Wheel-Running Exercise Alleviates Anxiety-Like Behavior via Down-Regulating S-Nitrosylation of Gephyrin in the Basolateral Amygdala of Male Rats.

Physical exercise has beneficial effect on anxiety disorders, but the underlying molecular mechanism remains largely unknown. Here, it is demonstrated that physical exercise can downregulate the S-nitrosylation of gephyrin (SNO-gephyrin) in the basolateral amygdala (BLA) to exert anxiolytic effects. It is found that the level of SNO-gephyrin is significantly increased in the BLA of high-anxiety rats and a downregulation of SNO-gephyrin at cysteines 212 and 284 produced anxiolytic effect. Mechanistically, inhibition of SNO-gephyrin by either Cys212 or Cys284 mutations increased the surface expression of GABAAR γ2 and the subsequent GABAergic neurotransmission, exerting anxiolytic effect in male rats. On the other side, overexpression of neuronal nitric oxide synthase in the BLA abolished the anxiolytic-like effects of physical exercise. This study reveals a key role of downregulating SNO-gephyrin in the anxiolytic effects of physical exercise, providing a new explanation for protein post-translational modifications in the brain after exercise.

Mitochondrial fission drives neuronal metabolic burden to promote stress susceptibility in male mice.

Neurons are particularly susceptible to energy fluctuations in response to stress. Mitochondrial fission is highly regulated to generate ATP via oxidative phosphorylation; however, the role of a regulator of mitochondrial fission in neuronal energy metabolism and synaptic efficacy under chronic stress remains elusive. Here, we show that chronic stress promotes mitochondrial fission in the medial prefrontal cortex via activating dynamin-related protein 1 (Drp1), resulting in mitochondrial dysfunction in male mice. Both pharmacological inhibition and genetic reduction of Drp1 ameliorates the deficit of excitatory synaptic transmission and stress-related depressive-like behavior. In addition, enhancing Drp1 fission promotes stress susceptibility, which is alleviated by coenzyme Q10, which potentiates mitochondrial ATP production. Together, our findings unmask the role of Drp1-dependent mitochondrial fission in the deficits of neuronal metabolic burden and depressive-like behavior and provides medication basis for metabolism-related emotional disorders.

BNIP3L/NIX-mediated mitophagy alleviates passive stress-coping behaviors induced by tumor necrosis factor-α.

Recent studies based on animal models of various neurological disorders have indicated that mitophagy, a selective autophagy that eliminates damaged and superfluous mitochondria through autophagic degradation, may be involved in various neurological diseases. As an important mechanism of cellular stress response, much less is known about the role of mitophagy in stress-related mood disorders. Here, we found that tumor necrosis factor-α (TNF-α), an inflammation cytokine that plays a particular role in stress responses, impaired the mitophagy in the medial prefrontal cortex (mPFC) via triggering degradation of an outer mitochondrial membrane protein, NIP3-like protein X (NIX). The deficits in the NIX-mediated mitophagy by TNF-α led to the accumulation of damaged mitochondria, which triggered synaptic defects and behavioral abnormalities. Genetic ablation of NIX in the excitatory neurons of mPFC caused passive coping behaviors to stress, and overexpression of NIX in the mPFC improved TNF-α-induced synaptic and behavioral abnormalities. Notably, ketamine, a rapid on-set and long-lasting antidepressant, reversed the TNF-α-induced behavioral abnormalities through activation of NIX-mediated mitophagy. Furthermore, the downregulation of NIX level was also observed in the blood of major depressive disorder patients and the mPFC tissue of animal models. Infliximab, a clinically used TNF-α antagonist, alleviated both chronic stress- and inflammation-induced behavioral abnormalities via restoring NIX level. Taken together, these results suggest that NIX-mediated mitophagy links inflammation signaling to passive coping behaviors to stress, which underlies the pathophysiology of stress-related emotional disorders.

