Potentiation of acid-sensing ion channel activity by the activation of 5-HT₂ receptors in rat dorsal root ganglion neurons

Acid-sensing ion channels (ASICs), as key sensors for extracellular protons, are expressed in nociceptive sensory neurons and contribute to signalling pain caused by tissue acidosis. ASICs are also the subject of various factors. Here, we further provide evidence that the activity of ASICs is potentiated by the activation of 5-HT₂ receptors in rat dorsal root ganglion neurons. A specific 5-HT₂ receptor agonist, α-methyl-5-HT, dose-dependently enhanced proton-gated currents with an EC₅₀ of 0.13 ± 0.07 nM. The α-methyl-5-HT enhancing effect on proton-gated currents was blocked by cyproheptadine, a 5-HT₂ receptor antagonist, and removed by intracellular dialysis of either GDP-β-S or protein kinase C inhibitor GF109203X. Moreover, α-methyl-5-HT altered acid-evoked membrane excitability of rat DRG neurons and caused a significant increase in the amplitude of the depolarization and the number of spikes induced by acid stimuli. Finally, α-methyl-5-HT increased nociceptive responses to injection of acetic acid in rats. These results suggest that α-methyl-5-HT up-regulates the activity of ASICs via 5-HT₂ receptor and protein kinase C dependent signal pathways in rat primary sensory neurons and this potentiation contributed to acid- mediated pain in tissue injury and inflammation.     

Comparison of CGRP release from rat lymphocytes and dorsal root ganglia neurons

Calcitonin gene-related peptide (CGRP), a neuropeptide contained in sensory neuron, has been demonstrated to be synthesized and released by rat lyrnphocytes in our previous studies.In this study, some release property and molecular     

Physostigmine blocked nicotinic acetylcholine receptors in rat sympathetic ganglion neurons

目的：研究毒扁豆碱阻断交感节神经元烟碱受体的作用机理．方法：以培养的新生大鼠颈上交感节神经元为标本，使用全细胞膜片箝技术，观察毒扁豆碱对交感节烟碱受体选择性激动剂ＤＭＰＰ诱发电流的影响．结果：毒扁豆碱（５－２０μｍｏｌ·Ｌ－１）以浓度依赖性方式抑制ＤＭＰＰ诱发电流，促进诱发电流的衰减，其抑制作用没有电压依赖性，毒扁豆碱２００μｍｏｌ·Ｌ－１不能激活烟碱受体．结论：与骨骼肌烟碱受体相比，交感神经元烟碱受体表现出不同的药理学特性．毒扁豆碱通过作用于变构位点抑制交感神经元烟碱受体，不影响其开放的离子通道和激动剂结合位点．     

Adrenoceptor agonists inhibit calcium-dependent potentials in rat stelate ganglion neurons

探索肾上腺素能受体激动剂对大鼠星状神经节细胞的作用及其作用机制．方法：离体交感神经节细胞内生物电记录．结果：去甲肾上腺素，可乐定（１０－３０μｍｏｌ·Ｌ－１）可逆地抑制细胞动作电位；依钙后超极化电位；在含ＴＴＸ与ＴＥＡ克氏液中记录到的钙锋电位及快兴奋性突触后电位．结论：肾上腺素受体激动剂对三种钙电位及快兴奋性突触后电位的抑制作用可能是其在突触前膜对钙内流的抑制所致．     

Quercetin protects rat dorsal root ganglion neurons against high glucose-induced injury in vitro through Nrf-2/HO-1 activation and NF-κB inhibition

Aim:

To examine the effects of quercetin, a natural antioxidant, on high glucose (HG)-induced apoptosis of cultured dorsal root ganglion (DRG) neurons of rats.

Methods:

DRG neurons exposed to HG (45 mmol/L) for 24 h were employed as an in vitro model of diabetic neuropathy. Cell viability, reactive oxygen species (ROS) level and apoptosis were determined. The expression of NF-кB, IкBα, phosphorylated IкBα and Nrf2 was examined using RT PCR and Western blot assay. The expression of hemeoxygenase-1 (HO-1), IL-6, TNF-α, iNOS, COX-2, and caspase-3 were also examined.

Results:

HG treatment markedly increased DRG neuron apoptosis via increasing intracellular ROS level and activating the NF-κB signaling pathway. Co-treatment with quercetin (2.5, 5, and 10 mmol/L) dose-dependently decreased HG-induced caspase-3 activation and apoptosis. Quercetin could directly scavenge ROS and significantly increased the expression of Nrf-2 and HO-1 in DRG neurons. Quercetin also dose-dependently inhibited the NF-κB signaling pathway and suppressed the expression of iNOS, COX-2, and proinflammatory cytokines IL-6 and TNF-α.

