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Showing posts with label Progesterone. Show all posts
Showing posts with label Progesterone. Show all posts

Tuesday, 25 June 2019

Learning from GABAa Dysfunction in Huntington’s Disease – useful ideas for Autism therapies?



Today’s post is really for the regular readers of this blog who are interested in the GABA switch and Bumetanide. It is not light reading.  We see how advanced some Taiwanese researchers are in their understanding of GABAA dysfunctions in Huntington’s Disease.




Taipei 101, briefly the world’s tallest building


It is an excellent paper and much of it is applicable to autism. There are some omissions, but you will struggle to find a more complete paper.

They even go into the detail of altered the sub-unit expression of GABAA receptors that occurs as the disease progresses. I think that correcting sub-unit miss-expression has great potential in treating some autism.

Huntington’s is an inherited brain disorder that first manifests itself around the age of 40 and then progresses for the next 15 to 20 years.

Much autism is present prior to birth but there is a progression that occurs as the brain develops in early childhood. Some people do seem to be entirely typical at birth and only around 2 years old develop symptoms. After 5 years old you cannot really develop “autism”, just the symptoms might not get noticed till later in life.
Schizophrenia only develops in early to mid-adulthood.

It is surprising to many people that such varied disorders share some similar aspects of biology.

In terms of practical interventions, in today’s paper these include:       

·        Inhibition of NKCC1 (bumetanide)
·        Activation of KCC2 (N-Ethylmaleimide)
·        Enhancer of CKB (creatine)
·        Inhibitor of WNK/SPAK
·        Activation of extra-synaptic GABAa receptors (taurine, progesterone)
·        Activation of synaptic GABAa receptors (zolpidem, alprazolam)
·        Inhibition of GABA transport mechanism (Tiagabine)

One thing to note is that activating GABAa receptors may well have a negative effect in some people.

Sub-unit specific therapies, like very low dose clonazepam targeting α3, are not mentioned in this paper, nor is the role of GABAb on NKCC1/KCC2 expression.

We are familiar with Bumetanide as an NKCC1 blocker intervention in autism, but looking at the list there are other common autism therapies (creatine and taurine) and the female hormone progesterone. We come upon the beneficial effect of female hormones on a regular basis in this blog (estradiol, pregnenalone, progesterone …).  We even saw how a sub-SSRI dose of Prozac increases the amount of the neurosteroid 3α-hydroxy-5α-pregnan-20-one (Allo) that potently, positively, and allosterically modulates GABA action at GABAA receptors. Progesterone is converted to Allo in the body.
 
Here is the excellent paper on Huntington’s:-






                                                                                                               

An overview of the g-aminobutyric acid (GABA) signalling system. (a) GABA homeostasis is regulated by neurons and astrocytes. GABA is synthesized by GAD65/67 from glutamate in neurons, while astrocytic GABA is synthesized through MAOB. The release of GABA is mediated by membrane depolarization in neurons and Best1 in astrocytes. The reuptake of GABA is mediated through GAT1 in neurons and GAT3 in astrocytes. The metabolism of GABA is mediated by GABA-T in neurons and astrocytes. The reuptake of GABA in astrocytes is further transformed into glutamine via the TCA cycle and glutamine synthetase (GS). The glutamine is then transported to neurons and converted to glutamate for regeneration of GABA.



(b) GABAA receptors are heteropentameric complexes assembled from 19 different subunits. The compositions of different subunits determines the subcellular distributions and functional properties of the receptors. Phasic inhibition is mediated via the activation of synaptic GABAA receptors following brief exposure to a high concentration of extracellular GABA. Tonic inhibition is mediated via the activation of extrasynaptic GABAA receptors by a low concentration of ambient GABA.






c) The excitatory inhibitory response of GABA is driven by the chloride gradient across cell membranes, which can be determined via two cation–chloride cotransporters (NKCC1 and KCC2). The high expression of NKCC1 during the developmental stage maintains higher intracellular [Cl2] via chloride influx to the cell. The activation of GABAA receptors at an early developmental stage results in an outward flow of chloride and an excitatory GABAergic response. As neurons mature, the high expression of KCC2 maintains lower intracellular [Cl2] via chloride efflux out of the cell. The activation of GABAA receptors on mature neurons results in the inward flow of chloride and an inhibitory GABAergic response.



