Rapid and persistent loss of TXNIP in HT22 neuronal cells under carbonyl and hyperosmotic stress Publication date: January 2020 Source: Neurochemistry International, Volume 132 Author(s): Alcir Luiz Dafre, Ariana Ern Schmitz, Pamela Maher Abstract Thioredoxin interacting protein (TXNIP) binds to thioredoxin thereby limiting its activity, but it also promotes internalization of glucose transporters, participates in inflammasome activation, and controls autophagy. Published data and this work demonstrate that TXNIP responds to a number of apparently unrelated stresses, such as serum deprivation, pH change, and oxidative, osmotic and carbonyl stress. Interestingly, we noticed that hyperosmotic (NaCl) and carbonyl (methylglyoxal, MGO) stresses in HT22 neuronal cells produced a rapid loss of TXNIP (half-life ∼12 min), prompting us to search for possible mechanisms controlling this TXNIP loss, including pH change, serum deprivation, calcium metabolism and inhibition of the proteasome and other proteases, autophagy and MAPKs. None of these routes stopped the TXNIP loss induced by hyperosmotic and carbonyl stress. Besides transcriptional, translational and microRNA regulation, there is evidence indicating that mTOR and AMPK also control TXNIP expression. Indeed, AMPK-deficient mouse embryonic fibroblasts failed to respond to phenformin (AMPK activator) and compound C (AMPK inhibitor), while rapamycin induced a marked increase in TXNIP levels, confirming the known AMPK/mTOR control over TXNIP. However, the TXNIP loss induced by NaCl or MGO were observed even in AMPK deficient MEFs or after mTOR inhibition, indicating AMPK/mTOR does not participate in this rapid TXNIP loss. These results suggest that rapid TXNIP loss is a general and immediate response to stress that can improve energy availability and antioxidant protection, eventually culminating in better cell survival. Graphical abstract

Simultaneous quantification of dopamine, serotonin, their metabolites and amino acids by LC-MS/MS in mouse brain following repetitive transcranial magnetic stimulation Publication date: December 2019 Source: Neurochemistry International, Volume 131 Author(s): Eugenia Z. Poh, Dorothee Hahne, Jessica Moretti, Alan R. Harvey, Michael W. Clarke, Jennifer Rodger Abstract Repetitive Transcranial Magnetic Stimulation (rTMS) is a form of non-invasive brain stimulation that has shown therapeutic potential for various nervous system disorders. In addition to its modulatory effects on neuronal excitability, rTMS is capable of altering neurotransmitter (e.g., glutamate, GABA, dopamine and serotonin) concentrations in cortical and subcortical brain regions. Here we used a modified liquid chromatography coupled tandem mass spectrometry (LC-MS/MS) to quantify changes in 27 free amino acids and the monoamines: dopamine (DA), serotonin (5HT) and their metabolites (DOPAC, HVA; 5HIAA) in the mouse brain. Awake C57BL/6 J mice (either sex, 8–12 weeks old) received 10 Hz rTMS using two devices that can deliver low (LI-; 12 mT; custom built) or high (Fo8-; 1.2 T; MagVenture) intensity rTMS. Sham (unstimulated) mice were used as controls. Samples were collected immediately following a single session of rTMS or sham and processed for LC-MS/MS. The modified LC-MS/MS method used to detect DA, 5-HT and their metabolites showed good accuracy and precision with regression coefficients greater than 0.999, and an intra- and inter-day reproducibility with values < 13%. Fo8-rTMS induced a significant reduction in cortical 5-HT turnover rates, hippocampal DOPAC and an increase in striatal DOPAC concentrations. Fo8-rTMS also reduced concentrations of hippocampal α-aminoadipic acid, and striatal serine, threonine, sarcosine, aspartate and glutamate. There were no changes in the level of any compounds following LI-rTMS as compared to sham. The rapid change in monoamine turnover and amino acid concentrations following Fo8-rTMS but not LI-rTMS suggests that different stimulation parameters recruit different cellular mechanisms related to rTMS-induced plasticity. The described method can be used for the characterisation of trace levels of neurotransmitters and amino acids in brain tissue homogenates, providing a useful and precise tool to investigate localised neurotransmitter changes in animal models of health and disease.

