Other interesting Probiotic Bacteria for Cholesterol, Osteoporosis, Diabetes, Eczema, Asthma, Cancer and perhaps some Autism

  

In the next 30 to 50 years I think many common diseases will be, in part, treated by bacteria.  There is already a great deal of research to show that gut bacteria play a key role in both some diseases and the effectiveness of some therapies.

I was surprised to read that the effectiveness of some common existing cancer drugs appears to depend on the presence, or not, of specific gut bacteria.

Many gut bacteria have very specific, but different, effects on the immune system.  There may be no one-size-fits-all options and it is not a case of good bacteria and bad bacteria.  Too much of some “good” bacteria and they becomes “bad” bacteria.

Taking a pragmatic approach you can look at the effects of widely available probiotic bacteria and see if any might have a beneficial effect on a specific person’s autism.

We already saw in the trials that people made following Alli from Switzerland’s revelation about the two L.reuteri bacteria found in Biogaia Gastrus, that what is good for one person might not be effective in the next person.

In my case one of the L.reuteri bacteria in Biogaia Gastrus has a profound positive effect on allergy, and hence autism, while the second bacteria has negative behavioral effects.  Fortunately, the L.reuteri protectis bacteria in Biogaia Gastrus can be purchased separately.

Not surprisingly, companies are patenting the bacteria with research-proven therapeutic effects.  Many supplement companies are using the non-patented bacteria because they are cheaper.  Very often they do not specify exactly which sub-type of bacteria they use and you have no means of knowing whether they change the bacteria over time depending on pricing and availability.

Nonetheless if you skim through the probiotic bacteria research and anecdotal evidence there are some interesting options.

 

First a quick recap

So far in this blog we have seen some particularly interesting individual probiotic bacteria:-

Miyairi 588 from Japan produces butyric acid in the gut.  Butyric acid has been shown to have several interesting effects.  It improves immune health and for this reason is included in animal feed.  It has been shown to improve the integrity of gut to avoid “leaky gut”.  It is an HDAC inhibitor which means it may well have epigenetic effects.  It is an alternative to using butyrate supplements.

 Lactobacillus reuteri 17938 (Lactobacillus reuteri Protectis)

This bacteria is the one we are using and it has potent effects on my son’s summertime allergy that makes his autism much worse.

Lactobacillus reuteri ATCC PTA 6475

This is a potent anti-inflammatory bacteria, but its mode of action does not agree with my son, but it seems to do great things for many others.

Viviomixx and VSL#3

We saw that many people with IBS/IBD and some with autism find these two combination bacteria helpful.  Being a mixture of bacteria means that it may be only certain ingredients that have a helpful effect in a specific person with autism.

Many people with types of IBD/IBS do seem to respond well to the combined bacteria found in Viviomixx and VSL#3.

Some other interesting, commercially available, bacteria

I came across several interesting products. 

Lactobacillus reuteri NCIMB 30242

This bacteria is very well researched and has effects on some of comorbidities that effect some people with autism, such as vitamin D metabolism and calcium homeostasis.

As is often the case the benefits mainly relate to the immune system.  This particular bacteria reduces C-reactive protein (CRP) which is a commonly used marked for inflammation.  It reduces “bad” cholesterol and it has an odd effect on vitamin D making it interesting for people with reduced bone density.

I have no idea if it will help some people with autism, but it is very easy to find out since this patented bacteria is available in several products, targeted at your heart, GI or bones but also lightening your wallet.

Given how quick the L.reuteri protectis showed effect (1 day) I only intend to trial NCIMB 30242 for a few days.

Lactobacillus reuteri NCIMB 30242 research

L. reuteri NCIMB 30242: A Probiotic with Benefits

 Objectives

 The objective of this study was to evaluate the effects of probiotic bile salt hydrolase-active Lactobacillus reuteri NCIMB 30242 on cholesterol lowering, mechanism of action and gastrointestinal (GI) symptomatology in hypercholesterolemic adults.

Methods 127 subjects consumed either L. reuteri NCIMB 30242 or placebo capsules over a 9-week intervention period in a randomized controlled trial.

Results L. reuteri NCIMB 30242 capsules reduced LDL-cholesterol by 11.6% (P=0.001), total cholesterol by 9.1%, 

Conclusions L. reuteri NCIMB 30242 capsules should be considered as an adjunctive therapy for hypercholesterolemia and may be useful for promoting GI health.

  

Oral supplementation with probiotic L. reuteri NCIMB 30242 increases mean circulating 25-hydroxyvitamin D: a post hoc analysis of a randomized controlled trial.

L. reuteri NCIMB 30242 increased serum 25-hydroxyvitamin D by 14.9 nmol/L, or 25.5%, over the intervention period, which was a significant mean change relative to placebo of 17.1 nmol/L, or 22.4%, respectively (P = .003).

CONCLUSIONS:

To our knowledge, this is the first report of increased circulating 25-hydroxyvitamin D in response to oral probiotic supplementation.

  

Building healthy bones takes guts

Probioticboosts bone density in mice, could point way to new, natural treatments for osteoporosis

  

"We know that inflammation in the gut can cause bone loss, though it's unclear exactly why," said lead author Laura McCabe, a professor in MSU's departments of Physiology and Radiology. "The neat thing we found is that a probiotic can enhance bone density."

In the study, the male mice showed a significant increase in bone density after four weeks of treatment. There was no such effect when the researchers repeated the experiment with female mice, an anomaly they're now investigating.

Lactobacillus Reuteri NCIMB 30350

One reader of this blog is already a fan of Lactobacillus Reuteri NCIMB 30350 which comes from BioAmicus in Canada.

