Big heads, the Car wash, Transcranial pulse stimulation, GABA alpha 5 and Potassium channel Kv3.1

Today’s post is a review of some interesting new research that relates to the scope of this blog.  It ranges from training young people with autism/ID to work at the car wash, to more complex science.

Let’s start with the easiest paper. Somewhat bizarrely it was carried out in Japan by researchers from India. I am a fan of teaching kids to wash cars but I was surprised to see that it would be covered in a published research study.

One often forgotten item to teach teenagers and young adults with autism or ID is how to safely use public transport, so they might travel independently to and from any future job. We have had a lot of success with this recently. Monty, now aged 20, can get all the way from home to various different locations across the city using public transport, including changing buses and with journey times more than one hour.

 

Increasing car washing competency in adolescents with autism and intellectual disabilities: Researching visual task evaluation

This study looked at how well visual task evaluation helped teenagers with autism and intellectual disabilities become more competent at car washing. For disabled people to promote their independence and employment chances, car washing skills are crucial. The goal of this study was to ascertain whether training techniques that include visual task evaluation can improve car washing proficiency in teenagers with autism and intellectual disabilities. 30 participants, ranging in age from 12 to 18, participated in a pre-test/post-test design. Randomly chosen groups of participants were put into the evaluation group for the visual task or the control group. According to the findings, the visual task evaluation group outperformed the control group in terms of car washing ability. Adolescents with autism and intellectual disabilities can learn skills more quickly and become more independent by including visual task evaluation into their teaching strategies. These results demonstrate the potential for such treatments to enhance their quality of life and employment chances.

 

Car washing with a pressure washer is great fun for most people and washing a car thoroughly has many individual steps to master, so it is good practice.

  

Head size

It has been known for decades that big heads (macrocephaly) and small heads (microcephaly) are a tell-tale sign of a neurodevelopment problem. Normally, big heads are linked to intellectual disability, but very small heads are also a warning sign.

Readers may recall the Zika virus epidemic in Brazil in 2015. This mosquito-borne virus caused pregnant women to give birth to children with microcephaly. Zika virus infection caused intellectual disability in babies. The severity of the intellectual disability varied from mild to severe. Babies with Zika virus infection may have difficulty learning and communicating. They may also have problems with problem-solving and abstract thinking. Hearing and vision can be impaired and growth is retarded.  

Head size parts autism into two major subtypes

Essentially opposite paths in fetal brain development may explain two major subtypes of autism. In one of these subtypes, an unusually high number of excitatory neurons in a key brain region leads to large heads, or macrocephaly, which affects roughly 20 percent of people with autism; in the other, a decreased number of the same cells in that area leads to more typical head sizes, a new study finds. 

This fundamental biological difference suggests that “therapeutic avenues may be drastically different for these subtypes,” says lead investigator Flora Vaccarino, professor of neuroscience at Yale University. “That in turn could explain why drug treatments for autism so far are failing.”

 

The opposite brain development paths found in this research may both lead to autism because they are each a case of imbalance, says investigator Alexej Abyzov, associate professor of biomedical informatics at the Mayo Clinic in Rochester, Minnesota. 

The full paper:- 

Modeling idiopathic autism in forebrain organoids reveals an imbalance of excitatory cortical neuron subtypes during early neurogenesis

Idiopathic autism spectrum disorder (ASD) is highly heterogeneous, and it remains unclear how convergent biological processes in affected individuals may give rise to symptoms. Here, using cortical organoids and single-cell transcriptomics, we modeled alterations in the forebrain development between boys with idiopathic ASD and their unaffected fathers in 13 families. Transcriptomic changes suggest that ASD pathogenesis in macrocephalic and normocephalic probands involves an opposite disruption of the balance between excitatory neurons of the dorsal cortical plate and other lineages such as early-generated neurons from the putative preplate. The imbalance stemmed from divergent expression of transcription factors driving cell fate during early cortical development. While we did not find genomic variants in probands that explained the observed transcriptomic alterations, a significant overlap between altered transcripts and reported ASD risk genes affected by rare variants suggests a degree of gene convergence between rare forms of ASD and the developmental transcriptome in idiopathic ASD.

 

Head circumference at birth is a useful measurement, but what really matters is how it changes over time.  Hyperactive pro-growth signaling affects more than just brain growth, it also affects muscle development, which is easy to notice.  I have highlighted the graphic below several times in this blog and in my book.  It is a good summary of what is going on.

 

Kv3.1

Regular readers will know that I like ion channels. The reason is that dysfunctions in these channels really should be treatable.  Usually we are looking for channel blockers, but today with Kv3.1 we are looking for channel enhancers.

Ion channel enhancers increase the activity of ion channels without directly opening them. They do this by increasing the number of open channels, increasing the opening time of each channel, or decreasing the closing time of each channel.

  

Grant aids drug discovery for autism, schizophrenia

At the heart of the study is a type of inhibitory neuron called GABAergic interneurons, which connect brain regions, playing vital roles in coordinating high-frequency brain activity. As a potential source of the excitatory/inhibitory imbalance in ASD and schizophrenia, evidence now points to malfunction of a type of potassium channel, Kv3.1, special to GABAergic interneurons. Denton and his team will aim to develop Kv3.1 enhancers and test their efficacy in restoring the balance of neural activity in a mouse model of ASD. In latter stages of this work, they’ll focus on key brain areas, using various lab techniques to carefully fill in neurological details surrounding any targeted drug effects.

“This grant creates opportunities for developing critically needed tool compounds to explore the role of Kv3.1 potassium channels in autism spectrum disorder and schizophrenia,” said Denton, professor of Anesthesiology and Pharmacology. “These are some of the most challenging and costly disorders going, and we’re excited to have this opportunity to take this work forward.”

 

Japanese researchers from the RIKEN Brain Science Institute are also thinking along the lines of targeting Kv3.1 to “correct aberrant developmental trajectories”. 

K_v3.1 channels regulate the rate of critical period plasticity 

The emergent function of fast-spiking PV-cell circuits during postnatal life may hold the key to a deeper understanding of critical periods in brain development (Reh et al., 2020) and the etiology of related mental illnesses as well (Do KQ and Hensch, 2015). The human neocortex notably shows a decrease in K_v3.1b channel protein in schizophrenia, a deficit that is restored by anti-psychotic drugs (Yanagi et al., 2014). Moreover, individuals with a KCNC1 loss-of-function variant can present intellectual disability without seizure and epilepsy (Poirier et al., 2017; Park et al., 2019). Our work points toward a prophylactic psychiatry that may target these particular channels to correct aberrant developmental trajectories.

 

As with head size, the “when” is also important with correcting Kv3.1.  The idea is to intervene at a very early age to redirect the developmental trajectory, rather than just to improve today’s functioning.

The logical question is what drugs will Professor Denton come up with to explore the benefit of targeting Kv3.1.  Perhaps someone can beat him to it and save us all a couple of decades?