Salubrinal attenuates nitric oxide mediated PERK:IRE1α: ATF-6 signaling and DNA damage in neuronal cells Publication date: December 2019 Source: Neurochemistry International, Volume 131 Author(s): Sonam Gupta, Joyshree Biswas, Parul Gupta, Abhishek Singh, Shubhangini Tiwari, Amit Mishra, Sarika Singh Abstract The present study was conducted to investigate the effect of salubrinal on nitric oxide mediated endoplasmic reticulum stress signaling and neuronal apoptosis. Rotenone treatment to neuro2a cells caused significantly decreased cell viability, increased cytotoxicity, augmented nitrite levels, increased nitrotyrosine level and augmented level of key ER stress markers (GRP-78, GADD153 and caspase-12). These augmented levels of ER stress markers could be attenuated with pretreatment of nitric oxide synthase inhibitor-aminoguanidine as well as with salubrinal. The rotenone treatment to neuro2a cells also triggered the ER stress induced up regulation of various signaling factors of unfolded protein response involving pPERK, ATF4, p-IRE1α, XBP-1 and ATF-6. Pretreatment of salubrinal significantly attenuated the activation of transmembrane kinases (PERK and IRE1) and ATF6 and restored the rotenone induced altered level of other UPR related signaling factors. Rotenone induced dephosphorylation of eIF2α was also inhibited with salubrinal treatment. Biochemically rotenone treatment to neuro2a cells caused the reactive oxygen species generation, depleted mitochondrial membrane potential and increased intra cellular calcium level which was attenuated with salubrinal treatment. Rotenone treatment to neuro2a cells also caused neuronal apoptosis, DNA fragmentation and chromatin condensation which were attenuated with salubrinal treatment. In conclusion, the findings suggested that rotenone causes the augmented level of nitric oxide which contributes in ER stress and could be inhibited by both aminoguanidine and/or salubrinal treatment. Further, salubrinal treatment attenuates the nitric oxide induced ER stress axis PERK:IRE1α:ATF-6 and inhibits the DNA damage and neuronal apoptosis.

Mechanism of the neuroprotective effect of GLP-1 in a rat model of Parkinson's with pre-existing diabetes Publication date: December 2019 Source: Neurochemistry International, Volume 131 Author(s): Eman A. Elbassuoni, Rasha F. Ahmed Abstract Several studies have suggested the association between neurodegenerative diseases and diabetes mellitus (DM), DM causes cognitive impairment with age, but its effect is not well known in Parkinson's disease (PD). As a member of the incretin family, Glucagon-like peptide-1 (GLP-1) has glycemic regulation functions. It also exerts many additional effects on different tissues through its receptor's widespread expression. Objective our aim is to investigate the effect of pre-existing diabetes on the severity of PD in male albino rats, and to find out whether GLP-1 could improve PD symptoms in diabetic animals in addition to its hypoglycemic effect, and how it could do that. Methods 75 adult male albino rats were equally divided into: Control, Parkinson's, Diabetic Parkinson's, Diabetic Parkinson's + low dose exenatide (GLP-1 receptor agonist), Diabetic Parkinson's + high dose exenatide group. Blood glucose and insulin, striatal dopamine, some striatal oxidative stress and inflammatory markers, and the catalepsy score were measured. Results Pre-existing of diabetes before initiation of PD raises the severity of PD shown by the more significant increase in catalepsy score, and the more significant decrease in striatal dopamine level. GLP-1 effects extend beyond their hypoglycemic effects only since it has a direct anti-oxidant, and anti-inflammatory neuronal effect with increasing the striatal dopamine and improving the catalepsy score in a dose dependent manner. Conclusions Diabetes increases the severity of impairment in PD, and GLP-1 improve it through its direct neuronal effect in addition to its indirect effect through producing hypoglycemia. Graphical abstract

