Brain Iron Deficiency Research Articles

1001 Intravenous Ferric Carboxymaltose For Restless Legs Syndrome In Children And Adolescents

Abstract Introduction Substantial scientific evidence implicates brain iron deficiency in the pathophysiology of restless legs syndrome (RLS). Current clinical guidelines recommend oral and intravenous iron (IV) in the treatment of both adult and pediatric RLS but studies using ferric carboxymaltose (FCM) are lacking in children and adolescents. Methods Retrospective case series of children and adolescents with RLS treated with IV FCM who had serum ferritin levels <50 μg/L. Patients were offered a single dose of IV FCM, 15 mg/kg if weighting <50 kg or 750 mg if weighting >50 kg. Iron profile, serum ferritin, and severity assessment by the International Restless Legs Study Group severity scale (IRLS) were collected pre- and post-infusion. Clinical Global Impression Scale (CGI) was used instead of the IRLS for children. Phosphorus level and adverse effects were assessed post-infusion in all patients. Age and sex-matched children with RLS treated with oral iron supplementation (mean dosage 1.5±0.62 mg/kg/day) were included as a comparison group. Results Twenty-eight subjects (15 females, mean age 11.5 years, SD 4.23) and 24 controls were included. Baseline ferritin levels were not significantly different from those of controls but increased significantly from 13.9±7.02 to 112.9±12.00 μg/L after 8 weeks from infusion (p<0.000001), when they were also significantly higher than control values (34.2±21.64 μg/mL, p<0.000001). Transferrin saturation increased from 22.8±9.77% to 31.7±6.81% (p<0.0001), total iron binding capacity decreased from 366.7±51.32 to 302.0±37.83 μg/dL (p<0.0000035). RLS was reported to be resolved or improved in all children treated with IV iron (vs. 62.5% of controls) while none of them reported no change (vs. 37.5% of controls; Chi-square test 9.84, p<0.002). IRLS Score decreased in adolescents from 30.7±22.68 to 3.2±4.21 (p<0.000008) while CGI-I was “very much improved” in six children and “much improved” in four. Side effects were reported in 17.8% of patients treated IV and 20.8% controls (Chi-square 0.0169, p=0.897). FCM side effects included lightheadedness and gastrointestinal discomfort. Post IV phosphorus levels were normal in all participants. Conclusion This open-label, observational and retrospective study indicates that FCM IV infusion is an effective treatment for pediatric RLS with higher efficacy than oral iron supplementation. Support

Iron Replacement Therapy in Restless Legs Syndrome

The main purpose of this manuscript is to review the existing clinical evidence about when iron replacement therapy should be initiated for RLS/WED, and how it should be administered (oral vs. intravenous). Our knowledge of the pathophysiology of restless legs syndrome/Willis-Ekbom (RLS/WED) continues to grow, revealing the existence of brain iron deficiency as the most well-known biological alteration behind this disorder. Since iron replacement therapy in RLS/WED is one of the first-line available treatments, its indication should be considered periodically, at both the beginning and during the follow-up. There is more and more evidence sustaining that the proper use of iron replacement therapy may lead to a reduction in symptomatic drug requirements, and therefore minimize the risk of complications that may result from their long-term use. For all the above, treatment paradigms for RLS/WED, previously based on symptomatic drugs, are changing, giving iron replacement therapy the importance it merits. Thus, it is essential for neurologists to be updated on why, when, and how to use iron in RLS/WED.

Iron-deficiency and dopaminergic treatment effects on RLS-Like behaviors of an animal model with the brain iron deficiency pattern of the restless legs syndrome

