Showing posts with label CBD. Show all posts
Showing posts with label CBD. Show all posts

Wednesday 23 October 2019

GABAa receptor trafficking, Migraine, Pain, Light Sensitivity, Autophagy, Jacobsen Syndrome, Angelman Syndrome, GABARAP, TRPV1, PX-RICS, CaMKII and CGRP ... Oh and the "fever effect"

The mechanism controlling transporting just the “right” number of GABAA receptors

Today’s post is not for the faint-hearted.  It is another one that could just keep on rolling.  Ling will like it.

It again shows that GABAA receptors are at the centre of much autism, whether single gene or idiopathic. Today we highlight what can go wrong as these receptors are “transported”.

Today’s post also draws on several quite recent papers. It seeks to tie together some previous things mentioned in this blog like the symptoms of pain, particularly felt in the head, sensory sensitivity with dysfunction processes like autophagy and linking it all back to the GABAA receptor.  There is even a link at the end to the "fever effect", which occurs when a high temperature in some people causes a marked improvement in their autism symptoms.

We will come across some expensive drugs like Erenumab, the medical food PEA (Palmitoylethanolamide) and indeed Natasa’s favourite, CBD (Cannabidiol) and a newcomer CBDV (Cannabidivarin).   
We come across a protein called GABARAP (GABAA receptor associated protein) for the first time in this blog.  There is a vast amount in this blog about the GABAA receptor, how and why to modulate it. 

CaMKII makes an appearance, this is a protein kinase that is miss-regulated in much neurological disease. It changes the effect of many other proteins, acting just like a switch, by chemically adding phosphate groups to them. We have previously seen how important the protein kinases PKA, PKB and PKC are to autism.  Today add CaMKII to the list.

We come across another distinctive “face” of autism, this time it is Jacobsen syndrome, which I think is easily spotted by the trained eye, or some facial recognition software.  Jacobsen syndrome is a rare chromosomal disorder resulting from deletion of genes from chromosome 11 that includes band 11q24. This may include the gene that encodes the protein PX-RICS and, if so, it will lead to “autism”. Loss of that gene should be treatable with a GABA agonist.     

We also come back to that happy puppet syndrome (Angelman syndrome) which usually involves loss of the gene UBE3A, from chromosome 15. What I found interesting was that both Jacobsen syndrome and Angelman syndrome should share impaired GABAA receptor trafficking as a feature. They each have a different impediment that should reduce the number of functioning GABAA receptors. In the case of Angelman the impediment is CaMKII inhibition, in Jacobsen it is lack of the protein PX-RICS. Angelman syndrome may well respond to the same therapy as Jacobsen syndrome – a GABA agonist, of just a PAM (positive allosteric modulator, to “turn up the volume”).


GABARAP has multiple functions:

1.     Transport of freshly minted GABAA receptors

In order for newly minted GABAA receptors to get to their final destination it requires four “helpers”: GABARAP, PX-RICS, 14-3-3 and Dynactin.  In addition, you need a dose of CaMKII. If you lack any one of these four, you will end up with reduced expression of GABAA receptors. If CaMKII is overactivated you get too many GABAA receptors.

In Jacobsen Syndrome there is reduced GABAA receptor trafficking/transport, leading to reduced surface expression. (in effect not enough functioning GABAA receptors in situ).  In some people with this syndrome the part of their DNA which encodes PX-RICS is missing.  This lack of PX-RICS produces autism.  The autism-like behavioural abnormalities in PX-RICS-deficient mice are ameliorated by enhancing inhibitory synaptic transmission with a GABAAR agonist.

2.     GABARAP modulates TRPV1 expression

GABARAP also does something totally different, it modulates TRPV1 ion channels, that we have previously touched on in this blog.  This then triggers a cascade of effects relating to pain, neuralgia, migraine headaches, microglial activation, epilepsy and indeed longevity.

The simple function of TRPV1 is detection and regulation of body temperature. In addition, TRPV1 provides a sensation of scalding heat and pain. TRPV1 is also known as the capsaicin receptor.  Capsaicin is the active component of chilli peppers.
TRPV1 not only plays a role in pain, but is suggested to play a role in migraine. In migraine TRPV1 plays a role along with calcitonin gene-related peptide receptor (CGRPR). TRPV1 determines how much of the CGRPR protein is produced. CGRPR affects your metabolism broadly and as such plays a key role in longevity.  Ablation of select pain sensory receptors (TRPV1) or the inhibition of CGRP are associated with increased metabolic health and longevity.
Erenumab/Aimovig is a medication which targets CGRPR for the prevention of migraine. It was the first of the group of CGRPR antagonists to be FDA approved in 2018. It is a form of monoclonal antibody therapy in which antibodies are used to block the receptors for the protein CGRP, thought to play a major role in starting migraines.
Recent evidence suggests that TRPV1 may contribute to the onset and progression of some forms of epilepsy;  Cannabidivarin  (CBDV) and cannabidiol (CBD), activate and desensitize TRPV1.
TRPV1 also plays a crucial role in the activation of microglia. As the researchers put it “TRPV1 channels are critical brain inflammation detectorsmicroglia shifted toward an anti-inflammatory phenotype when TRPV1 is lacking.

So, if we jump a few steps forward we can see that desensitizing TRPV1 might be helpful for people with: -

·        Some epilepsy
·        Some neuralgia
·        Perhaps some with chronic migraine
·        People with activated microglia, which is most autism

We also can see that a dysfunction in GABARAP may itself contribute to worsening the above conditions via its effect on TRPV1.

Epilepsy is the most common neurological disorder, with over 50 million people worldwide affected. Recent evidence suggests that the transient receptor potential cation channel subfamily member 1 (TRPV1) may contribute to the onset and progression of some forms of epilepsy. V Since the two nonpsychotropic cannabinoids cannabidivarin (CBDV) and cannabidiol (CBD) exert anticonvulsant activity in vivo and produce TRPV1-mediated intracellular calcium elevation in vitro, we evaluated the effects of these two compounds on TRPV1 channel activation and desensitization and in an in vitro model of epileptiform activity. Patch clamp analysis in transfected HEK293 cells demonstrated that CBD and CBDV dose-dependently activate and rapidly desensitize TRPV1, as well as TRP channels of subfamily V type 2 (TRPV2) and subfamily A type 1 (TRPA1). TRPV1 and TRPV2 transcripts were shown to be expressed in rat hippocampal tissue. When tested on epileptiform neuronal spike activity in hippocampal brain slices exposed to a Mg2+-free solution using multielectrode arrays (MEAs), CBDV reduced both epileptiform burst amplitude and duration. The prototypical TRPV1 agonist, capsaicin, produced similar, although not identical effects. Capsaicin, but not CBDV, effects on burst amplitude were reversed by IRTX, a selective TRPV1 antagonist. These data suggest that CBDV antiepileptiform effects in the Mg2+-free model are not uniquely mediated via activation of TRPV1. However, TRPV1 was strongly phosphorylated (and hence likely sensitized) in Mg2+-free solution-treated hippocampal tissue, and both capsaicin and CBDV caused TRPV1 dephosphorylation, consistent with TRPV1 desensitization. We propose that CBDV effects on TRP channels should be studied further in different in vitro and in vivo models of epilepsy.

TRPV1 channels are critical brain inflammation detectors and neuropathic pain biomarkers in mice

The capsaicin receptor TRPV1 has been widely characterized in the sensory system as a key component of pain and inflammation. A large amount of evidence shows that TRPV1 is also functional in the brain although its role is still debated. Here we report that TRPV1 is highly expressed in microglial cells rather than neurons of the anterior cingulate cortex and other brain areas. We found that stimulation of microglial TRPV1 controls cortical microglia activation per se and indirectly enhances glutamatergic transmission in neurons by promoting extracellular microglial microvesicles shedding. Conversely, in the cortex of mice suffering from neuropathic pain, TRPV1 is also present in neurons affecting their intrinsic electrical properties and synaptic strength. Altogether, these findings identify brain TRPV1 as potential detector of harmful stimuli and a key player of microglia to neuron communication.

TRPV1 controls cortical microglia activation

In the healthy mature brain, microglial cells play a role in immune surveillance and ensure the maintenance of brain homeostasis. Upon injuries these cells shift to an activated state characterized by drastic changes in the cellular shape, functional behavior and by the release of different proinflammatory and immunoregulatory factors58,59. Conforming to the capsaicin-mediated induction of microglial chemotaxis29, we investigated whether TRPV1 stimulation regulates the morphology of microglial cells…. Thus, stimulation of TRPV1 induced a pro-inflammatory phenotype of microglia from WTs. Conversely, microglia shifted toward an anti-inflammatory phenotype when TRPV1 is lacking.

