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Current clamp

Alessandro Tozzi, Valentina Durante, Guendalina Bastioli, Petra Mazzocchetti, Salvatore Novello, Alessandro Mechelli, Michele Morari, Cinzia Costa, Andrea Mancini, Massimiliano Di Filippo, Paolo Calabresi
Among genetic abnormalities identified in Parkinson's disease (PD), mutations of the leucine-rich repeat kinase2 (LRRK2) gene, such as the G2019S missense mutation linked to enhanced kinase activity, are the most common. While the complex role of LRRK2 has not been fully elucidated, evidence that mutated kinase activity affects synaptic transmission has been reported. Thus, our aim was to explore possible early alterations of neurotransmission produced by the G2019S LRRK2 mutation in PD. We performed electrophysiological patch-clamp recordings of striatal spiny projection neurons (SPNs) in the G2019S-Lrrk2 knock-in (KI) mouse model of PD, in D1994S kinase-dead (KD), Lrrk2 knock-out (KO) and wild-type (WT) mice...
June 13, 2018: Neurobiology of Disease
Hiroyuki Kawai, Nozomi Asaoka, Takahito Miyake, Kazuki Nagayasu, Takayuki Nakagawa, Hisashi Shirakawa, Shuji Kaneko
Neurotropin (NTP) is a Japanese analgesic agent for treating neuropathic pain; however, its method of action remains unclear. This study examined the effects of NTP on the activity of small dorsal root ganglion (DRG) neurons using whole-cell patch clamp recordings. After 3 days of treatment, NTP decreased current injection-induced firing activity of cultured DRG neurons by raising the current threshold for action potential generation. Additionally, NTP increased the sustained component of voltage-gated potassium (Kv ) channel currents without affecting other K+ currents...
May 31, 2018: Journal of Pharmacological Sciences
Tao Jin, Bingxue Hu, Shanshan Chen, Qiang Wang, Xue Dong, Yin Zhang, Yongqiang Zhu, Zhao Zhang
FHND004 is a newly synthesized epidermal growth factor receptor (EGFR) inhibitor for the treatment of non-small cell lung cancer (NSCLC). The aim of the present study was to investigate the impacts of FHND004 on human ether-à-go-go- related gene (hERG) K+ channels and the molecular mechanisms underlying of its action. Whole-cell patch clamp recording was performed on wild type (WT), mutant hERG channels heterologously expressed in human embryonic kidney (HEK) 293 cells or I Kr endogenously expressed in HL-1 cells, respectively...
2018: Frontiers in Pharmacology
Xin-Lu Yin, Hui-Qun Jie, Min Liang, Li-Na Gong, Han-Wei Liu, Hao-Lai Pan, Ya-Zhi Xing, Hai-Bo Shi, Chun-Yan Li, Lu-Yang Wang, Shan-Kai Yin
In developing sensory systems, elaborate morphological connectivity between peripheral cells and first-order central neurons emerges via genetic programming before the onset of sensory activities. However, how the first-order central neurons acquire the capacity to interface with peripheral cells remains elusive. By making patch-clamp recordings from mouse brainstem slices, we found that a subset of neurons in the cochlear nuclei, the first central station to receive peripheral acoustic impulses, exhibits spontaneous firings (SFs) as early as at birth, and the fraction of such neurons increases during the prehearing period...
2018: Frontiers in Molecular Neuroscience
Zhao Han, Yu-Yan Gu, Ning Cong, Rui Ma, Fang-Lu Chi
We aimed to investigate the beneficial effect of Celastrol on inner ear stem cells and potential therapeutic value for hearing loss. The inner ear stem cells were isolated and characterized from utricular sensory epithelium of adult mice. The stemness was evaluated by sphere formation assay. The relative expressions of Atoh1, MAP-2 and Myosin VI were measured by RT-PCR and immunoblotting. The up-regulation of MAP-2 was also analysed with immunofluorescence. The in vitro neuronal excitability was interrogated by calcium oscillation...
June 14, 2018: Organogenesis
Eric Salter, Julia Sunstrum, Sara Matovic, Wataru Inoue
KEY POINTS: Glutamatergic synaptic inputs to corticotropin releasing hormone (CRH) secreting neurons in the paraventricular nucleus of the hypothalamus (PVN) are required for stress-induced activation of the hypothalamic-pituitary-adrenal (HPA) axis. These synapses also undergo stress-induced plasticity thereby influencing HPA axis stress adaptation. By using patch clamp electrophysiology, we show that, in adult non-stressed mice, action potentials at these glutamatergic afferents elicit multiquantal transmission to the postsynaptic PVN-CRH neurons (i...
June 14, 2018: Journal of Physiology
Junxiu Liu, Dawei Liu, Jasmine J Liu, Chang Zhao, Shuzhong Yao, Liang Hong
Activity of the voltage-gated Nav1.5 sodium channel has been reported to be involved in cell proliferation, cancer invasion and gene expression. In addition, eicosapentaenoic acid (EPA) has recently been suggested to inhibit ovarian cancer cell growth and suppress tumor metastasis. The present study aimed to explore the association between EPA, the Nav1.5 sodium channel and ovarian cancer cells. Using patch-clamp technique and RNA interference approaches, sodium currents were recorded in epithelial ovarian cancer cells, and it was confirmed that the Nav1...