BackgroundBrain iron deficiency (BID), especially for the substantia nigra (SN), without peripheral iron deficiency (ID) has been well documented as a ubiquitous finding for restless legs syndrome (RLS) patients. This close association suggests the biology of RLS BID can produce RLS symptoms. Association, however, cannot establish such a direct relationship. Instead, the BID of RLS could be experimentally produced to determine if it then produces significant RLS-like biological or behavioral features. Forward genetics approach led to identification from the BXD strains the BXD40 females (BXD40f) as a putative animal model for the RLS BID. The BXD40f on an iron-sufficient diet have a lower iron in the VMB (containing the SN) during the active but not inactive period. This was not found for the other BXD strains evaluated. The BXD40f on an ID diet uniquely have even greater reduced VMB but not peripheral iron, matching the RLS BID pathophysiology. A prior report found that the BXD40f on an iron-sufficient diet had an RLS-like behavior of increased activity occurring only in the last part of the active period that was not present in the other strains without the low VMB iron. This increased activity matches the circadian pattern of symptoms in RLS patients with increased urge or drive to move in the last part of the day. This study asks first: if you decrease the VMB iron by an iron deficient diet do the RLS-like behaviors worsen; and second will the dopaminergic treatments effective for RLS also reduce the worsened RLSlike behaviors. MethodsIn sum, 13 BXD40f mice post weaning were randomly assigned for 100 days to either a iron-sufficient diet (n = 6) or an ID diet (N = 7). They were then evaluated for 24-h activity in their home cage using implanted G2 EMitter telemetry device. At 3 h before the end of the active period IP doses were given every other day of either: saline (vehicle only), 12.5 mg levodopa, 25 mg levodopa, 0.5 mg quinpirole, or 1 0.0 mg quinpirole. ResultsThe ID compared to irons-sufficient diet produced earlier onset of the RLS-like behavior matching the earlier onset of symptoms with increasing severity of RLS. The dopaminergic treatments significantly reduced the RLS-like behavior. Added analyses of the RLS-like behaviors as decreased resting times showed similar results to activity increases. ConclusionsThese data demonstrate both that The BXD40f provide a useful animal model of RLS and also strongly support the hypothesis that the biology of RLS BID can produce RLS symptoms.

Developing a behavioral model of Restless Legs Syndrome utilizing mice with natural variances in ventral midbrain iron

BackgroundThe primary symptoms of Restless Legs Syndrome (RLS) are circadian-dependent, leading to increased activity or decreased rest, especially at night. The primary pathology in RLS is brain iron insufficiency despite normal systemic iron stores. Natural variances in brain and peripheral iron concentrations across recombinant inbred (RI) murine strains provide a biological model of RLS. The question is whether these RI mice strains show a behavioral analog to circadian-dependent clinical phenotype of RLS. MethodsThe home cage activity of eight female RI strains was measured over a 72-h period. The ratio of the average activity in the last 2 h of the active period relative to that in the total 12-h active period (late active period activity ratio, LAPAR) was the primary outcome variable. The relation of average LAPAR scores to measures of ventral midbrain (VMB) iron was evaluated across strains in this study. ResultsRI strain 40 (LAPAR = 1.28) and RI strain 21 (LAPAR = 1.02) were the only strains to show an increased activity in the last part of the active period. ANOVA showed the increased activity was significantly greater during the last 2 h compared to the preceding 10 h of the active phase only for the RI strain 40. Average LAPAR across the eight strains did not significantly correlate with the VMB iron content (r = −0.27, p < 0.10) but did correlate with changes in VMB iron with iron deficiency (r = 0.71, p < 0.05) and diurnal change in VMB iron (r = 0.65, p < 0.05). ConclusionThe female RI strain 40 mice exhibited a distinct end-of-active-period behavior analogous to circadian-dependent clinical phenotype of RLS.

Transcreal sonography as a novel neuroimaging tool to determine brain iron deficiency in restless legs syndrome: results in a chilean sample

Transcreal sonography as a novel neuroimaging tool to determine brain iron deficiency in restless legs syndrome: results in a chilean sample

Cord Blood-Derived Exosomal CNTN2 and BDNF: Potential Molecular Markers for Brain Health of Neonates at Risk for Iron Deficiency