Angelman syndrome

Angelman syndrome (Happy puppet syndrome) is a genetic disorder that mainly affects the nervous system. Symptoms include a small head and a specific facial appearance, severe intellectual disability, developmental disability, speaking problems, balance and movement problems, seizures, and sleep problems. Children usually have a happy personality and have a particular interest in water. The symptoms generally become noticeable by one year of age.  Angelman syndrome is typically due to a new mutation rather than one inherited from a person's parents. Angelman syndrome is due to a lack of function of part of chromosome 15 inherited from a person's mother. Most of the time, it is due to a deletion or mutation of the UBE3A gene.

CaMKII inhibition underlies Angelman Syndrome

CaMKII is a serine/threonine-specific protein kinase that is regulated by the Ca2+/calmodulin complex. CaMKII is involved in many signaling cascades and is thought to be an important mediator of learning and memory. CaMKII is also necessary for Ca2+ homeostasis and reuptake in cardiomyocytes, chloride transport in epithelia, positive T-cell selection, and CD8 T-cell activation.
Misregulation of CaMKII is linked to Alzheimer’s disease, Angelman syndrome, and heart arrhythmia.

Recent evidence for CaMKII dysregulation in psychiatric diseases is reviewed.
Changes in postsynaptic structure and function appear to be central to multiple diseases.
Altered regulation of the CaMKIIα gene promoter may be a common mechanism among diseases.
CaMKII dysregulation in diverse brain regions may account for myriad disorders.
Although it has been known for decades that hippocampal calcium/calmodulin (CaM)-dependent protein kinase II (CaMKII) plays an essential role in learning and memory consolidation, the roles of CaMKII in other brain regions are only recently being explored in depth. A series of recent studies suggest that CaMKII dysfunction throughout the brain may underlie myriad neuropsychiatric disorders, including drug addiction, schizophrenia, depression, epilepsy, and multiple neurodevelopmental disorders, perhaps through maladaptations in glutamate signaling and neuroplasticity. I review here the structure, function, subcellular localization, and expression patterns of CaMKII isoforms, as well as recent advances demonstrating that disturbances in these properties may contribute to psychiatric disorders.

A Novel Human CAMK2A Mutation Disrupts Dendritic Morphology and Synaptic Transmission, and Causes ASD-Related Behaviors

Characterizing the functional impact of novel mutations linked to autism spectrum disorder (ASD) provides a deeper mechanistic understanding of the underlying pathophysiological mechanisms. Here we show that a de novo Glu183 to Val (E183V) mutation in the CaMKIIα catalytic domain, identified in a proband diagnosed with ASD, decreases both CaMKIIα substrate phosphorylation and regulatory autophosphorylation, and that the mutated kinase acts in a dominant-negative manner to reduce CaMKIIα-WT autophosphorylation. The E183V mutation also reduces CaMKIIα binding to established ASD-linked proteins, such as Shank3 and subunits of l-type calcium channels and NMDA receptors, and increases CaMKIIα turnover in intact cells. In cultured neurons, the E183V mutation reduces CaMKIIα targeting to dendritic spines. Moreover, neuronal expression of CaMKIIα-E183V increases dendritic arborization and decreases both dendritic spine density and excitatory synaptic transmission. Mice with a knock-in CaMKIIα-E183V mutation have lower total forebrain CaMKIIα levels, with reduced targeting to synaptic subcellular fractions. The CaMKIIα-E183V mice also display aberrant behavioral phenotypes, including hyperactivity, social interaction deficits, and increased repetitive behaviors. Together, these data suggest that CaMKIIα plays a previously unappreciated role in ASD-related synaptic and behavioral phenotypes.
SIGNIFICANCE STATEMENT Many autism spectrum disorder (ASD)-linked mutations disrupt the function of synaptic proteins, but no single gene accounts for >1% of total ASD cases. The molecular networks and mechanisms that couple the primary deficits caused by these individual mutations to core behavioral symptoms of ASD remain poorly understood. Here, we provide the first characterization of a mutation in the gene encoding CaMKIIα linked to a specific neuropsychiatric disorder. Our findings demonstrate that this ASD-linked de novo CAMK2A mutation disrupts multiple CaMKII functions, induces synaptic deficits, and causes ASD-related behavioral alterations, providing novel insights into the synaptic mechanisms contributing to ASD.

Jacobsen Sydrome

The signs and symptoms of Jacobsen syndrome can vary. Most affected people have delayed development of motor skills and speech; cognitive impairment; and learning difficulties. Behavioral features have been reported and may include compulsive behavior; a short attention span; and distractibility. Many people with the condition are diagnosed with attention deficit-hyperactivity disorder (ADHD). The vast majority of people with Jacobsen syndrome also have a bleeding disorder called Paris-Trousseau syndrome, which causes abnormal bleeding and easy bruising. 

People with Jacobsen syndrome typically have distinctive facial features, which include small and low-set ears; wide-set eyes (hypertelorism) with droopy eyelids (ptosis); skin folds covering the inner corner of the eyes; a broad nasal bridge; down-turned corners of the mouth; a thin upper lip; and a small lower jaw (micrognathia). Affected people often have a large head (macrocephaly) and a skull abnormality called trigonocephaly, giving the forehead a pointed appearance.

The Autism-Related Protein PX-RICS Mediates GABAergic Synaptic Plasticity in Hippocampal Neurons and Emotional Learning in Mice

GABAergic dysfunction underlies many neurodevelopmental and psychiatric disorders. GABAergic synapses exhibit several forms of plasticity at both pre- and postsynaptic levels. NMDA receptor (NMDAR)–dependent inhibitory long-term potentiation (iLTP) at GABAergic postsynapses requires an increase in surface GABAARs through promoted exocytosis; however, the regulatory mechanisms and the neuropathological significance remain unclear. Here we report that the autism-related protein PX-RICS is involved in GABAAR transport driven during NMDAR–dependent GABAergic iLTP. Chemically induced iLTP elicited a rapid increase in surface GABAARs in wild-type mouse hippocampal neurons, but not in PX-RICS/RICS–deficient neurons. This increase in surface GABAARs required the PX-RICS/GABARAP/14–3-3 complex, as revealed by gene knockdown and rescue studies. iLTP induced CaMKII–dependent phosphorylation of PX-RICS to promote PX-RICS–14-3-3 assembly. Notably, PX-RICS/RICS–deficient mice showed impaired amygdala–dependent fear learning, which was ameliorated by potentiating GABAergic activity with clonazepam. Our results suggest that PX-RICS–mediated GABAAR trafficking is a key target for GABAergic plasticity and its dysfunction leads to atypical emotional processing underlying autism.

There is a growing consensus that autism arises from the atypical regulation of the excitation/inhibition balance within specific neural microcircuitry. In terms of neural inhibition, autism is closely related to dysfunctional inhibitory signaling mediated by the γ-aminobutyric acid (GABA) type A receptors (GABAARs). Impaired presynaptic release of GABA and postsynaptic trafficking of GABAARs lead to autistic-like social behavior in mouse models of autism. There is a significant reduction in the number of GABAARs and GABAergic activity in certain brain areas of autistic individuals. Genetic association studies have revealed that several GABAAR subunits are linked to an increased risk for autism. GABAAR–mediated signaling is thus essential for the proper regulation of the excitation/inhibition balance associated with socio-emotional cognition.

