Loss Of Inhibitory Signaling Research Articles

Helicobacter pylori Infection Increased Anti-dsDNA and Enhanced Lupus Severity in Symptomatic FcγRIIb-Deficient Lupus Mice.

The defect on Fc gamma receptor IIb (FcγRIIb), the only inhibitory FcγR, has been identified as one of the genetic factors increasing susceptibility to lupus. The prevalence of Helicobacter pylori (HP) and FcγRIIb dysfunction-polymorphisms are high among Asians, and their co-existence is possible. Unfortunately, the influence of HP against lupus progression in patients with lupus is still controversial. In this study, the interactions between these conditions were tested with HP infection in 24-week-old FcγRIIb-/- mice (symptomatic lupus). HP induced failure to thrive, increased stomach bacterial burdens and stomach injury (histology and cytokines) in both wild-type and FcγRIIb-/- mice. While the severity of HP infection, as determined by these parameters, was not different between both strains, antibodies production (anti-HP, anti-dsDNA and serum gammaglobulin) were higher in FcγRIIb-/- mice compared to wild-type. Accordingly, HP infection also accelerated the severity of lupus as determined by proteinuria, serum creatinine, serum cytokines, renal histology, and renal immune complex deposition. Although HP increased serum cytokines in both wild-type and FcγRIIb-/- mice, the levels were higher in FcγRIIb-/- mice. As such, HP also increased spleen weight and induced several splenic immune cells responsible for antibody productions (activated B cell, plasma cell and follicular helper T cell) in FcγRIIb-/- mice, but not in wild-type. These data describe the different systemic responses against localized HP infection from diverse host genetic background. In conclusion, the mutual interactions between HP and lupus manifestations of FcγRIIb-/-mice were demonstrated in this study. With the prominent immune responses from the loss of inhibitory signaling in FcγRIIb-/- mice, HP infection in these mice induced intense chronic inflammation, increased antibody production, and enhanced lupus severity. Thus, the increased systemic inflammatory responses due to localized HP inducing gastritis in some patients with lupus may enhance lupus progression. More studies are needed.

Activity of novel lipid glycine transporter inhibitors on synaptic signalling in the dorsal horn of the spinal cord.

Inhibitory neurotransmission plays an important role in controlling excitability within nociceptive circuits of the spinal cord dorsal horn. Loss of inhibitory signalling is thought to contribute to the development of pathological pain. Preclinical studies suggest that increasing inhibitory glycinergic signalling is a good therapeutic strategy for treating pain. One approach to increase synaptic glycine is to inhibit the activity of the glycine transporter 2 (GlyT2) on inhibitory nerve terminals. These transporters are involved in regulating glycine concentrations and recycling glycine into presynaptic terminals. Inhibiting activity of GlyT2 increases synaptic glycine, which decreases excitability in nociceptive circuits and provides analgesia in neuropathic and inflammatory pain models. We investigated the effects of reversible and irreversible GlyT2 inhibitors on inhibitory glycinergic and NMDA receptor-mediated excitatory neurotransmission in the rat dorsal horn. The effect of these drugs on synaptic signalling was determined using patch-clamp electrophysiology techniques to measure glycine- and NMDA-mediated postsynaptic currents in spinal cord slices in vitro. We compared activity of four compounds that increase glycinergic tone with a corresponding increase in evoked glycinergic postsynaptic currents. These compounds did not deplete synaptic glycine release over time. Interestingly, none of these compounds increased glycine-mediated excitatory signalling through NMDA receptors. The results suggest that these compounds preferentially inhibit GlyT2 over G1yT1 with no potentiation of the glycine receptor and without inducing spillover from inhibitory to excitatory synapses. GlyT2 inhibitors increase inhibitory neurotransmission in the dorsal horn and have potential as pain therapeutics. This article is part of a themed section on Recent Advances in Targeting Ion Channels to Treat Chronic Pain. To view the other articles in this section visit http://onlinelibrary.wiley.com/doi/10.1111/bph.v175.12/issuetoc.

New approaches to target glycinergic neurotransmission for the treatment of chronic pain

Inhibitory glycinergic neurotransmission in the spinal cord dorsal horn plays an important role in regulating nociceptive signalling by inhibiting neuronal excitation. Blocking glycinergic transmission in the dorsal horn causes normally innocuous stimuli to become painful (allodynia) and increases sensitivity to noxious stimuli (hyperalgesia). Loss of inhibitory signalling is thought to contribute to the development of pathological pain. Management of neuropathic pain with current therapeutics is challenging and there is a great need for more effective treatments. Preclinical studies using drugs that increase glycinergic signalling by potentiating glycine receptor activity or inhibiting transporter activity suggest that targeting this system is a good therapeutic strategy. The spatially restricted expression of glycine receptors and transporters is an advantage for targeting specific pathologies such as pain. However, until recently there have been few pharmacological modulators identified and most of which do not specifically target glycinergic signalling. This mini-review provides an overview of recent advances in the development of therapeutics and novel approaches that aim to increase glycinergic neurotransmission for the treatment of persistent pain.