A-Kinase Anchoring Protein 150 and Protein Kinase A Complex in the Basolateral Amygdala Contributes to Depressive-like Behaviors Induced by Chronic Restraint Stress.

BACKGROUND: The basolateral amygdala (BLA) has been widely implicated in the pathophysiology of major depressive disorder. A-kinase anchoring protein 150 (AKAP150) directs kinases and phosphatases to synaptic glutamate receptors, controlling synaptic transmission and plasticity. However, the role of the AKAP150 in the BLA in major depressive disorder remains poorly understood. METHODS: Depressive-like behaviors in C57BL/6J mice were developed by chronic restraint stress (CRS). Mice received either intra-BLA injection of lentivirus-expressing Akap5 short hairpin RNA or Ht-31, a peptide to disrupt the interaction of AKAP150 and protein kinase A (PKA), followed by depressive-like behavioral tests. Alpha-amino-3-hydroxy-5-methyl-4-isoxazole propionic acid glutamate receptor (AMPAR)-mediated miniature excitatory postsynaptic currents were recorded by whole-cell patch-clamp techniques. RESULTS: Chronic stress exposure induced depressive-like behaviors, which were accompanied by an increase in total and synaptic AKAP150 expression in the BLA. Accordingly, CRS facilitated the association of AKAP150 with PKA, but not of calcineurin in the BLA. Intra-BLA infusion of lentivirus-expressing Akap5 short hairpin RNA or Ht-31 prevented depressive-like behaviors and normalized phosphorylation of serine 845 and surface expression of AMPAR subunit 1 (GluA1) in the BLA of CRS mice. Finally, blockage of AKAP150-PKA complex signaling rescued the changes in AMPAR-mediated miniature excitatory postsynaptic currents in depressive-like mice. CONCLUSIONS: These results suggest that AKAP150-PKA directly modulates BLA neuronal synaptic strength, and that AKAP150-PKA-GluA1 streamline signaling complex is responsible for CRS-induced disruption of synaptic AMPAR-mediated transmission and depressive-like behaviors in mice.

Activity-Dependent Sulfhydration Signal Controls N-Methyl-D-Aspartate Subtype Glutamate Receptor-Dependent Synaptic Plasticity via Increasing d-Serine Availability.

AIMS: Reactive sulfur species, including hydrogen sulfide (H2S) and its oxydates, have been raised as novel redox signaling molecules. The present study aimed at examining whether endogenous sulfhydration signal is required for long-term potentiation (LTP), a cellular model for memory. RESULTS: In this study, we found that increased synaptic activity triggered sulfide generation and protein sulfhydration. Activity-triggered sulfide production was essential for N-methyl-D-aspartate subtype glutamate receptor (NMDAR)-dependent LTP via maintaining the availability of d-serine, a primary coagonist for synaptic NMDARs. Genetic knockdown of cystathionine ß-synthase, not cystathionine γ-lyase, impaired LTP. H2S increased NMDAR-dependent LTP via sulfhydration and disinhibition of serine racemase (SR), a main synthetase of d-serine. We found that polysulfides also increased NMDAR-dependent LTP and NMDAR activity. In aged rats, the level of H2S and SR sulfhydration decreased significantly. Exogenous supplement of H2S restored the sulfhydration of SR, followed by the improvement of age-related deficits in LTP. Furthermore, boost of H2S signal in vivo improves hippocampus-dependent memory. Innovation and Conclusion: Our results provide a direct evidence for the biological significance of endogenous sulfhydration signal in synaptic plasticity. Exogenous supplement of H2S could be considered as the new therapeutic approach for the treatment of neurocognitive dysfunction after aging. Antioxid. Redox Signal. 27, 398-414.

Dimethyl sulfide protects against oxidative stress and extends lifespan via a methionine sulfoxide reductase A-dependent catalytic mechanism.