PX-RICS, GABARAP and 14-3-3ζ/θ are localized in the specific dendritic compartments that are immunopositive for organelle markers for the endoplasmic reticulum (ER), ER exit sites and the trans-Golgi network. This structure, termed the dendritic satellite secretory pathway, is comprised of the dendritic ER and the Golgi outposts and is involved in the local synthesis, processing and transport of membrane-integral or secretory proteins in dendrites. The rapid increase in surface-expressed GABAARs after NMDA stimulation could be explained by the localization of the PX-RICS–dependent trafficking machinery in the dendritic secretory compartments.
Several lines of evidence suggest that the dysregulation of GABA signaling underlies atypical social behavior in autism However, there has been no report describing deficits in GABAergic plasticity that contribute to autistic features. The present study has shown that PX-RICS is essential for GABAergic iLTP and that loss of the PX-RICS function in mice leads to impaired cued fear learning. Cued fear learning is closely associated with GABAAR–mediated activity and plasticity in the amygdala and is inversely correlated with the severity of autistic symptoms. Considering all of these findings, we thus reason that PX-RICS–dependent GABAAR transport may play critical roles in emotional learning in the amygdala through the control of GABAergic synaptic plasticity and that the impairment of this transport mechanism may lead to improper socio-emotional processing, resulting in autistic-like atypical social behavior (Supplementary Fig. 7). Further elucidation of the functional link between GABAergic plasticity and socio-emotional learning could lead to a better understanding of autism pathogenesis and treatment. 
We have previously identified and characterized two splicing isoforms of GTPase-activating proteins specific for Cdc42 predominantly expressed in neurons of the cerebral cortex, amygdala and hippocampus: RICS (ARHGAP32 isoform 2) and PX-RICS (ARHGAP32 isoform 1) . RICS regulates NMDAR–mediated signaling at the postsynaptic density and axonal elongation at the growth cone. In contrast, PX-RICS forms an adaptor complex with GABARAP and 14-3-3ζ/θ to facilitate steady-state trafficking of the N-cadherin/β-catenin complex and GABAARs. PX-RICS is also responsible for autistic-like features observed in more than half of the patients with Jacobsen syndrome (JBS) [3]. Mice lacking PX-RICS/RICS show marked decreases in surface-expressed GABAARs and GABAAR–mediated inhibitory synaptic transmission, resulting in various autistic-like behaviors and autism-related comorbidities. Rare single-nucleotide variations in PX-RICS are also linked to non-syndromic autism, schizophrenia and alexithymia. These findings strongly suggest that dysfunction of PX-RICS–mediated GABAAR trafficking has severe effects on socio-emotional processing of the brain.
Our previous study described above showed that PX-RICS and other components of the GABAAR trafficking complex are required for constitutive transport of the receptor. In this study, we have focused on the role of PX-RICS in the activity–induced promotion of GABAAR trafficking during iLTP. Here we show that PX-RICS–mediated GABAAR trafficking is also involved in NMDAR activity–dependent trafficking of GABAARs and that PX-RICS is a key target of CaMKII for regulating GABAergic synaptic plasticity. Furthermore, we show that PX-RICS dysfunction in mice leads to impaired amygdala–dependent emotional learning, which manifests as autistic-like social behavior [3].

Supplementary Fig. 7. PX-RICS–mediated GABAAR trafficking underlies NMDAR–dependent GABAergic iLTP PX-RICS, GABARAP and 14-3-3s are assembled to form an adaptor complex that interconnects γ2-containing GABAARs (cargo) and dynein/dynactin (motor). Interaction
of PX-RICS with 14-3-3s depends on the phosphorylation activity of CaMKII, and this interaction is a critical regulatory point for GABAAR trafficking. When CaMKII activity is at a basal level, the PX-RICS–mediated trafficking complex has a role in steady-state transport of GABAARs to maintain the number of surface GABAARs as needed for proper synaptic inhibition.3 Neural activity that evokes moderate Ca2+ influx through NMDAR preferentially increases the activated form of CaMKII and elicits its translocation to inhibitory synapses, where it phosphorylates target proteins such as gephyrin and the GABAAR β3 subunit. Phosphorylated gephyrin and the GABAAR β3 subunit regulate the surface dynamics of GABAARs such as lateral diffusion and synaptic confinement. The present study has revealed that PXRICS
is a downstream CaMKII target associated with anterograde transport of
GABAARs. Enhanced PX-RICS phosphorylation increases the PX-RICS–14-3-3 complex and thereby drives de novo GABAAR surface expression, resulting in GABAergic iLTP. Dysfunction of this trafficking mechanism in the amygdala causes impaired GABAergic synaptic plasticity, which may contribute to deficits in socioemotional behavior as observed in PX-RICS/RICS–deficient mice and JBS patients with autism.

PX-RICS-deficient mice mimic autism spectrum disorder in Jacobsen syndrome through impaired GABAA receptor trafficking

Jacobsen syndrome (JBS) is a rare congenital disorder caused by a terminal deletion of the long arm of chromosome 11. A subset of patients exhibit social behavioural problems that meet the diagnostic criteria for autism spectrum disorder (ASD); however, the underlying molecular pathogenesis remains poorly understood. PX-RICS is located in the chromosomal region commonly deleted in JBS patients with autistic-like behaviour. Here we report that PX-RICS-deficient mice exhibit ASD-like social behaviours and ASD-related comorbidities. PX-RICS-deficient neurons show reduced surface γ-aminobutyric acid type A receptor (GABAAR) levels and impaired GABAAR-mediated synaptic transmission. PX-RICS, GABARAP and 14-3-3ζ/θ form an adaptor complex that interconnects GABAAR and dynein/dynactin, thereby facilitating GABAAR surface expression. ASD-like behavioural abnormalities in PX-RICS-deficient mice are ameliorated by enhancing inhibitory synaptic transmission with a GABAAR agonist. Our findings demonstrate a critical role of PX-RICS in cognition and suggest a causal link between PX-RICS deletion and ASD-like behaviour in JBS patients.


We now come back to TRPV1, which we saw is modulated by GABARAP.

GABAA receptor associated protein (GABARAP) modulates TRPV1 expression and channel function and desensitization

Transient receptor potential vanilloid (TRPV1) transduces noxious chemical and physical stimuli in high-threshold nociceptors. The pivotal role of TRPV1 in the physiopathology of pain transduction has thrust the identification and characterization of interacting partners that modulate its cellular function. Here, we report that TRPV1 associates with γ-amino butyric acid A-type (GABAA) receptor associated protein (GABARAP) in HEK293 cells and in neurons from dorsal root ganglia coexpressing both proteins. At variance with controls, GABARAP augmented TRPV1 expression in cotransfected cells and stimulated surface receptor clustering. Functionally, GABARAP expression attenuated voltage and capsaicin sensitivity of TRPV1 in the presence of extracellular calcium. Furthermore, the presence of the anchor protein GABARAP notably lengthened the kinetics of vanilloid-induced tachyphylaxia. Notably, the presence of GABARAP selectively increased the interaction of tubulin with the C-terminal domain of TRPV1. Disruption of tubulin cytoskeleton with nocodazole reduced capsaicin-evoked currents in cells expressing TRPV1 and GABARAP, without affecting the kinetics of vanilloid-induced desensitization. Taken together, these findings indicate that GABARAP is an important component of the TRPV1 signaling complex that contributes to increase the channel expression, to traffic and cluster it on the plasma membrane, and to modulate its functional activity at the level of channel gating and desensitization.

‘Entourage’ effectsof N‐palmitoylethanolamide and N‐oleoylethanolamide on vasorelaxation to anandamide occur through TRPV1 receptors

Age-Dependent Anti-seizure and Neuroprotective Effect of Cannabidivarin in Neonatal Rats

Neonatal seizures and seizures of infancy represent a significant cause of morbidity. 30–40% of infants and children with seizures will fail to achieve seizure remission with current anti-epileptic drug (AED) treatment. Moreover, pharmacotherapy during critical periods of brain development can adversely affect nervous system function. We, and others, have shown that early life exposure to AEDs including phenobarbital, phenytoin, and valproate are associated with induction of enhanced neuronal apoptosis during a confined period of postnatal development in rats. Thus, identification of new therapies for neonatal/infantile epilepsy syndromes that provide seizure control without neuronal toxicity is a high priority.
Current clinical trials report that modulation of the cannabinoid system with the phytocannabinoid cannabidiol exerts anti-seizure effects in children with epilepsy. While cannabidiol and the propyl analog cannabidivarin (CBDV) display anti-seizure efficacy in adult animal models of seizures/epilepsy, they remained unexplored in neonatal models. Therefore, we investigated the therapeutic potential of CBDV in multiple neonatal rodent seizure models. To evaluate the therapeutic potential of CBDV, we tested its anti-seizure efficacy in five models of neonatal seizures: pentylenetetrazole (PTZ), DMCM, hypoxia, kainate and NMDA-evoked spasms, each representing a different clinical seizure phenotype. We also evaluated the preclinical safety profile in the developing brain.
Postnatal day (P) 10 or P20 male, Sprague-Dawley rat pups were pretreated with CBDV or vehicle prior to chemically or hypoxia induced seizures. CBDV only displayed anticonvulsant effects in the P20 rat pups in the PTZ and DMCM models, with no effect on seizure severity or latency in the P10 animals. Therefore, we next measured the relative expression of known targets for CBDV (TRPV1, TRPA1) to determine a mechanism for which CBDV is anticonvulsant in P20, but not P10 animals. The P20 animals show increased expression of TRPV1 in key brain regions implicated in epileptogenic activity.
Together, these results indicate that modulation of the cannabinoid system in a receptor independent manner can provide seizure control in developing animals, but in an age specific manner. Further, during a developmentally sensitive neonatal period, drugs targeting the cannabinoid system do not induce neuronal apoptosis characteristic of many other AEDs. These results provide some of the first systemic, preclinical data evaluating CBDV in pediatric models of epilepsy.