Reducing GABAA-mediated inhibition improves forelimb motor function after focal cortical stroke in mice.

A deeper understanding of post-stroke plasticity is critical to devise more effective pharmacological and rehabilitative treatments. The GABAergic system is one of the key modulators of neuronal plasticity, and plays an important role in the control of “critical periods” during brain development. Here, we report a key role for GABAergic inhibition in functional restoration following ischemia in the adult mouse forelimb motor cortex. After stroke, the majority of cortical sites in peri-infarct areas evoked simultaneous movements of forelimb, hindlimb and tail, consistent with a loss of inhibitory signalling. Accordingly, we found a delayed decrease in several GABAergic markers that accompanied cortical reorganization. To test whether reductions in GABAergic signalling were causally involved in motor improvements, we treated animals during an early post-stroke period with a benzodiazepine inverse agonist, which impairs GABAA receptor function. We found that hampering GABAA signalling led to significant restoration of function in general motor tests (i.e., gridwalk and pellet reaching tasks), with no significant impact on the kinematics of reaching movements. Improvements were persistent as they remained detectable about three weeks after treatment. These data demonstrate a key role for GABAergic inhibition in limiting motor improvements after cortical stroke.

Glycinergic dysfunction in a subpopulation of dorsal horn interneurons in a rat model of neuropathic pain.

The development of neuropathic pain involves persistent changes in signalling within pain pathways. Reduced inhibitory signalling in the spinal cord following nerve-injury has been used to explain sensory signs of neuropathic pain but specific circuits that lose inhibitory input have not been identified. This study shows a specific population of spinal cord interneurons, radial neurons, lose glycinergic inhibitory input in a rat partial sciatic nerve ligation (PNL) model of neuropathic pain. Radial neurons are excitatory neurons located in lamina II of the dorsal horn, and are readily identified by their morphology. The amplitude of electrically-evoked glycinergic inhibitory post-synaptic currents (eIPSCs) was greatly reduced in radial neurons following nerve-injury associated with increased paired-pulse ratio. There was also a reduction in frequency of spontaneous IPSCs (sIPSCs) and miniature IPSCs (mIPSC) in radial neurons without significantly affecting mIPSC amplitude. A subtype selective receptor antagonist and western blots established reversion to expression of the immature glycine receptor subunit GlyRα2 in radial neurons after PNL, consistent with slowed decay times of IPSCs. This study has important implications as it identifies a glycinergic synaptic connection in a specific population of dorsal horn neurons where loss of inhibitory signalling may contribute to signs of neuropathic pain.

Sialic acids and autoimmune disease.

An important underlying mechanism that contributes to autoimmunity is the loss of inhibitory signaling in the immune system. Sialic acid-recognizing Ig superfamily lectins or Siglecs are a family of cell surface proteins largely expressed in hematopoietic cells. The majority of Siglecs are inhibitory receptors expressed in immune cells that bind to sialic acid-containing ligands and recruit SH2-domain-containing tyrosine phosphatases to their cytoplasmic tails. They deliver inhibitory signals that can contribute to the constraining of immune cells, and thus protect the host from autoimmunity. The inhibitory functions of CD22/Siglec-2 and Siglec-G and their contributions to tolerance and autoimmunity, primarily in the B lymphocyte context, are considered in some detail in this review. The relevance to autoimmunity and unregulated inflammation of modified sialic acids, enzymes that modify sialic acid, and other sialic acid-binding proteins are also reviewed.

New approaches to the management of schizophrenia: focus on aberrant hippocampal drive of dopamine pathways.

Schizophrenia is a disease affecting up to 1% of the population. Current therapies are based on the efficacy of chlorpromazine, discovered over 50 years ago. These drugs block dopamine D2-like receptors and are effective at primarily treating positive symptoms in a subset of patients. Unfortunately, current therapies are far from adequate, and novel treatments require a better understanding of disease pathophysiology. Here we review the dopamine, gamma-aminobutyric acid (GABA), and glutamate hypotheses of schizophrenia and describe a pathway whereby a loss of inhibitory signaling in ventral regions of the hippocampus actually drives a dopamine hyperfunction. Moreover, we discuss novel therapeutic approaches aimed at attenuating ventral hippocampal activity in a preclinical model of schizophrenia, namely the MAM GD17 rat. Specifically, pharmacological (allosteric modulators of the α5 GABAA receptor), neurosurgical (deep brain stimulation), and cell-based (GABAergic precursor transplants) therapies are discussed. By better understanding the underlying circuit level dysfunctions in schizophrenia, novel treatments can be advanced that may provide better efficacy and a superior side effect profile to conventional antipsychotic medications.