Methionine (Met) sulfoxide reductase A (MsrA) is a key endogenous antioxidative enzyme with longevity benefits in animals. Only very few approaches have been reported to enhance MsrA function. Recent reports have indicated that the antioxidant capability of MsrA may involve a Met oxidase activity that facilities the reaction of Met with reactive oxygen species (ROS). Herein, we used a homology modeling approach to search the substrates for the oxidase activity of MsrA. We found that dimethyl sulfide (DMS), a main metabolite that produced by marine algae, emerged as a good substrate for MsrA-catalytic antioxidation. MsrA bounds to DMS and promoted its antioxidant capacity via facilitating the reaction of DMS with ROS through a sulfonium intermediate at residues Cys72, Tyr103, and Glu115, followed by the release of dimethyl sulfoxide (DMSO). DMS reduced the antimycin A-induced ROS generation in cultured PC12 cells and alleviated oxidative stress. Supplement of DMS exhibited cytoprotection and extended longevity in both Caenorhabditis elegans and Drosophila. MsrA knockdown abolished the cytoprotective effect and the longevity benefits of DMS. Furthermore, we found that the level of physiologic DMS was at the low micromolar range in different tissues of mammals and its level decreased after aging. This study opened a new window to elucidate the biological role of DMS and other low-molecular sulfides in the cytoprotection and aging.

AMPK Mediates Glucocorticoids Stress-Induced Downregulation of the Glucocorticoid Receptor in Cultured Rat Prefrontal Cortical Astrocytes.

Chronic stress induces altered energy metabolism and plays important roles in the etiology of depression, in which the glucocorticoid negative feedback is disrupted due to imbalanced glucocorticoid receptor (GR) functions. The mechanism underlying the dysregulation of GR by chronic stress remains elusive. In this study, we investigated the role of AMP-activated protein kinase (AMPK), the key enzyme regulating cellular energy metabolism, and related signaling pathways in chronic stress-induced GR dysregulation. In cultured rat cortical astrocytes, glucocorticoid treatment decreased the level, which was accompanied by the decreased expression of liver kinase B1 (LKB1) and reduced phosphorylation of AMPK. Glucocorticoid-induced effects were attenuated by glucocorticoid-inducible kinase 1 (SGK1) inhibitor GSK650394, which also inhibited glucocorticoid induced phosphorylation of Forkhead box O3a (FOXO3a). Furthermore, glucocorticoid-induced down-regulation of GR was mimicked by the inhibition of AMPK and abolished by the AMPK activators or the histone deacetylase 5 (HDAC5) inhibitors. In line with the role of AMPK in GR expression, AMPK activator metformin reversed glucocorticoid-induced reduction of AMPK phosphorylation and GR expression as well as behavioral alteration of rats. Taken together, these results suggest that chronic stress activates SGK1 and suppresses the expression of LKB1 via inhibitory phosphorylation of FOXO3a. Downregulated LKB1 contributes to reduced activation of AMPK, leading to the dephosphorylation of HDAC5 and the suppression of transcription of GR.

Potentiation of Surface Stability of AMPA Receptors by Sulfhydryl Compounds: A Redox-Independent Effect by Disrupting Palmitoylation.

Sulfhydryl compounds such as dithiothreitol (DTT) and ß-mercaptoethanol (ß-ME) are widely used as redox agents. Previous studies in our group and other laboratory have reported the effect of sulfhydryl compounds on the function of glutamate receptor, including plasticity. Most of these findings have focused on the N-methyl-D-aspartic acid receptor, in contrast, very little is known about the effect of sulfhydryl compounds on α-amino-3-hydroxy-5-methylisoxazole-4-propionic acid receptor (AMPAR). Here, we observed that DTT (100 µM), ß-ME (200 µM) and L-cysteine (200 µM) significantly elevated the surface expression of AMPARs via reducing their palmitoylation in rat hippocampal slices in vitro. Increased surface stability of AMPARs was not be correlated with the altered redox status, because the chemical entities containing mercapto group such as penicillamine (200 µM) and 2-mercapto-1-methylimidazole (200 µM) exhibited little effects on the surface expression of AMPARs. Computing results of Asp-His-His-Cys (DHHC) 3, the main enzyme for palmitoylation of AMPARs, indicated that only the alkyl mercaptans with chain-like configuration, such as DTT and ß-ME, can enter the pocket of DHHC3 and disrupt the catalytic activity via inhibiting DHHC3 auto-palmitoylation. Collectively, our findings indicate a novel redox-independent mechanism underlay the multiple effects of thiol reductants on synaptic function.

