and structural connectivity are characteristic of autism, but the causes have remained elusive.

Nearly half of the neurons in the area studied are expected to be removed through pruning, a process that extends well after birth.  That is something that you didn’t see referenced in too many places trumpeting this study as ‘proof’ that autism was caused by disturbances in the prenatal environment.  I’m not coming down on the prenatal environment as a critical timeframe for autism pathogensesis, just the difficult to defend underlying notion that this is the only time the environment should be evaluated, or the idea that if something is initiated prenatally other timeframes are therefore, unimportant.

So, I’d read that microglia were actively involved in proper synaptic pruning, contingent on utilization of fractalkine, and then read that impaired synaptic apoptotic mechanisms could be participating in autism, with a consequence of an over abundance of neurons.

Then, I got myself a copy of Microglia and Memory: Modulation by Early-Life Infection, which is another study in a growing body of evidence that immune challenges early in life can have unpredictable physiological consequences.  (This is another very cool paper with Staci Bilbo as an author, whom I think is seriously onto something.)  This study, in particular, focused on interactions microglia and formation of memories.   Here is the abstract:

The proinflammatory cytokine interleukin-1ß (IL-1ß) is critical for normal hippocampus (HP)-dependent cognition, whereas high levels can disrupt memory and are implicated in neurodegeneration. However, the cellular source of IL-1ß during learning has not been shown, and little is known about the risk factors leading to cytokine dysregulation within the HP. We have reported that neonatal bacterial infection in rats leads to marked HP-dependent memory deficits in adulthood. However, deficits are only observed if unmasked by a subsequent immune challenge [lipopolysaccharide (LPS)] around the time of learning. These data implicate a long-term change within the immune system that, upon activation with the “second hit,” LPS, acutely impacts the neural processes underlying memory. Indeed, inhibiting brain IL-1ß before the LPS challenge prevents memory impairment in neonatally infected (NI) rats. We aimed to determine the cellular source of IL-1ß during normal learning and thereby lend insight into the mechanism by which this cytokine is enduringly altered by early-life infection. We show for the first time that CD11b+ enriched cells are the source of IL-1ß during normal HP-dependent learning. CD11b+ cells from NI rats are functionally sensitized within the adult HP and produce exaggerated IL-1ß ex vivo compared with controls. However, an exaggerated IL-1ß response in vivo requires LPS before learning. Moreover, preventing microglial activation during learning prevents memory impairment in NI rats, even following an LPS challenge. Thus, early-life events can significantly modulate normal learning-dependent cytokine activity within the HP, via a specific, enduring impact on brain microglial function.

Briefly, the authors infected rats four days after birth with e-coli, and then challenged them with LPS in adulthood to simulate the immune system to evaluate if memory formation was affected.   As further evidence of an immune mediated effect, prevention of microglial activation in adulthood was sufficient to attenuate the effect.  Clearly the effect on memory formation was based on the immune system.  (Note:  Most of the studies I’ve read would indicate [i.e., educated guess] that a four day old rat is brain developmentally similar to the third trimester of a human fetus.)  While a terrifying and beautiful expression of developmental programming in its own right, there isn’t much to speak towards synaptic pruning in this paper, except maybe, potentially, one part of their findings.

In our study, CX3CL1 did not differ by group, whereas its receptor was decreased basally in NI rats, implicating a change at the level of microglia.

This is where things get either highly coincidental, or connected.  CX3CL1 is another name for fractalkine, i.e., animals that were infected in early life had decreased expression of the receptor for fractalkine compared to placebo animals, i.e., fractalkine is the same chemical messenger found to be integral in the process of synaptic pruning in Synaptic pruning by microglia is necessary for normal brain development!  From a functionality standpoint, having less receptor is very similar to having less fractalkine; as the animals in Microglial Cx3cr1 knockout prevents neuron loss in a mouse model of Alzheimer’s disease tell us.

If, if synaptic apoptotic processes are impaired in autism, perhaps this is one mechanism of action. The timeline would involve a prenatal immune challenge, which causes a persistent decrease fractalkine receptor expression, which in turn, causes a consequent impairment in synaptic pruning through interference in microglial targeting.  There is near universal agreement that immune disturbances in utero are capable of altering developmental trajectory undesirably, and here, in an animal model, we have evidence that infections are capable of reducing availability of receptors of ligands known to play a critical role in synaptic pruning, the absence of which leads to conditions which are “consistent with a delay in brain circuit development at the whole animal level”. 

Only time, and more research, will tell if this is a pattern, a phantom, or a little of both.

–          pD



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