BioAmicus have had feedback from other customers who tried it having read the press reports on Lactobacillus Reuteri and autism.

 They told me:-

“The parents who have seen improvement with BioAmicus Reuteri note eye contact, social activity, language use, as well as improved instruction comprehension.”

They plan to make their own autism clinical trial.

                     https://bioamicus.com/autism-research/

Lactobacillus Johnsonii NCIMB 30351

The next interesting bacteria I came across is Lactobacillus Johnsonii.  There numerous strains.

This bacteria has been shown to be behind why children who live in a house with pet dog are protected from asthma.  Numerous studies like the auto immune disease asthma with increased incidence of autism.

The bacteria is protective against development of another auto immune disease, Type 1 diabetes.

Lactobacillus Johnsonii appears to mediate the effectiveness of some common cancer drugs.

BioAmicus have a Lactobacillus Johnsonii bacteria called NCIMB 30351 usually given to babies.

  

As some readers have already highlighted Lactobacillus bacteria can be used to make all kinds of yoghurt, kefir etc.  So you can grow your own at home to keep the cost down.

Lactobacillus johnsonii research

House dust exposure mediates gut microbiome Lactobacillus enrichment and airway immune defense against allergens and virus infection

Early-life exposure to dogs is protective against allergic disease development, and dog ownership is associated with a distinct milieu of house dust microbial exposures. Here, we show that mice exposed to dog-associated house dust are protected against airway allergen challenge. These animals exhibit reduced Th2 cytokine production, fewer activated T cells, and a distinct gut microbiome composition, highly enriched for Lactobacillus johnsonii, which itself can confer airway protection when orally supplemented as a single species. This study supports the possibility that host–environment interactions that govern allergic or infectious airway disease may be mediated, at least in part, by the impact of environmental exposures on the gastrointestinal microbiome composition and, by extension, its impact on the host immune response.

Research Shows How Household Dogs Protect Against Asthma, Infection

UCLA research suggests that gut bacteria could help prevent cancer

  

The Intestinal Microbiota Modulates the Anticancer Immune Effects of Cyclophosphamide

 Cyclophosphamide is one of several clinically important cancer drugs whose therapeutic efficacy is due in part to their ability to stimulate antitumor immune responses. Studying mouse models, we demonstrate that cyclophosphamide alters the composition of microbiota in the small intestine and induces the translocation of selected species of Gram-positive bacteria into secondary lymphoid organs. There, these bacteria stimulate the generation of a specific subset of “pathogenic” T helper 17 (pT_H17) cells and memory T_H1 immune responses. Tumor-bearing mice that were germ-free or that had been treated with antibiotics to kill Gram-positive bacteria showed a reduction in pT_H17 responses, and their tumors were resistant to cyclophosphamide. Adoptive transfer of pT_H17 cells partially restored the antitumor efficacy of cyclophosphamide. These results suggest that the gut microbiota help shape the anticancer immune response.

Lactobacillus Johnsonii N6.2 Mitigates the Development of Type 1 Diabetes in BB-DP Rat

Inhibition of Type 1 Diabetes Correlated to a Lactobacillus johnsonii N6.2-Mediated Th17 Bias

Although it is known that resident gut flora contribute to immune system function and homeostasis, their role in the progression of the autoimmune disease type 1 diabetes (T1D) is poorly understood. Comparison of stool samples isolated from Bio-Breeding rats, a classic model of T1D, shows that distinct bacterial populations reside in spontaneous Bio-Breeding diabetes-prone (BBDP) and Bio-Breeding diabetes-resistant animals. We have previously shown that the oral transfer of Lactobacillus johnsonii strain N6.2 (LjN6.2) from Bio-Breeding diabetes-resistant to BBDP rodents conferred T1D resistance to BBDP rodents, whereas Lactobacillus reuteri strain TD1 did not. In this study, we show that diabetes resistance in LjN6.2-fed BBDP rodents was correlated to a Th17 cell bias within the mesenteric lymph nodes. The Th17 bias was not observed in the non-gut–draining axillary lymph nodes, suggesting that the Th17 bias was because of immune system interactions with LjN6.2 within the mesenteric lymph node. LjN6.2 interactions with the immune system were observed in the spleens of diabetes-resistant, LjN6.2-fed BBDP rats, as they also possessed a Th17 bias in comparison with control or Lactobacillus reuteri strain TD1–fed rats. Using C57BL/6 mouse in vitro assays, we show that LjN6.2 directly mediated enhanced Th17 differentiation of lymphocytes in the presence of TCR stimulation, which required APCs. Finally, we show that footpad vaccination of NOD mice with LjN6.2-pulsed dendritic cells was sufficient to mediate a Th17 bias in vivo. Together, these data suggest an interesting paradigm whereby T1D induction can be circumvented by gut flora-mediated Th17 differentiation.

The effect of Lactobacillus johnsonii Ncc533 (La1) on the balance of Th1/Th2 cells in BALB/c mice

  

 Lactobacillus rhamnosus GG

  
This bacteria has numerous scientifically researched beneficial effects. Most recently it was shown to affect the expression of GABA receptors.  For some people with autism this might be beneficial. In particular it may reduce anxiety, since this was the effect noted in mouse research.

Lactobacillus rhamnosus GG (ATCC 53103) is a strain of L. rhamnosus that was isolated in 1983 from the intestinal tract of a healthy human being; filed for patent on 17 April 1985, by Sherwood Gorbach and Barry Goldin, and the 'GG' derives from the first letters of their surnames. 