Maternal iron deficiency anemia, obesity, and diabetes are prevalent during pregnancy. All are associated with neonatal brain iron deficiency (ID) and neurodevelopmental impairment. Exosomes are extracellular vesicles involved in cell–cell communication. Contactin-2 (CNTN2), a neural-specific glycoprotein, and brain-derived neurotrophic factor (BDNF) are important in neurodevelopment and found in exosomes. We hypothesized that exosomal CNTN2 and BDNF identify infants at risk for brain ID. Umbilical cord blood samples were measured for iron status. Maternal anemia, diabetes, and body mass index (BMI) were recorded. Cord blood exosomes were isolated and validated for the exosomal marker CD81 and the neural-specific exosomal marker CNTN2. Exosomal CNTN2 and BDNF levels were quantified by ELISA. Analysis of CNTN2 and BDNF levels as predictors of cord blood iron indices showed a direct correlation between CNTN2 and ferritin in all neonates (n = 79, β = 1.75, p = 0.02). In contrast, BDNF levels inversely correlated with ferritin (β = −1.20, p = 0.03), with stronger association in female neonates (n = 37, β = −1.35, p = 0.06), although there is no evidence of a sex-specific effect. Analysis of maternal risk factors for neonatal brain ID as predictors of exosomal CNTN2 and BDNF levels showed sex-specific relationships between infants of diabetic mothers (IDMs) and CNTN2 levels (Interaction p = 0.0005). While male IDMs exhibited a negative correlation (n = 42, β = −0.69, p = 0.02), female IDMs showed a positive correlation (n = 37, β = 0.92, p = 0.01) with CNTN2. A negative correlation between BNDF and maternal BMI was found with stronger association in female neonates (per 10 units BMI, β = −0.60, p = 0.04). These findings suggest CNTN2 and BNDF are respective molecular markers for male and female neonates at risk for brain ID. This study supports the potential of exosomal markers to assess neonatal brain status in at-risk infants.

Alcohol's Dysregulation of Maternal-Fetal IL-6 and p-STAT3 Is a Function of Maternal Iron Status.

Prenatal alcohol exposure (PAE) causes long-term growth and neurodevelopmental deficits that are worsened by maternal iron deficiency (ID). In our preclinical rat model, PAE causes fetal anemia, brain ID, and elevated hepatic iron via increased maternal and fetal hepcidin synthesis. These changes are normalized by a prenatal iron-fortified (IF) diet. Here, we hypothesize that iron status and PAE dysregulate the major upstream pathways that govern hepcidin production-EPO/BMP6/SMAD and IL-6/JAK2/STAT3. Pregnant, Long Evans rat dams consumed ID (2 to 6ppm iron), iron-sufficient (IS, 100ppm iron), or IF (500ppm iron) diets and received alcohol (5g/kg) or isocaloric maltodextrin daily from gestational days (GD) 13.5 to 19.5. Protein and gene expression were quantified in the 6 experimental groups at GD 20.5. PAE did not affect Epo or Bmp6 expression, but reduced p-SMAD1/5/8/SMAD1/5/8 protein ratios in both IS and ID maternal and fetal liver (all p's<0.01). In contrast, PAE stimulated maternal hepatic expression of Il-6 (p=0.03) and elevated p-STAT3/STAT3 protein ratios in both IS and ID maternal and fetal liver (all p's<0.02). PAE modestly elevated maternal Il-1β, Tnf-α, and Ifn-γ. Fetal cytokine responses to PAE were muted compared with dams, and PAE did not affect hepatic Il-6 (p=0.78) in IS and ID fetuses. Dietary iron fortification sharply attenuated Il-6 expression in response to PAE, with IF driving a 150-fold decrease (p<0.001) in maternal liver and a 10-fold decrease (p<0.01) in fetal liver. The IF diet also normalized p-STAT3/STAT3 ratios in both maternal and fetal liver. These findings suggest that alcohol-driven stimulation of the IL-6/JAK2/STAT3 pathway mediates the elevated hepcidin observed in the PAE dam and fetus. Normalization of these signals by IF suggests that dysregulated hepcidin is driven by alcohol's disruption of the IL-6/JAK2/STAT3 pathway. Prenatal dietary IF represents a potential therapeutic approach for PAE that warrants further investigation.

Drug Treatment of Restless Legs Syndrome in Older Adults.