Weight-based dosing of 10 mg/kg/day of CBDV for 12 weeks
Primary Outcome Measures  :
1.     Aberrant Behavior Checklist-Irritability Subscale (ABC-I) [ Time Frame: Change in ABC-I from Baseline to Week 12 (Change over 12 weeks) ]
Change in ABC-I from Baseline to Endpoint


Lack of Autophagy will reduce the number of GABAA receptors, by blocking GABARAP function

Regular readers will recall that one feature of autism and many other neurological diseases is a reduction in autophagy, which I likened to an intra-cellular garbage collection service. 

The very recent paper below shows that lack of autophagy blocks GABARAP from its job to transport freshly minted GABAA receptors.
If correct, this actually has very wide implications.

The disruption of MTOR-regulated macroautophagy/autophagy was previously shown to cause autistic-like abnormalities; however, the underlying molecular defects remained largely unresolved. In a recent study, we demonstrated that autophagy deficiency induced by conditional Atg7 deletion in either forebrain GABAergic inhibitory or excitatory neurons leads to a similar set of autistic-like behavioral abnormalities even when induced following the peak period of synaptic pruning during postnatal neurodevelopment. Our proteomic analysis and molecular dissection further revealed a mechanism in which the GABAA receptor trafficking function of GABARAP (gamma-aminobutyric acid receptor associated protein) family proteins was compromised as they became sequestered by SQSTM1/p62-positive aggregates formed due to autophagy deficiency. Our discovery of autophagy as a link between MTOR and GABA signaling may have implications not limited to neurodevelopmental and neuropsychiatric disorders, but could potentially be involved in other human pathologies such as cancer and diabetes in which both pathways are implicated.


You may have skipped to the conclusion to avoid all the science.

The conclusion is simple, you need to keep your GABAA receptors in tip top form if you want to avoid the symptoms of autism.

o   You need the right number of them
o   You need the right balance among the five constituent subunits
o   You need the correct level of chloride inside neurons so the receptors are not “working backwards”

All of the genes that encode proteins involved in the above are individually “autism genes”, because any one of them can disrupt the process.

Whether it is Dravet syndrome (GABAA receptor α2 subunit), Angelman syndrome, Jacobsen syndrome, Down syndrome or numerous other autism syndromes, not to mention idiopathic autism, check the above 3 bullet points.

Tune up/down your GABAA receptors!

Desensitizing TRPV1 looks interesting and not just for epilepsy.  TRPV1 appears to be essential for microglia in the in brain to be activated.  We know that in autism microglia in the brain are permanently activated, as if there was a threat.

I do think there is cross-talk (feedback loops etc) going on here, for example you can treat the severe epilepsy in Dravet syndrome by any of the following:-

·        KBr, to lower intracellular chloride
·        Low dose clonazepam to affect α subunits of GABAA receptors
·        CBD or CBDV to modify TRPV1

Note that Dravet syndrome is caused by a mutation in the gene that encodes the sodium ion channel Nav1.1, the dysfunction of GABAA receptors is a secondary effect. Also of interest is that the seizures that occur in Dravet syndrome are often triggered by hot temperatures or fever, so you can see how TRPV1 is indeed likely involved.  More generally in idiopathic autism, we have the "fever effect" when high temperatures trigger a reduction in autistic behaviors, making it the opposite of Dravet syndrome. 

On the one hand the biology behind the various problems may look horribly complicated and interwoven, the solutions appear to be much simpler and you have multiple options.

I await the results of the autism clinical trial of CBDV (Cannabidivarin) with interest.

Just impaired autophagy may lead to a reduction in GABAA receptors and the appearance of autistic features in an otherwise “normal” brain. This reminds us again of why autism is not a medical diagnosis, it is just a vague/subjective observation, which, in severe cases, should then trigger a thorough medical investigation.

Thursday 21 March 2019

PEA (Palmitoylethanolamide) Therapy for Autism? Targeting CB1 and CB2 without the need for Cannabis, plus PPARα and Microbiome changes

I was spoilt for choice this week; which half-finished post to complete and publish?  The deciding factor was seeing just how interested people seem to be in cannabis.  It is the big thing at Autism conventions, currently.

Italy is home to some unconventional medicine

Today we see how you can target these receptors without the need to use any kind of cannabis. 

I do spend time encouraging Monty’s 18-year-old big brother to avoid recreational drugs; so that is to declare my broader perspective on the issue. These drugs seem very popular among college students of every nationality, but according to my son particularly among those from countries where they cannot freely buy alcohol, like the US.

In today’s post we go back to look at PEA (Palmitoylethanolamide) which I proposed, and then rejected, as an autism therapy way back in 2014. PEA is widely used to treat neuropathic pain in Italy and Spain. PEA is a substance you already have in your body and it plays a role in inhibiting mast cells to degranulate, it is a PPARα agonist and it affects the Cannabinoid receptors CB1 and CB2.

I did recently come across PEA again when looking at case histories un-related to autism and I bought the identical product used in the case histories, and in the quantity required to make a genuine trial on myself.

My autism trial in 2014 did not show any effect, but I did not continue very long and I think I used the Dutch version, not the Italian product, naturally used in all the Italian research.

PEA does not cross the blood brain barrier well and is not absorbed well.  There are prodrugs of PEA that do cross the BBB, but these are not commercially available.

We have nonetheless seen many times that reducing inflammation in the periphery does, perhaps surprisingly, affect the brain.

Another point to mention, since the gut microbiome is perhaps even more fashionable as cannabis, is that PEA does indeed modify the microbiome.

My old post from 2014: -

Human Growth Factors, Autism and the Centenarian Nobel Laureate

It turns out that from 2015 onwards Italian researchers started to look at PEA as an autism therapy. They have tested it in two different mouse models of autism and they have published case histories in humans.

PEA is big in Italy because the underlying research is Italian and the Nobel Laureate in my 2014 post was an Italian.

Italian medicine – where anything goes?

Our reader Tatjana has an Italian autism doctor. The only Italian autism clinician/researcher I ever contacted was Antonio Persico, but he was one of the few that did not reply.

Today we get a chance to read Italian autism research and many Europeans will be surprised how "American" these Italian doctors seem to be. The long-time followers of alternative medical therapies may recall Dr Bradstreet, the American DAN doctor who died from gun shots, he had some research pals in Italy and they are among the authors of one of the case studies.

For our North American readers, in most of the world there is only one kind of doctor, an MD, and they tend to be much more tightly controlled than in the US. For example, in the United Kingdom you will not find a single DAN doctor, or MAPS doctor. There used to one DAN type doctor from New Zealand and he was banned from seeing patients with autism.

Italy seems to be more Californian, as you will see if you read the doctors’ full case histories.

Palmitoylethanolamide (PEA) in mouse models of Autism


·        Autistic-like behaviour of BTBR mice are reversed by PEA through PPAR-α activation.
·        PEA restores hippocampal BDNF signaling pathway and mitochondrial dysfunction.
·        PEA improves central and peripheral inflammatory state of BTBR mice.
·        PEA modulates gut microbiota composition in BTBR mice.