Chronic administration tetrahydroxystilbene glucoside promotes hippocampal memory and synaptic plasticity and activates ERKs, CaMKII and SIRT1/miR-134 in vivo.

ETHNOPHARMACOLOGICAL RELEVANCE: Polygonum multiflorum Thunb is a traditional Chinese medicine with anti-aging effect. 2,3,5,4'-tetrahydroxystilbene-2-O-ß-D-glucoside (TSG) is generally considered as the main active component in Polygonum multiflorum Thunb. However, the effect of TSG on memory in adult is unclear till now. AIM OF STUDY: 2,3,5,4'-tetrahydroxystilbene-2-O-ß-D-glucoside (TSG) is a polyphenols compound from Polygonum multiflorum Thunb. The present study aimed to evaluate the effect of chronic administration of TSG on hippocampal memory in normal mice. MATERIALS AND METHODS: Behavioral test, electrophysiology and golgi staining were used to evaluate the effect of TSG on hippocampus-dependent memory and synaptic plasticity. Western blotting was used to determine the expression of ERK1/2, CaMKII, and SIRT1. Real-time quantitative PCR was explored to measure miR-134. RESULTS: It was found that TSG enhanced hippocampus-dependent contextual fear memory and novel object recognition, facilitated hippocampal LTP and increased dendrite spine density in the CA1 region of hippocampus. TSG obviously promoted the phosphorylations of ERK1/2, CaMKII, CREB and the expression of BDNF in the hippocampus, with upregulation of silent information regulator 1 (SIRT1) and downregulation of miR-134. CONCLUSIONS: Chronic administration of TSG promotes hippocampal memory in normal mice, suggesting that supplementary of TSG might serve as an enhancement of memory.

Sulfite triggers sustained calcium overload in cultured cortical neurons via a redox-dependent mechanism.

Sulfite is a compound commonly used as preservative in foods and pharmaceuticals. Many studies have examined the neurotoxicity of sulfite, but its effect on neuronal calcium homeostasis has not yet been reported. Here, we observed the effect of sulfite on the cytosolic free calcium concentration ([Ca(2+)]i) in cultured cortical neurons using Fura-2/AM based calcium imaging technique. Sulfite (250-1000µM) caused a sustained increase in [Ca(2+)]i in the neurons via a dose-dependent manner. In Ca(2+)-free solution, sulfite failed to increase [Ca(2+)]i. After the depletion of the intracellular calcium store, the effect of sulfite on the [Ca(2+)]i was largely abolished. Pharmacological inhibition of phospholipase C (PLC)-inositol 1,4,5-triphosphate (IP3) signaling pathway blocked sulfite-induced increase of [Ca(2+)]i. Interestingly, antioxidants such as trolox and dithiothreitol, abolished the increase of [Ca(2+)]i induced by sulfite. Exposure to sulfite triggered generation of sulfur- and oxygen-centered free radicals in neurons and increased oxidative stress both in the cultured cortical neurons and the prefrontal cortex of rats. Furthemore, sulfite decreased cell viability in cultured cortical neurons via a calcium-dependent manner. Thus, our current study suggests that the redox-dependent calcium overload triggered by sulfite in cortical neuronsmay be involved in its neurotoxicity.

HFS-Triggered AMPK Activation Phosphorylates GSK3ß and Induces E-LTP in Rat Hippocampus In Vivo.