Conclusion:

Quercetin protects rat DRG neurons against HG-induced injury in vitro through Nrf-2/HO-1 activation and NF-κB inhibition, thus may be beneficial for the treatment of diabetic neuropathy.     

Dual effects of pentobarbital on rat sacral dorsal commissural neurons in vitro

Ｄｕａｌｅｆｅｃｔｓｏｆｐｅｎｔｏｂａｒｂｉｔａｌｏｎｒａｔｓａｃｒａｌｄｏｒｓａｌｃｏｍｍｉｓｓｕｒａｌｎｅｕｒｏｎｓｉｎｖｉｔｒｏ１ＰＡＮＧＺｈｉＰｉｎｇ，ＸＵＴｉａｎＬｅ２，ＨＵＧｕｏＹｕａｎ３，ＬＩＪｉＳｈｕｏ（Ｄｅｐａｒｔｍｅｎｔｏ...     

Downregulation of Kυ4.2 and Kυ4.3 channel gene expression in right ventricular hypertrophy induced by monocrotaline in rat

INTRODUCTION The calcium-independent transient outward potas-sium current (Ito) is activated by depolarization and playsa key role in controlling the amplitudes and cardiac ac-tion potential duration (APD). Ito contributesto the phase1 repolarization of action potential in myocytes, theprolongation of APD is the result of decrease in Ito den-sity[1]. Ito which is encoded by genes of Kv1.4, Kv4.2,and Kv4.3[2], based on differences in kinetics, as wellas recovery from inactivation…     

Alpha-cobratoxin inhibits T-type calcium currents through muscarinic M4 receptor and Gο-protein βγ subunits-dependent protein kinase A pathway in dorsal root ganglion neurons

The long-chain neurotoxic protein, alpha-cobratoxin (α-CTx), has been shown to have analgesic effects. However, the underlying mechanisms still remain unclear. In this study, we examined the effects of α-CTx on T-type calcium channel currents (T-currents) and elucidated the relevant mechanisms in mouse dorsal root ganglion (DRG) neurons. Our results showed that α-CTx reversibly inhibited T-currents in a dose-dependent manner. This inhibitory effect was blocked by the selective muscarinic M4 receptor antagonist tropicamide, while methyllycaconitine, a specific antagonist for the α7 subtype of nicotinic receptor had no effect. siRNA targeting the M4 receptor in small DRG neurons abolished α-CTx-induced T-current inhibition. Intracellular application of GDP-β-S or a selective antibody against the G(o)α-protein, as well as pretreatment of the cells with pertussis toxin, abolished the inhibitory effects of α-CTx. The M4 receptor-mediated response was blocked by dialyzing cells with QEHA peptide or anti-G(β) antibody. Pretreatment of the cells with protein kinase A (PKA) inhibitor H89 or intracellular application of PKI 6-22 abolished α-CTx-induced T-current inhibition in small DRG neurons, whereas inhibition of phosphatidylinositol 3-kinase or PKC elicited no such effects. In addition, α-CTx significantly increased PKA activity in DRG neurons, whereas pretreatment of the cells with tropicamide abolished this effect. In summary, our results suggest that activation of muscarinic M4 receptor by α-CTx inhibits T-currents via the G(βγ) of G(o)-protein and PKA-dependent pathway. This article is part of a Special Issue entitled 'Post-Traumatic Stress Disorder'.     

Effects of complete Freund＇s adjuvant on immunohistochemical distribution of IL-1β and IL-1R I in neurons and glia cells of dorsal root ganglion

AIM: To investigate the effects of complete Freund adjuvant (CFA) on inflammatory hyperalgesia and morphological change of the coexistence of interleukin-1 beta (IL-1beta) and type I IL-1 receptor (IL-1RI) in neurons and glia cells of rat dorsal root ganglion (DRG). METHODS: The pain-related parameters and the expression of IL-1RI and IL-1beta positive neurons and glia cells of DRG in normal saline (NS) and adjuvant-induced arthritic (AA) group were examined with pain behavior assessment methods and immunohistochemical assay, respectively. RESULTS: Five hours, 1 d, and 2 d after intra-articular injection of 50 microL CFA, tactile hyperalgesia induced by CFA was observed in the foot flexion and extension scores of the ipsilateral hindpaw of AA group. Three days after injection, the distribution of IL-1RI/IL-1beta double-stained coexisted neurons and glia cells were observed in ipsilateral DRG of both groups. The number of IL-1beta positive neurons, IL-1RI positive neurons, IL-1beta/IL-1RI double-stained neurons, and IL-1RI positive glia cells in ipsilateral DRG of the AA group were higher than that of NS group (P<0.05 or P<0.01). CONCLUSION: The coexistence of IL-1beta and IL-1RI in neurons and nonneuronal cells suggests an as yet unknown autocrine and/or paracrine function of IL-1beta in the DRG. The function was enhanced in articular arthritis induced by CFA and could play an important role in hyperalgesia under inflammatory conditions.     

