GABAergic signaling by cells of the immune system: more the rule than the exception.

Gamma-aminobutyric acid (GABA) is best known as an essential neurotransmitter in the evolved central nervous system (CNS) of vertebrates. However, GABA antedates the development of the CNS as a bioactive molecule in metabolism and stress-coupled responses of prokaryotes, invertebrates and plants. Here, we focus on the emerging findings of GABA signaling in the mammalian immune system. Recent reports show that mononuclear phagocytes and lymphocytes, for instance dendritic cells, microglia, T cells and NK cells, express a GABAergic signaling machinery. Mounting evidence shows that GABA receptor signaling impacts central immune functions, such as cell migration, cytokine secretion, immune cell activation and cytotoxic responses. Furthermore, the GABAergic signaling machinery of leukocytes is implicated in responses to microbial infection and is co-opted by protozoan parasites for colonization of the host. Peripheral GABA signaling is also implicated in inflammatory conditions and diseases, such as type 1 diabetes, rheumatoid arthritis and cancer cell metastasis. Adding to its role in neurotransmission, growing evidence shows that the non-proteinogenic amino acid GABA acts as an intercellular signaling molecule in the immune system and, as an interspecies signaling molecule in host-microbe interactions. Altogether, the data raise the assumption of conserved GABA signaling in a broad range of mammalian cells and diversification of function in the immune system.

Autoantibodies to synapsin I sequestrate synapsin I and alter synaptic function.

Synapsin I is a phosphoprotein that coats the cytoplasmic side of synaptic vesicles and regulates their trafficking within nerve terminals. Autoantibodies against Syn I have been described in sera and cerebrospinal fluids of patients with numerous neurological diseases, including limbic encephalitis and clinically isolated syndrome; however, the effects and fate of autoantibodies in neurons are still unexplored. We found that in vitro exposure of primary hippocampal neurons to patient's autoantibodies to SynI decreased the density of excitatory and inhibitory synapses and impaired both glutamatergic and GABAergic synaptic transmission. These effects were reproduced with a purified SynI antibody and completely absent in SynI knockout neurons. Autoantibodies to SynI are internalized by FcγII/III-mediated endocytosis, interact with endogenous SynI, and promote its sequestration and intracellular aggregation. Neurons exposed to human autoantibodies to SynI display a reduced density of SVs, mimicking the SynI loss-of-function phenotype. Our data indicate that autoantibodies to intracellular antigens such as SynI can reach and inactivate their targets and suggest that an antibody-mediated synaptic dysfunction may contribute to the evolution and progression of autoimmune-mediated neurological diseases positive for SynI autoantibodies.

Microglia-Triggered Plasticity of Intrinsic Excitability Modulates Psychomotor Behaviors in Acute Cerebellar Inflammation.

Cerebellar dysfunction relates to various psychiatric disorders, including autism spectrum and depressive disorders. However, the physiological aspect is less advanced. Here, we investigate the immune-triggered hyperexcitability in the cerebellum on a wider scope. Activated microglia via exposure to bacterial endotoxin lipopolysaccharide or heat-killed Gram-negative bacteria induce a potentiation of the intrinsic excitability in Purkinje neurons, which is suppressed by microglia-activity inhibitor and microglia depletion. An inflammatory cytokine, tumor necrosis factor alpha (TNF-α), released from microglia via toll-like receptor 4, triggers this plasticity. Our two-photon FRET ATP imaging shows an increase in ATP concentration following endotoxin exposure. Both TNF-α and ATP secretion facilitate synaptic transmission. Region-specific inflammation in the cerebellum in vivo shows depression- and autistic-like behaviors. Furthermore, both TNF-α inhibition and microglia depletion revert such behavioral abnormality. Resting-state functional MRI reveals overconnectivity between the inflamed cerebellum and the prefrontal neocortical regions. Thus, immune activity in the cerebellum induces neuronal hyperexcitability and disruption of psychomotor behaviors in animals.

Autoimmune Attack of the Neuromuscular Junction in Myasthenia Gravis: Nicotinic Acetylcholine Receptors and Other Targets.