Restless legs syndrome (RLS) has a high prevalence in the elderly and can impact sleep quality and sleep quantity, reduce quality of life (QoL), and increase the risk of falls during episodes of night-time ambulation. In patients unable to verbalize their sensory symptoms, certain behavioral cues may help with the diagnosis. A state of brain iron deficiency could play a central role in the pathophysiology of RLS and be upstream to a series of dysfunctions that are not limited to the dopaminergic system. Management should initially emphasize lifestyle modifications and reduction of all possible iatrogenic contributors while maintaining a state of normal-high peripheral iron stores. Oral iron, in patients with ferritin levels < 75μg/dL, appears to be effective, although iron infusions should be considered when more immediate benefit or oral iron have not been effective. When other attempts fail and patients continue to experience chronic RLS symptoms substantially interfering with QoL, pharmacological agents may present a favorable benefit versus risk profile. Such agents may include α-2-δ drugs or dopaminergic agents, after careful consideration of the risk of RLS augmentation with the latter class. In patients with established RLS augmentation from the use of dopaminergic drugs, the addition of α-2-δ agents or low-dose opioids, with subsequent slow tapering of dopaminergic agents, is recommended. With any of these agents, caution should be made with regard to the risk of drug-drug interactions and altered pharmacokinetics in this fragile population. Although showing excellent long-term safety data in non-elderly adults with RLS, studies are needed to ascertain that such treatments are effective and well tolerated in older adults.

Adenosine mechanisms and hypersensitive corticostriatal terminals in restless legs syndrome. Rationale for the use of inhibitors of adenosine transport.

Our working hypothesis is that a hypoadenosinergic state is a main pathogenetic factor that determines the sensory-motor symptoms and hyperarousal of restless legs syndrome (RLS). We have recently demonstrated that brain iron deficiency (BID) in rodents, a well-accepted animal model of RLS, is associated with a generalized downregulation of adenosine A1 receptors (A1R) in the brain and with hypersensitivity of corticostriatal glutamatergic terminals. Here, we first review the experimental evidence for a pivotal role of adenosine and A1R in the control of striatal glutamatergic transmission and the rationale for targeting putative downregulated striatal A1R in RLS patients, which is supported by recent clinical results obtained with dipyridamole, an inhibitor of the nucleoside transporters ENT1 and ENT2. Second, we perform optogenetic-microdialysis experiments in rats to demonstrate that A1R determine the sensitivity of corticostriatal glutamatergic terminals and the ability of dipyridamole to counteract optogenetically-induced corticostriatal glutamate release in both animals with BID and controls. Thus, a frequency of optogenetic stimulation that was ineffective at inducing cortico-striatal glutamate release in control rats became effective with the local perfusion of a selective A1R antagonist. Furthermore, in animals with and without BID, the striatal application of dipyridamole blocked the optogenetic-induced glutamate release and decreased basal levels of glutamate, which was counteracted by the A1R antagonist. The results support the clinical application of ENT1 inhibitors in RLS.

The neurophysiology of hyperarousal in restless legs syndrome: Hints for a role of glutamate/GABA.

Restless legs syndrome (RLS) is a common sensory-motor circadian disorder, whose basic components include urge to move the legs, unpleasant sensory experience, and periodic leg movements during sleep, all associated with an enhancement of the individual's arousal state. Brain iron deficiency (BID) is considered to be a key initial pathobiological factor, based on alterations of iron acquisition by the brain, also moderated by genetic factors. In addition to the well-known dopaminergic involvement in RLS, previous studies pointed out that BID brings also a hyperglutamatergic state that influences a dysfunctional cortico-striatal-thalamic-cortical circuit in genetically vulnerable individuals. However, the enhancement of arousal mechanisms in RLS may also be explained by functional changes of the ascending arousal systems and by deficitary GABA-mediated inhibitory control. Very recently, it was also suggested that BID induces a hypoadenosinergic state in RLS, thus possibly providing a link for a putative unified pathophysiological mechanism accounting for both hyperarousal and sensory-motor signs. Consequently, RLS might be viewed as a multitransmitter neurochemical disorder, globally resulting in enhanced excitability and decreased inhibition. In this framework, understanding the complex interaction of different neuronal circuits in generating the symptoms of RLS is mandatory both for a better diagnostic refinement and for an innovative therapeutic support. Notably, multiple neurotransmission dysfunction, either primary or triggered by BID, may also bridge the gap between RLS and other chronic pain disorders. This chapter summarizes the current experimental and clinical findings into a heuristic model of the electrophysiology and neurochemistry underlying RLS.

