Functional Conservation and Genetic Divergence of Chordate Glycinergic Neurotransmission: Insights from Amphioxus Glycine Transporters

Glycine is an important neurotransmitter in vertebrates, performing both excitatory and inhibitory actions. Synaptic levels of glycine are tightly controlled by the action of two glycine transporters, GlyT1 and GlyT2, located on the surface of glial cells and neurons, respectively. Only limited information is available on glycinergic neurotransmission in invertebrates, and the evolution of glycinergic neurotransmission is poorly understood. Here, by combining phylogenetic and gene expression analyses, we characterized the glycine transporter complement of amphioxus, an important invertebrate model for studying the evolution of chordates. We show that amphioxus possess three glycine transporter genes. Two of these (GlyT2.1 and GlyT2.2) are closely related to GlyT2 of vertebrates, whereas the third (GlyT) is a member of an ancestral clade of deuterostome glycine transporters. GlyT2.2 expression is predominantly non-neural, whereas GlyT and GlyT2.1 are widely expressed in the amphioxus nervous system and are differentially expressed, respectively, in neurons and glia. Vertebrate glycinergic neurons express GlyT2 and glia GlyT1, suggesting that the evolution of the chordate glycinergic system was accompanied by a paralog-specific inversion of gene expression. Despite this genetic divergence between amphioxus and vertebrates, we found strong evidence for conservation in the role glycinergic neurotransmission plays during larval swimming, the implication being that the neural networks controlling the rhythmic movement of chordate bodies may be homologous.

Glycine Transporters and Receptors as Targets for Analgesics

The suitability of modulating glycinergic neurotransmission for the treatment of inflammatory and chronic pain has gained widespread recognition, with glycine receptors (GlyRs) and glycine transporters (GlyT1 and GlyT2) now considered key therapeutic targets [...]

Functional Conservation and Genetic Divergence of Chordate Glycinergic Neurotransmission: Insights from Amphioxus Glycine Transporters

Glycine is an important neurotransmitter in vertebrates, performing both excitatory and inhibitory actions. Synaptic levels of glycine are tightly controlled by the action of two glycine transporters, GlyT1 and GlyT2, located on the surface of glial cells and glycinergic or glutamatergic neurons, respectively. Glycinergic neurotransmission in invertebrates has so far only been investigated in a very limited number of species, and, although it was suggested that its functions are to some extent conserved with vertebrates, the evolution of glycinergic neurotransmission remains very poorly understood. Here, by combining phylogenetic and gene expression analyses, we characterized the glycine transporter complement of amphioxus, an important invertebrate model for studying the evolution of chordates. We show that amphioxus possesses three glycine transporter genes, two of which (GlyT2.1 and GlyT2.2) are closely related to GlyT2 of vertebrates, while the other (GlyT) is a member of an ancestral clade of deuterostome glycine transporters. While expression of GlyT2.2 is predominantly non-neural, GlyT and GlyT2.1 are widely expressed in the amphioxus nervous system and are characterized by differential expression in neurons and glia, respectively. However, in vertebrates, glycinergic neurons express GlyT2 and glia GlyT1, suggesting that the evolution of the chordate glycinergic system was accompanied by complex genetic remodeling leading to the paralog-specific inversion of gene expression. Albeit this genetic divergence between amphioxus and vertebrates, we found strong evidence for a general conservation of the role of glycinergic neurotransmission during larval swimming, allowing us to hypothesize that the neural networks controlling the rhythmic movement of chordate bodies are homologous.

Glycinergic Signaling in Macrophages and Its Application in Macrophage-Associated Diseases

Accumulating evidences support that amino acids direct the fate decision of immune cells. Glycine is a simple structural amino acid acting as an inhibitory neurotransmitter. Besides, glycine receptors as well as glycine transporters are found in macrophages, indicating that glycine alters the functions of macrophages besides as an inhibitory neurotransmitter. Mechanistically, glycine shapes macrophage polarization via cellular signaling pathways (e.g., NF-κB, NRF2, and Akt) and microRNAs. Moreover, glycine has beneficial effects in preventing and/or treating macrophage-associated diseases such as colitis, NAFLD and ischemia-reperfusion injury. Collectively, this review highlights the conceivable role of glycinergic signaling for macrophage polarization and indicates the potential application of glycine supplementation as an adjuvant therapy in macrophage-associated diseases.