An excerpt showing data on sub-unit misexpression in different parts of the brain at different stages of the disease



5.2. Modulation of chloride homeostasis via cation – chloride cotransporters
Emerging evidence suggests that chloride homeostasis is a therapeutic target for HD. Pharmacological agents that target cation–chloride cotransporters (i.e. NKCC1 or KCC2) therefore might be used to treat HD (figure 3b). Of note, dysregulation of cation–chloride cotransporters and GABA polarity was associated with several neuropsychiatric disorders [70,134–139] (reviewed in [27,140]). Such abnormal excitatory GABAA receptor neurotransmission can be rescued by bumetanide, an NKCC1 inhibitor that decreases intracellular chloride concentration. Bumetanide is an FDA-approved diuretic agent that has been used in the clinic. It attenuates many neurological and psychiatric disorders in preclinical studies and some clinical trials for traumatic brain injury, seizure, chronic pain, cerebral infarction, Down syndrome, schizophrenia, fragile X syndrome and autism (reviewed in [141]). Daily intraperitoneal injections of bumetanide also restored the impaired motor function of HD mice. The effect of bumetanide is likely to be mediated by NKCC1 because genetic ablation of NKCC1 in the striatum also rescued the motor deficits in R6/2 mice. This study uncovered a previously unrecognized depolarizing or excitatory action of GABA in the aberrant motor control in HD. In addition, chronic treatment with bumetanide also improved the impaired memory in R6/2 mice [69], supporting the importance of NKCC1 in HD pathogenesis. Owing to the poor ability of bumetanide to pass through the blood–brain barrier, further optimization of bumetanide and other NKCC1 inhibitors is warranted [142,143]. Disruption of KCC2 function is detrimental to inhibitory transmission and agents to activate KCC2 function would be beneficial in HD. However, no agonist of KCC2 has been described until very recently [144,145]. A new KCC2 agonist (CLP290) has been shown to facilitate functional recovery after spinal cord injury [145]. It would be of great interest to evaluate the effect of KCC2 agonists on HD progression. Another KCC2 activator, CLP257, was found to increase the cell surface expression of KCC2 in a rat model of neuropathic pain [146]. Post-translational modification of KCC2 by kinases may modulate the function of KCC2. The WNK/ SPAK kinase complex, composed of WNK (with no lysine) and SPAK (SPS1-related proline/alanine-rich kinase), is known to phosphorylate and stimulate NKCC1 or inhibit KCC2 [147]. Thus, compounds that inhibit WNK/SPAK kinases will result in KCC2 activation and NKCC1 inhibition. Some compounds have been noted as potential inhibitors of WNK/SPAK kinases and need to be further tested for their effects on cation –chloride cotransporters [148–150]. An alternative mechanism to activate KCC2 is manipulation of its interacting proteins (e.g. CKB [65,66]). Because CKB could activate the function of KCC2 [65,66], CKB enhancers may increase the function of KCC2. In HD, reduced expression and activity of CKB is associated with motor deficits and hearing impairment [68,88]. Enhancing CKB activity by creatine supplements ameliorated the motor deficits and hearing impairment of HD mice. It is worthwhile to further investigate the interaction of KCC2 and CKB in GABAergic neurotransmission and motor deficits in HD. The depolarizing GABA action with altered expression levels of NKCC1 or KCC2 is associated with neuroinflammation in HD brains [32,69]. Blockade of TNF-a using Xpro1595 (a dominant negative inhibitor of soluble TNF-a) [151] in vivo led to significant beneficial effects on disease progression in HD mice [152] and reduced the expression of NKCC1. It would be of great interest to test the effect of other anti-inflammatory agents [153] on the function and expression of NKCC1 and GABAergic inhibition. Neuroinflammation is implicated in most neurodegenerative diseases, including Alzheimer’s disease and Parkinson’s disease [154,155], and the interaction of cation–chloride cotransporters and neuroinflammation in GABAergic neurotransmission may also play a critical role in other neurodegenerative diseases.






