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Thursday, 18 May 2017

Amino Acids in Autism


Amino Acids (AAs) are very important to health and it is important that all 20 are within the reference ranges, or there can be serious consequences.  Inborn errors of amino acid metabolism do exist and there are metabolic disorders which impair either the synthesis and/or degradation of amino acids.
It has been suggested that a lack of certain amino acids might underlie some people’s autism. This seems to be the basis of one new autism drug, CM-AT, being developed in the US, but this idea remains somewhat controversial.

In those people who have normal levels of amino acids, potential does exist to modify their level for some therapeutic effect. 

Examples include:-

·        Using histidine to inhibit mast cells de-granulating and so reducing symptoms of allergy

·       Using the 3 branch chained AAs to reduce the level of the AA, phenylanine, which can drive movement disorders/tics

·       Methionine seems to promote speech in regressive autism, but for no known reason.

·        Some AAs, such as leucine, activate mTOR. It is suggested that others (histidine, lysine and threonine) can inhibit it, which might have a therapeutic benefit in those with too much mTOR signaling.

·        D-Serine, synthesized in the brain by from L-serine, serves as a neuromodulator by co-activating NMDA receptors.  D-serine has been suggested for the treatment of negative symptoms of schizophrenia

·        Aspartic acid is an NMDA agonist

·       Threonine is being studied as a possible therapy for Inflammatory Bowel Disease (IBD), because it may increase intestinal mucin synthesis.


Amino acids, the building blocks for proteins

To make a protein, a cell must put a chain of amino acids together in the right order. It makes a copy of the relevant DNA instruction in the cell nucleus, and takes it into the cytoplasm, where the cell decodes the instruction and makes many copies of the protein, which fold into shape as they are produced.

There are 20 standard or “canonical” amino acids, which can be thought of as protein building blocks.
Humans can produce 10 of the 20 amino acids; the others must be supplied in the food and are called “essential”. The human body does not store excess amino acids for later use, so these amino acids must be in your food every day.

The 10 amino acids that we can produce are alanine, asparagine, aspartic acid, cysteine, glutamic acid, glutamine, glycine, proline, serine and tyrosine. Tyrosine is produced from phenylalanine, so if the diet is deficient in phenylalanine, tyrosine will be required as well.

The essential amino acids (marked * below) are arginine, histidine, isoleucine, leucine, lysine, methionine, phenylalanine, threonine, tryptophan, and valine.

The three so-called branched-chain amino acids (BCAAs) are leucine, isoleucine and valine

The so-called aromatic amino acids (AAAs) are histidine, phenylanine, tryptophan and tyrosine

When plasma levels of BCAAs increase, this reduces the absorption of aromatic AAs; so the level of tryptophan, tyrosine, and phenylalanine will fall and this directly affects the synthesis and release of serotonin and catecholamines.
Many sportsmen, and indeed soldiers, take BCAA supplements in an attempt to build stronger muscles, but within the brain this will cause a cascade of other effects.
In people with tardive dyskinesia, which is a quite common tic disorder found in schizophrenia and autism, taking phenylalanine may make their tics worse.  It seems that taking BCAA supplements may make their tics reduce, because reducing the level of phenylalanine will impact dopamine (a catecholamine). Most movement disorders ultimately relate to dopamine.



In effect, BCAA supplements affect the synthesis and release of serotonin and catecholamines.  This might be good for you, or might be bad for you; it all depends where you started from.

   Alanine
   Arginine *
   Asparagine
   Aspartic acid
   Cysteine
   Glutamic acid
   Glutamine
   Glycine
   Histidine * Aromatic
   Isoleucine * BCAA
   Leucine * BCAA
   Lysine *
   Methionine *
   Phenylalanine *  Aromatic
   Proline
   Serine
   Threonine *
   Tryptophan * Aromatic
   Tyrosine  Aromatic
   Valine
*  BCAA


Blood levels of the BCAAs are elevated in people with obesity and those with insulin resistance, suggesting the possibility that BCAAs contribute to the pathogenesis of obesity and diabetes.  BCAA-restricted diets improve glucose tolerance and promote leanness in mice.