The patent refers to a strain of "L. acidophilus GG" with American Type Culture Collection (ATCC) accession number 53103; later reclassified as a strain of L. rhamnosus. The patent claims the L. rhamnosus GG (ATCC 53103) strain is acid- and bile-stable, has a great avidity for human intestinal mucosal cells, and produces lactic acid. Since the discovery of the L. rhamnosus GG (ATCC 53103) strain, it has been studied extensively on its various health benefits and currently L. rhamnosus GG (ATCC 53103) strain is the world's most studied probiotic bacterium with more than 800 scientific studies.

The genome sequence of Lactobacillus rhamnosus GG (ATCC 53103) has been decoded.

Medical research and use

While Lactobacillus rhamnosus GG (ATCC 53103) is able to survive the acid and bile of the stomach and intestine, is claimed to colonize the digestive tract, and to balance intestinal microflora, evidence suggests that Lactobacillus rhamnosus is likely a transient inhabitant, and not autochthonous. Regardless, it is considered a probiotic useful for treatment of various maladies, as it works on many levels. Most of the molecular mechanisms are not known, however.

Peanut allergy

Research is showing that L. rhamnosus as a probiotic could stop allergic reactions to peanuts in 80% of children.

Diarrhea

Lactobacillus rhamnosus GG has been shown beneficial in the prevention of rotavirus diarrhea in children. The prevention and treatment of various types of diarrhea has been shown both in children and in adults.

Respiratory tract infections

L. rhamnosus GG may reduce the risk of obtaining respiratory tract infections in children that attend daycare.

Atopic dermatitis, eczema

Lactobacillus rhamnosus GG also has shown potential in treatment and primary prevention of atopic dermatitis, but the results of intervention trials have been mixed. A clinical trial with seven-year follow-up shows L. rhamnosus GG is useful in the prevention of atopic dermatitis in children at high risk of allergy.

Urogenital tract

The clinical health effects of L. rhamnosus GG have been widely studied. Both L. rhamnosus GG and L. rhamnosus GR-1 appear to protect the urogenital tract by excreting biosurfactants to inhibit the adhesion of vaginal and urinary pathogens.

Intestinal tract permeability

L. rhamnosus has been found to reduce intestinal permeability in children who suffer from irritable bowel syndrome, and it also has been found to counter alcohol-related intestinal permeability.

Gastrointestinal carriage of VRE

In 2005, L. rhamnosus GG was first used successfully to treat gastrointestinal carriage of vancomycin-resistant Enterococcus (VRE) in renal patients.

Anxiety

Research published in the Proceedings of the National Academy of Sciences on August 29, 2011 reported this bacterium may have an effect on GABA neurotransmitter receptors. Mice who were fed L. rhamnosus JB-1 had less anxiety and had different levels of a brain-chemical sensor and stress hormones.

This paper was mentioned previously in this blog

Ingestion of Lactobacillus strain regulates emotional behavior and central GABA receptor expression in a mouse via the vagus nerve

There is increasing, but largely indirect, evidence pointing to an effect of commensal gut microbiota on the central nervous system (CNS). However, it is unknown whether lactic acid bacteria such as Lactobacillus rhamnosus could have a direct effect on neurotransmitter receptors in the CNS in normal, healthy animals. GABA is the main CNS inhibitory neurotransmitter and is significantly involved in regulating many physiological and psychological processes. Alterations in central GABA receptor expression are implicated in the pathogenesis of anxiety and depression, which are highly comorbid with functional bowel disorders. In this work, we show that chronic treatment with L. rhamnosus (JB-1) induced region-dependent alterations in GABA_B1b mRNA in the brain with increases in cortical regions (cingulate and prelimbic) and concomitant reductions in expression in the hippocampus, amygdala, and locus coeruleus, in comparison with control-fed mice. In addition, L. rhamnosus (JB-1) reduced GABA_Aα2 mRNA expression in the prefrontal cortex and amygdala, but increased GABA_Aα2 in the hippocampus. Importantly, L. rhamnosus (JB-1) reduced stress-induced corticosterone and anxiety- and depression-related behavior. Moreover, the neurochemical and behavioral effects were not found in vagotomized mice, identifying the vagus as a major modulatory constitutive communication pathway between the bacteria exposed to the gut and the brain. Together, these findings highlight the important role of bacteria in the bidirectional communication of the gut–brain axis and suggest that certain organisms may prove to be useful therapeutic adjuncts in stress-related disorders such as anxiety and depression.

Weight loss

Research published in the British Journal of Nutrition in 2013 suggests that Lactobacillus rhamnosus CGMCC 1.3724 may increase weight loss in women who are dieting. The research was initiated after several studies showed that the gut bacteria in obese individuals differs significantly from those in thin people. Women in the study lost nearly twice the weight that the placebo group lost. No difference was observed in men, however.

Risks

The use of L. rhamnosus GG for probiotic therapy has been linked with very rare cases of sepsis in certain risk groups, primarily those with a weakened immune system and infants. Ingestion of L. rhamnosus GG is, nevertheless, considered to be safe, and data from Finland show a significant growth in the consumption of L. rhamnosus GG at the population level has not led to an increase in the number of Lactobacillus bacteraemia cases.

Probiotic Lactobacillus Probiotic rhamnosus downregulates FCER1 and HRH4 expressionin human mast cells

Abstract

AIM: To investigate the effects of four probiotic bacteria and their combination on human mast cell gene expression using microarray analysis.