Association Study of the SLC1A2 (rs4354668), SLC6A9 (rs2486001) and SLC6A5 (rs2000959) Polymorphisms in Major Depressive Disorder

Background: In recent years, a growing body of evidence highlights a causal link between glutamate and depression. The membrane excitatory amino acid transporter 2 (EAAT2), encoded by SLC1A2 is responsible for the uptake and redistribution of most of the synaptic glutamate. Glycine, an inhibitory neurotransmitter, acts as an obligatory co-agonist of N-methyl-D-aspartate (NMDA) receptors and modulates excitatory neurotransmission. The clearance of synaptic glycine is performed by glycine transporters encoded by SLC6A9 and SLC6A5. Higher synaptic glycine and glutamate levels could enhance the activation of NMDA receptors, therefore counteract the hypofunction of glutamate neurotransmission described in major depressive disorder (MDD). The aim of the study was to assess whether polymorphisms of SCL1A2, SCL6A5 and SCL6A9 play a role in the development of MDD and its clinical picture in the Polish population. Methods: The study group consisted of 161 unrelated Caucasian patients with MDD, and 462 healthy controls. Polymorphisms of SLC1A2, SLC6A5 and SLC6A9 were genotyped with PCR-RLFP and TaqMan assays. The relationship between the studied single nucleotide polymorphisms (SNPs) was assessed based on the comparison of genotype and allele distribution in the study and the control group. The research also evaluated the relationships between the studied polymorphisms and clinical variables such as age of disease onset, number of episodes, duration of depression or severity of symptoms.Results: We found a statistically significant association between SLC1A2 rs4354668 polymorphism and MDD development (the frequency of rs4354668 CC genotype and allele C was 2-fold higher in patients than in the control). Such associations were not detected for SLC6A5 and SLC6A9 polymorphisms. No statistically significant impact of the studied SNPs on clinical variables of MDD was also observed. Conclusions: The results of the current study indicate an association of rs4354668 polymorphism in SCL1A2 with depression development in Polish population. Further studies with larger samples should be performed in the future to clarify the current findings.

Inhibition of Glycine Re-Uptake: A Potential Approach for Treating Pain by Augmenting Glycine-Mediated Spinal Neurotransmission and Blunting Central Nociceptive Signaling

Among the myriad of cellular and molecular processes identified as contributing to pathological pain, disinhibition of spinal cord nociceptive signaling to higher cortical centers plays a critical role. Importantly, evidence suggests that impaired glycinergic neurotransmission develops in the dorsal horn of the spinal cord in inflammatory and neuropathic pain models and is a key maladaptive mechanism causing mechanical hyperalgesia and allodynia. Thus, it has been hypothesized that pharmacological agents capable of augmenting glycinergic tone within the dorsal horn may be able to blunt or block aberrant nociceptor signaling to the brain and serve as a novel class of analgesics for various pathological pain states. Indeed, drugs that enhance dysfunctional glycinergic transmission, and in particular inhibitors of the glycine transporters (GlyT1 and GlyT2), are generating widespread interest as a potential class of novel analgesics. The GlyTs are Na+/Cl−-dependent transporters of the solute carrier 6 (SLC6) family and it has been proposed that the inhibition of them presents a possible mechanism by which to increase spinal extracellular glycine concentrations and enhance GlyR-mediated inhibitory neurotransmission in the dorsal horn. Various inhibitors of both GlyT1 and GlyT2 have demonstrated broad analgesic efficacy in several preclinical models of acute and chronic pain, providing promise for the approach to deliver a first-in-class non-opioid analgesic with a mechanism of action differentiated from current standard of care. This review will highlight the therapeutic potential of GlyT inhibitors as a novel class of analgesics, present recent advances reported for the field, and discuss the key challenges associated with the development of a GlyT inhibitor into a safe and effective agent to treat pain.