In the brain, BCAAs have two important influences on the production of neurotransmitters. As nitrogen donors, they contribute to the synthesis of excitatory glutamate and inhibitory gamma-aminobutyric acid (GABA) They also compete for transport across the blood-brain barrier (BBB) with tryptophan (the precursor to serotonin), as well as tyrosine and phenylalanine (precursors for catecholamines)Ingestion of BCAAs therefore causes rapid elevation of the plasma concentrations and increases uptake of BCAAs to the brain, but diminishes tryptophan, tyrosine, and phenylalanine uptake. The decrease in these aromatic amino acids directly affects the synthesis and release of serotonin and catecholamines. The reader is referred to Fernstrom (2005) for a review of the biochemistry of BCAA transportation to the brain. Oral BCAAs have been examined as treatment for neurological diseases such as mania, motor malfunction, amyotrophic lateral sclerosis, and spinocerebral degeneration. Excitotoxicity as a result of excessive stimulation by neurotransmitters such as glutamate results in cellular damage after traumatic brain injury (TBI). However, because BCAAs also contribute to the synthesis of inhibitory neurotransmitters, it is unclear to what extent the role of BCAAs in synthesis of both excitatory and inhibitory neurotransmitters might contribute to their potential effects in outcomes of TBI.

A list of human studies (years 1990 and beyond) evaluating the effectiveness of BCAAs in providing resilience or treating TBI or related diseases or conditions (i.e., subarachnoid hemorrhage, intracranial aneurysm, stroke, anoxic or hypoxic ischemia, epilepsy) in the acute phase is presented in Table 8-1; this also includes supporting evidence from animal models of TBI. The occurrence or absence of adverse effects in humans is included if reported by the authors.

Cell Signaling

Leucine indirectly activates p70 S6 kinase as well as stimulates assembly of the eIF4F complex, which are essential for mRNA binding in translational initiation. P70 S6 kinase is part of the mammalian target of rapamycin complex (mTOR) signaling pathway.



The present study provides the first evidence that mTOR signalling is enhanced in response to an acute stimulation with the proteinogenic amino acid, leucine, within cultured human myotubes. While these actions appear transient at the leucine dose utilised, activation of mTOR and p70S6K occurred at physiologically relevant concentrations independently of insulin stimulation. Interestingly, activation of mTOR signalling by leucine occurred in the absence of changes in the expression of genes encoding both the system A and system L carriers, which are responsible for amino acid transport. Thus, additional analyses are required to investigate the molecular mechanisms controlling amino acid transporter expression within skeletal muscle. Of note was the increased protein expression of hVps34, a putative leucine-sensitive kinase which intersects with mTOR. These results demonstrate the need for further clinical analysis to be performed specifically investigating the role of hVps34 as a nutrient sensing protein for mTOR signalling.

Skeletal muscle mass is determined by the balance between the synthesis and degradation of muscle proteins. Several hormones and nutrients, such as branched-chain amino acids (BCAAs), stimulate protein synthesis via the activation of the mammalian target of rapamycin (mTOR).
BCAAs (i.e., leucine, isoleucine, and valine) also exert a protective effect against muscle atrophy. We have previously reported that orally administered BCAA increases the muscle weight and cross-sectional area (CSA) of the muscle in rats



3.4. BCAAs in Brain Functions
BCAAs may also play important roles in brain function. BCAAs may influence brain protein synthesis and production of energy and may influence synthesis of different neurotransmitters, that is, serotonin, dopamine, norepinephrine, and so forth, directly or indirectly. Major portion of dietary BCAAs is not metabolized by liver and comes into systemic circulation after a meal. BCAAs and aromatic AA, such as tryptophan (Trp), tyrosine (Tyr), and phenylalanine (Phe), share the same transporter protein to transport into brain. Trp is the precursor of neurotransmitter serotonin; Tyr and Phe are precursors of catecholamines (dopamine, norepinephrine, and epinephrine). When plasma concentration of BCAAs increases, the brain absorption of BCAAs also increases with subsequent reduction of aromatic AA absorption. That may lead to decrease in synthesis of these related neurotransmitters [3]. Catecholamines are important in lowering blood pressure. When hypertensive rats were injected with Tyr, their blood pressure dropped markedly and injection with equimolar amount of valine blocks that action [49]. In vigorous working persons, such as in athletes, depletion of muscle and plasma BCAAs is normal. And that depletion of muscle and plasma BCAAs may lead to increase in Trp uptake by brain and release of serotonin. Serotonin on the other hand leads to central fatigue. So, supplementation of BCAAs to vigorously working person may be beneficial for their performance and body maintenance