METHODS: Human peripheral-blood-derived mast cells were stimulated with Lactobacillus rhamnosus (L. rhamnosus) GG (LGG^®), L. rhamnosus Lc705 (Lc705), Propionibacterium freudenreichii ssp. shermanii JS (PJS) and Bifidobacterium animalis ssp. lactis Bb12 (Bb12) and their combination for 3 or 24 h, and were subjected to global microarray analysis using an Affymetrix GeneChip^® Human Genome U133 Plus 2.0 Array. The gene expression differences between unstimulated and bacteria-stimulated samples were further analyzed with GOrilla Gene Enrichment Analysis and Visualization Tool and MeV Multiexperiment Viewer-tool.

RESULTS: LGG and Lc705 were observed to suppress genes that encoded allergy-related high-affinity IgE receptor subunits α and γ (FCER1A and FCER1G, respectively) and histamine H4 receptor. LGG, Lc705 and the combination of four probiotics had the strongest effect on the expression of genes involved in mast cell immune system regulation, and on several genes that encoded proteins with a pro-inflammatory impact, such as interleukin (IL)-8 and tumour necrosis factor alpha. Also genes that encoded proteins with anti-inflammatory functions, such as IL-10, were upregulated.

CONCLUSION: Certain probiotic bacteria might diminish mast cell allergy-related activation by downregulation of the expression of high-affinity IgE and histamine receptor genes, and by inducing a pro-inflammatory response.

Bifidobacterium Infantis 35624 

Bifidobacterium infantis 35624  is marketed in the US by Proctor & Gamble, while in Europe it is sold by the Irish developer.

It is well researched and does have effects beyond the gut.

Bifidobacterium infantis 35624 modulates host inflammatory processes beyond the gut

Certain therapeutic microbes, including Bifidobacteria infantis (B. infantis) 35624 exert beneficial immunoregulatory effects by mimicking commensal-immune interactions; however, the value of these effects in patients with non-gastrointestinal inflammatory conditions remains unclear. In this study, we assessed the impact of oral administration of B. infantis 35624, for 6‒8 weeks on inflammatory biomarker and plasma cytokine levels in patients with ulcerative colitis (UC) (n = 22), chronic fatigue syndrome (CFS) (n = 48) and psoriasis (n = 26) in three separate randomized, double-blind, placebo-controlled interventions. Additionally, the effect of B. infantis 35624 on immunological biomarkers in healthy subjects (n = 22) was assessed. At baseline, both gastrointestinal (UC) and non-gastrointestinal (CFS and psoriasis) patients had significantly increased plasma levels of C-reactive protein (CRP) and the pro-inflammatory cytokines tumor necrosis factor α (TNF-α) and interleukin-6 (IL-6) compared with healthy volunteers. B. infantis 35624 feeding resulted in reduced plasma CRP levels in all three inflammatory disorders compared with placebo. Interestingly, plasma TNF-α was reduced in CFS and psoriasis while IL-6 was reduced in UC and CFS. Furthermore, in healthy subjects, LPS-stimulated TNF-α and IL-6 secretion by peripheral blood mononuclear cells (PBMCs) was significantly reduced in the B. infantis 35624-treated groups compared with placebo following eight weeks of feeding. These results demonstrate the ability of this microbe to reduce systemic pro-inflammatory biomarkers in both gastrointestinal and non-gastrointestinal conditions. In conclusion, these data show that the immunomodulatory effects of the microbiota in humans are not limited to the mucosal immune system but extend to the systemic immune system.

The research highlighted by Proctor & Gamble is here:-

http://www.bifantis.com/probiotic-scientific-data.php

The product is sold as Alflorex in Europe and Align in the US.

               http://www.alimentaryhealth.ie/products/Alflorex

Conclusion

One big issue with all probiotics is just how potent they are when you actually consume them, rather than when they are manufactured.

Most people are taking probiotics for very general reasons, but people with IBS/IBD are a group who have very specific problems.  VSL#3 and Viviomixx do seem to be the probiotics of choice among those with IBS/IBD.

For allergy and atopic dermatitis some people clearly benefit from specific probiotics such as Bifidobacterium lactis BB-12 and Lactobacillus GG, but not all people respond.

Lowering cholesterol by probiotic is very easy to verify, so I presume it really must work in some cases.

Generally reducing colic, reflux, gas etc. in babies is a claim made for numerous probiotics.

You could spend a vast amount of money on probiotics for autism and it really is only worth using one(s) that have a genuine impact.

It would be useful to collect some data on what dosage is required when somebody actually does respond behaviourally to a probiotic.  Thanks to Alli and other readers I think we have the data on Biogaia products.

So far only one reader has given feedback on Lactobacillus Reuteri NCIMB 30350 (Bioamicus), but it was positive. The people at BioAmicus in Canada are very interested to know if their products are effective in some autism.

There are many people in the US using Culturelle for kids with autism, but I did not see any rave reviews.  Probably it is used for GI problems rather than to improve autism itself.

It does depend a lot where you live, how easy it is to access specific probiotics at a reasonable price.  Some are much cheaper in the US and some cheaper in Europe.

My current list of potentially interesting probiotics is:-

I really never expected to be writing about the merits of probiotics. It was a big surprise to learn that Miyiari 588 is put in animal feed to improve immune health via increasing the SCFA (short chained fatty acid) butyric acid.  Butyric acid is relevant to autism.  It was a bigger surprise to see L.reuteri Protectis reduce my son’s troublesome pollen allergy and changed the colour of his nose.

It is worthwhile doing some experimentation to see what, if anything, actually is helpful.