June 12, 2018: International Journal of Oncology
Guo-Sheng Xiao, Yan-Hui Zhang, Yan Wang, Hai-Ying Sun, Clive M Baumgarten, Gui-Rong Li
BACKGROUND AND PURPOSE: Adrenergic regulation of cell volume-regulated chloride current (ICl.vol ) is species-dependent. The present study investigates the mechanism underlying adrenergic regulation of ICl.vol in human atrial myocytes. EXPERIMENTAL APPROACH: A conventional whole-cell patch voltage-clamp technique was utilized to record membrane current in human atrial myocytes. ICl.vol was evoked by hyposmotic bath solution (0.6-times isosmotic, 0.6T). KEY RESULTS: ICl...
June 13, 2018: British Journal of Pharmacology
Ruxiu Liu, Jie Li, Yu Liu, Jie Peng, Xuanke Guan
Objective: We investigated the role of astragaloside in the treatment of sick sinus syndrome (SSS). Methods: Neonatal New Zealand rabbits were selected for the study. Rabbit sinoatrial node (SAN) cells were isolated by the method of dual enzymatic digestion and differential adherence. The injury model was prepared through simulated ischemia and reperfusion (I/R), and changes in the pacemaker current ( I f ) were recorded using the whole-cell patch-clamp technique. The proteins F-actin and vinculin were examined between various groups of SAN cells using a microplate reader and laser scanning confocal microscopy...
2018: Frontiers in Pharmacology
Hussam Hayani, Inseon Song, Alexander Dityatev
The neural extracellular matrix (ECM) is enriched with hyaluronic acid, chondroitin sulfate proteoglycans (CSPGs) and the glycoprotein tenascin-R, which play important roles in synaptic plasticity, as shown by studies of the CA1 region of the hippocampus. However, ECM molecules are strongly expressed in the CA2 region, which harbors a high number of fast-spiking interneurons (FSIs) surrounded by a particularly condensed form of ECM, perineuronal nets. Despite this intriguing peculiarity, the functional role of ECM in the CA2 region is mostly unknown...
2018: Frontiers in Cellular Neuroscience
Colline Sanchez, Christine Berthier, Bruno Allard, Jimmy Perrot, Clément Bouvard, Hidekazu Tsutsui, Yasushi Okamura, Vincent Jacquemond
Ion channel activity in the plasma membrane of living cells generates voltage changes that are critical for numerous biological functions. The membrane of the endoplasmic/sarcoplasmic reticulum (ER/SR) is also endowed with ion channels, but whether changes in its voltage occur during cellular activity has remained ambiguous. This issue is critical for cell functions that depend on a Ca2+ flux across the reticulum membrane. This is the case for contraction of striated muscle, which is triggered by opening of ryanodine receptor Ca2+ release channels in the SR membrane in response to depolarization of the transverse invaginations of the plasma membrane (the t-tubules)...
June 13, 2018: Journal of General Physiology
Daniel B Yaeger, Emma J Coddington
Neurons in the medullary reticular formation are involved in the control of postural and locomotor behaviors in all vertebrates. Reticulospinal neurons in this brain region provide one of the major descending projections to the spinal cord. Although neurons in the newt medullary reticular formation have been extensively studied using in vivo extracellular recordings, little is known of their intrinsic biophysical properties or of the underlying circuitry of this region. Using whole-cell patch-clamp recordings in brain slices containing the rostromedial reticular formation (rmRF) from adult male newts, we observed spontaneous miniature outward currents (SMOCs) in approximately two-thirds of neurons...
June 13, 2018: Journal of Neurophysiology
Min-Chul Shin, Kiku Nonaka, Toshitaka Yamaga, Masahito Wakita, Hironari Akaike, Norio Akaike
The current electrophysiological study investigated the functional roles of high- and low-voltage-activated calcium (Ca2+ ) channel subtypes on glutamatergic small mossy fiber nerve terminals (SMFTs) that synapse onto rat hippocampal CA3 neurons. Experiments combining both the 'synapse bouton' preparation and single-pulse focal stimulation technique were performed using the conventional whole-cell patch configuration under voltage-clamp conditions. Nifedipine, at a high concentration, and BAY-K8644 (BAY-K) inhibited and facilitated the glutamatergic excitatory postsynaptic currents (eEPSCs) that were evoked by 0...
June 13, 2018: Journal of Neurophysiology
Ming-Xiao Wang, Xiao-Tong Su, Peng Wu, Zhong-Xiuzi Gao, Wen-Hui Wang, Olivier Staub, Dao-Hong Lin
Kir4.1/5.1 heterotetramer participates in generating the negative cell membrane potential in distal convoluted tubule (DCT) and plays a critical role in determining the activity of Na-Cl co-transporter (NCC). Kir5.1 contains a phosphothreonine-motif at its c-terminus (AA249-252). Co-immunoprecipitation showed that Nedd4-2 was associated with Kir5.1 in HEK293 cells cotransfected with Kir5.1 or Kir4.1/Kir5.1. GST-pull-down further confirmed the association between Nedd4-2 and Kir5.1. Ubiquitination assay showed that Nedd4-2 increased the ubiquitination of Kir4...