BACKGROUND: The AMP-activated protein kinase (AMPK) is a sensor of cellular energy and nutrient status, with substantial amount of cross talk with other signaling pathways, including its phosphorylation by Akt, PKA, and GSK3ß. AIMS: Various signaling pathways and energy-consuming transport of glutamate receptors subunits are required in synaptic plasticity. However, it is unknown which energy sensors integrate the signaling pathways in these processes. In this article, we elucidated the role of AMPK activation and GSK3ß phosphorylation after HFS during the inducement of early-phase long-term potentiation (E-LTP). METHODS: Synaptic LTP in vivo was induced by high-frequency stimulation (HFS at 200 Hz at a 5-s interval). In addition, phosphorylation of AMPK and glycogen synthase kinase 3ß (GSK3ß) were measured using Western blotting. The amount of hippocampal AMP, ADP and ATP was measured by HPLC. RESULTS: We showed that the phosphorylation of AMPK and GSK3ß was significantly increased by HFS in vivo. HFS-induced AMPK activation occurred via increased (AMP + ADP)/ATP ratio and activation of Ca(2+) /calmodulin-dependent kinase kinase beta (CaMKKß). Pharmacological inhibition of AMPK by compound C (CC) prevented HFS-induced inhibitory phosphorylation of GSK3ß and the induction of LTP in dentate gyrus (DG) area in vivo. CONCLUSIONS: Our findings reveal that HFS-triggered AMPK activation phosphorylates GSK3ß and induces E-LTP in vivo.

Resveratrol promotes cellular glucose utilization in primary cultured cortical neurons via calcium-dependent signaling pathway.

BACKGROUND AND PURPOSE: Impairment of glucose utilization contributes to neuronal degeneration of Alzheimer's disease patients. Cellular glucose utilization can be regulated by calcium-dependent signaling pathways. Resveratrol (RSV) is a plant-derived polyphenol with multiple beneficial effects, including neuroprotection and metabolic improvement. Here, we investigated the effect of RSV on neuronal calcium signal and glucose utilization. EXPERIMENTAL METHODS: Primary culture of cortical neurons, calcium imaging, 2-NBDG assay and western blotting were employed to investigate RSV-mediated effects on neuronal calcium signal and glucose utilization. RESULTS: RSV elevated intracellular calcium in cortical neurons via modulation of secondary messenger system including nitrous oxide, cGMP and cAMP. Secondarily, a calcium-dependent enhancement of neuronal glucose utilization after RSV treatment was observed. The effects on neuronal glucose utilization are largely dependent on RSV-induced calcium-dependent AMP-activated protein kinase activation. CONCLUSION: Our findings show that activation of calcium-dependent signaling pathways by RSV may convey improvements of neuronal glucose utilization.

Resveratrol preconditioning increases methionine sulfoxide reductases A expression and enhances resistance of human neuroblastoma cells to neurotoxins.

Methionine sulfoxide reductases A (MsrA) has been postulated to act as a catalytic antioxidant system involved in the protection of oxidative stress-induced cell injury. Recently, attention has turned to MsrA in coupling with the pathology of Parkinson's disease, which is closely related to neurotoxins that cause dopaminergic neuron degeneration. Here, we firstly provided evidence that pretreatment with a natural polyphenol resveratrol (RSV) up-regulated the expression of MsrA in human neuroblastoma SH-SY5Y cells. It was also observed that the expression and nuclear translocation of forkhead box group O 3a (FOXO3a), a transcription factor that activates the human MsrA promoter, increased after RSV pretreatment. Nicotinamide , an inhibitor of silent information regulator 1 (SIRT1), prevented RSV-induced elevation of FOXO3a and MsrA expression, indicating that the effect of RSV was mediated by a SIRT1-dependent pathway. RSV preconditioning increased methionine sulfoxide(MetO)-reducing activity in SH-SY5Y cells and enhanced their resistance to neurotoxins, including chloramine-T and 1-methyl-4-phenyl-pyridinium. In addition, the enhancement of cell resistance to neurotoxins caused by RSV preconditioning can be largely prevented by MsrA inhibitor dimethyl sulfoxide. Our findings suggest that treatment with polyphenols such as RSV can be used as a potential regulatory strategy for MsrA expression and function.