The nicotinic acetylcholine receptor (nAChR) family, the archetype member of the pentameric ligand-gated ion channels, is ubiquitously distributed in the central and peripheral nervous systems, and its members are the targets for both genetic and acquired forms of neurological disorders. In the central nervous system, nAChRs contribute to the pathological mechanisms of neurodegenerative disorders, such as Alzheimer and Parkinson diseases. In the peripheral nerve-muscle synapse, the vertebrate neuromuscular junction, "classical" myasthenia gravis (MG) and other forms of neuromuscular transmission disorders are antibody-mediated autoimmune diseases. In MG, antibodies to the nAChR bind to the postsynaptic receptors and activate the classical complement pathway culminating in the formation of the membrane attack complex, with the subsequent destruction of the postsynaptic apparatus. Divalent nAChR-antibodies also cause internalization and loss of the nAChRs. Loss of receptors by either mechanism results in the muscle weakness and fatigability that typify the clinical manifestations of the disease. Other targets for antibodies, in a minority of patients, include muscle specific kinase (MuSK) and low-density lipoprotein related protein 4 (LRP4). This brief Review analyzes the current status of muscle-type nAChR in relation to the pathogenesis of autoimmune diseases affecting the peripheral cholinergic synapse.

Blocking Tumor Necrosis Factor-Alpha Expression Prevents Blast-Induced Excitatory/Inhibitory Synaptic Imbalance and Parvalbumin-Positive Interneuron Loss in the Hippocampus.

Traumatic brain injury (TBI) is a major cause of neurological disorder and death in civilian and military populations. It comprises two components-direct injury from the traumatic impact and secondary injury from ensuing neural inflammatory responses. Blocking tumor necrosis factor-alpha (TNF-α), a central regulator of neural inflammation, has been shown to improve functional recovery after TBI. However, the mechanisms underlying those therapeutic effects are still poorly understood. Here, we examined effects of 3,6'-dithiothalidomide (dTT), a potentially therapeutic TNF-α inhibitor, in mice with blast-induced TBI. We found that blast exposure resulted in elevated expression of TNF-α, activation of microglial cells, enhanced excitatory synaptic transmission, reduced inhibitory synaptic transmission, and a loss of parvalbumin-positive (PV+) inhibitory interneurons. Administration of dTT for 5 days after the blast exposure completely suppressed blast-induced increases in TNF-α transcription, largely reversed blasted-induced synaptic changes, and prevented PV+ neuron loss. However, blocking TNF-α expression by dTT failed to mitigate blast-induced microglial activation in the hippocampus, as evidenced by their non-ramified morphology. These results indicate that TNF-α plays a major role in modulating neuronal functions in blast-induced TBI and that it is a potential target for treatment of TBI-related brain disorders.

Myasthenia gravis: the role of complement at the neuromuscular junction.

Generalized myasthenia gravis (gMG) is a rare autoimmune disorder characterized by skeletal muscle weakness caused by disrupted neurotransmission at the neuromuscular junction (NMJ). Approximately 74-88% of patients with gMG have acetylcholine receptor (AChR) autoantibodies. Complement plays an important role in innate and antibody-mediated immunity, and activation and amplification of complement results in the formation of membrane attack complexes (MACs), lipophilic proteins that damage cell membranes. The role of complement in gMG has been demonstrated in animal models and patients. Studies in animals lacking specific complement proteins have confirmed that MAC formation is required to induce experimental autoimmune MG (EAMG) and NMJ damage. Complement inhibition in EAMG models can prevent disease induction and reverse its progression. Patients with anti-AChR+ MG have autoantibodies and MACs present at NMJs. Damaged NMJs are associated with more severe disease, fewer AChRs, and MACs in synaptic debris. Current MG therapies do not target complement directly. Eculizumab is a humanized monoclonal antibody that inhibits cleavage of complement protein C5, preventing MAC formation. Eculizumab treatment improved symptoms compared with placebo in a phase II study in patients with refractory gMG. Direct complement inhibition could preserve NMJ physiology and muscle function in patients with anti-AChR+ gMG.