Example of a treatable Amino Acid variant of Autism


Autism Spectrum Disorders (ASD) are a genetically heterogeneous constellation of syndromes characterized by impairments in reciprocal social interaction. Available somatic treatments have limited efficacy. We have identified inactivating mutations in the gene BCKDK (Branched Chain Ketoacid Dehydrogenase Kinase) in consanguineous families with autism, epilepsy and intellectual disability (ID). The encoded protein is responsible for phosphorylation-mediated inactivation of the E1-alpha subunit of branched chain ketoacid dehydrogenase (BCKDH). Patients with homozygous BCKDK mutations display reductions in BCKDK mRNA and protein, E1-alpha phosphorylation and plasma branched chain amino acids (BCAAs). Bckdk knockout mice show abnormal brain amino acid profiles and neurobehavioral deficits that respond to dietary supplementation. Thus, autism presenting with intellectual disability and epilepsy caused by BCKDK mutations represents a potentially treatable syndrome.

The data suggest that the neurological phenotype may be treated by dietary supplementation with BCAAs. To test this hypothesis, we studied the effect of a chow diet containing 2% BCAAs or a BCAA-enriched diet, consisting of 7% BCAAs, on the neurological phenotypes of the Bckdk−/− mice. Mice raised on the BCAA-enriched diet were phenotypically normal. On the 2% BCAA diet, however, Bckdk−/− mice had clear neurological abnormalities not seen in wild-type mice, such as seizures and hindlimb clasping, that appeared within 4 days of instituting the 2% BCAA diet (Fig. 3B). These neurological deficits were completely abolished within a week of the Bckdk−/− mice starting the BCAA-enriched diet, which suggests that they have an inducible yet reversible phenotype (Fig. 3C).

Our experiments have identified a Mendelian form of autism with comorbid ID and epilepsy that is associated with low plasma BCAAs. Although the incidence of this disease among patients with autism and epilepsy remains to be determined, it is probably quite a rare cause of this condition. We have shown that murine Bckdk−/− brain has a disrupted amino acid profile, suggesting a role for the BBB in the pathophysiology of this disorder. The mechanism by which abnormal brain amino acid levels lead to autism, ID, and epilepsy remains to be investigated. We have shown that dietary supplementation with BCAAs reverses some of the neurological phenotypes in mice. Finally, by supplementing the diet of human cases with BCAAs, we have been able to normalize their plasma BCAA levels (table S10), which suggests that it may be possible to treat patients with mutations in BCKDK with BCAA supplementation.


(Look at the three red rows, the BCAAs, all lower than the reference range, before supplementation)


Threonine, Mucin and Akkermansia muciniphila in Autism
Mucins are secreted as principal components of mucus by mucous membranes, like the lining of the intestines.  People with Inflammatory Bowel Disease (IBD) have mucus barrier changes.

The low levels of the mucolytic bacterium Akkermansia muciniphila found in children with autism, apparently suggests mucus barrier changes.

The amino acid Threonine is a component of mucin and Nestle have been researching for some time the idea of a threonine supplement to treat Inflammatory Bowel Disease (IBD), being a serious Swiss company they publish their research.      

Threonine Requirement in Healthy Adult Subjects and in Patients With Crohn's Disease and With Ulcerative Colitis Using the Indicator Amino Acid Oxidation (IAAO) Methodology

Threonine is an essential amino acid which must be obtained from the diet. It is a component of mucin. Mucin, in turn, is a key protein in the mucous membrane that protects the lining of the intestine.

Inflammatory bowel disease (IBD) is a group of inflammatory conditions that affect the colon and small intestine. IBD primarily includes ulcerative colitis (UC) and Crohn's disease (CD). In UC, the inflammation is usually in the colon whereas in CD inflammation may occur anywhere along the digestive tract. Studies in animals have shown that more threonine is used when there is inflammation in the intestine.

The threonine requirement in healthy participants and in IBD patients will be determined using the indicator amino acid oxidation method. The requirement derived in healthy participants will be compared to that derived in patients with IBD.