There are sound reasons why some people with autism may respond to one of the above bacteria.  As of now though, Biogaia is the probiotic of choice to try first, since many people with autism respond well.

All positive and negative feedback on these, or any other probiotic bacteria is very welcome.

Excitotoxicity triggered by GABAa dysfunction

  

This blog, as you will have noticed, does rather meander through science of autism.  As a result there are some gaps and unanswered questions.

The blog talks a lot about the neurotransmitter GABA and the excitatory/inhibitory imbalance.  We have ended up with some therapies based on this that do seem to help many people.

The opposing (excitatory) neurotransmitter is glutamate which affects the NMDA, AMP and mGlu receptors.

It appears that in autism there is an unusually high level of glutamate, but another issue looks likely to be at specific receptors, for example mGluR5

Two routes to autism phenotypes via mGluR5

This does get very complicated and lacks any immediate therapies. 

One very interesting insight was that you can repurpose the existing cheap generic GABA_B drug Baclofen to treat NMDAR-hypofunction. 

This seems to work really well at low doses with many people with Asperger’s.  People with more severe autism do not seem to respond to low doses, however some do to higher doses.  The more potent version R Baclofen is a research drug.

GABAb-mediated rescue of altered excitatory–inhibitory balance, gamma synchrony and behavioral deficits following constitutive NMDAR-hypofunction

Reduced N-methyl-D-aspartate-receptor (NMDAR) signaling has been associated with schizophrenia, autism and intellectual disability. NMDAR-hypofunction is thought to contribute to social, cognitive and gamma (30–80 Hz) oscillatory abnormalities, phenotypes common to these disorders.

Constitutive NMDAR-hypofunction caused a loss of E/I balance, with an increase in intrinsic pyramidal cell excitability and a selective disruption of parvalbumin-expressing interneurons. Disrupted E/I coupling was associated with deficits in auditory-evoked gamma signal-to-noise ratio (SNR). Gamma-band abnormalities predicted deficits in spatial working memory and social preference, linking cellular changes in E/I signaling to target behaviors. The GABA_B-receptor agonist baclofen improved E/I balance, gamma-SNR and broadly reversed behavioral deficits.

Excitotoxicity

We have touched on this subject on a few occasions but today, excitotoxicity is the focus of this post.

  

Excitotoxicity looks likely to be present in much autism and helps to connect all the various dysfunctions that we can read about in the literature.

It is a little scary because you cannot know to what extent this process is reversible.  It looks like in milder cases it should be treatable, whereas in extreme cases damage will be irreversible.

Excitotoxicity is the pathological process by which nerve cells are damaged or killed by excessive stimulation by neurotransmitters, particularly glutamate. This occurs when receptors for the excitatory neurotransmitter glutamate (glutamate receptors) such as the NMDA receptor and AMPA receptor are overactivated by glutamatergic storm. 

Unfortunately you can trigger glutamate excitotoxity via a dysfunction in GABA_A receptors.

For example if you severely inhibit GABA_A receptors you kill brain cells, but it was the reaction in glutamate signaling that did the damage.  GABA is supposed to be inhibitory; in some autism it is not and then Glutamate gets out of balance.  This does lead to excess firing of neurons, which seems to degrade cognition, but it will tend towards glutamate excitotoxity.

When you see the cascade of events triggered by glutamate excitotoxity you will see how this really helps to explain biological finding in autism, even mitochondrial dysfunctions.

You can then trace this all back to the faulty GABA switch caused by too little KCC2 and too much NKCC1.

Then you can look at other neurological conditions that feature glutamate excitotoxity, like traumatic brain injury and neuropathic pain, and you see that the research shows low expression of KCC2.

This then suggests that much of autism would have been prevented if you could increase KCC2.  You would not just fix the E/I imbalance but you would avoid all the damage done by excitotoxity.

Just how early you would have to correct KCC2 expression is not clear.  For sure it is a case of better late than never, but how much damage caused by excitotoxicity is reversible?

Good News

The good news is that because KCC2 underexpression is a feature of many conditions there is plenty of research money being spent looking for answers.  When they find a solution for increasing KCC2 to treat neuropathic pain, or spinal cord injury (SCI), the drug can be simply re-purposed for autism.

The French government is funding research into increasing KCC2 to treat SCI.  They are starting with serotin  5-HT_2A receptor agonists.  Regular readers without any memory loss may recall that back in the 1960 Lovaas was giving LSD to people with autism at UCLA.  LSD is a potent 5-HT_2A receptor agonist.  The French are also looking at BDNF to upregulate KCC2 and then they plan to have a blind test where they try all the chemicals they have in their library.  The French are of course doing their trials in test tubes.

When I looked at this subject a while back, I looked for existing therapies that are known to be safe and should be effective.

Treating KCC2 Down-Regulation in Autism, Rett/Down Syndromes, Epilepsy and Neuronal Trauma ?

My conclusion then was that intranasal insulin was the best choice.