SDF-1α/CXCL12 enhances GABA and glutamate synaptic activity at serotonin neurons in the rat dorsal raphe nucleus

The serotonin (5-hydroxytryptamine; 5-HT) system has a well-characterized role in depression. Recent reports describe comorbidities of mood-immune disorders, suggesting an immunological component may contribute to the pathogenesis of depression as well. Chemokines, immune proteins which mediate leukocyte trafficking, and their receptors are widely distributed in the brain, mediate neuronal patterning, and modulate various neuropathologies. The purpose of this study was to investigate the neuroanatomical relationship and functional impact of the chemokine stromal cell-derived factor-1α/CXCL12 and its receptor, CXCR4, on the serotonin dorsal raphe nucleus (DRN) system in the rat using anatomical and electrophysiological techniques. Immunohistochemical analysis indicates that over 70% of 5-HT neurons colocalize with CXCL12 and CXCR4. At a subcellular level, CXCL12 localizes throughout the cytoplasm whereas CXCR4 concentrates to the outer membrane and processes of 5-HT neurons. CXCL12 and CXCR4 also colocalize on individual DRN cells. Furthermore, electrophysiological studies demonstrate CXCL12 depolarization of 5-HT neurons indirectly via glutamate synaptic inputs. CXCL12 also enhances the frequency of spontaneous inhibitory and excitatory postsynaptic currents (sIPSC and sEPSC). CXCL12 concentration-dependently increases evoked IPSC amplitude and decreases evoked IPSC paired-pulse ratio selectively in 5-HT neurons, effects blocked by the CXCR4 antagonist AMD3100. These data indicate presynaptic enhancement of GABA and glutamate release at 5-HT DRN neurons by CXCL12. Immunohistochemical analysis further shows CXCR4 localization to DRN GABA neurons, providing an anatomical basis for CXCL12 effects on GABA release. Thus, CXCL12 indirectly modulates 5-HT neurotransmission via GABA and glutamate synaptic afferents. Future therapies targeting CXCL12 and other chemokines may treat serotonin related mood disorders, particularly depression experienced by immune-compromised individuals.     

Activation of Cl− channels by avermectin in rat cultured hippocampal neurons

Summary The actions of the insecticide avermectin (AVM) were studied in rat cultured hippocampal neurons with patch-clamp techniques. Application of micromolar concentrations of AVM to voltage-clamped cells gave rise to whole-cell currents, which showed a slow time-course of activation in the order of 10 s, and wash-out periods of typically 20 min. Dose-response curves revealed a half-maximally activating AVM concentration (EC₅₀) of 2.0±0.6 M and a Hill coefficent of 1.5±0.9. The current activated by AVM was carried predominantly by Cl^– ions, as demonstrated by ion-substitution experiments. The Cl^– channel blocker picrotoxinin (100 M) substantially but transiently reduced the AVM response. Outsideout patch recording showed that AVM opened Cl^– channels with a conductance of 40±12 pS. The open-time distribution was characterized by two time constants of 11 ms and 259 ms. It is suggested that AVM directly activates Cl^– channels in mammalian central neurons, which resemble the channels activated by the physiological transmitters GABA and glycine.Correspondence to: J. Bormann at the above address     

The acute and long‐term L‐DOPA effects are independent from changes in the activity of dorsal raphe serotonergic neurons in 6‐OHDA lesioned rats

Background and Purpose

L‐DOPA is still the most efficacious pharmacological treatment for Parkinson''s disease. However, in the majority of patients receiving long‐term therapy with L‐DOPA, its efficacy is compromised by motor complications, notably L‐DOPA‐induced dyskinesia. Evidence suggests that the serotonergic system is involved in the therapeutic and the side effects of L‐DOPA. Here, we investigate if long‐term L‐DOPA treatment alters the activity of the dorsal raphe nucleus (DRN) and its responses to serotonergic drugs.

Experimental Approach

We measured the responses of serotonergic neurons to acute and chronic L‐DOPA treatment using in vivo electrophysiological single unit‐extracellular recordings in the 6‐OHDA‐lesion rat model of Parkinson''s disease.

Key Results

The results showed that neither acute nor chronic L‐DOPA administration (6 mg·kg⁻¹ s.c.) altered the properties of serotonergic‐like neurons. Furthermore, no correlation was found between the activity of these neurons and the magnitude of L‐DOPA‐induced dyskinesia. In dyskinetic rats, the inhibitory response induced by the 5‐HT_1A receptor agonist 8‐OH‐DPAT (0.0625–16 μg·kg⁻¹, i.v.) was preserved. Nonetheless, L‐DOPA impaired the ability of the serotonin reuptake inhibitor fluoxetine (0.125–8 mg·kg⁻¹, i.v) to inhibit DRN neuron firing rate in dyskinetic animals.

