Abstract:
:Small interfering RNA (siRNA) directs the targeted destruction of mRNA encoding a specific protein, in a process known as RNA interference (RNAi). This stops translation of the targeted mRNA into protein, effectively silencing the gene. RNAi is a recent discovery, identified in mammalian cells in 2001, but it has rapidly advanced into a practical technique and is being used increasingly to investigate mammalian gene function. Tools are available to induce RNAi in cell lines, intact tissue preparations and even in vivo. Depending on the method used, loss of gene expression may be transient or sustained, enabling a wide range of functions to be investigated. RNAi therefore offers a powerful technique that can be used to produce targeted knockout of ion channel genes in mammalian cells. Its applications potentially include identification of ion channel function in health and disease, identification of novel channel genes and drug target validation. This paper outlines our current understanding of siRNA and the experimental requirements for producing efficient RNAi and gene silencing. Effective RNAi requires an appropriate siRNA sequence to be designed and an efficient method for delivering the siRNA to the cells of interest. Since not all potential siRNA sequences are effective, it is also important to verify the loss of gene expression by measuring the level of channel protein remaining. Limitations of the methods available for delivering siRNA are one of the main obstacles to producing efficient RNAi, especially in intact tissue preparations. Here we describe an in vitro method for targeted RNAi against the TASK-1 potassium channel gene in an isolated vascular preparation, using a DNA construct to direct the expression of siRNA, along with a non-viral method for transfecting cells within the vessel. Successful silencing of the TASK-1 gene is verified by immunostaining with an antibody directed against the TASK-1 protein.
journal_name
J Pharmacol Toxicol Methodsjournal_title
Journal of pharmacological and toxicological methodsauthors
Gurney AM,Hunter Edoi
10.1016/j.vascn.2004.08.012keywords:
subject
Has Abstractpub_date
2005-05-01 00:00:00pages
253-62issue
3eissn
1056-8719issn
1873-488Xpii
S1056-8719(05)00010-9journal_volume
51pub_type
杂志文章abstract:INTRODUCTION:Telemetry beagle dogs are the most frequently used species in cardiovascular telemetry assessments. However, beagle dogs may not be always suitable for all of the tests. Recently minipigs have received increased attention for these studies. Differences between the two species regarding the response of thei...
journal_title:Journal of pharmacological and toxicological methods
pub_type: 杂志文章
doi:10.1016/j.vascn.2014.03.172
更新日期:2014-07-01 00:00:00
abstract:INTRODUCTION:MEKK1 is a 196-kDa serine-threonine kinase activated in response to a variety of stimuli, including EGF, lysophosphatidic acid, osmotic stress, UV light, and microtubule toxins. However, there are few reports about the expression level of MEKK1 in cancers. METHODS:In this report, a direct competitive ELIS...
journal_title:Journal of pharmacological and toxicological methods
pub_type: 杂志文章
doi:10.1016/j.vascn.2004.08.004
更新日期:2005-03-01 00:00:00
abstract:INTRODUCTION:The combined evaluation of physiology and behaviour allows a complete and more comprehensive pre-clinical assessment of central nervous system (CNS) function. An integrated video-telemetric electroencephalography (Video-tEEG) system, which allows the simultaneous and continuous recording of EEG and video i...
journal_title:Journal of pharmacological and toxicological methods
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doi:10.1016/j.vascn.2010.04.004
更新日期:2010-07-01 00:00:00
abstract:INTRODUCTION:Recently we have demonstrated that to the choice of tissue type is important in identifying I(Kr) and I(Ks)-induced prolongation of the action potential. However, the differential sensitivity of cardiac tissues to other ionic current blockers or modulators is relatively unknown. The aim of the present stud...
journal_title:Journal of pharmacological and toxicological methods
pub_type: 杂志文章
doi:10.1016/j.vascn.2007.06.005
更新日期:2008-01-01 00:00:00
abstract:INTRODUCTION:The quantification of [(3)H]noradrenaline spillover from electrically stimulated, sympathetic nerves is a widely used method to study presynaptic effects of hormones, transmitters and drugs. Although a straightforward approach, the execution of the experiments is not trivial. This holds true mainly for a r...
journal_title:Journal of pharmacological and toxicological methods
pub_type: 杂志文章
doi:10.1016/s1056-8719(02)00170-3
更新日期:2001-11-01 00:00:00
abstract::The use of animal models is an essential part of medical research and drug development. The essential skills required to be able to do such research includes experimental design, statistical analysis and the actual handling and treating of the animals (in vivo skills). The number of students in the U.K. receiving trai...
journal_title:Journal of pharmacological and toxicological methods
pub_type: 杂志文章
doi:10.1016/j.vascn.2011.07.002
更新日期:2011-09-01 00:00:00
abstract:INTRODUCTION:In the present study, a model was developed to determine the effect of secretase inhibition on beta-amyloid peptide (Abeta) levels in the cerebrospinal fluid (CSF) of freely moving adult rats. METHODS:Rats were chronically implanted with a cannula into the cisterna magna and CSF samples were collected at ...