PKCε-dependent H-Ras activation encompasses the recruitment of the RasGEF SOS1 and of the RasGAP neurofibromin in the lipid rafts of embryonic neurons Publication date: December 2019 Source: Neurochemistry International, Volume 131 Author(s): Sophia Karouzaki, Charoula Peta, Emmanouella Tsirimonaki, Dimitra Mangoura Abstract The spatial organization of plasma membrane proteins is a key factor in the generation of distinct signal outputs, especially for PKC/Ras/ERK signalling. Regulation of activation of the membrane-bound Ras, critical for neuronal differentiation and highly specialized functions, is controlled by exchanges in nucleotides catalyzed by nucleotide exchange factors (GEFs) for GTP loading and Ras activation, and by Ras GTPase Activated Proteins (RasGAPs) that lead to activation of the intrinsic GTPase activity of Ras and thus its inactivation. PKCs are potent Ras activators yet the mechanistic details of these interactions, or the involvement of specific PKC isoforms are now beginning to be addressed. Even less known is the topology where RasGAPs terminate Ras activation. Towards this aim, we isolated lipid rafts from chick embryo neural tissue and primary neuronal cultures when PKCε is the prominent isoform and in combination with in vitro kinase assays, we now show that, in response the PKCε-specific activating peptide ψεRACK, an activated PKCε is recruited to lipid rafts; similar mobility was established when PKCε was physiologically activated with the Cannabinoid receptor 1 (CB1) agonist methanandamide. Activation of H-Ras for both agents was then established for the first time using in vivo RasGAP activity assays, which showed similar temporal profiles of activation and lateral mobility. Moreover, we found that the GEF SOS1, and the major neuronal RasGAP neurofibromin, a specific PKCε substrate, were both transiently significantly enriched in the rafts. Finally, our in silico analysis revealed a highly probable, conserved palmitoylation site adjacent to a CARC motif on neurofibromin, both of which are included only in the RasGAP related domain type I (GRDI) with the known high H-RasGAP activity. Taken together, these results suggest that PKCε activation regulates the spatial plasma membrane enrichments of both SOS1 and neurofibromin, thus controlling the output of activated H-Ras available for downstream signalling in neurons.

The effect of coniferaldehyde on neurite outgrowth in neuroblastoma Neuro2a cells Publication date: December 2019 Source: Neurochemistry International, Volume 131 Author(s): Jieun Jeon, Huiyoung Kwon, Eunbi Cho, Kyung Sook Kim, Jeanho Yun, Young Choon Lee, Dong Hyun Kim Abstract Neurite outgrowth is the differentiation process by which neurons establish synapses. In the dentate gyrus of the hippocampus, new neurons are constantly produced and undergo neurite outgrowth to form synapses, and this process is involved in cognitive ability. Therefore, if an agent could modulate neurite outgrowth, it could potentially be developed as a compound for modulating cognitive ability. In this study, we examined whether coniferaldehyde, a natural compound, regulates neurite outgrowth in Neuro2a cells. We ascertained morphological changes and measured the percentage of neurite-bearing cells and neurite lengths. Coniferaldehyde significantly increased the percentage of neurite-bearing cells, and the length of neurites in a concentration-dependent manner, without inducing cell death. We then have identified that, coniferaldehyde activates the extracellular signals-regulated Kinase 1 and 2 (ERK1/2), and further noted that, U0126, an ERK1/2 inhibitor, blocks coniferaldehyde-facilitated neurite outgrowth. Moreover, Subchronic administration of CA enhanced learning and memory, and increased neurite length of newborn neurons in the hippocampus. These results suggest that coniferaldehyde induces neurite outgrowth by a process possibly mediated by ERK1/2 signaling and enhances learning and memory.