Kidins220 and tumour development: Insights into a complexity of cross-talk among signalling pathways (Review).

The mechanistic complexes of kinase D-interacting substrate of 220 kDa/ankyrin repeat-rich membrane spanning (Kidins220/ARMS) bind and integrate a variety of cellular cues to mediate neuronal activities such as neuronal differentiation, survival, and cytoskeleton remodelling by interacting with a variety of binding partners. Accumulated evidence has also indicated its role in the regulation of vascular development. Mice with Kidins220 knockdown phenotypically present with cardiovascular abnormalities. Kidins220 also contributes to immunomodulation in combination with B cells and T cells. Moreover, emerging evidence has revealed that this protein regulates many crucial cellular processes and thus has been implicated in an increasing number of malignancies. Here, we review recent advances in our understanding of Kidins220 and its role in cancer development. Further investigation is warranted to shed light on the role played by Kidins220 in the dynamic arrangement of the cytoskeleton and epithelial-mesenchymal transition, and its implication in tumourigenesis and cancer progression.

Myasthenia gravis: recent advances in immunopathology and therapy.

INTRODUCTION: Myasthenia gravis is the most frequent acquired disorder of neuromuscular transmission. In the majority of cases, pathogenic antibodies against components of the postsynaptic muscle endplate membrane can be detected. In recent years there have been significant advances in the pathophysiological understanding and therapy of the disease. Areas covered: PubMed searches were conducted for the term 'myasthenia gravis' cross-referenced with the terms 'immunology', 'subgroups', 'antibody', 'ocular', 'thymoma', 'treatment' and 'thymectomy'. Additionally, we summarized the current state of immunopathology and therapy. Expert commentary: Immunological research defined new target antigens at the postsynaptic neuromuscular junction which along with clinical features allow a refined definition of disease subgroups. Overall the prognosis of myasthenia gravis with best possible symptomatic, immunosuppressive and supportive treatment is good but new immunomodulatory treatment options are developed for patients who do not respond well to the first line therapy. For most patients individually adapted long-term drug therapy is needed.

Evidence for TNFα action on excitatory and inhibitory neurotransmission in the central amygdala: a brain site influenced by stress.

Anxiety-like responses to stress are accompanied by elevation of brain cytokine-mRNAs. Because cytokines microinjected into central-amygdala (CeA) substitute for stress in a behavioral paradigm, the possibility was raised that cytokines increased by stress influence behavior by affecting CeA-neural activity. Previously, cytokines increased firing-rate of CeA-neurons comparable to that induced by corticotropin-releasing factor (CRF). In this investigation, tumor-necrosis-factor-α (TNFα) increased amplitude, but not frequency of mEPSCs from CeA-neurons. Additionally, TNFα decreased the threshold for triggering action potentials from CeA-neurons without altering membrane-properties during current-clamp recording. Glutamate-receptor-antagonist blockade of mEPSCs and the TNFα-induced reduction in firing threshold implicated glutamate in these changes. A phosphatidyl-inositol-3-kinase-antagonist prevented the TNFα-induced increased in amplitude of mEPSCs, documenting a TNFα intracellular influence. Additionally, TNFα increased frequency, but not amplitude of mIPSCs. CRF-receptor-antagonists were found to prevent the TNFα-induced increase in mIPSC-frequency, without altering the TNFα-induced amplitude increase in mEPSCs or the reduced threshold for action-potentials by TNFα. To clarify how TNFα was increasing CRF-release in the presence of tetrodotoxin, the possibility tested was whether preventing glial-activation would prevent this elevated mIPSC-frequency blocked by CRF-receptor antagonists. Minocycline, which blocks glial activation, prevented the TNFα-induced increase in mIPSC-frequency - a finding consistent with glia contributing to the CRF-involvement in this TNFα action. To fully understand the means by which a CRF1-receptor-antagonist and minocycline prevent TNFα from increasing mIPSC-frequency will require further clarification. Nonetheless, these data provide convincing evidence that release of TNFα by stress could alter neural activity of CeA-neurons by influencing GABA-and glutamate function.