Iron uptake at the blood-brain barrier is influenced by sex and genotype.

Iron transport to the brain is a critically important and highly regulated process necessary for proper brain function. This review aims to summarize iron uptake mechanisms in the brain and the importance of sex and genotype on this uptake. In restless legs syndrome (RLS), brain iron uptake has been hypothesized to be dysregulated, leading to the clinically observed brain iron deficiency, so this review specifically comments on this disorder. The review covers transferrin-bound transport and the more recently discovered role of ferritin in brain iron delivery. Studies on the impact of sex, MEIS1 (associated with RLS), and H63D gene variants have revealed critical regulatory mechanisms and driving factors that significantly impact iron transport that may considerably affect the clinical treatment of RLS. RLS has provided insights into the gaps in knowledge underlying brain iron uptake mechanisms and their regulation, which in turn have provided insights into the pathophysiology of RLS.

Examining Consequence of Brain Iron Deficiency in the Absence of Anemia

Examining Consequence of Brain Iron Deficiency in the Absence of Anemia

Reticulocyte hemoglobin content as an early predictive biomarker of brain iron deficiency.

Fetal and neonatal brain iron content is compromised at the time of anemia, suggesting that screening for iron deficiency by measuring hemoglobin is inadequate to protect the brain. Reticulocyte hemoglobin (Ret-He) reflects iron-deficient (ID) erythropoiesis prior to anemia. At postnatal day (P), 10 and 20 iron-sufficient rat pups were fostered to ID dams to produce a postnatal ID (PNID) group, which was compared to 20 iron-sufficient (IS) pups fostered by IS dams. Pups were assessed from P13 to P15 for hemoglobin, hematocrit, reticulocyte count, and Ret-He. Hippocampal iron status was assessed by transferrin receptor-1 (Tfrc-1) and divalent metal transporter-1 (Slc11a2) mRNA expression. At P13, brain iron status was similar between groups; only Ret-He was lower in the PNID group. At P14, the PNID group had lower Ret-He, hematocrit, mean corpuscular volume (MCV), and reticulocyte percentage (RET%). Tfrc-1 expression was increased, consistent with brain iron deficiency. Both Ret-He and MCV correlated with brain iron status at P14 and P15. Ret-He was the only red cell marker affected prior to the onset of brain ID. The clinical practice of using anemia as the preferred biomarker for diagnosis of iron deficiency may need reconsidering.

Endothelial cells are critical regulators of iron transport in a model of the human blood–brain barrier

Iron delivery to the brain is essential for multiple neurological processes such as myelination, neurotransmitter synthesis, and energy production. Loss of brain iron homeostasis is a significant factor in multiple neurological disorders. Understanding the mechanism by which the transport of iron across the blood–brain barrier (BBB) is regulated is crucial to address the impact of iron deficiency on brain development and excessive accumulation of iron in neurodegenerative diseases. Using induced pluripotent stem cell (iPSC)-derived brain endothelial cells (huECs) as a human BBB model, we demonstrate the ability of transferrin, hepcidin, and DMT1 to impact iron transport and release. Our model reveals a new function for H-ferritin to transport iron across the BBB by binding to the T-cell immunoglobulin and mucin receptor 1. We show that huECs secrete both transferrin and H-ferritin, which can serve as iron sources for the brain. Based on our data, brain iron status can exert control of iron transport across the endothelial cells that constitute the BBB. These data address a number of pertinent questions such as how brain iron uptake is regulated at the regional level, the source of iron delivery to the brain, and the clinical strategies for attempting to treat brain iron deficiency.

Pharmaceutical iron formulations do not cross a model of the human blood-brain barrier.