Autism spectrum disorders (ASD) are a group of heterogeneous neurodevelopmental conditions characterized by impaired social interaction, and repetitive stereotyped behaviours. Interestingly, functional and inflammatory gastrointestinal diseases are often reported as a comorbidity in ASDs, indicating gut-brain axis as a novel emerging approach. Recently, a central role for peroxisome-proliferator activated receptor (PPAR)-α has been addressed in neurological functions, associated with the behaviour. Among endogenous lipids, palmitoylethanolamide (PEA), a PPAR-α agonist, has been extensively studied for its anti-inflammatory effects both at central and peripheral level.
Based on this background, the aim of this study was to investigate the pharmacological effects of PEA on autistic-like behaviour of BTBR T+tf/J mice and to shed light on the contributing mechanisms.
Our results showed that PEA reverted the altered behavioural phenotype of BTBR mice, and this effect was contingent to PPAR-α activation. Moreover, PEA was able to restore hippocampal BDNF signaling pathway, and improve mitochondrial dysfunction, both pathological aspects, known to be consistently associated with ASDs. Furthermore, PEA reduced the overall inflammatory state of BTBR mice, reducing the expression of pro-inflammatory cytokines at hippocampal, serum, and colonic level. The analysis of gut permeability and the expression of colonic tight junctions showed a reduction of leaky gut in PEA-treated BTBR mice. This finding together with PEA effect on gut microbiota composition suggests an involvement of microbiota-gut-brain axis.
In conclusion, our results demonstrated a therapeutic potential of PEA in limiting ASD symptoms, through its pleiotropic mechanism of action, supporting neuroprotection, anti-inflammatory effects, and the modulation of gut-brain axis.


Autism spectrum disorder (ASD) is a condition defined by social communication deficits and repetitive restrictive behaviors. Association of the fatty acid amide palmitoylethanolamide (PEA) with the flavonoid luteolin displays neuroprotective and anti-inflammatory actions in different models of central nervous system pathologies. We hypothesized that association of PEA with luteolin might have therapeutic utility in ASD, and we employed a wellrecognized autism animal model, namely sodium valproate administration, to evaluate cognitive and motor deficits.


Two sets of experiments were conducted. In the first, we investigated the effect of association of ultra micronized PEA with luteolin, coultra micronized PEALUT® (coultraPEALUT®) in a murine model of autistic behaviors, while in the second, the effect of coultraPEALUT® in a patient affected by ASD was examined.


CoultraPEALUT® treatment ameliorated social and nonsocial behaviors in valproic acidinduced autistic mice and improved clinical picture with reduction in stereotypes in a 10yearold male child.


These data suggest that ASD symptomatology may be improved by agents documented to control activation of mast cells and microglia. CoultraPEALUT® might be a valid and safe therapy for the symptoms of ASD alone or in combination with other used drugs.

Palmitoylethanolamide (PEA) in human case studies

Introduction. Autism spectrum disorders are defined by behavioral and language atypias. Growing body of evidence indicates inflammatory mediators may contribute to the condition. Palmitoylethanolamide (PEA) is naturally occurring and has been available as a nonprescription medical food supplement in Europe since 2008. PEA has been tested in thousands of human subjects without any noted significant side effects. Here we report the first cases of the administration of PEA to two children with autism. Case Presentations. The first 13-year-old male child (Subject 1) presented with a total IgE of 572 IU/mL (nl < 200) and with low mature CD57+ natural killer cell counts (32 cells/µL; nl = 60–300 cells/µL) and with significant eczema and allergic stigmata. Expressive language, as measured by mean length of utterance, and overall autism severity as measured by the Childhood Autism Rating Scale, Second Edition, improved significantly. Atopic symptoms diminished. No side effects were reported. The second male child, age 15 (Subject 2), also displayed noticeable and rapid improvements in cognitive, behaviors, and sociability. Conclusion. Currently, there is no definitive treatment for autism condition. Palmitoylethanolamide could be an effective treatment for autism syndrome. We propose appropriate double-blind clinical trials to further explore palmitoylethanolamide efficacy and safety.

2.2. Subject 1

He is a 13-year-old autistic male with a well-documented history of significant atopic illnesses. He was originally diagnosed with autism by a child neurologist at age of 21 months following a significant regression in child development noted after a viral-like illness with associated high fevers (up to 45.5°C) at age of 15 months. Chronic urticaria, eczema, allergic rhinitis, and asthma have been treated with a variety of antihistamines, montelukast, and topical and systemic steroids since age of 2 years with only temporary beneficial effects. Serum food allergy testing revealed IgE mediated responses to the following: corn, peanut, soybean, wheat, milk, rice, egg, and numerous other dietary proteins. Both skin prick testing and serum IgE inhalant allergy testing revealed similar patterns of significant reactions to most antigens tested including dust mites and nearly all grasses, trees, weeds, molds, and both domestic and farm animals. Both traditional desensitization with injectable antigens and sublingual immunotherapies have failed to reduce allergic stigmata and had no noticeable effects on the course of the child's autism. Total serum IgE testing on numerous occasions has been significantly elevated and at the time PEA was introduced; it was 572 IU/mL (nl ≤ 200). Despite the high total levels of IgE and multiple IgE reactants, blood eosinophils have remained in the normal range throughout. Serum vitamin D-OH25 levels remain deficient despite extensive efforts at oral supplementation, inferring a defect in absorption of fat-soluble nutrients. Serum vitamin D-OH25 levels were at 21 ng/mL at the time PEA was started.
Cellular immune markers reflected specific abnormalities with deficient mature CD57+ natural killer (CD57+ NK) cell counts 32 cells/μL (nl = 60–300 cells/μL), with a total white count of 6.3 × 103/μL. However, the percentage of lymphocytes was high at 58%, while neutrophils were underrepresented at 33%, and absolute lymphocytes were also elevated at 3.5 × 103 cells/μL. All other obtained typical cell indices were in the normal range.
The Childhood Autism Rating Scale, Second Edition (CARS-2) [21], was administered as part of a routine evaluation of the child's severity prior to a change that is his academic placement. Prior to PEA, CARS-2 scoring placed the child at 43.5 total (82nd percentile) with a verbal communication subscale of 3 out 4. A speech assessment was performed including a mean length of utterance (MLU) measurement. MLU is based on the linguistic concept of morphemes as the smallest component of speech [6]. Speech therapists use MLU as a routine assessment and, in this case, MLU was obtained prior to a change in school placement. MLU was observed to be 3.0 prior to PEA and this corresponded to an age equivalent of approximately 34-35 months (just under a 3-year-old level of speech).

2.3. Post-PEA Supplementation (Subject 1)

After the practitioner reviewed the medical literature and the therapeutic rationale with his parents, they treated the child with Normast 600 mg tablets (Epitech Group Srl, Milano, Italy). Normast is available without prescription in Italy and Spain and is classified as “Food for Special Medical Purposes” by the Health Authorities of European Union member states according to standards set forth under European Commission Directive 1999/21/EC.
Initially, 1/2 tablet (300 mg) was given orally with water twice daily on an empty stomach to this boy (a child capable of swallowing tablets). After a week with no observable negative effects, the dose was increased to 1 (600 mg) tablet twice daily. Subject 1's parents returned for assessment after one month of oral supplementation with PEA. They reported “remarkable” changes in his behavior and his expressive language. Specifically, they noted he was spontaneously joining a conversation and commenting on various things happening in the home. The parents also reported the schoolteacher and speech therapist inquired about what had changed and noted similar positive changes at school. Equally noteworthy was the significant reduction in tantrums, outburst, self-talking, and stereotypies. Further, nose-picking, asthmatic cough, allergy stigmata (dark circles under eyes and nasal itching), nasal edema, and both skin eczema and urticaria diminished clinically after a month of PEA administration. No adverse effects were noted.
Following these comments, we re-administered the CARS-2 test and noted a total score of 32 (representing the 28th percentile) whereas the starting score was 43.5 (net change 11.5 points and 54 percentiles less). The subscale for expressive language changed to 2 out 4 and represented significant and noticeable changes. Due to these observations, we suggested repeating the MLU assessment for further definition of the degree of language changes. Reassessment of MLU also changed significantly to 5.4 (approximately 58 month's age-equivalency). This represented a 2-year age gain in expressive language in only one month of supplementation with PEA.
Total serum IgE testing after PEA was introduced remained essentially unchanged at 567 IU/mL (nl < 200). Serum vitamin D-OH25 levels rose to 46 ng/mL after PEA was started (possibly due to better absorption). CD57+ natural killer (CD57+ NK) cell counts increased after PEA to 52 cells/μL (nl = 60–300 cells/μL), with a total white count remaining similar at 5.6 × 103/μL. Noticeably the percentage of lymphocytes reduced to 44%, and the other cell indices were also in the normal range.