If you look up Kv3.1 or the gene that encodes it called KCNC1 you can read all about it.

https://www.genecards.org/cgi-bin/carddisp.pl?gene=KCNC1

 

As expected, there is no shortage of channel blockers – Nifedipine (used a calcium channel blocker), Miconazole (an antifungal), Capsaicin (an active component of chili peppers), Fluoxetine (better known as Prozac, which is vitamin P to many people) plus many more.

Professor Denton is hunting for a channel enhancer.  Keep an eye on what he comes up with. He has $2.7 million over 4 years to play with. 

 

Transcranial pulse stimulation

Many autism parents do not like drug therapies, but often like the idea of zapping the brain from outside. I liked the idea of Photo biomodulation (PBMT) a form of light therapy that utilizes light sources including lasers or LEDs.

 

Low Level Laser Therapy (LLLT) for Autism – seems to work in Havana

Home/Clinic based Photobiomodulation/Laser Therapy in Autism - acting on Light Sensitive Ion Channels, Mitochondria, Lymph Nodes and more

 

You could potentially do Low Level Laser Therapy (LLLT) at home.

Professor Manual Casanova is a fan of transcranial magnetic stimulation (TMS).

Today’s paper below is about transcranial pulse stimulation, which I suppose we can just call TPS.

Transcranial pulse stimulation (TPS) is a non-invasive brain stimulation technique that uses pulsed electrical or magnetic fields to stimulate the brain. It is a relatively new technique, but it has the potential to be used for a variety of purposes, including:

• Treating neurological disorders such as Parkinson's disease, Alzheimer's disease, and depression
• Enhancing cognitive function, such as memory and attention
• Improving mood and well-being
• Reducing pain
• Promoting neuroplasticity, the ability of the brain to change and adapt

 

 

Effects of transcranial pulse stimulation on autism spectrum disorder: a double-blind, randomized, sham-controlled trial

 

Transcranial pulse stimulation has been proven effective to improve cognition, memory and depressive symptoms of Alzheimer’s disease, but supporting evidence on other neurological diseases or neuropsychiatric disorders remains limited. This study aimed to investigate the effects of transcranial pulse stimulation on the right temporoparietal junction, which is a key node for social cognition for autism spectrum disorder, and to examine the association between transcranial pulse stimulation and executive and social functions. This double-blinded, randomized, sham-controlled trial included 32 participants (27 males), aged 12–17 years with autism spectrum disorder. All eligible participants were randomized into either the verum or sham transcranial pulse stimulation group, on a 1:1 ratio, based on the Childhood Autism Rating Scale screening score. Sixteen participants received six verum transcranial pulse stimulation sessions (energy level: 0.2–0.25 mJ/mm2; pulse frequency: 2.5–4.0 Hz, 800 pulse/session) in 2 weeks on alternate days. The remaining 16 participants received sham transcranial pulse stimulation. The primary outcome measure included Childhood Autism Rating Scale score changes, evaluated by parents, from baseline to 3-month follow-ups. Secondary outcomes included a self-reported questionnaire responded to by parents and cognitive tests responded to by participants. A licensed mental health professional evaluated clinical global impression severity, improvement, efficacy and total score. Results revealed significant interactions in Childhood Autism Rating Scale and other secondary outcomes. Significant group and time effects were found in most secondary outcomes. Additionally, significant differences were found between the transcranial pulse stimulation and sham transcranial pulse stimulation groups in Childhood Autism Rating Scale and clinical global impression improvement and total score immediately after 2 weeks of transcranial pulse stimulation intervention (all P < 0.05), and effects were sustainable at 1- and 3-month follow-up, compared with baseline. The effect size of Childhood Autism Rating Scale (d = 0.83–0.95) and clinical global impression improvement (d = 4.12–4.37) were large to medium immediately after intervention and sustained at 1-month post-stimulation; however, the effects were reduced to small at 3-month post-stimulation (d = 2.31). These findings indicated that transcranial pulse stimulation over right temporoparietal junction was effective to reduce the core symptoms of autism spectrum disorder, as evidenced by a 24% reduction in the total Childhood Autism Rating Scale score in the verum transcranial pulse stimulation group. Additionally, the clinical global impression total score was reduced by 53.7% in the verum transcranial pulse stimulation group at a 3-month follow-up, compared with the baseline. Participants in the verum transcranial pulse stimulation group had shown substantial improvement at 1- and 3-month follow-ups, compared with baseline, although some of the neuropsychological test results were deemed statistically insignificant. Future replication of this study should include a larger sample derived from multi-nations to determine transcranial pulse stimulation as an alternative top-on treatment option in neuropsychiatry

 

TPS looks pretty impressive, based on the above study. TPS is available today, but it does need a lot of visits to the therapist. The effects are not permanent so you would have to keep going back for more.

People are doing transcranial direct current stimulation (tDCS) at home. 

People are zapping their brains at home to improve focus and clear brain fog. But is it safe?

For any kind of zapping therapy to be viable, it would have to be possible to do it yourself at home.

 

Targeting alpha 5 subunit of GABA_A receptors

Some earlier posts in this blog did get rather complicated.  One field that I looked at in rather painful detail was the GABA_A receptor. Some readers of this blog have children whose autism is entirely caused by a defect in this receptor, many other readers just see the effects of a GABA_A malfunction caused by a problem with NKCC1/KCC2 expression resulting from the GABA developmental switch failing to occur.

I looked to me that targeting alpha 3 and alpha 5 subunits could well enhance cognition.

Alpha 3 is targeted by low dose Clonazepam, thanks to Professor Catterall.

Alpha 5 was targeted to treat Down syndrome, using a new drug called Basmisanil (an inverse agonist of alpha 5 subunit of GABA_A). That work failed. I wrote about Cardiazol/ Pentylenetetrazol (PTZ) a drug that was widely used in the 1930s in mental hospitals to trigger seizures that were supposed to treat people with schizophrenia.  At much lower doses, it found a new purpose decades ago as an ingredient in cough medicine. 

The alpha 5 subunit is one of several subunits that can make up a GABA_A receptor. GABA_A receptors containing the alpha 5 subunit are thought to be involved in cognitive function, learning and memory, and mood regulation.

PTZ has been shown to block the action of GABA at alpha 5-containing GABAa receptors in animal studies.  

Variable Expression of GABRA5 and Activation of α5 -  a Modifier of Cognitive Function in Autism?

 

Sodium Benzoate and GABRA5 - Raising Cognitive Function in Autism 

Cardiazol, a failed Schizophrenia treatment from the 1930s, repurposed at low doses as a Cognitive Enhancer in Down Syndrome and likely some Autism

 

The logical human trial would be to use the cough mixture, Cardiazole that is already used in children. 