The pronociceptive role of 5-HT6 receptors in ventrolateral orbital cortex in a rat formalin test model Publication date: December 2019 Source: Neurochemistry International, Volume 131 Author(s): Yu-Xiang Zhang, Mei Yang, Feng Liang, Shao-Qing Li, Jing-Si Yang, Fu-Quan Huo, Chun-Xia Yan Abstract Recent studies have shown the 5-HT6 receptors are expressed in regions which are important in pain processing such as the cortex, amygdala, thalamus, PAG, spinal cord and dorsal root ganglia (DRG), suggesting a putative role of 5-HT6 receptors in pain modulation. The ventrolateral orbital cortex (VLO) is part of an endogenous analgesic system, consisting of the spinal cord – thalamic nucleus submedius (Sm) – VLO – periaqueductal gray (PAG) – spinal cord loop. The present study assessed the possible role of 5-HT6 receptors in the VLO in formalin-induced inflammatory pain model. Firstly we found that microinjection of selective 5-HT6 receptor agonists EMD-386088 (5 μg in 0.5 μl) and WAY-208466 (8 μg in 0.5 μl) both augmented 5% formalin-induced nociceptive behavior. Microinjection of selective 5-HT6 receptor antagonist SB-258585 (1,2 and 4 μg in 0.5 μl) significantly reduced formalin-induced flinching. Besides, the pronociceptive effects of EMD-386088 and WAY-208466 were dramatically reduced by SB-258585, implicating 5-HT6 receptor mechanisms in mediating these responses. In addition, the pronociceptive effect of EMD-386088 was also prevented by the adenylate cyclase (AC) inhibitor SQ-22536 (2 nmol in 0.5 μl) and the protein kinase A (PKA) inhibitor H89 (10 nmol in 0.5 μl), respectively. We further confirmed the above results with quantification of spinal c-fos expression. Taken together, our results suggested that 5-HT6 receptors play a pronociceptive role in the VLO in the rat formalin test due to its activation of AC - PKA pathway. Therefore, cerebral cortical 5-HT6 receptors could be a new target to develop analgesic drugs.

Histamine H1 and H3 receptor activation increases the expression of Glucose Transporter 1 (GLUT-1) in rat cerebro-cortical astrocytes in primary culture Publication date: December 2019 Source: Neurochemistry International, Volume 131 Author(s): Juan Parra-Abarca, Nayeli Rivera-Ramírez, Luis-Fernando Villa-Maldonado, Ubaldo García-Hernández, Penélope Aguilera, José-Antonio Arias-Montaño Abstract Astrocytes take up glucose via the 45 kDa isoform of the Glucose Transporter 1 (GLUT-1), and in this work we have investigated whether histamine regulates GLUT-1 expression in rat cerebro-cortical astrocytes in primary culture. Cultured astrocytes expressed histamine H1 and H3 receptors (H1Rs and H3Rs) as evaluated by radioligand binding. Receptor functionality was confirmed by the increase in the intracellular concentration of Ca2+ (H1R) and the inhibition of forskolin-induced cAMP accumulation (H3R). Quantitative RT-PCR showed that histamine and selective H1R and H3R agonists (1 h incubation) significantly increased GLUT-1 mRNA to 153 ± 7, 163 ± 2 and 168 ± 13% of control values, respectively. In immunoblot assays, incubation (3 h) with histamine or H1R and H3R agonists increased GLUT-1 protein levels to 224 ± 12, 305 ± 11 and 193 ± 13% of control values, respectively, an action confirmed by inmunocytochemistry. The effects of H1R and H3R agonists were blocked by the selective antagonists mepyramine (H1R) and clobenpropit (H3R). The pharmacological inhibition of protein kinase C (PKC) prevented the increase in GLUT-1 protein induced by either H1R or H3R activation. Furthermore, histamine increased ERK-1/2 phosphorylation, and the effect of H1R and H3R activation on GLUT-1 protein levels was reduced or prevented, respectively, by MEK-1/2 inhibition. These results indicate that by activating H1Rs and H3Rs histamine regulates the expression of GLUT-1 by astrocytes. The effect appears to involve the phospholipase C (PLC) → diacylglycerol (DAG)/Ca2+→ PKC and PLC → DAG/Ca2+ → PKC → MAPK pathways.