Conclusions and Implications

Although serotonergic neurons are involved in the dopaminergic effects of L‐DOPA, we provide evidence that the effect of L‐DOPA is not related to changes of the activity of DRN neurons. Rather, L‐DOPA might reduce the efficacy of drugs that normally enhance the extracellular levels of serotonin.

Linked Articles

This article is part of a themed section on Updating Neuropathology and Neuropharmacology of Monoaminergic Systems. To view the other articles in this section visit http://onlinelibrary.wiley.com/doi/10.1111/bph.v173.13/issuetoc

Abbreviations

8‐OH‐DPAT

8‐hydroxy‐2‐(dipropylamino)tetralin

AIM

abnormal involuntary movements

DRN

dorsal raphe nucleus

L‐DOPA

L‐3,4‐dihydroxyphenylalanine

LID

L‐DOPA‐induced dyskinesia

6‐OHDA

6‐hydroxydopamine

PD

Parkinson''s disease

SSRI

selective serotonin reuptake inhibitor

     

Regulating effect of carbonic anhydrase (CA) inhibitor on aquaporin-1 (AQP1) water channel gene expression in rat kidney

AIM: To investigate whether the aquaporin-1 (AQR-1) geneexpression can be regulated by acetazolamide, a carboincanhydrase inhibitor in rat kidney. Carbonic anhydrase inhibitor isone of the diuretics with the site of action on renal proximaltubules. And AQP-1 wate channel is located on the epithelialcells of the renal proximal tubules with the function of facilitatingwater transport across proximal tubules epithelia. METHODS:The kidneys were excised from immature (20 d) female rat at     

Anti—inflammatory effects of aqueous extract from Dichroa febrifuga root in rat liver

AIM:To study the anti-inflammatory effects of aqueous extract from Dichroa febrifuga root (AEDF)for suppres-sion in the process of lipopolysaccharide(LPS)-induced sepsis in the rat liver.METHODS:The inhibitory effect of AEDF on the alteration of inflammatory proteins was investigated by Westen blot and immunohistochemical analysis.RESULTS:Western blot analysis showed that the level of nuclar factor(NF)-kBp65 was markedly up-regulated and (I)-kBα was down-regulatedf by LPS (8mg/kg) challenge,However,AEDF 100mg/kg inhibited induction of NF-kBp65 and degradation of I-kBα in the liver of LPS-challenged rats. Immunohistochemical analysis showed that while the expression of the NF-kBp65,tumor necrosis factor (TNF-α）,and inducible nitric oxide synthase (iNOS) tende to increase ,that of I-kBα was decreased in the hepatocytes of rats chaallenged with LPS,A slight decline of NF-kBp65,TNF-α,and iNOS,but an increase of I-kBα were observed in the hepatocytes of the rats pretreated with AEDF.CONCLUSION:AED may act as a therapeutic agent for inflammatory disease through a regulation of inflammation-related proteins.     

Role of delayed rectifier K channel currents in β-amyloid （1-40）-induced caspase-3 activation and apoptosis in primary cultured cortical neurons

AIM: To investigate the effect of chronic exposure to/]-amyloid（1-40)(Aβ1-40) on delayed rectifier K^ currents (IK) in primary cultured ratcortical neurons and the effect of IK on caspase-3 activation and neurons apoptosis induced by Aβ1-40. METHODS: K^ currents were recorded using whole-cell patch clamp techniques; cell sviability rate and apoptosis were studied using MTT and Hoechst     

Kinetic properties of nicotinic receptors in cultured rat sympathetic neurons from superior cervical ganglia

目的：研究培养的新生大鼠颈上神经节交感神经元烟碱受体的动力学特性．方法：膜片箝技术的全细胞记录方法，记录不同浓度烟碱诱发的电流，使用Ｃｌａｒｋ方程对烟碱作用的量效曲线进行拟合．结果：１０，２０，４０，８０和１６０μｍｏｌ·Ｌ－１烟碱诱发电流的幅度分别为：０９１±００８，１５６±０１４，２５３±０２７，３９３±０４６和４５７±０５５ｎＡ（ｎ＝１５），经Ｃｌａｒｋ方程拟合，得到Ｈ＝１０９７，Ｅｍａｘ＝５９５８ｎＡ，Ｋ＝７３０６１μｍｏｌ·Ｌ－１，将Ｈ＝１时拟合得到的Ｅｍａｘ（６５１３ｎＡ）和Ｋ值（６１４５７μｍｏｌ·Ｌ－１）代入Ｃｌａｒｋ方程，所计算出的理论值与相应浓度烟碱诱发电流的实测值基本相符．结论：烟碱与交感神经元烟碱受体作用的动力学特性符合一个作用位点的反应模型．