“We actual have quite a few readers from India and that is the only other country using this drug.  In India the producer is Nicholas Piramal and the brand name is Cardiazol Dicodid, it cost 30 US cents for 10ml.  So for less than $1, or 70 rupees, you might have a few months of cognitive enhancement, that is less than some people pay for 1 minute of ABA therapy.

If a few drops of this children’s cough medicine improves cognition please lets us all know.”

 

Back to recent research on alpha 5 that caught my attention.

 

An alpha 5-GABAa receptor positive allosteric modulator attenuates social and cognitive deficits without changing dopamine system hyperactivity in an animal model for autism

 Autism Spectrum Disorders (ASD) are characterized by core behavioral symptoms in the domains of sociability, language/communication, and repetitive or stereotyped behaviors. Deficits in the prefrontal and hippocampal excitatory/inhibitory balance due to a functional loss of GABAergic interneurons are proposed to underlie these symptoms. Increasing the postsynaptic effects of GABA with compounds that selectively modulate GABAergic receptors could be a potential target for treating ASD symptoms. In addition, deficits in GABAergic interneurons have been linked to dopamine (DA) system dysregulation, and, despite conflicting evidence, abnormalities in the DA system activity may underly some ASD symptoms. Here, we investigated whether the positive allosteric modulator of α5-containing GABA_A receptors (α5-GABA_ARs) SH-053-2’F-R-CH3 (10 mg/kg) attenuates behavioral abnormalities in a rat model for autism based on in utero VPA exposure. We also evaluated if animals exposed to VPA in utero present changes in the ventral tegmental area (VTA) DA system activity using in vivo electrophysiology and if SH-053-2’F-R-CH3 could attenuate these changes. In utero VPA exposure caused male and female rats to present increased repetitive behavior (self-grooming) in early adolescence and deficits in social interaction in adulthood. Male, but not female VPA rats, also presented deficits in recognition memory as adults. SH-053-2’F-R-CH3 attenuated the impairments in sociability and cognitive function in male VPA-exposed rats without attenuating the decreased social interaction in females. Male and female adult VPA-exposed rats also showed an increased VTA DA neuron population activity, which was not changed by SH-053-2’F-R-CH3. Despite sex differences, our findings indicate α5-GABA_ARs positive allosteric modulators may effectively attenuate some core ASD symptoms

 

Fine tuning alpha 5, perhaps you need more, perhaps less?

 

Neurobiology and Therapeutic Potential of α5-GABA Type A Receptors

Despite being a genetically heterogeneous disorder, the potential utility for mechanism-based GABAAR pharmacologic treatment with ASDs is supported by shared pathologies both in patients and related mouse models.

  

PAM α5 GABAAR Therapeutic Applications

Neurodevelopmental Disorders

Mouse models of neurodevelopmental disorders that present with insufficient inhibitory tone show improvement with positive modulators of GABAAR signaling. In the Scn1a+/− mouse model of Dravet syndrome, a severe childhood epileptic encephalopathy syndrome with hyperactivity and autism behaviors, abnormal social behaviors and fear memory deficits were rescued following treatment with a benzodiazepine, clonazepam (Han et al., 2014). In an ASD mouse model with reduced GABAAR-mediated inhibition, the BTBR T+tf/J mouse, the α2,3 and 5 PAM L-838,417, improved deficits in social interaction, repetitive behaviors, and spatial learning (Han et al., 2014).

 

Postweaning positive modulation of α5GABAA receptors improves autism‐like features in prenatal valproate rat model in a sex‐specific manner 

Autism spectrum disorder (ASD), as a common neurodevelopmental disorder that encompasses impairments in social communication and interaction, as well as repetitive and restrictive behavior, still awaits an effective treatment strategy. The involvement of GABAergic neurotransmission, and especially a deficit of GABA A receptors that contain the α5 subunits, were implicated in pathogenesis of ASD. Therefore, we tested MP‐III‐022, a positive allosteric modulator (PAM) selective for α5GABAA receptors, in Wistar rats prenatally exposed to valproic acid, as an animal model useful for studying ASD. Postweaning rats of both sexes were treated for 7 days with vehicle or MP‐III‐022 at two doses pharmacokinetically determined as selective, and thereafter tested in a behavioral battery (social interaction test, elevated plus maze, spontaneous locomotor activity, and standard and reverse Morris water maze). Additional rats were used for establishing a primary neuronal culture and performing calcium imaging, and determination of hippocampal mRNA levels of GABRA5, NKCC1, and KCC2. MP‐III‐022 prevented impairments in many parameters connected with social, repetitive and restrictive behavioral domains. The lower and higher dose was more effective in males and females, respectively. Intriguingly, MP‐III‐022 elicited certain changes in control animals similar to those manifested in valproate animals themselves. Behavioral results were mirrored in GABA switch and spontaneous neuronal activity, assessed with calcium imaging, and also in expression changes of three genes analyzed. Our data support a role of α5GABAA receptors in pathophysiology of ASD, and suggest a potential application of selective PAMs in its treatment, that needs to be researched in a sex‐specific manner. Lay Summary In rats prenatally exposed to valproate as a model of autism, a modulator of α5GABAA receptors ameliorated social, repetitive and restrictive impairments, and, intriguingly, elicited certain autism‐like changes in control rats. Behavioral results were mirrored in GABA switch and spontaneous neuronal activity, and partly in gene expression changes. This shows a role of α5GABAA receptors in pathophysiology of ASD, and a potential application of their selective modulators in its treatment.

 

Note the researchers actually know about the GABA switch and so measured mRNA levels of NKCC1 and KCC2.

Note also that the lower dose of MP‐III‐022 was more effective in males and the higher dose in females.

We even have the recent associated PhD thesis from Anja Santrač:-

 

The influence of positive modulation of GABAA receptors containing the alpha5 subunit on behavioral changes of mice and rats in models of autistic disorders

The role of α5 GABAA receptors in learning and memory is well known. Therefore, we decided to examine the effect of the selective positive allosteric modulator (PAM) MP-III-022 on learning and memory of healthy animals, as well as GABRA5 expression. After demonstrating the needed tolerability and potential procognitive effects, the ligand would be used in an animal model of autism spectrum disorders (ASD). ASD is a neurodevelopmental disorder that encompasses impairments in social communication and interaction, as well as repetitive and restrictive behavior, still without an effective treatment. In this context, animal models that imitate specific disease’s symptoms are an excellent tool of translational research. Some of the most frequently used models are BTBR T+ tf/J mouse strain (BTBR) and valproate prenatal model (VPA). Our experiments have shown that the variability of α5GABAA receptors’ roles depends on its level of expression and localization, on the type and protocol of cognitive tasks, the timing of testing and intensity of pharmacological modulation. Obtained results proved potential beneficial effects of MP-III-022 in cognitive tasks. The BTBR model failed to express sufficient face validity, while VPA demonstrated adequate face validity and in part construct validity. Thus, we decided to subacutely apply MP-III-022 to juvenile VPA rats. In control animals, treatment led to GABRA5 decrease and to impairments similar to ones seen in ASD, suggesting the possible role of this receptor in the pathogenesis of the disease. Most importantly, our results demonstrated the potential of α5 GABAA receptor PAMs in secondary prevention and treatment of ASD, with the caveat that the drug development program would require adaptations tailored to sex-specific differences revealed.