LAIR and collagens in immune regulation

The immune system constantly receives opposing signals that on the one side activate immune cells allowing them to eradicate diseased cells and pathogens, and on the other side inhibit these same cells to limit and ultimately terminate an immune response. A correct balance is crucial for effective defence against pathogens without affecting healthy tissues. One of the mechanisms of negative regulation in the immune system is provided by specialized inhibitory receptors that upon interaction with their ligands attenuate activation signals initiated by activating receptors. Loss of inhibitory signaling is often associated with autoreactivity and unchecked inflammatory responses, illustrating the essential role these systems play in immune regulation. This thesis is focused on the function of such an inhibitory receptor called leukocyte-associated immunoglobulin-like receptor (LAIR)-1, which is expressed on most immune cells in peripheral blood. Although the inhibitory potential and mechanisms of LAIR-1 inhibition are well established, the actual function of the receptor in vivo remains unknown due the lack of animal models and the unknown identity of the LAIR-1 ligand. In the first part we identified and characterized mouse and rat LAIR-1 orthologues of human LAIR-1. Like human LAIR-1, mouse LAIR-1 is expressed on immune cells and functions as an inhibitory receptor. Furthermore we show that human and mouse LAIR-1 bind to similar, but not identical, cytoplasmic signalling effector molecules. In the second part we identified collagens as ligands for the LAIR-1 molecules. Both transmembrane and extracellular matrix collagens interact with LAIR-1 and the interaction results in inhibition of immune cell functions via LAIR-1. Our data reveal a novel mechanism of peripheral immune regulation by inhibitory immune receptors binding to extracellular matrix collagens. Additionally, we studied the functional interaction of LAIR-1 and collagens by using synthetic trimeric collagen peptides overlapping the entire human collagen II and III proteins. We show that LAIR-1 binds to multiple sites on these collagens and that the relative content of collagen-specific glycine-proline-hydroxyproline triplets in the proteins is associated with increased LAIR-1 binding. Besides the transmembrane LAIR-1, humans encode soluble LAIR-2 receptors. For the first time we provide evidence that LAIR-2 is expressed in humans as a soluble collagen receptor and that the molecule can block the collagen/LAIR-1 interaction. Hence, we hypothesize that soluble LAIR-2 may function as a natural competitor for LAIR-1 in vivo, thereby regulating its inhibitory potential. Further studies should aim at dissecting the in vivo role of LAIR-1 and LAIR-2 in normal physiology and disease. Furthermore, detailed studies on the biochemical interaction between collagens and LAIR may allow rational design of new and potent research tools and could result in the development of therapeutics which exploits collagen/LAIR interactions.

Plexin C1, A Receptor for Semaphorin 7A, Inactivates Cofilin and Is a Potential Tumor Suppressor for Melanoma Progression

Melanocytes are progenitor cells for melanoma, which arises through step-wise progression from dysplastic to invasive, to metastatic tumor. Our previous data showed that semaphorin 7A (Sema7A), a protein involved in axon guidance, stimulates melanocyte adhesion and dendricity through opposing actions of beta1-integrin and Plexin C1 receptors. We now show that Plexin C1 is diminished or absent in human melanoma cell lines; analysis of tissue microarrays of nevi, melanoma, and metastatic melanoma showed a decrease in Plexin C1 expression in metastatic melanoma, and an inverse correlation of Plexin C1 expression with depth of invasion. We examined the signaling intermediates of Sema7A and downstream targets of Plexin C1 in human melanocytes. Sema7A activated mitogen-activated protein kinase and inactivated cofilin, an actin-binding protein involved in cell migration. When Plexin C1 expression was silenced, Sema7A failed to phosphorylate cofilin, indicating that cofilin is downstream of Plexin C1. Further, Lim kinase II, a protein that phosphorylates cofilin, is upregulated by Sema7A in a Plexin C1-dependent manner. These data identify Plexin C1 as a potential tumor suppressor protein in melanoma progression, and suggest that loss of Plexin C1 expression may promote melanoma invasion and metastasis through loss of inhibitory signaling on cofilin activation.

Ablation of Neurons Expressing Agouti-Related Protein ActivatesFosand Gliosis in Postsynaptic Target Regions

We have developed a mouse model in which a specific population of inhibitory neurons can be selectively ablated by the action of diphtheria toxin (DT). The model involves targeting the human DT receptor to the agouti-related protein (Agrp) locus so that systemic administration of DT kills all of the AgRP-expressing neurons, resulting in starvation of the mice. Ablation of AgRP neurons results in robust (5- to 10-fold) activation of Fos gene expression in many brain regions that are innervated by AgRP neurons, including the arcuate nucleus (ARC), the paraventricular nucleus, the medial preoptic area, the lateral septum, and nucleus of the solitary tract. As expected, there is robust increase in GFAP staining (astrocytes) as well as IBA1 and CD11b staining (microglia) in the ARC in response to AgRP neuron ablation. There is also a dramatic increase of these markers in most, but not all, postsynaptic targets of AgRP axons. We used a genetic approach to reduce melanocortin signaling, which attenuated Fos activation in some brain regions after ablation of AgRP neurons. We suggest that loss of inhibitory signaling onto target neurons results in unopposed excitation that is responsible for the activation of Fos and that dysregulation of these neuronal circuits is responsible for starvation. Furthermore, glial cell activation in target areas of AgRP neurons appears to be a result of excitotoxicity.