Each participant will take part in two x 3 day study periods. The first two days are called adaptation days where the subjects will consume a liquid diet specially designed for him. The diet will be consumed at home. It contains all vitamins, minerals, protein and all other nutrients required. On the third day, the participant will come to the Hospital for Sick Children in Toronto. Subjects will consume hourly meals for a total of 8 meals and a stable isotope 13C-phenylalanine. Breath and urine samples will be collected to measure the oxidation of phenylalanine from which the threonine requirement will be determined. 



We determined whether the steady-state levels of intestinal mucins are more sensitive than total proteins to dietary threonine intake. For 14 d, male Sprague-Dawley rats (158 ± 1 g, n = 32) were fed isonitrogenous diets (12.5% protein) containing 30% (group 30), 60% (group 60), 100% (control group), or 150% (group 150) of the theoretical threonine requirement for growth. All groups were pair-fed to the mean intake of group 30. The mucin and mucosal protein fractional synthesis rates (FSR) did not differ from controls in group 60. By contrast, the mucin FSR was significantly lower in the duodenum, ileum, and colon of group 30 compared with group 100, whereas the corresponding mucosal protein FSR did not differ. Because mucin mRNA levels did not differ between these 2 groups, mucin production in group 30 likely was impaired at the translational level. Our results clearly indicate that restriction of dietary threonine significantly and specifically impairs intestinal mucin synthesis. In clinical situations associated with increased threonine utilization, threonine availability may limit intestinal mucin synthesis and consequently reduce gut barrier function.
  


It has been proposed that excessive mucin degradation by intestinal bacteria may contribute to intestinal disorders, as access of luminal antigens to the intestinal immune system is facilitated. However, it is not known whether all mucin-degraders have the same effect. For example A. muciniphila may possess anti-inflammatory properties, as a high proportion of the bacteria has been correlated to protection against inflammation in diseases such as type 1 diabetes mellitus, IBD, atopic dermatitis, autism , type 2 diabetes mellitus, and.



Gastrointestinal disturbance is frequently reported for individuals with autism. We used quantitative real-time PCR analysis to quantify fecal bacteria that could influence gastrointestinal health in children with and without autism. Lower relative abundances of Bifidobacteria species and the mucolytic bacterium Akkermansia muciniphila were found in children with autism, the latter suggesting mucus barrier changes. 

Previous studies in rats by MacFabe et al. have shown that intraventricular administration of propionate induces behaviors resembling autism (e.g., repetitive dystonic behaviors, retropulsion, seizures, and social avoidance) (12, 13). We have also reported increased fecal propionate concentrations in ASD children compared with that in controls in the same fecal samples (25). However, the abundance of a key propionate-producing bacterium, Prevotella sp., was not significantly different between the study groups. This suggests that other untargeted bacteria, such as those from Clostridium cluster IX, which also includes major propionate producers (24), may be responsible for the observed differences in fecal propionate concentrations. Moreover, it is possible that the activities of the bacteria responsible for producing propionate, rather than bacterial numbers, have been altered. Other factors, such as differences in GI function that change GI transit time in ASD children, should also be considered.
In summary, the current findings of depleted populations of A. muciniphila and Bifidobacterium spp. add to our knowledge of the changes in the GI tracts of ASD children. These findings could potentially guide implementation of dietary/probiotic interventions that impact the gut microbiota and improve GI health in individuals with ASD.


Conclusion
I think that modifying levels of amino acids can have merit for some people, but it looks like another case for personalized medicine, rather than the same mix of powders given to everyone.
Threonine is interesting given the incidence of Inflammatory Bowel Disease (IBD) in autism.  IBD mainly describes ulcerative colitis and Crohn's disease.
The research into Threonine, is being funded by Nestle, the giant Swiss food company, who fortunately do publish their research.
The trial in the US of CM-AT is unusual because no results have ever been published in the literature, so we just have press releases. It likely that CM-AT is a mixture of pancreatic enzymes from pigs and perhaps some added amino acids.



This 14-week, double-blind, randomized, placebo-controlled Phase 3 study is being conducted to determine if CM-AT may help improve core and non-core symptoms of Autism. CM-AT, which has been granted Fast Track designation by FDA, is designed to enhance protein digestion thereby potentially restoring the pool of essential amino acids. Essential amino acids play a critical role in the expression of several genes important to neurological function and serve as precursors to key neurotransmitters such as serotonin and dopamine.