Tetrahydroxystilbene glucoside, a plant-derived cognitive enhancer, promotes hippocampal synaptic plasticity.

Plant or food derived polyphenols have received a great deal of attention due to their biological properties including anti-oxidative effects, neuroprotection and memory enhancement. Here, we investigated the roles of 2,3,5,4'-tetrahydroxystilbene-2-O-ß-d-glucoside (TSG), an active component of the rhizome extracted from Polygonum multiflorum, in the regulation of hippocampal synaptic plasticity in normal mice as well as the underlying mechanisms. It was shown that TSG promoted the differentiation of PC12 cells and increased the intracellular calcium level in hippocampal neurons. TSG facilitated high-frequency stimulation (HFS)-induced hippocampal long-term potentiation (LTP) in a bell-shaped manner. The facilitation of LTP induction by TSG required calcium/calmodulin-dependent protein kinase II (CaMKII) and extracellular signal-regulated kinase (ERK) activation. Taken together, our data demonstrate that TSG promotes LTP induction and this effect may contribute to the enhancement of learning and memory seen in animal models.

The flavonoid baicalein promotes NMDA receptor-dependent long-term potentiation and enhances memory.

BACKGROUND AND PURPOSE: There is growing interest in the physiological functions of flavonoids, especially in their effects on cognitive function and on neurodegenerative diseases. The aim of the current investigation was to evaluate the role of the flavonoid baicalein in long-term potentiation (LTP) in the hippocampal CA1 region and cognitive behavioural performance. EXPERIMENTAL APPROACH: Effects of baicalein on LTP in rat hippocampal slices were investigated by electrophysiological methods. Phosphorylation of Akt (at Ser(473)), the extracellular signal-regulated kinase (ERK1/2) and the transcription factor cAMP response element-binding protein (CREB) (at Ser(133)) were analysed by Western blot. Fear conditioning was used to determine whether baicalein could improve learning and memory in rats. KEY RESULTS: Baicalein enhanced the N-methyl-d-aspartate glutamate receptor-dependent LTP in a bell-shaped concentration-dependent manner. Addition of the lipoxygenase metabolites 12(S)-HETE and 12(S)-HPETE did not reverse these effects of baicalein. Baicalein treatment enhanced phosphorylation of Akt during induction of LTP with the same bell-shaped dose-response curve. LTP potentiation induced by baicalein was blocked by inhibitors of phosphoinositide 3-kinase. CREB phosphorylation was also increased in the CA1 region of baicalein-treated slices. Baicalein-treated rats performed significantly better than controls in a hippocampus-dependent contextual fear conditioning task. Furthermore, baicalein treatment selectively increased the phosphorylation of Akt and CREB in the CA1 region of hippocampus, but not in the prefrontal cortex, after fear conditioning training. CONCLUSIONS AND IMPLICATIONS: Our results demonstrate that the flavonoid baicalein can facilitate memory, and therefore it might be useful in the treatment of patients with memory disorders.

HPLC and LC-MS analysis of sinomenine and its application in pharmacokinetic studies in rats.