Figure 2. Molecular mechanism(s) underlying the abnormal GABAAergic system in HD. (a) In the normal condition, adult neurons express high KCC2 and few NKCC1 to maintain the lower intracellular chloride concentration, which results in an inward flow of chloride when GABAA receptors are activated. Astrocytes function normally for the homeostasis of glutamate, potassium and glutamate/GABA-glutamine cycle. (b) In Huntington’s disease, reduced GABAA receptor-mediated neuronal inhibition is associated with enhanced NKCC1 expression and a decreased expression in KCC2 and membrane localized GABAA receptors. The dysregulated GABAAergic system might be caused by mutant HTT, excitotoxicity, neuroinflammation or other factors. Mutant HTT in neurons alters the transcription of genes (GABAAR and KCC2) through interactions with transcriptional activators (SP1) and repressors (REST/NRSF). Mutant HTT in neurons also disrupts the intracellular trafficking of GABAARs to the cellular membrane. HD astrocytes have impaired homeostasis of extracellular potassium/glutamate (due to deficits of astrocytic Kir4.1 channel and glutamate transporters, Glt-1) and cause neuronal excitability, which might be related to the changes of KCC2, NKCC1 and GABAAR. The activity of KCC2 could be affected through its interacting proteins, such as CKB and mHTT. Neuroinflammation, which is evoked by the interaction of HD astrocyte and microglia, enhances NKCC1 expression in neurons at the transcriptional level through an NF-kB-dependent pathway. HD astrocytes also have compromised astrocytic metabolism of glutamate/GABA–glutamine cycle that contributes to lower GABA synthesis.


Notably, neuroinflammation and the GABA neurotransmitter system are reciprocally regulated in the brain (reviewed in [104,105]). Specifically, neuroinflammation induces changes in the GABA neurotransmitter system, such as reduced GABAA receptor subunit expression, while activation
of GABAA receptors likely antagonizes inflammation.

TNF-a, a proinflammatory cytokine, induces a downregulation of the surface expression of GABAARs containing a1, a2, b2/3 and g2 subunits and a decrease in inhibitory synaptic strength in a cellular model of hippocampal neuron culture [106]. The same group further demonstrated that protein phosphatase 1-dependent trafficking of GABAARs was involved in the TNF-a evoked downregulation of GABAergic neurotransmission [107]. Upregulation of TNF-a also negatively impacts the expression of GABAAR a2 subunit mRNA and thus decreases the presynaptic inhibition in the dorsal root ganglion in a rat experimental neuropathic painmodel [108]. Conversely, blockade of central GABAARs in mice by aGABAAR antagonist increased both the basal and restraint stress-induced plasma IL-6 levels [109]. Inhibition of GABAAR activation by picrotoxin increased the nuclear translocation of NF-kB in acute hippocampal slice preparations [110]. Collectively, neuroinflammation
weakens the inhibitory synaptic strength in neurons, at least partly, through the reduction of GABAARs.

The reduced expression and function of GABAARs may further increase inflammatory responses. It remains elusive whether the same mechanism occurs in the inflammatory environment in HD brains.


hyperexcitability resulting from deficiency of astrocytic Kir4.1 might have also contributed to neuronal NKCC1 upregulation and altered GABAergic signalling in HD brains.




Figure 3. Strategy to target (a) GABAAR and (b) cation–chloride cotransporters as potential therapeutic avenues. (a) The GABAergic system is influenced directly by agents that (1) target synaptic GABAAR, (2) increase tonic GABA current or interfere with synaptic GABA concentrations via a reduction of GABA reuptake (3), and (4) block GABA metabolism.