 

Good job Anja. For our Serbian speaking readers, here is the link to her thesis:-

https://nardus.mpn.gov.rs/bitstream/handle/123456789/21424/Disertacija_13513.pdf?sequence=1&isAllowed=y

Perhaps we should connect her with Professor Ben-Ari?

  

Conclusion

Fine tuning alpha 5 subunits of GABA_A receptors really should be followed up.  I think you need both options - a little bit more and a little bit less. It did not work for Roche in Down syndrome, but the potential remains.

Kv3.1 is another focused target for research, that very likely will become actionable. 

Transcranial pulse stimulation, like all the other zapping therapies, looks interesting, but it needs to be packaged in way that can actually be implemented every day at home.

In the meantime, at least getting your kid to wash the car is something we can all do.

Cesarian Delivery and Autism – another inconvenient truth?

Brasil is the C-section capital of the world, with rates in the public sector of 35–45%, and 80–90% in the private sector.

A recent study from the Karolinska Institute in Sweden, analysing 61 previous studies, has again shown a connection between birth by Cesarian Section and an increased risk of autism or indeed ADHD. 

C-sections account for just 16% of births in Sweden, but 32% in North America.

This of course prompted a reaction to reassure future mothers that they have nothing to fear, from experts in obstetrics who of course know nothing about the etiology of autism.  Mothers should be reassured, but trashing the study helps nobody.  Instead of a 1% risk of non-trivial autism, it rises to 1.3%. You still have more than a 98% chance of having a neurotypical child, all other factors being equal.  Without a medically necessary C-section, death is a real possibility.

It was a couple of years ago that the Karolinska Institute highlighted the fact that those with severe autism currently have a life expectancy of under 40 years.  Another inconvenient truth.

Association of Cesarean Delivery With Risk of Neurodevelopmental and Psychiatric Disorders in the Offspring 

Question  Is birth by cesarean delivery associated with an increased risk of neurodevelopmental and psychiatric disorders in the offspring compared with birth by vaginal delivery?

Findings  In this systematic review and meta-analysis of 61 studies comprising more than 20 million deliveries, birth by cesarean delivery was significantly associated with autism spectrum disorder and attention-deficit/hyperactivity disorder.

Meaning  The findings suggest that understanding the potential mechanisms behind these associations is important, especially given the increase in cesarean delivery rates for nonmedical reasons.

Abstract

Importance  Birth by cesarean delivery is increasing globally, particularly cesarean deliveries without medical indication. Children born via cesarean delivery may have an increased risk of negative health outcomes, but the evidence for psychiatric disorders is incomplete. 

Conclusions and Relevance  The findings suggest that cesarean delivery births are associated with an increased risk of autism spectrum disorder and attention-deficit/hyperactivity disorder, irrespective of cesarean delivery modality, compared with vaginal delivery. Future studies on the mechanisms behind these associations appear to be warranted. 

Very many things are known to slightly increase the odds of a person having autism and the more risk factors you have the more severely autistic you may be.  This ranges from maternal stress (anything from experiencing a hurricane, work stress, life trauma) to maternal/paternal age, obesity, gestational diabetes, alcohol/drug abuse, illness during pregnancy etc. This combines with whatever is in the parents’ DNA and random mutations that are bound to occur.    

A more rational reaction might be to investigate further why there might be a link and how you could counter any risk to children born by cesarian section.  You only have to read the existing research, or this blog.

There are 2 very good reasons why there should be a link between autism and C section, both have been covered in this blog.

1.     The microbiome comes from the mother. Science is only recently starting to understand the role of bacteria in health, but we know that it plays a key role in conditioning/calibrating the immune system of babies.  Once the immune system has been calibrated it is set for life.  Early exposure to bacteria is necessary and humans evolved to expect it.  If your immune system is over/under sensitive there will be consequences. Birth via C-section avoids exposure to bacteria in the birth canal, unless the newly arrived baby is “seeded” with bacteria from the mother. Mother’s milk is another key source of transferring the mother’s microbiome to the baby. 

2.     We saw that the birthing hormone Oxytocin plays a key role in triggering the “GABA switch” in new-borns. This is the process which transforms immature neurons with high chloride to mature neurons with low chloride shortly after birth.  During natural birth there is a surge in the hormone Oxytocin that is transferred to the baby, this causes the chloride transporter KCC2 to be further expressed and the “opposing” transporter NKCC1 to fade away.  In many people with severe autism their neurons remain in the immature state their entire life.  Just as you can replace the bacteria transfer lost in birth via C-section, there would be absolutely no reason why you could not replicate the surge in Oxytocin to "flip the GABA switch".

The recent study showed that elective C-sections (where the baby is in perfect health and not distressed) are associated with the elevated risk of both Autism and ASD.

Regular readers of this blog would probably be surprised if C-section did not increase autism prevalence.

The important thing is to acknowledge this likely connection and mitigate it, rather than try and fault the numerous studies that have shown the same effect.

The same of course applies to reducing the very small risk from vaccines, rather than construct new studies in a contrived way to show there is zero risk.   If you can safely and cheaply reduce the risk of a negative reaction to vaccines, why wouldn’t you?  Just follow Johns Hopkins example and give Ibuprofen or Montelukast (Singular) for a few days before and after and remember to never give Paracetamol/Acetaminophen (Tylenol) in response to fever after a vaccine. Paracetamol/ Acetaminophen reduces the body’s key antioxidant GSH just when the baby/child may need its neuroprotection most.

Some conditions are associated with preterm births, a good example is Cerebral Palsy (CP), which is twice as common in babies born very early. CP is rarely genetic and is usually considered to be caused by a complication during pregnancy, birth or shortly thereafter. I think you would find a correlation between C-sections and CP, but in this case I doubt you would find it in elective C-sections.   In other words C-sections do not “cause” CP, but they may be associated with it. The ID/MR often found in CP might be elevated by C-section and, if so, would be treatable.

Conclusion

In order to halt the rise in incidence of the disabling kinds of autism there should be steps taken to reduce some of the very many factors that are driving the increase, albeit each one sometimes by a tiny amount.

This would be a good application of all those thousands of autism research papers, many of which have shown what factors contribute to increased risk, that now sit gathering dust.

We are not at the stage of wide scale gene editing, but many simple steps can be taken today to improve future health.  This does not mean do not vaccinate, or avoid medically necessary C-sections; vaccinations and C-sections have saved millions of lives. But, why would you not want to take a good thing and make it even better?  That is what we humans tend to be good at, like the Swedes and their Volvos.