Pathogenic and physiological autoantibodies in the central nervous system.

In this article, we review the current knowledge on pathological and physiological autoantibodies directed toward structures in the central nervous system (CNS) with an emphasis on their regulation and origin. Pathological autoantibodies in the CNS that are associated with autoimmunity often lead to severe neurological deficits via inflammatory processes such as encephalitis. In some instances, however, autoantibodies function as a marker for diagnostic purposes without contributing to the pathological process and/or disease progression. The existence of naturally occurring physiological autoantibodies has been known for a long time, and their role in maintaining homeostasis is well established. Within the brain, naturally occurring autoantibodies targeting aggregated proteins have been detected and might be promising candidates for new therapeutic approaches for neurodegenerative disorders. Further evidence has demonstrated the existence of naturally occurring antibodies targeting antigens on neurons and oligodendrocytes that promote axonal outgrowth and remyelination. The numerous actions of physiological autoantibodies as well as their regulation and origin are summarized in this review.

Neural antigen-specific autoimmune disorders.

Neural-specific autoantibodies have been documented and their diagnostic utility validated in diseases affecting the neuraxis from cerebral cortex to the somatic, autonomic, and enteric nervous system and skeletal muscle. These neurological disorders occur both idiopathically and in a paraneoplastic context. Molecular identification of the antigens has expedited development of confirmatory and high-throughput tests for serum and cerebrospinal fluid, which permit early diagnosis and reveal the underlying molecular pathogenic mechanisms. The autoantibodies are classifiable on the basis of antigen location: intracellular (nuclear or cytoplasmic) or plasma membrane. Immunohistopathological studies of patients' biopsied and autopsied tissues suggest that effector T cells mediate the autoimmune neurological disorders for which defining autoantibodies recognize intracellular antigens. Antigens within intact cells are inaccessible to circulating antibody, and the associated neurological deficits rarely improve with antibody-depleting therapies. Tumoricidal therapies may arrest neurological progression, but symptom reversal is rare. In contrast, autoantibodies specific for plasma membrane antigens have pathogenic potential, and the associated neurological deficits are often amenable to antibody-depleting immunotherapy, such as plasma exchange and anti-B-cell monoclonal antibody therapy. These reversible neurological disorders are frequently misdiagnosed as neurodegenerative. The focus of this review is the immunobiology, pathophysiology, and clinical spectrum of autoimmune neurological disorders accompanied by neural-specific IgGs.

Brain IL-6 elevation causes neuronal circuitry imbalances and mediates autism-like behaviors.

Abnormal immune responses have been reported to be associated with autism. A number of studies showed that cytokines were increased in the blood, brain, and cerebrospinal fluid of autistic subjects. Elevated IL-6 in autistic brain has been a consistent finding. However, the mechanisms by which IL-6 may be involved in the pathogenesis of autism are not well understood. Here we show that mice with elevated IL-6 in the brain display many autistic features, including impaired cognitive abilities, deficits in learning, abnormal anxiety traits and habituations, as well as decreased social interactions. IL-6 elevation caused alterations in excitatory and inhibitory synaptic formations and disrupted the balance of excitatory/inhibitory synaptic transmissions. IL-6 elevation also resulted in an abnormal change in the shape, length and distributing pattern of dendritic spines. These findings suggest that IL-6 elevation in the brain could mediate autistic-like behaviors, possibly through the imbalances of neural circuitry and impairments of synaptic plasticity.

Antibodies interfering with the type 3 muscarinic receptor pathway inhibit gastrointestinal motility and cholinergic neurotransmission in Sjögren's syndrome.