5.1. Modulating the GABAA receptor as a therapeutic target

In view of the presently discovered HD-related deficit in the GABA system, the question arises whether HD patients can benefit from drugs that stimulate the GABA system (figure 3a). HD patients suffer from motor abnormalities and
non-motor symptoms, including cognitive deficits, psychiatric symptoms, sleep disturbance, irritability, anxiety, depression and an increased incidence of seizures [74,77,116,117].
Seizures are a well-established part of juvenile HD but no more prevalent in adult-onset HD than in the general population [73,74,118]. Several pharmacological compounds can enhance inhibitory GABAergic neurotransmission by targeting GABAAR and thereby producing sedative, anxiolytic, anticonvulsant and muscle-relaxant effects. A recent study demonstrated that zolpidem, a GABAAR modulator that enhances GABA inhibition mainly via the a1-containing GABAA receptors, corrected sleep disturbance and electroencephalographic abnormalities in symptomatic HD mice (R6/2) [119]. Alprazolam, a benzodiazepine-activating GABA receptor, reversed the dysregulated circadian rhythms and improved cognitive performance of HD mice (R6/2) [120].
In addition, progesterone, a positive modulator of GABAAR, significantly reversed the behavioural impairment in a 3-nitropropionic acid (3-NP)-induced HD rat model [121]. Apart from modulating the activity of the GABAergic system by interfering directly with the receptor, pharmacological agents can also interfere with synaptic GABA concentrations. Tiagabine, a drug that specifically blocks the GABA transporter (GAT1) to increase synaptic GABA level,was found to improve motor performance and extend survival inN171-82Q and R6/2 mice [122]. It is also worth evaluating whether vigabatrin, a GABA-T inhibitor that blocksGABAcatabolismin neurons and astrocytes [123], plays a role in the compromised astrocytic glutamate–GABA–glutamine cycling [56]. Interestingly, taurine exerted GABAA agonistic and antioxidant activities in a 3-NP HD model and improved locomotor deficits and increased GABA levels [124]. However, several early studies failed to provide the expected benefits of GABA analogues in slowing disease progression in HD patients [125–127]. For example, gaboxadol, an agonist for the extrasynaptic d-containing GABAA receptor, failed to improve the decline in cognitive and motor functions of five HD patients during a short two-week trial, but it caused side effects at the maximal dose [125]. Interestingly, although treatment with muscimol (a potent agonist of GABA receptors) did not improve motor or cognitive deficits in 10HDpatients, it did ameliorate chorea in the most severely hyperkinetic patient [126]. The therapeutic failure of GABA stimulation in early clinical trials does not argue against the importance of GABAergic deficits in HD pathogenesis. The alteration of GABAergic circuits plays a primary role or is a compensatory response to excitotoxicity, and it may contribute to HD by disrupting the balance between the excitation and inhibition systems and the overall functions of neuronal circuits. Because the subunits of the GABAA receptor are brain region- or neuron subtypespecific, the choice of drugs may have distinct effects on the brain region or neuronal population targeted [128–130]. For example, the expression of GABAAR subunits is differentially altered in MSNs and other striatal interneurons in HD 54,60]. The early involvement of D2-expressing MSNs can cause chorea [131], while dysfunctional PV-expressing interneurons can cause dystonia in HD patients [132]. Specific alteration in neuronal populations and receptor subtypes during HD progression needs to be taken into consideration when treating the dysfunction of GABAergic circuitry.
Notably, striatal tonic inhibition mediated by the dcontaining GABAARs may have neuroprotective effects against excitotoxicity in the adult striatum [63]. Because the reductions in d-containing GABAARs and tonic GABA currents in D2-expressing MSNs have been observed in early HD [32,39,40,54,61], it would be of great interest to evaluate the effects of several available compounds, such as alphaxalone and ganaxolone [133], that target d-containing GABAARs, in animal models of HD.





(b) GABAAR-mediated signalling in HD neurons is depolarizing due to the high intracellular chloride concentration caused by high NKCC1 expression and low KCC2 expression. Rescuing the function of cation–chloride cotransporters can occur via (1) inhibition of NKCC1 activity using bumetanide, (2, 3) increase in KCC2 function using a KCC2 activator or CKB enhancer, and (4) inhibitors of WNK/SPAK kinases.