Excitoxicity in Autism

Excitotoxicity in the Pathogenesis ofAutism

Autism is a debilitating neurodevelopment disorder characterized by stereotyped interests and behaviours, and abnormalities in verbal and non-verbal communication. It is a multifactorial disorder resulting from interactions between genetic, environmental and immunological factors. Excitotoxicity and oxidative stress are potential mechanisms, which are likely to serve as a converging point to these risk factors. Substantial evidence suggests that excitotoxicity, oxidative stress and impaired mitochondrial function are the leading cause of neuronal dysfunction in autistic patients. Glutamate is the primary excitatory neurotransmitter produced in the CNS, and overactivity of glutamate and its receptors leads to excitotoxicity. The over excitatory action of glutamate, and the glutamatergic receptors NMDA and AMPA, leads to activation of enzymes that damage cellular structure, membrane permeability and electrochemical gradients. The role of excitotoxicity and the mechanism behind its action in autistic subjects is delineated in this review

The influx of intracellular calcium triggers the induction of inducible nitric oxide (iNOS) and phosphorylation of protein kinase C. Increased iNOS enhances nitric oxide (NO•) production in excess, whereas protein kinase C activates phospholipase A2 which in turn results in the generation of pro-inflammatory molecules The subsequent generation of free radicals can inhibit oxidative phosphorylation and damage mitochondrial enzymes involved in the electron transport chain, which mitigate energy production .

Reactive intermediates such as peroxynitrates and other peroxidation products hamper the normal function of mitochondrial enzymes by impairing oxidative phosphorylation and inhibiting complex II of the electron transport chain. Moreover, lipid peroxidation products, such as 4-hydroxynonenal (4-HNE) can interact with synaptic protein and impair transport of glucose and glutamate, thereby decreasing energy production and increasing excitotoxic sensitivity

Overstimulation of the glutamate receptors, NMDA and AMPA, leads to the release of other excitotoxins resulting in the accumulation of glutamate. Indeed, excess glutamate concentrations results in an increase in calcium levels in the cytosol. This effect is attributed to the fact that excessive glutamate allows calcium channel to open for longer periods of time, leading to increased influx of calcium into cells. Calcium triggers inducible nitric oxide and protein kinase C that produce free radicals, ROS and arachidonic aid. Generation of these oxidants results in mitochondrial dysfunction and accumulation of pro-inflammatory molecules and finally cell death. Free radicals interact with the mitochondrial and cellular membrane to form lipid peroxidation. 4-HNE is a major destructive product of this process. Lipid peroxidation prevents the dephosphorylation of excessively phosphorylated tau protein, significantly interfering with microtubule function. It has also been shown to inhibit glutathione reductase needed to convert oxidised glutathione to its functional reduced form

The mechanism responsible for excitotoxicity and neuronal cell death is diverse. Experimental studies have shown that the apoptotic and/or necrotic cell death may be due to the severity of NMDA damage or can be dependent on receptor subunit composition of neurons (Bonfoco et al. 1995; Portera-Cailliau et al. 1997). Pathological events related to this mode of action can be loss of cellular homoeostasis with acute mitochondrial dysfunction leading to hindrance in ATP production. Moreover, glutamatergic insults can cause cell death by the action of one or more molecular pathways which involves the action of signaling molecules such as cysteine proteases, mitochondrial endonucleases, peroxynitrite, PARP-1 and GAPDH in the excitotoxic neurodegeneration pathway.

Intracellular calcium levels also rely on voltage-dependent calcium channels and Na exchangers . The Na?/Ca2? exchanger is a bi-directional membrane ion transporter, which during membrane depolarisation or the opening of the gated sodium channels, transports sodium out of the cell and calcium into the cell. AMPA-type glutamate receptors are highly permeable to calcium and its over expression can lead to excitotoxicity. The Ca2? permeability capability of AMPA-type glutamate receptors relies on the presence or the absence of the GluR2 subunit in the receptor complex. Reduced GluR2 expression permits the construction of AMPA receptors with high Ca2? permeability and contributes to neuronal defect and excitotoxicity. Another mechanism is the release of calcium from internal stores such as the endoplasmic reticulum and mitochondria. It results in mitochondrial dysfunction, reduction in ATP synthesis and ROS generation.

Voltage gated channels found in dendrites and cell bodies of neurons modulate neuronal excitability and calcium-regulated signaling cascades (Dolmetsch et al. 2001; Catterall et al. 2005). Point mutations in the gene encoding the L-type voltage-gated channels Ca v1.2 (CACNA1C) and Ca v1.4. (CACNA1F) prevent voltage-dependent inactivation of these genes. This causes the channel to open for longer time, leading to excessive influx of calcium.

Conclusion

Autism is a multifactorial disorder characterized by neurobehavioral and neurological dysfunction. Excitotoxicity is the major neurobiological mechanism that modulates diverse risk factors associated with autism. It is triggered by potential mutation in ion channels and signalling pathways, viral and bacterial pathogens, toxic metals and free radical generation. Over expression of glutamate receptors and increased glutamate levels leads to increased calcium influx and oxidative stress and progressive cellular degeneration and cell death. Genetic defect, such as mutation in voltage gated or ligand channels that regulate neuronal excitability leads to defect in synaptic transmission and excitotoxic condition in autism. Mutation in BKCa and Ca v1.2 channels also results in excess calcium influx Sodium, potassium and chloride channels also play important roles in maintaining homoeostasis of neuronal cells, and decreased channel activity leads to destabilization of membrane potential and excitotoxicity. Moreover, over expression of BDNF results in hyperexcitability. Excessive BDNF and NMDA receptor activity increases the neurotransmitter release and excitotoxic vulnerability. Given that autism is a multifaceted disorder with multiple risk factors, more precise studies are needed to explore the signalling pathways that influence emergence of excitotoxicity in ASDs.

Some relevant reading for those interested:-

GABAergic/glutamatergic imbalance relative to excessive neuroinflammation in autism spectrum disorders

Abstract

Background

Autism spectrum disorder (ASD) is characterized by three core behavioral domains: social deficits, impaired communication, and repetitive behaviors. Glutamatergic/GABAergic imbalance has been found in various preclinical models of ASD. Additionally, autoimmunity immune dysfunction, and neuroinflammation are also considered as etiological mechanisms of this disorder. This study aimed to elucidate the relationship between glutamatergic/ GABAergic imbalance and neuroinflammation as two recently-discovered autism-related etiological mechanisms.