2.4. Subject 2

He is an Italian 15-year-old male, whom developed normally for the first 2 years, but near 3 years of age, the child developed a progressive loss of previously acquired language, eye contact, and social interaction. Ultimately, this was diagnosed with autism spectrum disorder. At 6 years of age, he experienced his first partial complex epileptic episode, described as the sudden onset of unconsciousness with resultant loss of motor control. The child fell and struck his head although no apparent concussion occurred. He was eventually treated with sodium valproate. The last episode of epilepsy was in March 2006. No recurrent seizures have been reported. After this event the family initiated dietary restriction by eliminating gluten, casein, yeast, sugar, and soy, and they also began supplementation of a multivitamin and additional folic acid. He started diet restriction and vitamins since 2007 at age of 8 years and never stopped. Following these changes the child's overall tone and energy improved and sensory integration also became more appropriate.
The Autism Treatment Evaluation Checklist (ATEC) [22] test was administered as part of a routine evaluation of the child's severity prior to a change in his academic placement. The ATEC has been shown to correlate with the CARS test in domains of sensory/cognitive awareness, speech/language, and sociability [23]. Prior to PEA supplementation the ATEC total scoring was in the mild to moderate range, specifically 25 (a lower number is better): speech 6 of 28 points, sociability 3 of 40 points, sensory/cognitive 1 point (normal), and overall health/behaviors 15 of 75 points. Total serum IgE testing was in the normal range at 127 IU/mL (nl < 200). Serum vitamin D-OH25 levels were normal at 45.10 ng/mL at the time PEA was started. We conducted the observations until three months after treatment. The boy never stopped diet restrictions and multivitamin supplementation since 2007.

2.5. PEA Supplementation

After practitioner NA reviewed the medical literature and the therapeutic rationale with his parents, they treated the child with Normast 600 mg tablets (Epitech Group Srl, Milano, Italy). After three months of supplementation, ATEC total score reduced by half (12 (after) versus 25 (before) points). The individual scores in the separate domains were speech 4 points, sociability 1 point, sensory/cognitive 0 points, and overall health/behaviors 7 points (Figure 1, Table 1). Also shown for comparison in (Figure 1, Table 1) are the child's improvements in aggression and cognitive and behavioral skills. Language showed mild improvements. In addition, in this case, total serum IgE reduced by half (66.0 post compared to 127 IU/mL pre-PEA). No adverse effects were noted (i.e., nausea, fever, hyperactivity, sleep disorders, skin rash, allergic reactions, and gastrointestinal problems).

Dynamics on a scale of Autism Treatment Evaluation Checklist (ATEC) in Subject 2 autistic patient before (baseline) PEA supplementation and three months later. The purpose of the ATEC is to measure change in an individual due to various interventions, that is, the difference between the initial (baseline) ATEC scores and later ATEC scores

4. Conclusion

In this original report of two cases, we show PEA-mediated effects may be beneficial for treating core symptoms of autism. PEA is a well-studied, apparently safe even in the pediatric population, anti-inflammatory capable of regulating mast cells and modulating immune chemistry. It is manufactured in the EU and prepared under strict standards according to Good Manufacturing Practice (GMP). PEA also appears to be an atypical endocannabinoid, with additional anti-inflammatory effects mediated via the PPAR pathway. In the United States, recent prevalence data indicate greater than 2% of boys may have ASDs [35, 36]. Given the urgent need for safe and effective strategies for treating ASDs, we propose appropriate double-blind controlled clinical trials to explore further the potential for PEA efficacy and its safety.


This study is dedicated to the memory of Professor James Jeffrey Bradstreet who passed away on June 19, 2015. His contribution to this work is fundamental.

Patient History

The participating physician who brought to our attention the patient for this study is part of our group's collaborative clinical network. This male child was born at term with natural spontaneous delivery [39 weeks + 6 days with an Apgar index between 8 and 10] and was found to have Tetralogy of Fallot, which was corrected surgically at the age of 3 months without complications. As is typical with autism, physical, and mental development appeared normal during the first few years of life. At the age of 28 months febrile episodes occurred, which were treated with antibiotics and nonsteroidal anti-inflammatory drugs. In April 2005, the parents reported three febrile episodes in succession characterized by inflammation of the upper respiratory tract; these were treated with oral antibiotics and nonsteroidal anti-inflammatory drugs and with paracetamol to control hyperpyrexia. At this time, over the course of 30 days, behavioral changes suddenly emerged: the child began to show rejection behavior of strangers with weeping and escape responses toward other children of same age; hyperactivity with constant running; changed eating behavior with refusal of food and limiting the choice only to 3–4 kind of aliments; up to 7–8 bowel movements per day; loss of acquired verbal skills, avoidance of gazing at people, and appearance of motor stereotypes. Approximately 30 days after onset of the first episode of fever, the child was admitted to the day hospital at the Pediatric Neuropsychiatry Clinic of Saint Gerardo Hospital—Monza, where a diagnosis of “delay of verbalization and socialization” with suspected autism was made. The child lives with his family in a provincial town, far from sources of industrial pollution. The mother (36 years old at time of childbirth) did not take any medications during pregnancy with the exception of supplementation with folic acid, iron, calcium, and magnesium; amniocentesis was performed at week 16 without complications and the remainder of the pregnancy followed a regular course. A genetics analysis of the patient was not performed, although there was no family history of autism.

Protocol with Co‐UltraPEA‐LUT®

The child was 10 years old when this study began. After diagnosis of ASD the child underwent 6 months of psychotherapy with his parents, without benefit. Psychomotor and speech therapy produced some benefit on behavior. A glutenfree/caseinfree diet and vitamin supplements led to a rapid improvement in sleep–wake behavior and eye contact. However, vitamin supplements in particular, at doses far exceeding their daily recommended allowance, were suspended almost immediately due to worsening of hyperactivity and stereotypical behaviors. From time to time, the patient underwent homotoxicologicalhomeopathic therapy with favorable results, especially with regard to mercury detoxification and treatment with probiotics.
Given the involvement of brain inflammation in the development of ASD, as well as the presence of activated mast cells 19, we undertook a therapy with the anti-inflammatory and mast cell modulating agent coultraPEALUT®, at a dose of 700 mg+70 mg b.i.d. (Glialia® microgranules, Epitech Group SpA, Italy) for 1 year. During the study period (May 2012–2013), the child continued the gluten and caseinfree diet. The patient was evaluated before treatment, after ~5 months and ~12 months using the ATEC questionnaire ( and by a quarterly questionnaire administered to the child's teachers for the assessment of motor stereotypic behaviors ( The ATEC questionnaire is sensitive enough to measure changes in the child's condition and consists of four subgroup scales: Scale I. Speech/Language/Communication (14 items—scores can differ from 0 to 28), Scale II. Sociability (20 items—scores can differ from 0 to 40), Scale III. Sensory/Cognitive Awareness (18 items—scores can differ from 0 to 36), and Scale IV. Health/Physical Behavior (25 items—scores can differ from 0 to 75). The four subgroup scores can be utilized to calculate a total score (total scores can range from 0 to 180). The scores are estimated according to the reply and corresponding subscale. The higher the subscale and total score, the more impaired is the patient. The lower the subscale and total score, the less impaired the patient. The stereotyping behavior was evaluated, before, during, and after 12 months by means of a fivestep scale of intensity as follows:
0 = symptom or behavior not present; 1 = rarely present and not linked to particular situations; 2 = present only in precise moments (e.g., before sleeping); 3 = present if the child is not actively involved in the environmental situation; 4 = very present and difficult to interrupt.
In addition to cognitive, behavioral, and motor symptoms, the child experienced an elevated frequency of nighttime enuresis, independent of time of year, general conditions or diet. This problem is often present in autistic children over five years of age 43.

Clinical data

Clinical Assessment of an Autistic Patient Treated with Co‐ultraPEA‐LUT®

CoultraPEALUT® treatment was well tolerated by the patient. Evaluation using the ATEC test revealed a decrease of scores (both total score and subgroup scores), indicative of an improved behavioral outcome of about 23%. The most evident effect over time appeared in the sociability subgroup, where the score decreased from 24 to 13 (Table 1). CoultraPEALUT® reduced most indices of hyperactivity, as demonstrated by reduction in motor stereotypies which from the very beginning were present only rarely or at precise moments (Table 2).