5.2. Modulation of chloride homeostasis via cation–chloride cotransporters

Emerging evidence suggests that chloride homeostasis is a therapeutic target for HD. Pharmacological agents that target cation–chloride cotransporters (i.e.NKCC1 orKCC2) therefore might be used to treat HD (figure 3b). Of note, dysregulation of cation–chloride cotransporters and GABA polarity was associated with several neuropsychiatric disorders [70,134–139] (reviewed in [27,140]). Such abnormal   receptor neurotransmission can be rescued by bumetanide, an NKCC1 inhibitor that decreases intracellular chloride concentration. Bumetanide is an FDA-approved diuretic agent that has been used in the clinic. It attenuates many neurological and psychiatric disorders in preclinical studies and some clinical trials for traumatic brain injury, seizure, chronic pain, cerebral infarction, Down syndrome, schizophrenia, fragile X syndrome and autism (reviewed in [141]). Daily intraperitoneal injections of bumetanide also restored the impaired motor function ofHDmice (R6/2, Y-T Hsu,Y-GChang, Y-CLi, K-YWang, H-MChen, D-J Lee, C-HTsai, C-C Lien,YChern 2018, personal communication). The effect of bumetanide is likely to be mediated by NKCC1 because genetic ablation of NKCC1 in the striatum also rescued the motor deficits in R6/2 mice (Y-T Hsu, Y-G Chang, Y-C Li, K-Y Wang, H-M Chen, D-J Lee, C-H Tsai, C-C Lien, Y Chern 2018, personal communication). This study uncovered a previously unrecognized depolarizing or excitatory action of GABA in the aberrant motor control in HD. In addition, chronic treatment with bumetanide also improved the impaired memory in R6/2 mice [69], supporting the importance of NKCC1 in HD pathogenesis. Owing to the poor ability of bumetanide to pass through the blood–brain barrier, further optimization of bumetanide and other NKCC1 inhibitors is warranted [142,143].
Disruption of KCC2 function is detrimental to inhibitory transmission and agents to activate KCC2 function would be beneficial in HD. However, no agonist of KCC2 has been described until very recently [144,145]. A new KCC2 agonist (CLP290) has been shown to facilitate functional recovery after spinal cord injury [145]. It would be of great interest to evaluate the effect of KCC2 agonists on HD progression. Another KCC2 activator, CLP257, was found to increase the cell surface expression of KCC2 in a rat model of neuropathic pain [146]. Post-translational modification of KCC2 by kinases may modulate the function of KCC2. The WNK/SPAK kinase complex, composed of WNK (with no lysine) and SPAK (SPS1-related proline/alanine-rich kinase), is known to phosphorylate and stimulate NKCC1 or inhibit KCC2 [147]. Thus, compounds that inhibit WNK/SPAK kinases will result in KCC2 activation and NKCC1 inhibition.
Some compounds have been noted as potential inhibitors of WNK/SPAK kinases and need to be further tested for their effects on cation–chloride cotransporters [148–150]. An alternative mechanism to activate KCC2 is manipulation of its interacting proteins (e.g. CKB [65,66]). Because CKB could activate the function of KCC2 [65,66], CKB enhancers may increase the function of KCC2. In HD, reduced expression and activity of CKB is associated with motor deficits and hearing impairment [68,88]. Enhancing CKB activity by creatine supplements ameliorated the motor deficits and hearing impairment of HD mice. It is worthwhile to further investigate the interaction of KCC2 and CKB in GABAergic neurotransmission and motor deficits in HD. The depolarizing GABA action with altered expression levels of NKCC1 or KCC2 is associated with neuroinflammation in HD brains [32,69]. Blockade of TNF-a using Xpro1595 (a dominant negative inhibitor of soluble TNF-a) [151] in vivo led to significant beneficial effects on disease progression in HD mice [152] and reduced the expression of NKCC1It would be of great interest to test the effect of other anti-inflammatory agents [153] on the function and expression of NKCC1 and GABAergic inhibition. Neuroinflammation is implicated in most neurodegenerative diseases, including Alzheimer’s disease and Parkinson’s disease [154,155], and the interaction of cation–chloride cotransporters and neuroinflammation in GABAergic neurotransmission may also play a critical role in other neurodegenerative diseases.




Discovery of Novel SPAK Inhibitors That Block WNK Kinase Signaling to Cation Chloride Transporters

Upon activation by with-no-lysine kinases, STE20/SPS1-related proline–alanine-rich protein kinase (SPAK) phosphorylates and activates SLC12A transporters such as the Na+-Cl cotransporter (NCC) and Na+-K+-2Cl cotransporter type 1 (NKCC1) and type 2 (NKCC2); these transporters have important roles in regulating BP through NaCl reabsorption and vasoconstriction. SPAK knockout mice are viable and display hypotension with decreased activity (phosphorylation) of NCC and NKCC1 in the kidneys and aorta, respectively. Therefore, agents that inhibit SPAK activity could be a new class of antihypertensive drugs with dual actions (i.e., NaCl diuresis and vasodilation). In this study, we developed a new ELISA-based screening system to find novel SPAK inhibitors and screened >20,000 small-molecule compounds. Furthermore, we used a drug repositioning strategy to identify existing drugs that inhibit SPAK activity. As a result, we discovered one small-molecule compound (Stock 1S-14279) and an antiparasitic agent (Closantel) that inhibited SPAK-regulated phosphorylation and activation of NCC and NKCC1 in vitro and in mice. Notably, these compounds had structural similarity and inhibited SPAK in an ATP-insensitive manner. We propose that the two compounds found in this study may have great potential as novel antihypertensive drugs.


Chemical library screening for WNK signalling inhibitors using fluorescence correlation spectroscopy.