Methods

Twenty autistic patients aged 3 to 15 years and 19 age- and gender-matched healthy controls were included in this study. The plasma levels of glutamate, GABA and glutamate/GABA ratio as markers of excitotoxicity together with TNF-α, IL-6, IFN-γ and IFI16 as markers of neuroinflammation were determined in both groups.

Results

Autistic patients exhibited glutamate excitotoxicity based on a much higher glutamate concentration in the autistic patients than in the control subjects. Unexpectedly higher GABA and lower glutamate/GABA levels were recorded in autistic patients compared to control subjects. TNF-α and IL-6 were significantly lower, whereas IFN-γ and IFI16 were remarkably higher in the autistic patients than in the control subjects.

Conclusion

Multiple regression analysis revealed associations between reduced GABA level, neuroinflammation and glutamate excitotoxicity. This study indicates that autism is a developmental synaptic disorder showing imbalance in GABAergic and glutamatergic synapses as a consequence of neuroinflammation.

Keywords: Autism, Glutamate excitotoxicity, Gamma aminobutyric acid (GABA), Glutamate/GABA, Tumor necrosis factor-α, Interleukin-6, Interferon-gamma, Interferon-gamma-inducible protein 16

Postmortem brain abnormalities of the glutamate neurotransmitter system in autism.

CONCLUSIONS:

Subjects with autism may have specific abnormalities in the AMPA-type glutamate receptors and glutamate transporters in the cerebellum. These abnormalities may be directly involved in the pathogenesis of the disorder.

Pathophysiologyof traumatic brain injury

General pathophysiology of traumatic brain injury

The first stages of cerebral injury after TBI are characterized by direct tissue damage and impaired regulation of CBF and metabolism. This ‘ischaemia-like’ pattern leads to accumulation of lactic acid due to anaerobic glycolysis, increased membrane permeability, and consecutive oedema formation. Since the anaerobic metabolism is inadequate to maintain cellular energy states, the ATP-stores deplete and failure of energy-dependent membrane ion pumps occurs. The second stage of the pathophysiological cascade is characterized by terminal membrane depolarization along with excessive release of excitatory neurotransmitters (i.e. glutamate, aspartate), activation of N-methyl-d-aspartate, α-amino-3-hydroxy-5-methyl-4-isoxazolpropionate, and voltage-dependent Ca²⁺- and Na⁺-channels. The consecutive Ca²⁺- and Na⁺-influx leads to self-digesting (catabolic) intracellular processes. Ca²⁺ activates lipid peroxidases, proteases, and phospholipases which in turn increase the intracellular concentration of free fatty acids and free radicals. Additionally, activation of caspases (ICE-like proteins), translocases, and endonucleases initiates progressive structural changes of biological membranes and the nucleosomal DNA (DNA fragmentation and inhibition of DNA repair). Together, these events lead to membrane degradation of vascular and cellular structures and ultimately necrotic or programmed cell death (apoptosis).

Excitotoxicity and oxidative stress

TBI is primarily and secondarily associated with a massive release of excitatory amino acid neurotransmitters, particularly glutamate.⁸⁵⁴ This excess in extracellular glutamate availability affects neurons and astrocytes and results in over-stimulation of ionotropic and metabotropic glutamate receptors with consecutive Ca²⁺, Na⁺, and K⁺-fluxes.²²⁷³ Although these events trigger catabolic processes including blood–brain barrier breakdown, the cellular attempt to compensate for ionic gradients increases Na⁺/K⁺-ATPase activity and in turn metabolic demand, creating a vicious circle of flow–metabolism uncoupling to the cell.¹⁶⁵⁰

Oxidative stress relates to the generation of reactive oxygen species (oxygen free radicals and associated entities including superoxides, hydrogen peroxide, nitric oxide, and peroxinitrite) in response to TBI. The excessive production of reactive oxygen species due to excitotoxicity and exhaustion of the endogenous antioxidant system (e.g. superoxide dismutase, glutathione peroxidase, and catalase) induces peroxidation of cellular and vascular structures, protein oxidation, cleavage of DNA, and inhibition of the mitochondrial electron transport chain.³¹¹⁶⁰ Although these mechanisms are adequate to contribute to immediate cell death, inflammatory processes and early or late apoptotic programmes are induced by oxidative stress.¹¹

Knocking down of the KCC2 in rat hippocampal neurons increases intracellular chloride concentration and compromises neuronal survival

Non-technical summary

‘To be, or not to be’– thousands of neurons are facing this Shakespearean question in the brains of patients suffering from epilepsy or the consequences of a brain traumatism or stroke. The destiny of neurons in damaged brain depends on tiny equilibrium between pro-survival and pro-death signalling. Numerous studies have shown that the activity of the neuronal potassium chloride co-transporter KCC2 strongly decreases during a pathology. However, it remained unclear whether the change of the KCC2 function protects neurons or contributes to neuronal death. Here, using cultures of hippocampal neurons, we show that experimental silencing of endogenous KCC2 using an RNA interference approach or a dominant negative mutant reduces neuronal resistance to toxic insults. In contrast, the artificial gain of KCC2 function in the same neurons protects them from death. This finding highlights KCC2 as a molecule that plays a critical role in the destiny of neurons under toxic conditions and opens new avenues for the development of neuroprotective therapy.