Based on the study I referred to early this year:-


·        Amino acids, his, lys and thr, inhibited mTOR pathway in antigen-activated mast cells

·     Amino acids, his, lys and thr inhibited degranulation and cytokine production of mast cells

·     Amino acid diet reversed mTOR activity in the brain and behavioral deficits in allergic and BTBR mice.

in my post:



I for one will be evaluating both lysine and threonine, having already found a modest dose histidine very beneficial in allergy (stabilizing mast cells).




11 comments:

  1. Thanks Peter for this post, it is very interesting and particularly helpful to me as I’ve been trying to sum up AAs use in autism and explain recent strange behavior of my son possibly linked to a supplement containing some AAs.

    It is still not entirely clear to me. There is a paper by Prof. Evangeliou about BCAAs as an adjunctive therapy to ketogenic diet, in which they described that the addition of BCAAs reduced seizures in children with intractable epilepsy on KD. They presented several possible explanations including “ketotic action” of BCCAs, but also suggested that “BCAAs decreasing the effects of glutamatergic neurotransmission could facilitate those of GABAergic neurotransmission”. Is a trial and error needed once again to see if BCAAs could be beneficial via these mechanisms or those proposed by Tyler or the effect is just opposite because of mTOR activation?

    https://www.ncbi.nlm.nih.gov/pubmed/19687389

    As discussed here before, I’ve been trialling exogenous ketones (beta hydroxybutyrate) after trying on myself. One of the supplement available is a protein/BHB bar containing: L-taurine, L-tyrosine, L-leucine and also calcium. I am not sure if I can attribute behavior to this product, but M. had a few days of unusual irritability and sleep issues recently. He didn’t react this way to other forms of BHB and it was on holidays where a number of other things could be involved. But my question is: should I expect a bad reaction to any of these AAs for any reason? Unfortunately the company does not provide the full details about the AAs dose, but the amount of calcium was 50% RDA. I have never given calcium supplement to my son, but he had a clear, negative reaction to vit. D.

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    1. Agnieszka, it looks like another case where responsible trial and error may deliver benefits. In the TBI study I posted in a comment below

      https://www.ncbi.nlm.nih.gov/pmc/articles/PMC5323875/

      it is clear that moderate TBI caused a transient change in brain AAs, but a severe TBI caused a long lasting change. There are some parallels between TBI and more severe autism, although there are big differences.

      I think AAs are just one part of a much wider metabolic disturbance. If they can be used to tune the brain a little , without side effects and avoiding feedback loops, that would be another step forward.

      In the US you can buy a wide range of AAs as cheap bulk powders, so you can cheaply make your own customized AA mix. The good thing is that doses are of the same magnitude as the RDA.

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    2. Leucine is paradoxical as it is clearly an anabolic amino acid as it stimulates mtor, but it also promotes autophagy in one study I read, which is unintuitive since mtor up regulation generally suppresses autophagy.

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    3. Tyler, I am concerned about leucine in BCAAs, I know that it promotes autophagy, but do you think that in spite of the fact it stimulates mtor is harmless even in the long run? could he be having a bad reaction to leucine in terms of behavior?
      Valentina

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    4. Leucine is in ample amounts in just about any type of animal protein. Because it is so ubiquitous, it along with sugars is likely why they are the two main nutritional signals for insulin release. Some bcaa formulations will have a leucine, valine, isoleurine ratio of 9:1:1 or higher and that is not what you want. Leucine is also an essential amino acid which means you cannot live without it. Cottage cheese has copious amounts of leucine, but it is high in aromatic amino acids which is why supplementing bcaa's directly makes more sense than simply eating cottage cheese.

      In general, if you want to limit leucine intake, which I think is a bad idea in general, you will want to simply have a low protein diet which is almost always a terrible idea as well unless your goal is to have them be weak, frail, and sickly all the time, and especially when they get older. Muscle mass is especially important for the immune system since it is where your body gets the amino acids it needs for synthesizing proteins to fight off infections. Lack of muscle means you are more vulnerable to being overcome by disease. During the black death, one of the reasons it killed so many people was that europe's population was on the brink of starvation most of the time as only the nobles ever had enough protein in thein diet to keep their immune system in good enough shape to fight off disease.