Table 1. ATEC scores in response to PEALUT® treatment in a 10yearold child with ASD
Autism treatment evaluation checklist
Time (months)
I. Speech
II. Sociability
III. Sensory/Cognitive
IV. Health/Physical/Behavior

Table 2. Motor stereotypy changes in response to PEALUT® treatment in a 10yearold child with ASD
Motor stereotypies
Time (months)
Head banging
Flapping tremor
Fingers movements
Walking on Tip‐Toes
Skill stereotypies (e.g., quickly turning a knife on a table)
Vocal stereotypy (unmotivated screaming, frog noise, grunts)
Clinical improvement, as highlighted by the questionnaire results, coincided with significant progress in cognitive behavior as observed by parents and teachers. The patient is, in fact, now more able to understand simple commands and execute them with ease; eye contact is much improved and the child's behavior far more affectionate. Unfortunately, no significant progress in speech profile has been observed until now.
In the described case report, chronic coultraPEALUT® reduced behavioral alterations of a child with ASD. Our observations are consistent with the results obtained after treatment with ultra micronized PEA, administered alone, in two children suffering from autism spectrum disorder 59. It is important to point out that children with autism experience a higher incidence of anxiety, particularly social anxiety, than children with other developmental disorders or with a normal development 60. Anxiety in children with ASD is related to sensory overresponsivity and gastrointestinal problems, suggesting that these pathological states represent interconnected phenomena, sharing common underlying mechanisms mediated by mast cell and microglia activation. Our data suggest that ASD symptomatology may be improved by agents documented to control activation of mast cells and microglia. These results, while pointing to coultraPEALUT® as a valid and safe treatment in the autism symptoms—alone or in combination with other drugs—clearly need to be confirmed in a larger number of cases. The proofofprinciple study presented here encourages further investigations.

Palmitoylethanolamide modes of action

·        PPARα agonist

·        Indirect activation of cannabinoid receptors CB1 and CB2, which will affect microglial activation (M0, M1 M2) as explained in this previous post:
·        Modification of gut microbiota

·        Mast cells stabilization, via PPARα

The effects of the PEA are due to its interaction with several pathways: at first, it reduces, via the peroxisome proliferator-activated receptor alpha (PPARα), the recruitment and activation of mast cells at sites of nerve injury and the release of pro-inflammatory mediators from these cells [3, 4]; secondly, it inhibits the microglia activation and the recruitment of mast cells into spinal cord after peripheral nerve injury, as well as following spinal neuroinflammation or spinal cord injury [5, 6]. In the beginning, PEA was also supposed to be an agonist of the cannabinoid type II receptor (CB2) [7]; subsequently, in their research, Sugiura et al. have demonstrated that PEA has just a very low affinity for this receptor [8], clarifying why CB2 antagonists do not inhibit some of its anti-inflammatory effects [9]. Anyhow, PEA indirectly activates CB2 and the cannabinoid receptor type 1 (CB1) [10], down-modulating fatty acid amide hydrolase (FAAH), the enzyme responsible of the degradation of the anandamide (AEA), a CB1 agonist [11].
Several studies focused on the use of PEA in a multitude of chronic pain conditions. For example, it can have a beneficial effect like adjuvant for the treatment of the low back pain [12] or it was used alone for chronic pain management in critically ill older patients, where the use of traditional analgesics can lead to high risk of adverse effect [13]. Encouraging results have been shown in the treatment of non-surgical radiculopathies with an ultra-micronized formulation of PEA [14] and the combination therapy with alpha-lipoic acid to reduce chronic prostatitis/chronic pelvic pain syndrome [15].

Now to Cannabis

The autism research on cannabis is rather light on the science, but it does exist.

Cannabinoid receptors

Before the 1980s, it was often speculated that cannabinoids produced their physiological and behavioral effects via nonspecific interaction with cell membranes, instead of interacting with specific membrane-bound receptors. The discovery of the first cannabinoid receptors in the 1980s helped to resolve this debate.[7] These receptors are common in animals, and have been found in mammalsbirdsfish, and reptiles. At present, there are two known types of cannabinoid receptors, termed CB1 and CB2,[1] with mounting evidence of more.[8] The human brain has more cannabinoid receptors than any other G protein-coupled receptor (GPCR) type.[9]

Cannabinoid receptor type 1

 CB1 receptors are found primarily in the brain, more specifically in the basal ganglia and in the limbic system, including the hippocampus [1] and the striatum. They are also found in the cerebellum and in both male and female reproductive systems. CB1 receptors are absent in the medulla oblongata, the part of the brain stem responsible for respiratory and cardiovascular functions. CB1 is also found in the human anterior eye and retina.[10]

Cannabinoid receptor type 2

CB2 receptors are predominantly found in the immune system, or immune-derived cells[11] with the greatest density in the spleen. While found only in the peripheral nervous system, a report does indicate that CB2 is expressed by a subpopulation of microglia in the human cerebellum.[12] CB2 receptors appear to be responsible for the anti-inflammatory and possibly other therapeutic effects of cannabis seen in animal models.[11]

Cannabidiol (CBD) is non-psychotropic. Recent evidence shows that the compound counteracts cognitive impairment associated with the use of cannabis.[15] Cannabidiol has little affinity for CB1 and CB2 receptors but acts as an indirect antagonist of cannabinoid agonists.[16] 

There is now strong evidence that CBD treats chronic pain, and it is used by many in the form of a topical oil treatment

Tetrahydrocannabinol (THC) is one of at least 13 cannabinoids identified in cannabis. THC is the principal psychoactive constituent of cannabis. With chemical name (−)-trans-Δ⁹-tetrahydrocannabinol, the term THC also refers to cannabinoid isomers.
The actions of THC result from its partial agonist activity at the cannabinoid receptor CB1 (Ki = 10 nM[19]), located mainly in the central nervous system, and the CB2 receptor (Ki = 24 nM[19]), mainly expressed in cells of the immune system.[20]The psychoactive effects of THC are primarily mediated by the activation of cannabinoid receptors, which result in a decrease in the concentration of the second messenger molecule cAMP through inhibition of adenylate cyclase.[21

We know that we can use CB1 and CB2 to shift microglia away from the M2 state. In the post below:

I extracted this section to remind readers: -

Molecules to Skew M2 Polarization of Microglia                                                                                                                   

7.1. Endocannabinoids and Cannabinoid Receptors

Based on these results, it is strongly suggested that 2-AG-CB1 axis contributes to polarization and maintenance of M1 microglia, while 2-AG-CB2 axis acts as a switch from M1 to M2 polarization of microglia (Figure 4). CB2 agonists are known to induce phosphorylation of AMP-activated protein kinase (AMPK), suggesting that the CB2 plays an important role in AMPK-mediated anti-oxidative and cytoprotective effects [119,120,121]. Furthermore, 2-AG is reported to activate PPAR-γ in M2 macrophages [122]. Thus, AMPK may be one of key signal molecules for the switch to M2 polarization. Besides endocannabinoids, adiponectin and ghrelin can induce down-stream signal transduction of their receptors via AMPK and therefore these molecules may be involved in skewing M2 polarization of microglia

Very recent cannabis in autism literature: -

Real life Experience of Medical Cannabis Treatment in Autism: Analysis of Safety and Efficacy

There has been a dramatic increase in the number of children diagnosed with autism spectrum disorders (ASD) worldwide. Recently anecdotal evidence of possible therapeutic effects of cannabis products has emerged. The aim of this study is to characterize the epidemiology of ASD patients receiving medical cannabis treatment and to describe its safety and efficacy. We analysed the data prospectively collected as part of the treatment program of 188 ASD patients treated with medical cannabis between 2015 and 2017. The treatment in majority of the patients was based on cannabis oil containing 30% CBD and 1.5% THC. Symptoms inventory, patient global assessment and side effects at 6 months were primary outcomes of interest and were assessed by structured questionnaires. After six months of treatment 82.4% of patients (155) were in active treatment and 60.0% (93) have been assessed; 28 patients (30.1%) reported a significant improvement, 50 (53.7%) moderate, 6 (6.4%) slight and 8 (8.6%) had no change in their condition. Twenty-three patients (25.2%) experienced at least one side effect; the most common was restlessness (6.6%). Cannabis in ASD patients appears to be well tolerated, safe and effective option to relieve symptoms associated with ASD.