Perhaps take your C-section with a generous smear of Mum's bacteria and a shot of synthetic oxytocin?  

There will be more on Cerebral Palsy in a later post on D-NAC (Dendrimer N-Acetyl Cysteine). 

                                                               

Ketones and Autism Part 1 - Ketones, Epilepsy, GABA and Gut Bacteria

Source: The Gut Microbiota Mediates the Anti-Seizure Effects of the Ketogenic Diet

Today’s post is the first in a short series about ketones, that looks into very specific areas of the science.

There was an earlier post on the Ketogenic Diet (KD).

The Clever Ketogenic Diet for some Autism

We already know, anecdotally, that some people with autism respond well to the Ketogenic Diet (KD) and some to just ketone supplements. We also know from some very small clinical trials that a minority of those with autism benefit from the KD. 

What percentage of people with autism respond to ketones?

This is the important question and the simple answer appears to be a minority, albeit a significant minority. A lot depends on what you mean by autism and what you judge to be a response. From reading up on the subject I would estimate that about 20% respond well, but which 20%?

Why do some people with autism respond to ketones and how do you maximize the effect?

There is quite a lot of useful information in the literature, but it is clear that most people do not respond to ketones, so you really need to know if your case is one of the minority that do respond, before getting too carried away with changing diet. From research on people wanting to lose weight there are plenty of practical tips to maximize ketones, even drinking coffee produces more ketones; increasing the hours you fast each day also helps. 

If you have a healthy, sporty, child with autism you might want more ketones but you do not want to lose weight.

Do you have a responder? 

There does not yet appear to be a way to predict who will respond well to ketones, other than those people who have seizures. 

You can read all about ketones or just try them out. To try them out, a good place to start is with the exogenous ketone supplements mainly sold to people trying to lose weight. It looks like 10ml of C8 oil and 10ml of BHB ester is a good place to start and soon you may produce enough ketones to establish if someone is a responder, then you just have to figure out why they are a responder and then to maximize this effect, which might involve things other than ketone supplements.  

You can measure ketones in urine and so you can see whether you have reached the level found in trials that produces a positive effect in some people. If you can establish that you have indeed produced this level of ketones and you see no effect, then it is time to cross ketones and the ketogenic diet off your to-do list.  

If you are fortunate to find a responder, then you can move forward with trying to maximize the benefit. 

Ketone supplements

Ketone supplements can be extremely expensive when you use the recommended dosage and may contain quite large amounts of things that you might not want (sodium, potassium and calcium for example).  Always read the labels.

It is clear that some of these products are much more effective at producing ketones than others. For a change, you can measure their effectiveness and avoid wasting your money. Ketone testing strips are inexpensive. 

Ketone Posts 

·        Part1 - Ketones, epilepsy, GABA and gut bacteria

·        Ketones as a fuel, Alzheimer’s and GLUT1 deficiency

·        Ketones and microglia

·        Ketones as HDAC inhibitors and epigenetic modifiers

·        Ketones, exercise and BDNF

·        Maximizing ketones through diet, fasting, exercise, coffee and supplementation

Ketones, epilepsy, GABA and gut bacteria

Today’s post just looks at the effect of ketones on the neurotransmitter GABA, which should be inhibitory, but in much autism is actually excitatory. The GABA switch failed to flip just after birth and neurons remain in an immature state, due to too many NKCC1 chloride transporters and too few KCC2 chloride transporters. With too much chloride inside neurons GABA has an excitatory effect on neurons causing them to fire when they should not.

In epilepsy, too much excitation from Glutamate and too little inhibition from GABA may lead to seizures. So, in some types of epilepsy you want to increase GABA and reduce Glutamate, i.e. you want to increase the GABA/Glutamate ratio.

In autism it is not clear that increasing the GABA/Glutamate ratio is going to help, it all depends on the kind of autism. Much of this blog is about changing the effect of GABA (flipping the GABA switch) and not changing the amount of it.

In some people with autism and epilepsy ketones resolve the seizures but do not improve the autism.

In other people with autism and no seizures, ketones improve their autism. This may, or may not, be due to the effect ketones have on GABA.  

Two issues are looked into in this post: -

·        Do ketones affect NKCC1/KCC2 expression and hence the effect of GABA in the brain?

·        Do ketones affect the amount of GABA and Glutamate in key parts of the brain?

The good news is that research into epilepsy shows that ketones do indeed have an effect on GABA, but it is highly disputed whether they modify NKCC1/KCC2 expression and the GABA switch from immature to mature neurons. 

The KD and Epilepsy

The KD has been used to treat epilepsy for almost a century, but until very recently nobody really knew why it worked.

A recent very thoughtful study at UCLA has shown that the KD mediates its anti-seizure effects via changes to the bacteria in the gut. The researchers identified the two bacteria and then showed that, at least in mice, the same anti-seizure effect provided by the KD could be provided just by adding these two bacteria (i.e. no need to follow the ketogenic diet).

Gut bacteria play critical role in anti-seizure effects of ketogenic diet, UCLA scientists report

UCLA scientists have identified specific gut bacteria that play an essential role in the anti-seizure effects of the high-fat, low-carbohydrate ketogenic diet. The study, published today in the journal Cell, is the first to establish a causal link between seizure susceptibility and the gut microbiota — the 100 trillion or so bacteria and other microbes that reside in the human body’s intestines.

The ketogenic diet has numerous health benefits, including fewer seizures for children with epilepsy who do not respond to anti-epileptic medications, said Elaine Hsiao, UCLA assistant professor of integrative biology and physiology in the UCLA College, and senior author of the study. However, there has been no clear explanation for exactly how the diet aids children with epilepsy.

Researchers in Hsiao’s laboratory hypothesized that the gut microbiota is altered through the ketogenic diet and is important for the diet’s anti-seizure effects. Hsiao’s research team conducted a comprehensive investigation into whether the microbiota influences the ability of the diet to protect against seizures and if so, how the microbiota achieves these effects.

In a study of mice as a model to more thoroughly understand epilepsy, the researchers found that the diet substantially altered the gut microbiota in fewer than four days, and mice on the diet had significantly fewer seizures.

To test whether the microbiota is important for protection against seizures, the researchers analyzed the effects of the ketogenic diet on two types of mice: those reared as germ-free in a sterile laboratory environment and mice treated with antibiotics to deplete gut microbes.

“In both cases, we found the ketogenic diet was no longer effective in protecting against seizures,” said lead author Christine Olson, a UCLA graduate student in Hsiao’s laboratory. “This suggests that the gut microbiota is required for the diet to effectively reduce seizures.”

The biologists identified the precise order of organic molecules known as nucleotides from the DNA of gut microbiota to determine which bacteria were present and at what levels after the diet was administered. They identified two types of bacteria that were elevated by the diet and play a key role in providing this protection: Akkermansia muciniphila and Parabacteroides species.

With this new knowledge, they studied germ-free mice that were given these bacteria.