Iron status influences non-alcoholic fatty liver disease in obesity through the gut microbiome

Background The gut microbiome and iron status are known to play a role in the pathophysiology of non-alcoholic fatty liver disease (NAFLD), although their complex interaction remains unclear. Results Here, we applied an integrative systems medicine approach (faecal metagenomics, plasma and urine metabolomics, hepatic transcriptomics) in 2 well-characterised human cohorts of subjects with obesity (discovery n = 49 and validation n = 628) and an independent cohort formed by both individuals with and without obesity (n = 130), combined with in vitro and animal models. Serum ferritin levels, as a markers of liver iron stores, were positively associated with liver fat accumulation in parallel with lower gut microbial gene richness, composition and functionality. Specifically, ferritin had strong negative associations with the Pasteurellaceae, Leuconostocaceae and Micrococcaea families. It also had consistent negative associations with several Veillonella, Bifidobacterium and Lactobacillus species, but positive associations with Bacteroides and Prevotella spp. Notably, the ferritin-associated bacterial families had a strong correlation with iron-related liver genes. In addition, several bacterial functions related to iron metabolism (transport, chelation, heme and siderophore biosynthesis) and NAFLD (fatty acid and glutathione biosynthesis) were also associated with the host serum ferritin levels. This iron-related microbiome signature was linked to a transcriptomic and metabolomic signature associated to the degree of liver fat accumulation through hepatic glucose metabolism. In particular, we found a consistent association among serum ferritin, Pasteurellaceae and Micrococcacea families, bacterial functions involved in histidine transport, the host circulating histidine levels and the liver expression of GYS2 and SEC24B. Serum ferritin was also related to bacterial glycine transporters, the host glycine serum levels and the liver expression of glycine transporters. The transcriptomic findings were replicated in human primary hepatocytes, where iron supplementation also led to triglycerides accumulation and induced the expression of lipid and iron metabolism genes in synergy with palmitic acid. We further explored the direct impact of the microbiome on iron metabolism and liver fact accumulation through transplantation of faecal microbiota into recipient’s mice. In line with the results in humans, transplantation from ‘high ferritin donors’ resulted in alterations in several genes related to iron metabolism and fatty acid accumulation in recipient’s mice. Conclusions Altogether, a significant interplay among the gut microbiome, iron status and liver fat accumulation is revealed, with potential significance for target therapies.

A System for Assessing Dual Action Modulators of Glycine Transporters and Glycine Receptors

Reduced inhibitory glycinergic neurotransmission is implicated in a number of neurological conditions such as neuropathic pain, schizophrenia, epilepsy and hyperekplexia. Restoring glycinergic signalling may be an effective method of treating these pathologies. Glycine transporters (GlyTs) control synaptic and extra-synaptic glycine concentrations and slowing the reuptake of glycine using specific GlyT inhibitors will increase glycine extracellular concentrations and increase glycine receptor (GlyR) activation. Glycinergic neurotransmission can also be improved through positive allosteric modulation (PAM) of GlyRs. Despite efforts to manipulate this synapse, no therapeutics currently target it. We propose that dual action modulators of both GlyTs and GlyRs may show greater therapeutic potential than those targeting individual proteins. To show this, we have characterized a co-expression system in Xenopus laevis oocytes consisting of GlyT1 or GlyT2 co-expressed with GlyRα1. We use two electrode voltage clamp recording techniques to measure the impact of GlyTs on GlyRs and the effects of modulators of these proteins. We show that increases in GlyT density in close proximity to GlyRs diminish receptor currents. Reductions in GlyR mediated currents are not observed when non-transportable GlyR agonists are applied or when Na+ is not available. GlyTs reduce glycine concentrations across different concentration ranges, corresponding with their ion-coupling stoichiometry, and full receptor currents can be restored when GlyTs are blocked with selective inhibitors. We show that partial inhibition of GlyT2 and modest GlyRα1 potentiation using a dual action compound, is as useful in restoring GlyR currents as a full and potent single target GlyT2 inhibitor or single target GlyRα1 PAM. The co-expression system developed in this study will provide a robust means for assessing the likely impact of GlyR PAMs and GlyT inhibitors on glycine neurotransmission.