Tamoxifen promotes white matter recovery and cognitive functions in male mice after chronic hypoperfusion Publication date: December 2019 Source: Neurochemistry International, Volume 131 Author(s): Yuxue Chen, Yeye Tian, Hao Tian, Qibo Huang, Yongkang Fang, Wei Wang, Yue Wan, Dengji Pan, Minjie Xie Abstract Cerebral white matter lesions (WMLs) induced by chronic cerebral hypoperfusion are one of the major components of stroke pathology and closely associated with cognitive impairment. However, the repair and related pathophysiology of white matter after brain injury remains relatively elusive and underexplored. Successful neuroregeneration is a method for the potential treatment of central nervous system (CNS) disorders. A non-steroidal estrogen receptor modulator, Tamoxifen, is an effective inhibitor of cell-swelling-activated anion channels and can mimic neuroprotective effects of estrogen in experimental ischemic stroke. However, its remains unclear whether Tamoxifen has beneficial effects in the pathological process after WMLs. In the present study, we investigated the efficacy of Tamoxifen on multiple elements of oligovascular niche of the male C57BL/6 mice brain after bilateral carotid artery stenosis (BCAS) - induced WMLs. Tamoxifen was injected intraperitoneally once daily from 1 day after BCAS until 1 day before sacrificed. Following chronic hypoperfusion, BCAS mice presented white matter demyelination, loss of axon-glia integrity, activated inflammatory response, and cognitive impairments. Tamoxifen treatment significantly facilitated functional restoration of working memory impairment in mice after white matter injury, thus indicating a translational potential for this estrogen receptor modulator given its clinical safety and applicability for WMLs, which lack of currently available treatments. Furthermore, Tamoxifen treatment reduced microglia activation and inflammatory response, favored microglial polarization toward to the M2 phenotype, enhanced oligodendrocyte precursor cells proliferation and differentiation, and promoted remyelination after chronic hypoperfusion. Together, our data indicate that Tamoxifen could alleviate white matter injury and play multiple targets protective effects following chronic hypoperfusion, which is a promising candidate for the therapeutic target for ischemic WMLs and other demyelination diseases associated cognitive impairment.

The P2Y14 receptor in the trigeminal ganglion contributes to the maintenance of inflammatory pain Publication date: December 2019 Source: Neurochemistry International, Volume 131 Author(s): Jiu Lin, Yan-yan Zhang, Fei Liu, Xin-yi Fang, Meng-ke Liu, Chao-lan Huang, Hang Wang, Da-qing Liao, Cheng Zhou, Jie-fei Shen Abstract P2Y purinergic receptors expressed in neurons and satellite glial cells (SGCs) of the trigeminal ganglion (TG) contribute to inflammatory and neuropathic pain. P2Y14 receptor expression is reported in the spinal cord, dorsal root ganglion (DRG), and TG. In present study, the role of P2Y14 receptor in the TG in inflammatory orofacial pain of Sprague-Dawley (SD) rats was investigated. Peripheral injection of complete Freund's adjuvant (CFA) induced mechanical hyperalgesia with the rapid upregulation of P2Y14 receptor, glial fibrillary acidic protein (GFAP), interleukin-1β (IL-1β), tumor necrosis factor-α (TNF-α), C–C chemokine CCL2, phosphorylated extracellular signal-regulated kinase 1/2 (p-ERK1/2), and phosphorylated p38 (p-p38) proteins in the TG. Furthermore, immunofluorescence staining confirmed the CFA-induced upregulation of P2Y14 receptor. Double immunostaining showed that P2Y14 receptor colocalized with glutamine synthetase (GS) and neuronal nuclei (NeuN). Finally, trigeminal injection of a selective antagonist (PPTN) of P2Y14 receptor attenuated CFA-induced mechanical hyperalgesia. PPTN also decreased the upregulation of the GFAP, IL-1β, TNF-α, CCL2, p-ERK1/2, and p-p38 proteins. Our findings showed that P2Y14 receptor in TG may contribute to orofacial inflammatory pain via regulating SGCs activation, releasing cytokines (IL-1β, TNF-α, and CCL2), and phosphorylating ERK1/2 and p38.