New understanding of brainchemistry could prevent brain damage after injury

Sciences de la vie, de la santé et des écosystèmes : Neurosciences (Blanc SVSE 4) 2010
Projet KCC2-SCI

The potassium-chloride transporter KCC2 : a new target for the treatment of neurological diseases

A decrease in synaptic inhibition –disinhibition- appears to be an important substrate in several neuronal disorders, such as spinal cord injury (SCI), neuropathic pain... Glycine and GABA are the major inhibitory transmitters in the spinal cord. An important emerging mechanism by which the strength of inhibitory synaptic transmission can be controlled is via modification of the intracellular concentration of chloride ions ([Cl-]i) to which receptors to GABA/glycine are permeable. Briefly, a low [Cl-]i is a pre-requisite for inhibition to occur and is maintained in healthy neurons by cation-chloride co-transporters (KCC2) in the plasma membrane, which extrude Cl-. We showed recently (Nature Medicine, accepted for publication) that these transporters are down-regulated after SCI, thereby switching the action of GABA and glycine from inhibition to excitation; this can account for both SCI-induced spasticity and chronic pain. KCC2 transporters therefore appear as a new target to restore inhibition within neuronal networks in pathological conditions. The present project aims at reducing spasticity and chronic pain after SCI by up-regulating KCC2. 
An important part will consist in identifying new compounds that increase the cell surface expression and/or the functionality of KCC2. Two strategies are considered. 1) Serotonin and BDNF will be tested on the basis of preliminary experiments and/or previous reports in other areas of the central nervous system indicating that these two compounds may affect the expression of KCC2. 2)Testing a large amount of compounds available in a library (“blind test”) to sort out KCC2-modulating molecules. This task can only be done in vitro on an assay that enables to easily visualize and quantify cell surface expression of KCC2, in response to these molecules (HEK293 cells). The few compounds isolated at the end of this task will then be tested on cultures of motoneurons (both mouse motoneurons and human motoneurons derived from induced pluripotent cells) and characterized further (potential toxicity, ability to cross the Brain Blood Barrier and effect on internalization and endocytosis of KCC2). 
The selected candidate compounds will enter into the in vivo validation phase aimed at increasing the expression of KCC2 following spinal cord injury (SCI; both contusion and complete spinal cord transection). The selected hits will be applied by intrathecal injections in SCI rats and their effects on KCC2 expression in the plasma membrane of motoneurons will be tested by means of western blots and immunohistochemistry. Their efficacy in increasing the cell-surface expression of KCC2 will also be tested electrophysiologically in vitro (i.e. their ability to hyperpolarize ECl). Functionally, their efficacy in reducing both SCI-induced spasticity and chronic pain will be assessed. 
Genetic tools will be used to increase the expression of KCC2 in some spinal neurons. This task will be done in collaboration with teams in the USA. Lentiviral vectors aimed at increasing KCC2 in the host cells, after parenchymal injection, have been developed in San Diego. A transgenic mouse model with a conditional tamoxifen-induced overexpression of KCC2 has been developed in Pittsburgh. The rationale for this part of the project is to use these genetic tools in the chronic phase of SCI to reduce spasticity and chronic pain. 
The last part of the project will focus on more fundamental issues regarding the relationship between the SCI-induced downregulation of KCC2 and the development of spasticity and chronic pain. 
The significance of the expected results goes far beyond the scope of SCI, since altered chloride homeostasis resulting from mutation or dysfunction of cation-chloride cotransporters has been implicated in various neurological disorders such as, for instance, ischemic seizures neonatal seizures and temporal lobe epilepsy. 

KCC2 escape from neuropathic pain

Activationof 5-HT2A receptors upregulates the function of the neuronal K-Cl cotransporter KCC2.

 In healthy adults, activation of γ-aminobutyric acid (GABA)(A) and glycine receptors inhibits neurons as a result of low intracellular chloride concentration ([Cl(-)](i)), which is maintained by the potassium-chloride cotransporter KCC2. A reduction of KCC2 expression or function is implicated in the pathogenesis of several neurological disorders, including spasticity and chronic pain following spinal cord injury (SCI). Given the critical role of KCC2 in regulating the strength and robustness of inhibition, identifying tools that may increase KCC2 function and, hence, restore endogenous inhibition in pathological conditions is of particular importance. We show that activation of 5-hydroxytryptamine (5-HT) type 2A receptors to serotonin hyperpolarizes the reversal potential of inhibitory postsynaptic potentials (IPSPs), E(IPSP), in spinal motoneurons, increases the cell membrane expression of KCC2 and both restores endogenous inhibition and reduces spasticity after SCI in rats. Up-regulation of KCC2 function by targeting 5-HT(2A) receptors, therefore, has therapeutic potential in the treatment of neurological disorders involving altered chloride homeostasis. However, these receptors have been implicated in several psychiatric disorders, and their effects on pain processing are controversial, highlighting the need to further investigate the potential systemic effects of specific 5-HT(2A)R agonists, such as (4-bromo-3,6-dimethoxybenzocyclobuten-1-yl)methylamine hydrobromide (TCB-2).

Conclusion

Very little is certain in autism, in great part because only about 200 brains have ever been examined post mortem.  There are many theories, but very many more sub-types of autism.

GABA_A dysfunction due to the faulty GABA switch never increasing KCC2 expression in the first weeks of life, triggering glutamate excitotoxicity and all that follows would go a long way to explaining my son’s type of autism. It might well explain 30+% of all autism.

Clearly other causes of excess glutamate would lead to a similar result.