“We found we could restore seizure protection if we gave these particular types of bacteria together,” Olson said. “If we gave either species alone, the bacteria did not protect against seizures; this suggests that these different bacteria perform a unique function when they are together.”

The researchers measured levels of hundreds of biochemicals in the gut, blood and hippocampus, a region of the brain that plays an important role in spreading seizures in the brain. They found that the bacteria that were elevated by the ketogenic diet alter levels of biochemicals in the gut and the blood in ways that affect neurotransmitters in the hippocampus.

How do the bacteria do this? “The bacteria increased brain levels of GABA — a neurotransmitter that silences neurons — relative to brain levels of glutamate, a neurotransmitter that activates neurons to fire,” said co-author Helen Vuong, a postdoctoral scholar in Hsiao’s laboratory.

“This study inspires us to study whether similar roles for gut microbes are seen in people that are on the ketogenic diet,” Vuong said.

“The implications for health and disease are promising, but much more research needs to be done to test whether discoveries in mice also apply to humans,” said Hsiao, who is also an assistant professor of medicine in the David Geffen School of Medicine at UCLA.

On behalf of the Regents of the University of California, the UCLA Technology Development Group has filed a patent on Hsiao’s technology that mimics the ketogenic diet to provide seizure protection. It has exclusively licensed it to a start-up company Hsiao has helped to launch that will examine the potential clinical applications of her laboratory’s findings.

Here is Hsiao’s new start-up:

Bloom Science launches to treat epilepsy with ketogenic diet-associated gut microbes

“We are hacking the ketogenic diet to identify microbes that have therapeutic potential for the treatment of epilepsy,” says Bloom CEO Tony Colasin.

The San Diego-based start-up isn’t announcing how much seed funding it raised, but Colasin plans to move fast and get a product on the market in a mere two to three years. That’s because Bloom will first develop a medical food based on the two kinds of bacteria identified in Hsiao’s study. Bloom also has plans to optimize the bacterial strains for specific kinds of epilepsy to develop a traditional approved drug.

The way the bacteria help control seizures actually appears to be similar to the mechanism of many commercial ant epilepsy drugs. The microbes’ metabolism increases the ratio of inhibitory to excitatory neurotransmitters in the brain—specifically, higher levels of gamma-aminobutyric acid (GABA) relative to levels of glutamate.

Bloom is also considering how the microbiome benefits of the ketogenic diet could be helpful in other neurological conditions, including autism, depression, and Parkinson’s disease. “It is early days, but we are excited about the potential,” Colasin says.

Here is the full paper: -

The Gut Microbiota Mediates the Anti-Seizure Effects of the Ketogenic Diet

The ketogenic diet (KD) is used to treat refractory epilepsy, but the mechanisms underlying its neuroprotective effects remain unclear. Here, we show that the gut microbiota is altered by the KD and required for protection against acute electrically induced seizures and spontaneous tonic-clonic seizures in two mouse models. Mice treated with antibiotics or reared germ free are resistant to KD-mediated seizure protection. Enrichment of, and gnotobiotic co-colonization with, KD-associated Akkermansia and Parabacteroides restores seizure protection. Moreover, transplantation of the KD gut microbiota and treatment with Akkermansia and Parabacteroides each confer seizure protection to mice fed a control diet. Alterations in colonic lumenal, serum, and hippocampal metabolomic profiles correlate with seizure protection, including reductions in systemic gamma-glutamylated amino acids and elevated hippocampal GABA/glutamate levels. Bacterial cross-feeding decreases gamma-glutamyltranspeptidase activity, and inhibiting gamma-glutamylation promotes seizure protection in vivo. Overall, this study reveals that the gut microbiota modulates host metabolism and seizure susceptibility in mice.

Achieving the Gut Bacteria changes of the KD without the diet

Since the ketogenic diet (KD) is very restrictive, it would be much more convenient to achieve the GABA/Glutamate effect in a simpler way. You cannot currently just buy these two bacteria as a supplement.  There would seem to be 2 other obvious options: -

·        Take exogenous ketone supplements and hope this causes the same gut bacterial changes produced by the KD. No evidence exists.

·        Use other known methods to increase to increase Akkermansia muciniphila and Parabacteroides species in a similar way to that likely being developed by Bloom Sciences in San Diego. Bloom are developing a medical food to achieve this, so it will require a prescription and it will be costly. 

Increasing Akkermansia muciniphila can be achieved using fructooligosaccharides (FOS). FOS is included in many types of formula milk for babies and is sold as a supplement.

Metformin, a drug used to treat type 2 diabetes, greatly increases Akkermansia muciniphila. Vancomycin, the antibiotic that stays in the gut, also greatly increases Akkermansia muciniphila, but it is also going to wipe out many bacteria.

The research shows you also need the second bacteria Parabacteroides, of which there are many types. These bacteria are found in high levels in people following a Mediterranean type diet but it can be increased using Resistant Starch Type 4. This type of starch has been chemically modified to resist digestion. This starch is sold as food ingredient to add to bakery products.

http://www.ingredion.us/ingredients/na/versafibe-2470-dietary-fiber-22457b00.html

http://resistantstarchfoundation.com/resistant-starch-type-4/

Strategies to promote abundance of Akkermansia muciniphila, an emerging probiotics in the gut, evidence from dietary intervention studies

Viable A. muciniphila and fructooligosaccharides contently promote A. muciniphila. 

Metformin and vancomycin also significantly promote A. muciniphila.

https://biofoundations.org/increasing-and-maintaining-akkermansia-muciniphila-for-a-healthy-gut-microbiome/

“Akkermansia can also be increased by consuming polyphenol-rich foods, including:

·         pomegranate (attributed to ellagitannins and their metabolites)

·      grape polyphenols (grape seed extract) (proanthocyanidin-rich extracts may increase mucus secretion, therefore creating a favorable environment for Akkermansia to thrive) 

·   cranberries “ 

Two Healthy Diets Modulate Gut Microbial Community Improving Insulin Sensitivity in a Human Obese Population

The abundance of Parabacteroides distasonis (P = .025) and Faecalibacterium prausnitzii (P = .020) increased after long-term consumption of the Med diet and the LFHCC diet, respectively.

Beneficial modulation of the gut microbiota

Resistant starches types 2 and 4 have differential effects on the composition of the fecal microbiota in human subjects.

Abstract

BACKGROUND:

To systematically develop dietary strategies based on resistant starch (RS) that modulate the human gut microbiome, detailed in vivo studies that evaluate the effects of different forms of RS on the community structure and population dynamics of the gut microbiota are necessary. The aim of the present study was to gain a community wide perspective of the effects of RS types 2 (RS2) and 4 (RS4) on the fecal microbiota in human individuals.