OBJECTIVE: In primary Sjögren's syndrome (SS), impairment of the gastrointestinal (GI) tract is common, and includes reduced esophageal motor function, delayed gastric emptying, and abnormalities in colonic motility; the pathogenesis is as yet unknown. We undertook this study to investigate the role of functional antibodies to the type 3 muscarinic receptor (M3R) in GI dysfunction associated with primary SS. METHODS: Muscle strip and whole-organ functional assays were used to determine whether IgG with anti-M3R activity from patients with primary SS disrupted neurotransmission in tissue from throughout the mouse GI tract. Specificity of the autoantibody for the M3R was determined using knockout mice that were deficient in the expression of muscarinic receptor subtypes. RESULTS: Functional antibodies to the M3R inhibited neuronally mediated contraction of smooth muscle from throughout the GI tract and disrupted complex contractile motility patterns in the colon. The autoantibodies were not active on tissue from mice that lacked the M3R, providing compelling evidence of the direct interaction of patient autoantibodies with the M3R. CONCLUSION: Our results indicate that anti-M3R autoantibodies have the potential to mediate multiple dysfunctions of the GI tract in primary SS, ranging from reduced esophageal motor activity to altered colonic motility. We hypothesize that altered GI motility forms part of a broader autonomic dysfunction mediated by pathogenic anti-M3R autoantibodies in primary SS.

CX3CL1-induced modulation at CA1 synapses reveals multiple mechanisms of EPSC modulation involving adenosine receptor subtypes.

We characterized the role of adenosine receptor (AR) subtypes in the modulation of glutamatergic neurotransmission by the chemokine fractalkine (CX3CL1) in mouse hippocampal CA1 neurons. CX(3)CL1 causes a reversible depression of excitatory postsynaptic current (EPSC), which is abolished by the A(3)R antagonist MRS1523, but not by A(1)R (DPCPX) or A(2A)R (SCH58261) antagonists. Consistently, CX3CL1-induced EPSC depression is absent in slices from A(3)R(-/-) but not A(1)R(-/-) or A(2A)R(-/-) mice. Further, A(3)R stimulation causes similar EPSC depression. In cultured neurons, CX3CL1-induced depression of AMPA current shows A(1)R-A(3)R pharmacology. We conclude that glutamatergic depression induced by released adenosine requires the stimulation of different ARs.

Autoantibodies to glutamate receptor GluRepsilon2 in a patient with limbic encephalitis associated with relapsing polychondritis.

Limbic encephalitis is a rare central nervous system (CNS) manifestation of relapsing polychondritis (RP). Vasculitis is assumed to be the cause of CNS involvement in RP. Several studies, however, have described CNS involvement in RP with no evidence of vasculitis but with a more nonspecific inflammatory picture. We report a patient with limbic encephalitis associated with RP who presented with anti-glutamate receptor (GluR) epsilon2 (NR2B) autoantibodies in his cerebrospinal fluid and sera. Brain MRI showed a high signal intensity lesion in the medial temporal lobe and progressive atrophy without multifocal abnormality on fluid-attenuated inversion recovery scanning. Our patient's results raise the interesting possibility that anti-GluRepsilon2 (NR2B) antibodies function in the development of limbic encephalitis in certain patients with RP.

Allogeneic hematopoietic cell transplantation for refractory myasthenia gravis.

OBJECTIVE: To describe a patient with intractable myasthenia gravis (MG) who was treated with a matched sibling peripheral blood stem cell transplantation. DESIGN: Case report. Patient A 17-year-old boy with MG diagnosed at 11 months of age who was previously treated with pyridostigmine, intravenous immunoglobulin, corticosteroids, thymectomies, azathioprine, mycophenolate mofetil, plasmaphereses, rituximab, and high-dose cyclophosphamide. RESULTS: The patient underwent a reduced-toxicity conditioning with intravenous busulfan, fludarabine, and alemtuzumab, followed by a peripheral blood stem cell infusion from his HLA-matched sibling. Before transplantation, the patient was receiving frequent plasmaphereses, intravenous immunoglobulin, and pyridostigmine. He had ophthalmoplegia, oropharyngeal and limb muscle involvement, and limited mobility. At 40 months posttransplantation, his oropharyngeal and skeletal muscle weakness has completely resolved, he is not taking any medications for MG, and he is an avid athlete. However, his ophthalmoplegia persists, and his anti-acetylcholine receptor antibody levels remain elevated. CONCLUSIONS: Following allogeneic hematopoietic stem cell transplantation, the presence of anti-acetylcholine receptor antibodies was not sufficient for inducing symptoms of MG. This confirms that additional immune mechanisms are important in pathogenesis of this disease. Allogeneic transplantation may be a therapeutic option for patients with severe, refractory MG. However, little is known about the long-term efficacy of allogeneic transplantation for this disease, and long-term follow-up is warranted.