Side Effects

The most common side effects, reported at six months by 23 patients (25.2%, with at least one side effect) were: restlessness (6 patients, 6.6%), sleepiness (3, 3.2%), psychoactive effect (3, 3.2%), increased appetite (3, 3.2%), digestion problems (3, 3.2%), dry mouth (2, 2.2%) and lack of appetite (2, 2.2%).
Out of 23 patients who discontinued the treatment, 17 (73.9%) had responded to the follow-up questionnaire at six months. The reasons for the treatment discontinuation were: no therapeutic effect (70.6%, twelve patients) and side effects (29.4%, five patients). However, 41.2% (seven patients) of the patients who discontinued the treatment had reported on intentions to return to the treatment.
Cannabis as a treatment for autism spectrum disorders patients appears to be well-tolerated, safe and seemingly effective option to relieve symptoms, mainly: seizures, tics, depression, restlessness and rage attacks. The compliance with the treatment regimen appears to be high with less than 15% stopping the treatment at six months follow-up. Overall, more than 80% of the parents reported at significant or moderate improvement in the child global assessment.
The exact mechanism of the cannabis effects in patients with ASD is not fully elucidated. Findings from ASD animal models indicate a possible dysregulation of the endocannabinoid (EC) system11,12,13,14,15,16 signalling behaviours, a dysregulation that was suggested to be also present in ASD patients17. Mechanism of action for the effect of cannabis on ASD may possibly involve GABA and glutamate transmission regulation. ASD is characterized by an excitation and inhibition imbalance of GABAergic and glutamatergic signalling in different brain structures18. The EC system is involved in modulating imbalanced GABAergic19 and glutamatergic transmission20.
Other mechanism of action can be through oxytocin and vasopressin, neurotransmitters that act as important modulators of social behaviours21. Administration of oxytocin to patients with ASD has been shown to facilitate processing of social information, improve emotional recognition, strengthen social interactions, reduce repetitive behaviours22 and increase eye gaze23. Cannabidiol was found to enhance oxytocin and vasopressin release during activities involving social interaction16.
Two main active ingredients (THC and CBD) can have different psychoactive action mechanisms. THC was previously shown to improve symptoms characteristic to ASD patients in other treated populations. For example, patients reported lower frequency of anxiety, distress and depression24, following THC administration, as well as improved mood and better quality of life in general25. In patients suffering from anxiety, THC led to improved anxiety levels compared to placebo26 and in dementia patients, it led to reduction in nocturnal motor activity,violence27,28 behavioural and severity of behavioural disorders29. Moreover, cannabis was shown to enhances interpersonal communication30 and decrease hostile feelings within small social groups31.
In our study we have shown that a CBD enriched treatment of ASD patients can potentially lead to an improvement of behavioural symptoms. These findings are consistent with the findings of two double-blind, placebo-controlled crossover studies demonstrating the anxiolytics properties of CBD in patients with anxiety disorder32,33. In one, CBD had a significant effect on increased brain activity in the right posterior cingulate cortex, which is thought to be involved in the processing of emotional information32, and in the other, simulated public speaking test was evaluated in 24 patients with social anxiety disorder. The CBD treated group had significantly lower anxiety scores than the placebo group during simulated speech, indicating reduction in anxiety, cognitive impairment, and discomfort factors33.
What does the Harvard Medical School have to say about tinkering with CBD?

Cannabidiol (CBD) — what we know andwhat we don’t

CBD is commonly used to address anxiety, and for patients who suffer through the misery of insomnia, studies suggest that CBD may help with both falling asleep and staying asleep.
CBD may offer an option for treating different types of chronic pain. A study from the European Journal of Pain showed, using an animal model, CBD applied on the skin could help lower pain and inflammation due to arthritis. Another study demonstrated the mechanism by which CBD inhibits inflammatory and neuropathic pain, two of the most difficult types of chronic pain to treat. More study in humans is needed in this area to substantiate the claims of CBD proponents about pain control.

Is cannabidiol safe?

Side effects of CBD include nausea, fatigue and irritability. CBD can increase the level in your blood of the blood thinner coumadin, and it can raise levels of certain other medications in your blood by the exact same mechanism that grapefruit juice does. A significant safety concern with CBD is that it is primarily marketed and sold as a supplement, not a medication. Currently, the FDA does not regulate the safety and purity of dietary supplements. So, you cannot know for sure that the product you buy has active ingredients at the dose listed on the label. In addition, the product may contain other (unknown) elements. We also don’t know the most effective therapeutic dose of CBD for any particular medical condition.

The bottom line on cannabidiol

Some CBD manufacturers have come under government scrutiny for wild, indefensible claims, such that CBD is a cure-all for cancer, which it is not. We need more research but CBD may be prove to be an option for managing anxiety, insomnia, and chronic pain. Without sufficient high-quality evidence in human studies we can’t pinpoint effective doses, and because CBD is currently is mostly available as an unregulated supplement, it’s difficult to know exactly what you are getting. If you decide to try CBD, talk with your doctor — if for no other reason than to make sure it won’t affect other medications you are taking.

CBD for Anxiety


Preclinical evidence conclusively demonstrates CBD’s efficacy in reducing anxiety behaviors relevant to multiple disorders, including PTSD, GAD, PD, OCD, and SAD, with a notable lack of anxiogenic effects. CBD’s anxiolytic actions appear to depend upon CB1Rs and 5-HT1ARs in several brain regions; however, investigation of additional receptor actions may reveal further mechanisms. Human experimental findings support preclinical findings, and also suggest a lack of anxiogenic effects, minimal sedative effects, and an excellent safety profile. Current preclinical and human findings mostly involve acute CBD dosing in healthy subjects, so further studies are required to establish whether chronic dosing of CBD has similar effects in relevant clinical populations. Overall, this review emphasizes the potential value and need for further study of CBD in the treatment of anxiety disorders.


I think I would put PEA and CBD in the same category of possibly beneficial therapies.

THC, the key psychoactive constituent in cannabis, will of course improve features of some people’s autism, as would LSD.

Indeed, one post that I never completed was all about the new idea of micro dosing LSD. I thought it would appeal to our Dutch Aspie readers.

First ever trials on theeffects of micro dosing LSD set to begin

Some Silicon Valley creatives are vocal proponents of small daily doses of psychedelics to boost creativity and mental health. But can micro dosing with LSD really live up to the claims?
Participants will take the capsules over a short period, fill out questionnaires and conduct cognitive tests online during the course of the trial.
“The most exciting part of this is the trial’s design, developed by a collaborator in the US,” explained Dr Erritzoe. “Due to the nature of the design, we’re not expecting the findings to be as robust as a large-scale lab-based clinical trials, but if people follow the manual, we should see some interesting results.”
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Several of you have enquired about the closing date of the study. We don’t have an exact date yet, but we guarantee that the option to sign up will be open at least until 31st of October 2019.
Furthermore, we would like to let you know that Reddit user /u/Greg_5656 has created an accurate ‘how-to’ video on setting up the project. The video goes through the same steps as the manual, making it easier to follow along. Thank you Greg!

The Swiss have their own LSD clinical trial running: -

Will PEA give a benefit in your case of autism? Maybe it will, but I think you will need to give it a try for several weeks and make sure you use a product that contains what is says on the label.

I did make my brief trial of PEA for autism before these case studies were published. Perhaps I should have persevered longer? Our reader Kei in Spain tells us that the French Bumetanide researchers have responders who needed 3 months of therapy to show the effect. I am not that patient; two weeks is a long time for me.  

Is PEA effective in its original application to treat neuropathic pain? Only sometimes; it works often enough for the Italians and Spanish, but it is no panacea. I think it is worth a try for neuropathy, there is a lot of evidence to support it.

There is no doubt that psychoactive substances will improve how some people with psychiatric disorders feel.  In the 1960s at UCLA they were trialing LSD on people with autism.

The problem with these psychoactive substances is that long term use definitely can cause permanent damage. That is part of the reason why they were banned.

If you have untreatable epilepsy, it may be that a psychoactive substance can save your life, and so the choice is clear.

Should the clinic at Johns Hopkins, that treats extreme cases of autism with electro-convulsive therapy (ECT), be able to prescribe psychoactive substances like LSD and THC? I would say definitely yes.

Is Cannabidiol (CBD) going to help your case of autism? I think if anxiety is part of the problem, it very likely will help.  Is it going to raise cognition like Bumetanide does in some people? I don’t think so.

Is Cannabidiol (CBD) a game changing autism therapy? I don’t think so, but every little does help.

Can Aspies make effective use of micro-dosing to manage any troublesome symptoms? Quite possibly.

Will big time CBD responders also respond to PEA? That, I would like to know.

Clearly some people taking PEA for neuropathy might get a similar benefit from CBD. If they took THC they would probably forget all about the pain.

I think the prodrug of PEA that crosses the blood brain barrier is interesting and I hope the Italians pursue it.