METHODS AND FINDINGS:

Ten human subjects consumed crackers for three weeks each containing either RS2, RS4, or native starch in a double-blind, crossover design. Multiplex sequencing of 16S rRNA tags revealed that both types of RS induced several significant compositional alterations in the fecal microbial populations, with differential effects on community structure. RS4 but not RS2 induced phylum-level changes, significantly increasing Actinobacteria and Bacteroidetes while decreasing Firmicutes. At the species level, the changes evoked by RS4 were increases in Bifidobacterium adolescentis and Parabacteroides distasonis, while RS2 significantly raised the proportions of Ruminococcus bromii and Eubacterium rectale when compared to RS4. The population shifts caused by RS4 were numerically substantial for several taxa, leading for example, to a ten-fold increase in bifidobacteria in three of the subjects, enriching them to 18-30% of the fecal microbial community. The responses to RS and their magnitudes varied between individuals, and they were reversible and tightly associated with the consumption of RS.

CONCLUSION:

Our results demonstrate that RS2 and RS4 show functional differences in their effect on human fecal microbiota composition, indicating that the chemical structure of RS determines its accessibility by groups of colonic bacteria. The findings imply that specific bacterial populations could be selectively targeted by well-designed functional carbohydrates, but the inter-subject variations in the response to RS indicates that such strategies might benefit from more personalized approaches.

Ketones and NKCC1/KCC2  

The next part of this post does get complicated and so it will be of interest to a smaller number of readers; the question is whether ketones play a role in the (miss) expression of those two critical chloride transporters NKCC1 and KCC2. If ketones play a role, then they might have therapeutic potential in all those people with autism and Down Syndrome who respond to bumetanide.

Some researchers think ketones play such a role but the bumetanide for autism researchers disagree.

The ongoing disagreement in the research is about the role played by a lack of ketones in why in immature neurons GABA is excitatory. Regular readers will know that in a large group of autism the GABA switch never flips and so neurons remain in the immature state that was only supposed to last weeks after birth. The debate in the research is to what extent ketone bodies play a role.

We know that in much autism and indeed in those with other problems like neuropathic pain, there can be elevated chloride due to over-expression of NKCC1 (through which chloride enters) and under-expression of KCC2 (through which chloride ions exit neurons). 

GABA action in immature neocortical neurons directly depends on the availability of ketone bodies. 

Abstract

In the early postnatal period, energy metabolism in the suckling rodent brain relies to a large extent on metabolic pathways alternate to glucose such as the utilization of ketone bodies (KBs). However, how KBs affect neuronal excitability is not known. Using recordings of single NMDA and GABA-activated channels in neocortical pyramidal cells we studied the effects of KBs on the resting membrane potential (E(m)) and reversal potential of GABA-induced anionic currents (E(GABA)), respectively. We show that during postnatal development (P3-P19) if neocortical brain slices are adequately supplied with KBs, E(m) and E(GABA) are both maintained at negative levels of about -83 and -80 mV, respectively. Conversely, a KB deficiency causes a significant depolarization of both E(m) (>5 mV) and E(GABA) (>15 mV). The KB-mediated shift in E(GABA) is largely determined by the interaction of the NKCC1 cotransporter and Cl(-)/HCO3 transporter(s). Therefore, by inducing a hyperpolarizing shift in E(m) and modulating GABA signaling mode, KBs can efficiently control the excitability of neonatal cortical neurons.

In the early postnatal period, energy metabolism in the suckling rodent brain relies to a large extent on metabolic pathways alternate to glucose such as the utilization of ketone bodies (KBs). However, how KBs affect neuronal excitability is not known. Using recordings of single NMDA and GABA activated channels in neocortical pyramidal cells we studied the effects of KBs on the resting membrane potential (Em) and reversal potential of GABA-induced anionic currents (EGABA), respectively. We show that during postnatal development (P3–P19) if neocortical brain slices are adequately supplied with KBs, Em and EGABA are both maintained at negative

levels of about )83 and )80 mV, respectively. Conversely, a KB deficiency causes a significant depolarization of both Em (>5 mV) and EGABA (>15 mV). The KB-mediated shift in EGABA is largely determined by the interaction of the NKCC1 cotransporter and Cl)/HCO3 transporter(s). Therefore, by inducing a hyperpolarizing shift in Em and modulating GABA signaling mode, KBs can efficiently control the excitability of neonatal cortical neurons. Keywords: cortex, development, energy substrates, GABA, ketone bodies, resting potential.

showed that in the presence of KBs, values of EGABA in neocortical pyramidal neurons were close to Em, and did not change significantly during postnatal development, being maintained at about )80 mV (see Fig. 3). We cannot exclude the possibility that these values may differ in dendritic (Gulledge and Stuart 2003) or axonal (Price and Trussell 2006; Trigo et al. 2007; Khirug et al. 2008) compartments, an issue for future studies. Additionally, in this study we have limited our investigations to pyramidal cells, and the effects of KBs on interneurons remain to be explored. Nevertheless, the present observations suggest that energy substrates in the developing brain are an important issue to consider when studying neonatal neuronal excitability. Indeed, the most straightforward explanation for the difference between the results of the current study and those of previous studies of the development of neonatal GABA signaling lies in the fact that the brain of the suckling rodent relies strongly on KBs (Cremer and Heath 1974; Dombrowski et al. 1989; Hawkins et al. 1971; Lockwood and Bailey 1971; Lust et al. 2003; Page et al. 1971; Pereira de Vasconcelos and Nehlig 1987; Schroeder et al. 1991; Yeh and Zee 1976). Glucose utilization is limited at this age (Dombrowski et al. 1989; Nehlig, 1997; Nehlig et al. 1988; Prins 2008) because of the delayed maturation of the glycolytic enzymatic system (Dombrowski et al. 1989; Land et al. 1977; Leong and Clark 1984; Prins 2008). Use of glucose as the sole energy substrate caused an increase in neonatal neuronal [Cl)]i in our experiments, similar to that observed previously, while the addition of KBs resulted in a hyperpolarizing shift in both Em and EGABA. These results highlighted the need for caution in the interpretation of results obtained from neonatal brain slices superfused with standard ACSF. The cation chloride cotransporters NKCC1 and KCC2have been suggested to be the main regulators of neuronal Cl) homeostasis both during development (Farrant and Kaila 2007; Fiumelli and Woodin 2007) and in pathology (Galanopoulou, 2007; Kahle and Staley, 2008; Kahle et al. 2008).Although the possible contribution of anion exchangers to neuronal Cl) homeostasis has been noted previously (Farrant and Kaila 2007; Hentschke et al. 2006; Hubneret al. 2004; Pfeffer et al. 2009), they have not attracted the same degree of attention. Results from our study demonstrate, however, that the Cl)/HCO)3 transporter system is strongly involved in the KB-mediated regulation of [Cl)]i during postnatal development. Within this family, the Na-dependent Cl)/HCO)3 transporter (NDCBE), is of particular interest as it is expressed in the cortex (Chen et al. 2008) and has a strong dependence on ATP for its action (Chen et al. 2008; Davis et al. 2008; Romero et al. 2004). In addition, the sodium driven chloride bicarbonate exchanger(NCBE),(Giffardet al. 2003; Hubner et al. 2004; Lee et al. 2006) was expressed in the brain early during prenatal development and its expression preceded that of KCC2 (Hubner et al. 2004).