WNKs (with-no-lysine kinases) are the causative genes of a hereditary hypertensive disease, PHAII (pseudohypoaldosteronism type II), and form a signal cascade with OSR1 (oxidative stress-responsive 1)/SPAK (STE20/SPS1-related proline/alanine-rich protein kinase) and Slc12a (solute carrier family 12) transporters. We have shown that this signal cascade regulates blood pressure by controlling vascular tone as well as renal NaCl excretion. Therefore agents that inhibit this signal cascade could be a new class of antihypertensive drugs. Since the binding of WNK to OSR1/SPAK kinases was postulated to be important for signal transduction, we sought to discover inhibitors of WNK/SPAK binding by screening chemical compounds that disrupt the binding. For this purpose, we developed a high-throughput screening method using fluorescent correlation spectroscopy. As a result of screening 17000 compounds, we discovered two novel compounds that reproducibly disrupted the binding of WNK to SPAK. Both compounds mediated dose-dependent inhibition of hypotonicity-induced activation of WNK, namely the phosphorylation of SPAK and its downstream transporters NKCC1 (Na/K/Cl cotransporter 1) and NCC (NaCl cotransporter) in cultured cell lines. The two compounds could be the promising seeds of new types of antihypertensive drugs, and the method that we developed could be applied as a general screening method to identify compounds that disrupt the binding of two molecules.







N-Ethylmaleimide increases KCC2 cotransporter activity by modulating transporter phosphorylation


K+/Cl cotransporter 2 (KCC2) is selectively expressed in the adult nervous system and allows neurons to maintain low intracellular Cl levels. Thus, KCC2 activity is an essential prerequisite for fast hyperpolarizing synaptic inhibition mediated by type A γ-aminobutyric acid (GABAA) receptors, which are Cl-permeable, ligand-gated ion channels. Consistent with this, deficits in the activity of KCC2 lead to epilepsy and are also implicated in neurodevelopmental disorders, neuropathic pain, and schizophrenia. Accordingly, there is significant interest in developing activators of KCC2 as therapeutic agents. To provide insights into the cellular processes that determine KCC2 activity, we have investigated the mechanism by which N-ethylmaleimide (NEM) enhances transporter activity using a combination of biochemical and electrophysiological approaches. Our results revealed that, within 15 min, NEM increased cell surface levels of KCC2 and modulated the phosphorylation of key regulatory residues within the large cytoplasmic domain of KCC2 in neurons. More specifically, NEM increased the phosphorylation of serine 940 (Ser-940), whereas it decreased phosphorylation of threonine 1007 (Thr-1007). NEM also reduced with no lysine (WNK) kinase phosphorylation of Ste20-related proline/alanine-rich kinase (SPAK), a kinase that directly phosphorylates KCC2 at residue Thr-1007. Mutational analysis revealed that Thr-1007 dephosphorylation mediated the effects of NEM on KCC2 activity. Collectively, our results suggest that compounds that either increase the surface stability of KCC2 or reduce Thr-1007 phosphorylation may be of use as enhancers of KCC2 activity.


                                                                  


Tiagabine (trade name Gabitril) is n anticonvulsant medication produced by Cephalon that is used in the treatment of epilepsy. The drug is also used off-label in the treatment of anxiety disorders and panic disorder.

Tiagabine is approved by U.S. Food and Drug Administration (FDA) as an adjunctive treatment for partial seizures in individuals of age 12 and up. It may also be prescribed off-label by physicians to treat anxiety disorders and panic disorder as well as neuropathic pain (including fibromyalgia). For anxiety and neuropathic pain, tiagabine is used primarily to augment other treatments. Tiagabine may be used alongside selective serotonin reuptake inhibitorsserotonin-norepinephrine reuptake inhibitors, or benzodiazepines for anxiety, or antidepressantsgabapentin, other anticonvulsants, or opioids for neuropathic pain.[4]
Tiagabine increases the level of γ-aminobutyric acid (GABA), the major inhibitory neurotransmitter in the central nervous system, by blocking the GABA transporter 1 (GAT-1), and hence is classified as a GABA reuptake inhibitor (GRI).


Conclusion

Today’s post shows how you need to read well beyond the autism research, not to miss something useful.

Some of today’s suggested therapies for Huntington’s are likely to help some types of autism, but some will certainly have a negative effect in some people.  For example, increasing the amount of GABA in the CNS would do my son no good at all.

The emerging field of drugs that enhance KCC2 should be very beneficial to all those with autism who are bumetanide responders.

Enhancing CKB with creatine is interesting. Creatine is a muscle building supplement used by body builders and some DAN doctors. It does have interactions at high doses.







Saturday, 16 December 2017

Turner Syndrome, Estradiol and Autism-lessons from the X Chromosome

This post is best read if you have reviewed the earlier ones regarding the estradiol/testosterone disturbances in autism and how they govern the RORα “switch” that then triggers a torrent of other dysfunctions. So the hormonal disturbance, if present, is a key point at which to make a potent intervention. 