June 13, 2018: American Journal of Physiology. Renal Physiology
Wei Wang, Yi Guo, Liang He, Chengzhi Chen, Jing Luo, Yuanlin Ma, Jie Li, Yong Yang, Qin Yang, Chao Du, Yanke Zhang, Zhonggui Li, Xin Xu, Xin Tian, Xuefeng Wang
miRNA-137 is an extremely abundant miRNA in the central nervous system and is thought to be closely related to synaptic plasticity. Here, we report a previously unrecognized role of miRNA-137 in epilepsy. The expression of miRNA-137 was decreased both in patients with temporal lobe epilepsy (TLE) and in two different mouse models of epilepsy. Overexpression of miRNA-137 induced by an intrahippocampal injection of a specific agomir prolonged the latency to spontaneous recurrent seizures (SRSs) and reduced seizure severity in a mouse model of pilocarpine-induced epilepsy...
June 9, 2018: Neuropharmacology
Jennifer M Egan, Colleen A Peterson, W Mark Fry
The sodium leak channel NALCN is poorly understood, but is reported as a Na+ -permeable, nonselective cation leak channel which regulates resting membrane potential and electrical excitability. Previous work has indicated that NALCN currents can be stimulated by activation of several G protein coupled receptors, including the M3 muscarinic receptor. We undertook a study using voltage clamp electrophysiology to investigate NALCN currents. We compared currents elicited from untransfected control HEK239 cells in response to M3R agonists muscarine or Oxotremorine M to currents elicited from cells transfected with M3R only or the M3R plus NALCN and cDNA encoding accessory proteins UNC-80 and Src...
June 2018: Biochimie Open
Chen-Wei Tsai, Ming-Feng Tsai
The mitochondrial calcium uniporter is a multisubunit Ca2+ channel that mediates mitochondrial Ca2+ uptake, a cellular process crucial for the regulation of oxidative phosphorylation, intracellular Ca2+ signaling, and apoptosis. In the last few years, genes encoding uniporter proteins have been identified, but a lack of efficient tools for electrophysiological recordings has hindered quantitative analysis required to determine functional mechanisms of this channel complex. Here, we redirected Ca2+ -conducting subunits (MCU and EMRE) of the human uniporter to the plasma membrane of Xenopus oocytes...
June 11, 2018: Journal of General Physiology
Guozhen Cui, Qiqi Xin, Hisa Hui Ling Tseng, Maggie PuiMan Hoi, Yan Wang, Binrui Yang, InLeng Choi, Yuqiang Wang, Rong Yuan, Keji Chen, Weihong Cong, Simon MingYuen Lee
We previously reported a novel danshensu derivative (R)-(3,5,6-Trimethylpyrazinyl) methyl-2-acetoxy-3-(3,4-diacetoxyphenyl) propanoate (ADTM) that exhibited promising cardiovascular protective activities, such as antioxidant and antiplatelet activities, as well as arterial relaxation. Particularly, ADTM treatment for 24 h exhibited anti-oxidative activity and effectively protected against acute myocardial infarction (MI) in a rat model. Here, we further investigated the pharmacological actions of 14 days of treatment with ADTM in alleviating and restoring the MI size by stimulating revascularization...
June 8, 2018: Pharmacological Research: the Official Journal of the Italian Pharmacological Society
Arsen S Hunanyan, Ashley R Helseth, Elie Abdelnour, Bassil Kherallah, Monisha Sachdev, Leeyup Chung, Melanie Masoud, Jordan Richardson, Qiang Li, J Victor Nadler, Scott D Moore, Mohamad A Mikati
OBJECTIVE: Na+ /K+ -ATPase dysfunction, primary (mutation) or secondary (energy crisis, neurodegenerative disease) increases neuronal excitability in the brain. To evaluate the mechanisms underlying such increased excitability we studied mice carrying the D801N mutation, the most common mutation causing human disease, specifically alternating hemiplegia of childhood (AHC) including epilepsy. Because the gene is expressed in all neurons, particularly γ-aminobutyric acid (GABA)ergic interneurons, we hypothesized that the pathophysiology would involve both pyramidal cells and interneurons and that fast-spiking interneurons, which have increased firing rates, would be most vulnerable...
June 11, 2018: Epilepsia
Sohyeon Moon, Hee Jung Baik
We investigated the effects of flumazenil, aminophylline, and ephedrine on the excitatory amino acid transporter type 3 (EAAT3) activity and the interaction with propofol. EAAT3 was expressed in the Xenopus oocytes. L-Glutamate-induced membrane currents were measured using the two-electrode voltage clamp at various drug concentrations. Oocytes were preincubated with protein kinase C- (PKC-) activator, or inhibitor, and phosphatidylinositol 3-kinase (PI3K) inhibitor. To study the interaction with propofol, oocytes were exposed to propofol, propofol + aminophylline, or ephedrine...
2018: BioMed Research International
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