      The most practical and safe way to restrict overconsumption of leucine or any other mtor stimulating food is to fast. Unfortunately there are no studies on fasting for children and the animal research is inconclusive as there is evidence both ways that fasting children is good or bad. Fasting is definit Ely good for adults but what is good for adults is not always good for children.

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  2. I have already supplemented Lysine daily for quite a while now, but for different reasons. Supplemental lysine is thought to act as a benzo booster and many people supplement it under the hopes of needing to use less benzos because they think it will make using benzos less addictive. So in a way Lysine could potentially give a boost to GABAA subunit interventions such as low-dose clonazepam. GABAA signaling seems to show up impaired most of the time in studies that look at the subject, so Lysine is supplemented for that reason because it is pretty unclear what causes GABAA dysfunction (a lot of research and many hypotheses and no clear answers yet) so you could say this is part of an "everything and the kitchen sink" approach in this particular domain. Other uses I came across for Lysine were to inhibit various types of retroviruses such as the various herpes viruses since Lysine (if my memory serves correctly) is antagonistic to Arginine which seems to act as a stimulant for retrovirus infection. Also, I don't supplement threonine directly but have been doing Magnesium-Threonate for some time now, even though threonate is better thought of as a metabolite of Vitamin C.

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  3. Some very interesting research concerning the striatum (caudate nuclean and putamen) came out today which mirrors one consistent finding in several autism studies I have read recently:

    Press Release:

    https://www.sciencedaily.com/releases/2017/05/170518104045.htm

    Paper:

    http://www.apa.org/pubs/journals/releases/emo-emo0000331.pdf

    What they found is that people who have anxiety over making decisions concerning an uncertain future. An enlarged striatum is one of the most common brain abnormalities in idiopathic autism and many people with autism prefer and do better academically in a "predictable" environment. (Speculating here) It could be that this research directly explains the need for sameness among high-functioning and low-functioning autistics alike not to mention other existing research in OCD and autism showing an enlarged striatum tracks with severity in repetitive behaviors.

    A hyperactive amygdala could in part be the result of being driven extra hard from a hyperfunctioning striatum which is attuned to reacting to too many hypothetical predictive fears which to a normal person might just seem like irrational phobias. The need for sameness could be explained as the need to reduce potential possible outcomes that the autistic person is alerted to.

    Therapy wise, it might be that a brute force approach to dampening down the striatum as a whole might help in this regard except that under the assumption of the solution being a drug-based solution, the dosage would need to be perfect for the individual (i.e. some doctor prescribing a dosage straight off of some medical chart would not do).

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  4. Here is another article for those interested in how changes in brain amino acids (AAs) may affect behavior. This study shows how traumatic brain injury affects AAs.

    Severity of experimental traumatic brain injury modulates changes in concentrations of cerebral free amino acids
    https://www.ncbi.nlm.nih.gov/pmc/articles/PMC5323875/

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  5. Thankyou Peter once again for the last 2 posts, I can give my opinion since I follow this therapy since almost 5 months, BCAAs are extremely helpful for my son,I couldn´t have left risperidone without them.There are still madness episodes from time to time, but are predictible because his behavior starts to change little by little at least 1 hour before the episode, which is like a dopamine storm invading his brain.About L serine, the experience is being better than expected because I discovered that if he takes it before bed time, the quality of sleep improves a lot.
    Valentina

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  6. Hi Peter, I had sought suggestions for my 23 yr old with very blistery skin, brain fog, rages, and had had a second regression when very little.
    We started 2 g of histidine this week (spread across the day) and I do think overall his mood has been better and I even think his skin is starting to be less inflammed.
    That said, we are not on a low histimine diet.
    His rages, though fewer, seem worse and more intense. Could the histidine actually make responses to normal or high histimine foods worse?

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    1. I would give it more time to see if histidine helps or not. There may be external effects that vary over time, which is very common in allergy. So you could try it for a month, give it break for month, then try again to decide if it helps or not.

      If you have autism, mast cell issues (allergy, skin rashes etc) and rages, I think it highly likely that Verapamil will help a great deal.

      You only need a low histamine diet if you are histamine intolerant.

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