Beauty is in the eye of the beholder


In the mass media it is now popular to dismiss the fact that autism is far more prevalent on boys than girls. In the scientific literature, fortunately, they stick to the facts and much is written about the sex differences in autism.
As we have seen in earlier posts, females have some natural defences against autism. They have two X chromosomes and of course they have those all-important neuroprotective female hormones (estrogen/estradiol, progesterone etc.). In effect, the more female you are, the more protection you have against idiopathic autism and any X-chromosome linked single gene autism. So a girl with Fragile-X syndrome is likely to be far less affected than her brother with same condition.
Recall that we all have 23 pairs of chromosomes and that the 23rd set contains two Xs in girls and in boys one X and one Y. The girls’ “spare” X chromosome is also what gives them their feminine features.  

It is interesting to look what happens to females who lack part of their second set of X- chromosomes. This diagnosis is called Turner Syndrome. As you might have guessed people with Turner Syndrome have much lower levels of female hormones and a higher incidence of autism, although some people find this controversial. The autism-like characteristics of TS include:-

·      Impairments in social functioning

·      Impairments in face and emotion processing

·      Spatial executive deficits

·      Poor social coping skills and increased immaturity

·      Hyperactivity and impulsivity

Turner syndrome occurs in 50 per 100,000 live-born females. Autism occurs about ten times for frequently, so about 500 per 100,000 live-born females.  Turner syndrome provides the extreme case of what happens when females have too little estrogen/estradiol.
I think you will find a large group of females with idiopathic autism (no identified genetic defects) have/had low levels of estradiol. I think this is the reason that facial recognition studies show that some females with idiopathic autism look different, (as do many boys, of course). We already know that most single gene types of autism produce tell-tale signs, often on the face (big ears, wide face, big/small head etc).

I am not suggesting that there is anything wrong with looking different; rather it may be a useful diagnostic tool and not an expensive or invasive one. Physical variation has long been used to identify genetic syndromes, before genetic testing became widely available.

Physical variation inside your head
We saw in an earlier post that MRI scans of the autistic brains actually do often show subtle differences, particularly when you use software to read them, rather than the naked eye. Traditionally doctors say that MRIs are “normal” in autism and cannot be used to diagnose it. Yet in a recent studies machine reading of MRIs was able to identify 70%-96% of autism cases.  Some of these are scans taken before birth.

This is interesting, because ultimately you might bypass the current very slow and subjective observational diagnosis process.




MRIs show a brain anomaly in nearly 70 percent of babies at high risk of developing the condition who go on to be diagnosed, laying the groundwork for a predictive aid for pediatricians and the search for a potential treatment



Predicting the future with brain imaging

In a new study, Emerson et al. show that brain function in infancy can be used to accurately predict which high-risk infants will later receive an autism diagnosis. Using machine learning techniques that identify patterns in the brain’s functional connections, Emerson and colleagues were able to predict with greater than 96% accuracy whether a 6-month-old infant would develop autism at 24 months of age. These findings must be replicated, but they represent an important step toward the early identification of individuals with autism before its characteristic symptoms develop.


MRI scanners are very widely used, but you do have to keep very still inside when they are in operation. The even harder part is the reading of the data. It is clear that some standardized machine reading (A/I artificial intelligence) process is required to notice every possible variation. You could have a centralized location where you just submit your MRI data, the center gets to keep the data and learn from it; and you get their insight as to what differences there might be.

Facial Differences vs MRI Brain Differences
I like to keep things simple and under my control.  In the short term we have to settle for facial differences, since any well-managed MRI process will be decades away.

Hormonal Variation in Autism
Hormonal differences were one of the key areas I identified years ago in this blog. Big/small heads result from disturbances in pro-growth signalling pathways. We should expect variations in bone-age, early/late onset of puberty and indeed big variations in height and weight.

In Turner Syndrome, the girls tend to be very short and they are often treated with growth hormones, as well as female/feminizing hormones.  
Great caution has to be taken when treating children with any hormones. When children are treated, it is for serious reasons like not achieving puberty, or having a serious growth delay (being very short).

Hormone Therapy During Pregnancy
In some countries hormones are given during pregnancy although I think this would be seen as odd/risky in some advanced countries.

We have already seen that couples who have difficulty producing a child often have a family history that includes autism. It was proposed by one serious fertility expert that what helps prevent miscarriage also helps prevent autism. This did sound odd when I first read about, but when you look in more depth there is a basis for this idea.
That expert has these two websites:-



Progesterone supplements have been recommended for more than 50 years for women struggling with infertility, but research now shows they can also help prevent miscarriage.