AIM: To improve and validate analytical methods based on HPLC and liquid chromatography coupled to electrospray ionization tandem mass spectrometry (LC-ESI-MS/MS) for the quantitative measurement of sinomenine in rat plasma and brain tissue. METHODS: The separation of analytes and the internal standard (IS), chloramphenicol, was performed on an Agilent TC-C18 column (250×4.6 mm, 5 µm). Blood samples were measured with a Surveyor photodiode array (PDA) detector at a wavelength of 263 nm. The LCQ DECA XP(Plus) mass spectrometer was operated in the multiple reactions monitoring mode using positive electrospray ionization, and the transition from the precursor ion (m/z 279) to the product ion (m/z 224) for sinomenine was measured in brain tissue. RESULTS: Measurements were linear over the concentration range of 0.1-100 µg/mL for sinomenine in plasma and over the range of 0.01-5.00 µg/g for sinomenine in brain tissue. The intra- and inter-day variabilities were less than 10% of the relative standard deviation (RSD), and the extraction and recovery of sinomenine was 72.48%-80.26% from plasma and 73.75%-80.26% from brain tissue. The limit of quantification (LOQ) was 0.1 µg/mL for plasma, and 0.01 µg/g for brain tissue. Identification of sinomenine was reproducible at 0.5, 5, and 50 µg/mL in the plasma and at 0.05, 0.50, and 2.00 µg/g in brain tissue. The concentration of sinomenine measured in brain tissue after a single ip dose had a neuroprotective effect on H2O2-induced injury in PC12 cells in vitro. CONCLUSION: Our methods offered a sensitivity within a wide linear concentration range for sinomenine. These methods were successfully applied to evaluate sinomenine pharmacokinetics over time in rat brain tissue after a single ip dose of 30 mg/kg.

Differential effects of methionine and cysteine oxidation on [Ca2+] i in cultured hippocampal neurons.

Methionine and cysteine residues in proteins are the major targets of reactive oxygen species (ROS). The present work was designed to characterize the impact of methionine and cysteine oxidation upon [Ca(2+)](i) in hippocampal neurons. We investigated the effects of H(2)O(2) and chloramine T(Ch-T) agents known to oxidize both cysteine and methionine residues, and 5, 5'-dithio-bis (2-nitrobenzoic acid) (DTNB)--a cysteine-specific oxidant, on the intracellular calcium in hippocampal neurons. The results showed that these three oxidants, 1 mM H(2)O(2), 1 mM Ch-T, and 500 microM DTNB, induced an sustained elevation of [Ca(2+)](i) by 76.1 +/- 3.9%, 86.5 +/- 5.0%, and 24.4 +/- 3.2% over the basal level, respectively. The elevation induced by H(2)O(2) and Ch-T was significantly higher than DTNB. Pretreatment with reductant DTT at 1 mM for 10 min completely prevented the action of DTNB on [Ca(2+)](i), but only partially reduced the effects of H(2)O(2) and Ch-T on [Ca(2+)](i), the reductions were 44.6 +/- 4.2% and 29.6 +/- 6.1% over baseline, respectively. The elevation of [Ca(2+)](i) induced by H(2)O(2) and Ch-T after pretreatment with DTT were statistically higher than that induced by single administration of DTNB. Further investigation showed that the elevation of [Ca(2+)](i) mainly resulted from internal calcium stores. From our data, we propose that methionine oxidation plays an important role in the regulation of intracellular calcium and this regulation may mainly be due to internal calcium stores.

[Redox modulation of large conductance calcium-activated potassium channels in rat cultured trigeminal ganglion neurons].

AIM: To observe redox modulation of ion channel in trigeminal ganglion neurons by oxidants and reducing agents. METHODS: The effects of oxidants and reducing agents on maxi-conductance calcium-activated potassium channel in cultured rat trigeminal ganglion neurons by using whole-cell patch-clamp technique. RESULTS: Methionine-specific oxidant chloramine-T (Ch-T) 1 mmol/L slightly increased the current amplitude and this enhancement did not antagonized by DTT. In contrast, cysteine-specific reagent 5, 5'-dithio-bis(2-nitrobenzoic acid) (DTNB) 500 micromol/L significantly decreased current amplitude of BK(Ca) channels. The effect was reversed by the reducing agent 2 mmol/L 1, 4-dithio-DL-threitol (DTT). CONCLUSION: Reactive oxygen species were definitely involved in regulation of native neuronal function via redox modulation of BK(Ca) channels, which are suggested to play compensatory roles under oxidative stress-related conditions.