Whether iron formulations used therapeutically for a variety of conditions involving iron deficiency can deliver iron to the brain is a significant clinical question given the impact that iron loading has on the brain in neurodegenerative diseases. In this study, we examine the ability of 5 pharmaceutical iron formulations that are given intravenously for treatment of iron deficiency to cross an in vitro model of the blood-brain barrier. The model uses human brain endothelial cells derived from induced pluripotent stem cells. We report that, compared to the natural iron delivery proteins, transferrin and H-ferritin, the pharmaceutical iron formulations neither cross the blood-brain barrier model nor significantly load the endothelial cells with iron. Furthermore, we report that mimicking brain iron sufficiency or deficiency by exposing the endothelial cells to apo- or holo-transferrin does not alter the amount of iron compound transported by or loaded into the cells. Coupled with previous studies, we propose that pharmaceutical iron formulations must first be processed in macrophages to make iron bioavailable. The results of this study have significant clinical and mechanistic implications for the use of therapeutic iron formulations.

Growth Outcomes in Very Low Birth Weight Infants Receiving Erythropoietin and Aggressive Iron Therapy

Introduction: Very low birth weight (VLBW) infants are at risk for developing symptomatic anemia and iron deficiency anemia. Erythropoietin (EPO) has been used to stimulate erythropoiesis in at risk infants, and more recent literature suggests a possible neuroprotective effect. Iron is important for neurodevelopment, erythropoeisis and somatic growth in the preterm infant. Brain iron deficiency …

Treatment of restless legs syndrome/Willis-Ekbom disease with the non-selective ENT1/ENT2 inhibitor dipyridamole: testing the adenosine hypothesis

ObjectivesRecent animal models of restless legs syndrome (RLS) suggest that brain iron deficiency is associated with a hypoadenosinergic state, with downregulation of adenosine A1 receptors (A1R) in the striatum and cortex. We hypothesized that an increase in extracellular adenosine induced by inhibitors of adenosine transporters, such as the non-selective ENT1/ENT2 inhibitor dipyridamole, would result in an improvement in RLS symptoms. MethodsIn a prospective two-month open-label, non-placebo controlled clinical trial, 15 untreated idiopathic RLS patients began treatment with 100 mg dipyridamole (with uptitration to 400 mg if necessary). Multiple Suggested Immobilization Tests and polysomnography were performed at baseline and at eight weeks. Severity was assessed at four and eight weeks using the IRLS, and the CGI scales. The primary endpoint was therapeutic response (50% improvement in IRLS total score). ResultsThirteen patients completed the study. IRLS score improved from a mean (±S.D.) of 23.4 ± 4.6 at baseline to 10.7 ± 4.5 at eight weeks. Six out of 13 patients were full responders and four were partial responders. The mean (±S.D.) effective dose of dipyridamole at eight weeks was 281.8 ± 57.5 mg/day. Sleep variables also improved, and the mean (±S.D.) periodic leg movement index decreased from 26.7 ± 7.2 to 4.3 ± 1.9. Dipyridamole was generally well tolerated. Main side effects were abdominal cramps, diarrhea, dizziness, and flushing. ConclusionsThese preliminary results suggest that dipyridamole has significant therapeutic effects on both sensory and motor symptoms, as well as sleep. In addition, it provides evidence that hypoadenosinergic mechanisms play a central role in RLS. Classification of evidenceThe study provides class III evidence supporting the therapeutic effects of dipyridamole in RLS.

Approaches for Reducing the Risk of Early-Life Iron Deficiency-Induced Brain Dysfunction in Children.

Iron deficiency is the most common micronutrient deficiency in the world. Women of reproductive age and young children are particularly vulnerable. Iron deficiency in late prenatal and early postnatal periods can lead to long-term neurobehavioral deficits, despite iron treatment. This may occur because screening and treatment of iron deficiency in children is currently focused on detection of anemia and not neurodevelopment. Anemia is the end-stage state of iron deficiency. The brain becomes iron deficient before the onset of anemia due to prioritization of the available iron to the red blood cells (RBCs) over other organs. Brain iron deficiency, independent of anemia, is responsible for the adverse neurological effects. Early diagnosis and treatment of impending brain dysfunction in the pre-anemic stage is necessary to prevent neurological deficits. The currently available hematological indices are not sensitive biomarkers of brain iron deficiency and dysfunction. Studies in non-human primate models suggest that serum proteomic and metabolomic analyses may be superior for this purpose. Maternal iron supplementation, delayed clamping or milking of the umbilical cord, and early iron supplementation improve the iron status of at-risk infants. Whether these strategies prevent iron deficiency-induced brain dysfunction has yet to be determined. The potential for oxidant stress, altered gastrointestinal microbiome and other adverse effects associated with iron supplementation cautions against indiscriminate iron supplementation of children in malaria-endemic regions and iron-sufficient populations.