The role of cytokines in the regulation of neurotransmission.

Cytokines are highly inducible, secretory proteins that mediate intercellular communication in the immune system. They are grouped in several protein families, namely tumor necrosis factors, interleukins, interferons and colony-stimulating factors. In recent years, evidence has elucidated that some of these proteins as well as their receptors are also produced in the central nervous system (CNS) by specific neural cell lineages under physiological and pathological conditions. Cytokines regulate a variety of processes in the CNS, including neurotransmission. The current data let us to suggest that cytokines play an important role in the regulation of both excitatory and inhibitory neurotransmission in the CNS. This knowledge could be fundamental for the proposal of new therapeutic approaches to neurological and psychiatric disorders.

Anti-MuSK patient antibodies disrupt the mouse neuromuscular junction.

OBJECTIVE: A subset of myasthenia gravis patients that are seronegative for anti-acetylcholine receptor (anti-AChR) antibodies are instead seropositive for antibodies against the muscle-specific kinase (anti-MuSK-positive). Here, we test whether transfer of IgG from anti-MuSK-positive patients to mice confers impairment of the neuromuscular junction and muscle weakness. METHODS: IgG from anti-MuSK-positive myasthenia gravis patients or control IgG (seronegative for AChR and MuSK) was injected intraperitoneally (45 mg daily for 14 days) into 6-week-old female FVB/NJ and C57BL/6J mice. Changes at neuromuscular junctions in the tibialis anterior and diaphragm muscles were assessed by confocal fluorescent imaging of AChRs stained with fluorescent-alpha-bungarotoxin. Loss of function was assessed by electromyography. RESULTS: In experimental mice injected with anti-MuSK-positive patient IgG, postsynaptic AChR staining was reduced to as little as 22% of that seen in control mice. Experimental mice showed reduced apposition of the nerve terminal (labeled with antibodies against synaptophysin and neurofilament) and the postsynaptic AChR cluster (labeled with fluorescent-alpha-bungarotoxin). Mice injected with IgG from two of three anti-MuSK-positive patients lost weight and developed muscle weakness associated with a decremental electromyographic trace on repetitive nerve stimulation. INTERPRETATION: IgG from anti-MuSK-positive patients can cause myasthenia gravis when injected into mice. This may be explained by a progressive reduction in the density of postsynaptic AChR combined with changes in the nerve terminal and its relation to the postsynaptic structure.

Autoimmune disorders of neuromuscular transmission.

Myasthenia gravis and Lambert-Eaton syndrome are autoimmune disorders of the neuromuscular junction. The most common form of myasthenia gravis is associated with antibodies directed against the acetylcholine receptor on the postsynaptic membrane. In Lambert-Eaton syndrome, antibodies are directed against P/Q-type voltage-gated calcium channels on presynaptic cholinergic nerve terminals at the neuromuscular junction and in the autonomic nervous system. Lambert-Eaton syndrome may represent a paraneoplastic disease that is most commonly associated with small-cell lung carcinoma or an autoimmune disorder. In both myasthenia gravis and Lambert-Eaton syndrome, the approach to treatment includes symptomatic and immune therapy. Symptomatic therapy in both disorders includes acetylcholinesterase inhibitors. In Lambert-Eaton syndrome, agents that augment the quantal release of acetylcholine are also effective. Immune therapy includes immune suppression with various medications, short-term immune modulation with plasma exchange and intravenous immunoglobulin, and thymectomy in myasthenia gravis. When Lambert-Eaton syndrome is associated with cancer, the disease may improve or remit with effective treatment of the underlying malignancy. Current treatment options will be summarized for both disorders.