In neonatal neocortical neurons the interaction of NKCC1 and the Cl)/HCO3) transporter(s) maintained [Cl)]i, with KCC2 playing a less significant role at this stage. In the absence of KBs, when Cl)/HCO3) transporter(s) were less effective, the role of NKCC1 as a Cl) loader was especially noticeable and resulted in a depolarizing EGABA. During development the contribution of KCC2 to neocortical neuronal Cl) homeostasis is likely to increase (Stein et al. 2004; Zhang et al. 2006), and the balance between the actions of the different Cl) transporters in adults should be studied in the future. In humans, blood levels of KBs increase considerably during fasting, strenuous exercise, stress, or on the high-fat, low-carbohydrate ketogenic diet (KD) (Newburgh and Marsh 1920). A rapidly growing body of evidence indicates that the KD can have numerous neuroprotective effects (Gasior et al. 2006). During treatment with the KD, levels of KBs increase in both blood and brain, and cerebral metabolism adapts to preferentially use KBs as an alternate energy substrate to glucose (Kim do and Rho 2008). In children, the KD has been used as an effective treatment for medically refractory epilepsy (Freeman et al. 2007; Hartman and Vining 2007). However, despite nearly a century of use, the mechanisms underlying its clinical efficacy have proved elusive (Morris 2005; Bough and Rho 2007; Kim do and Rho 2008). Suckling rodents provide a natural model of the KD because of the high ketogenic ratio (Wilder and Winter 1922) of rodent milk (Page et al. 1971; Nehlig 1999). We propose that the KB-induced modulation of GABA-signaling may constitute a mechanism of anticonvulsive actions of the KD.

Now for the opposing views: -

Excitatory action of GABA on immature neurons is not due to absence of ketone bodies metabolites or other energy substrates

Summary

Brain slices incubated with glucose have provided most of our knowledge on cellular, synaptic, and network driven mechanisms. It has been recently suggested that γ‐aminobutyric acid (GABA) excites neonatal neurons in conventional glucose‐perfused slices but not when ketone bodies metabolites, pyruvate, and/or lactate are added, suggesting that the excitatory actions of GABA are due to energy deprivation when glucose is the sole energy source. In this article, we review the vast number of studies that show that slices are not energy deprived in glucose‐containing medium, and that addition of other energy substrates at physiologic concentrations does not alter the excitatory actions of GABA on neonatal neurons. In contrast, lactate, like other weak acids, can produce an intracellular acidification that will cause a reduction of intracellular chloride and a shift of GABA actions. The effects of high concentrations of lactate, and particularly of pyruvate (4–5 mm), as used are relevant primarily to pathologic conditions; these concentrations not being found in the brain in normal “control” conditions. Slices in glucose‐containing medium may not be ideal, but additional energy substrates neither correspond to physiologic conditions nor alter GABA actions. In keeping with extensive observations in a wide range of animal species and brain structures, GABA depolarizes immature neurons and the reduction of the intracellular concentration of chloride ([Cl⁻]_i) is a basic property of brain maturation that has been preserved throughout evolution. In addition, this developmental sequence has important clinical implications, notably concerning the higher incidence of seizures early in life and their long‐lasting deleterious sequels. Immature neurons have difficulties exporting chloride that accumulates during seizures, leading to permanent increase of [Cl⁻]_i that converts the inhibitory actions of GABA to excitatory and hampers the efficacy of GABA‐acting antiepileptic drugs.  

Depolarizing Actions of GABA in Immature Neurons Depend Neither on Ketone Bodies Nor on Pyruvate

GABA depolarizes immature neurons because of a high [Cl⁻]_i and orchestrates giant depolarizing potential (GDP) generation. Zilberter and coworkers (Rheims et al., 2009; Holmgren et al., 2010) showed recently that the ketone body metabolite dl-3-hydroxybutyrate (dl-BHB) (4 mm), lactate (4 mm), or pyruvate (5 mm) shifted GABA actions to hyperpolarizing, suggesting that the depolarizing effects of GABA are attributable to inadequate energy supply when glucose is the sole energy source. We now report that, in rat pups (postnatal days 4–7), plasma d-BHB, lactate, and pyruvate levels are 0.9, 1.5, and 0.12 mm, respectively. Then, we show that dl-BHB (4 mm) and pyruvate (200 μm) do not affect (i) the driving force for GABA_A receptor-mediated currents (DF_GABA) in cell-attached single-channel recordings, (2) the resting membrane potential and reversal potential of synaptic GABA_A receptor-mediated responses in perforated patch recordings, (3) the action potentials triggered by focal GABA applications, or (4) the GDPs determined with electrophysiological recordings and dynamic two-photon calcium imaging. Only very high non physiological concentrations of pyruvate (5 mm) reduced DF_GABA and blocked GDPs. Therefore, dl-BHB does not alter GABA signals even at the high concentrations used by Zilberter and colleagues, whereas pyruvate requires exceedingly high non-physiological concentrations to exert an effect. There is no need to alter conventional glucose enriched artificial CSF to investigate GABA signals in the developing brain.

Very recent research shows that ketones do not affect the expression of NKCC1 or KCC2. This would tend to support the argument of Ben Ari and Tyzio.

Ketogenic diet does not change NKCC1 and KCC2 expression in rat hippocampus

  

Nonetheless it seems that ketone bodies do indeed have an effect on GABA; they appear to change the hippocampal GABA/Glutamate ratio. 

So Tyzio might not be as right as he thought in his rebuttal paper when he said.

“suggesting, contrary to Zilberter and colleagues, that the antiepileptic actions of ketone bodies are not mediated by GABA signalling”

Tyzio is thinking about the resting membrane potential (Em) and reversal potential of GABA-induced anionic currents (EGABA).

At the end of that day a reduction in gamma-glutamylated amino acids, caused by changes in gut microbiota cause an increase in hippocampal GABA/Glutamate ratio. If you happen to have epilepsy this may mean less seizures.  

Conclusion

I think we can say with a fair degree of certainty that we now know why the ketogenic diet, and indeed the modified Atkins diet, greatly reduce seizures in many people with epilepsy. The diet changes the gut microbiota by increasing the amount of Akkermansia muciniphila and Parabacteroides species, the end result is an increase in GABA, the inhibitory neurotransmitter, inside the brain. In much epilepsy, more inhibition to neurons firing results is far less seizures.

GABA plays a key role in autism, albeit a complex one.

The ketone driven changes to GABA might explain why some people with autism respond to the KD or just ketone supplements, but ketones have many other effects relevant to autism that will be reviewed in later posts.