Pivotal Role of Adenosine Neurotransmission in Restless Legs Syndrome.

The symptomatology of Restless Legs Syndrome (RLS) includes periodic leg movements during sleep (PLMS), dysesthesias, and hyperarousal. Alterations in the dopaminergic system, a presynaptic hyperdopaminergic state, seem to be involved in PLMS, while alterations in glutamatergic neurotransmission, a presynaptic hyperglutamatergic state, seem to be involved in hyperarousal and also PLMS. Brain iron deficiency (BID) is well-recognized as a main initial pathophysiological mechanism of RLS. BID in rodents have provided a pathogenetic model of RLS that recapitulates the biochemical alterations of the dopaminergic system of RLS, although without PLMS-like motor abnormalities. On the other hand, BID in rodents reproduces the circadian sleep architecture of RLS, indicating the model could provide clues for the hyperglutamatergic state in RLS. We recently showed that BID in rodents is associated with changes in adenosinergic transmission, with downregulation of adenosine A1 receptors (A1R) as the most sensitive biochemical finding. It was hypothesized that A1R downregulation leads to hypersensitive striatal glutamatergic terminals and facilitation of striatal dopamine release. Hypersensitivity of striatal glutamatergic terminals was demonstrated by an optogenetic-microdialysis approach in the rodent with BID, indicating that it could represent a main pathogenetic factor that leads to PLMS in RLS. In fact, the dopaminergic agonists pramipexole and ropinirole and the α2δ ligand gabapentin, used in the initial symptomatic treatment of RLS, completely counteracted optogenetically-induced glutamate release from both normal and BID-induced hypersensitive corticostriatal glutamatergic terminals. It is a main tenet of this essay that, in RLS, a single alteration in the adenosinergic system, downregulation of A1R, disrupts the adenosine-dopamine-glutamate balance uniquely controlled by adenosine and dopamine receptor heteromers in the striatum and also the A1R-mediated inhibitory control of glutamatergic neurotransmission in the cortex and other non-striatal brain areas, which altogether determine both PLMS and hyperarousal. Since A1R agonists would be associated with severe cardiovascular effects, it was hypothesized that inhibitors of nucleoside equilibrative transporters, such as dipyridamole, by increasing the tonic A1R activation mediated by endogenous adenosine, could represent a new alternative therapeutic strategy for RLS. In fact, preliminary clinical data indicate that dipyridamole can significantly improve the symptomatology of RLS.

Targeting hypersensitive corticostriatal terminals in restless legs syndrome.

The first aim was to demonstrate a previously hypothesized increased sensitivity of corticostriatal glutamatergic terminals in the rodent with brain iron deficiency (BID), a pathogenetic model of restless legs syndrome (RLS). The second aim was to determine whether these putative hypersensitive terminals could constitute a significant target for drugs effective in RLS, including dopamine agonists (pramipexole and ropinirole) and α2 δ ligands (gabapentin). A recently introduced in vivo optogenetic-microdialysis approach was used, which allows the measurement of the extracellular concentration of glutamate upon local light-induced stimulation of corticostriatal glutamatergic terminals. The method also allows analysis of the effect of local perfusion of compounds within the same area being sampled for glutamate. BID rats showed hypersensitivity of corticostriatal glutamatergic terminals (lower frequency of optogenetic stimulation to induce glutamate release). Both hypersensitive and control glutamatergic terminals were significant targets for locally perfused pramipexole, ropinirole, and gabapentin, which significantly counteracted optogenetically induced glutamate release. The use of selective antagonists demonstrated the involvement of dopamine D4 and D2 receptor subtypes in the effects of pramipexole. Hypersensitivity of corticostriatal glutamatergic terminals can constitute a main pathogenetic mechanism of RLS symptoms. Selective D4 receptor agonists, by specifically targeting these terminals, should provide a new efficient treatment with fewer secondary effects. Ann Neurol 2017;82:951-960.