Tamoxifen, an estrogen receptor (ER) antagonist, is also used to treat infertility.
Estradiol is sometimes prescribed during pregnancy.
Testosterone is produced naturally during pregnancy.

All this is clearly beyond the scope of this blog, but perhaps altered female/male hormones during pregnancy might be a biomarker of some future autism and female hormones might be a protective therapy in the subgroup of pregnant mothers with low levels of these hormones and/or high levels of testosterone. Recall that human trials in the hospital ER have shown certain substances are highly neuroprotective (progesterone, atorvastatin etc) and when administered immediately after a traumatic brain injury markedly improve the outcome.                                         

Hormone Therapy for Autism
Hormone therapy in people with autism would be controversial, but we saw in an earlier post that via RORα the balance between testosterone and estradiol affects numerous biological relevant to autism.

Many pictures of girls/women with autism, that you can view online, suggest reduced levels of estradiol. Faces look more boy-like. Many males with autism are reported to have physical features of high testosterone and low estradiol. 
One example of many:-


Both faces in the above article show clear indications of autism. Since both young people do have autism, this should not surprise anyone.
My own conclusion is that if you have autism or Asperger’s, a little extra estradiol could therapeutic, particularly if you have physical features that reinforce this.
There are of course many males and females with autism who are physically indistinguishable from the rest of the world. The point of this post is to highlight that visible differences may help to define the sub-type of autism and indicate possibly effective therapies, that exist today.

Obesity and Estradiol
In an earlier post on estradiol, I pointed out that in males estradiol is made in your adipose (fat) tissue. In the US many people with autism are overweight, in part due to side effects from their likely un-needed psychiatric medications; this has the hidden benefit of increasing their estradiol levels, feminizing their behavior slightly and shifting RORalpha in the right direction.
This also means that losing weight should be helpful to obese females with estrogen receptor positive breast cancer.  Research does support this.


Asperger’s and too much Estradiol?
We saw in earlier posts that much autism is associated with reduced expression of estrogen receptor beta and low aromatase, so high testosterone and low estradiol.

We have seen on many occasions that when one extreme exists in autism, so usually does the other; so many big heads, but also some tiny ones, NMDAR hypofunction, but also hyperfunction.

There was a lot of talk a while back in the media about children undergoing therapy to change their gender, and it was highlighted that Asperger’s was much over-represented in this group. One expert got into trouble for suggesting that their autism was causing them to obsess about their identity and so mistakenly convince a boy that he would rather be a girl.  It seems that these days some clinicians are then all too willing to provide drug therapy and then operate on them, to make them female.  I do wonder if perhaps some of these boys with Asperger’s might have the other extreme of aromatise. That would give them too little testosterone and too much Estradiol.
I think measuring these hormones is quite a good idea, as I keep repeating, they go on to affect the critical “switch”  RORα, which then impacts a large number of biological processes implicated in autism.  In other words you can try to normalize a wide range of important autism variables, just be tweaking RORα, via estradiol/testosterone.

A boy with high testosterone, and so low estradiol, will likely exhibit physical signs of this, just like the girl with low estradiol. These are just pieces of the puzzle, in plain view, that can be used to understand each specific case of autism. And no machine reading of an MRI is required.






For those left wanting more:
A very thorough paper on Turner Syndrome:-

Turner syndrome (TS) is a neurogenetic disorder characterized by partial or complete monosomy-X. TS is associated with certain physical and medical features including estrogen deficiency, short stature and increased risk for several diseases with cardiac conditions being among the most serious. Girls with TS are typically treated with growth hormone and estrogen replacement therapies to address short stature and estrogen deficiency. The cognitive-behavioral phenotype associated with TS includes strengths in verbal domains with impairments in visual-spatial, executive function and emotion processing. Genetic analyses have identified the short stature homeobox (SHOX) gene as being a candidate gene for short stature and other skeletal abnormalities associated with TS but currently the gene or genes associated with cognitive impairments remain unknown. However, significant progress has been made in describing neurodevelopmental and neurobiologic factors underlying these impairments and potential interventions are on the horizon

We utilized an ultrasensitive assay to study estradiol levels in 34 girls with TS and 34 normal age-matched prepubertal girls between the ages of 5 and 12 years. The average estradiol level in the girls with TS (6.4 +/- 4.9 pmol/l estradiol equivalents) was significantly lower than in the normal prepubertal girls (12.7 +/- 10.8 pmol/l estradiol equivalents; p < 0.01). Girls with TS were significantly shorter, and weighed less than the normal prepubertal girls, as expected. The estradiol level was not significantly correlated with height, bone age,