journal_title:Journal of pharmacological and toxicological methods
pub_type: 杂志文章
doi:10.1016/j.vascn.2005.02.002
更新日期:2005-09-01 00:00:00
abstract:INTRODUCTION:cDNA microarray technology has greatly facilitated mechanistic studies in pharmacology and toxicology. A clean hybridization with minimal background is critical for successful microarray analysis and is highly desired. However, clean hybridization alone is not enough; verification is needed. METHODS:Total...
journal_title:Journal of pharmacological and toxicological methods
pub_type: 杂志文章
doi:10.1016/j.vascn.2004.02.001
更新日期:2004-07-01 00:00:00
abstract::Toxicogenomics, the application of genomic data to elucidate or predict an organism's response to a toxicant, can inform the drug development process in important ways. It is apparent that standardized approaches to many types of toxicogenomic questions are still being formulated. Specifically, a significant body of p...
journal_title:Journal of pharmacological and toxicological methods
pub_type: 杂志文章,评审
doi:10.1016/j.vascn.2005.05.006
更新日期:2006-01-01 00:00:00
abstract:INTRODUCTION:Multiple cardiac ion channels are prone to block by pharmaceutical compounds, and this can have large implications for cardiac safety. The effect of a compound on individual ion currents can now be measured in automated patch clamp screening assays. In-silico action potential models are proposed as one way...
journal_title:Journal of pharmacological and toxicological methods
pub_type: 杂志文章
doi:10.1016/j.vascn.2015.05.002
更新日期:2015-09-01 00:00:00
abstract:INTRODUCTION:Quantification of lactate dehydrogenase (LDH) release is a widely accepted assay for the quantitative determination of cell viability and late-stage apoptosis. Major disadvantages of commercially available LDH assay kits include proprietary formulations, limited options for optimization and high cost, all ...
journal_title:Journal of pharmacological and toxicological methods
pub_type: 杂志文章
doi:10.1016/j.vascn.2015.02.001
更新日期:2015-05-01 00:00:00
abstract::Hemodynamic and biochemical changes were studied on 36 white ELCO-rabbits, seven adult older than 150 days, seven immatures between 21 and 27 days, and seven neonatals between 7 and 14 days. Five supplementary hearts of each age group served for preischemic biochemical values. Protection during 60 min of global ischem...
journal_title:Journal of pharmacological and toxicological methods
pub_type: 杂志文章
doi:10.1016/1056-8719(93)90016-8
更新日期:1993-12-01 00:00:00
abstract:INTRODUCTION:Parts A and B of the ICH S7 guidelines on safety pharmacology describe the in vivo studies that must be conducted prior to first time in man administration of any new pharmaceutical. ICH S7A requires a consideration of the sensitivity and reproducibility of the test systems used. This could encompass maint...
journal_title:Journal of pharmacological and toxicological methods
pub_type: 杂志文章,多中心研究
doi:10.1016/j.vascn.2013.04.008
更新日期:2013-07-01 00:00:00
abstract::Electroencephalogram (EEG) data in nonclinical species can play a critical role in the successful evaluation of a compound during drug development, particularly in the evaluation of seizure potential and for monitoring changes in sleep. Yet, while non-invasive electrocardiogram (ECG) monitoring is commonly included in...
journal_title:Journal of pharmacological and toxicological methods
pub_type: 杂志文章,评审
doi:10.1016/j.vascn.2016.03.002
更新日期:2016-09-01 00:00:00
abstract::We have developed a method for the rapid isolation of hepatocyte nuclei, which employs gentle homogenization and centrifugation conditions, and involves minimal processing time. The purified nuclei were morphologically unaltered when observed by light and electron microscopy. No significant contamination from cytoplas...
journal_title:Journal of pharmacological and toxicological methods
pub_type: 杂志文章
doi:10.1016/s1056-8719(97)00082-8
更新日期:1997-11-01 00:00:00
abstract:INTRODUCTION:In order to investigate the pharmacokinetics of water-borne chemicals while eliminating exposures by other routes, a dermal exposure system was developed to expose the hand and forearm of human subjects. METHODS:The goal was, primarily, to study the dermal pharmacokinetics of methyl tertiary butyl ether (...
journal_title:Journal of pharmacological and toxicological methods
pub_type: 杂志文章
doi:10.1016/S1056-8719(03)00004-2
更新日期:2002-05-01 00:00:00
abstract:INTRODUCTION:Reversible protein cysteine oxidation is recognised as a pivotal post-transitional modification that transduces physiological as well as pathological signalling. Pharmacological interventions that target specific sources of oxidant formation are currently being trialled to ascertain their potential ability...
journal_title:Journal of pharmacological and toxicological methods
pub_type: 杂志文章
doi:10.1016/j.vascn.2013.07.001
更新日期:2013-11-01 00:00:00
abstract::This editorial prefaces the annual themed issue on safety pharmacology (SP) methods published in the Journal of Pharmacological and Toxicological Methods (JPTM). We highlight here the content derived from the recent 2016 Safety Pharmacology Society (SPS), Canadian Society of Pharmacology and Therapeutics (CSPT), and J...
journal_title:Journal of pharmacological and toxicological methods
pub_type: 社论
doi:10.1016/j.vascn.2017.04.012
更新日期:2017-09-01 00:00:00
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journal_title:Journal of pharmacological and toxicological methods
pub_type: 杂志文章
doi:10.1016/j.vascn.2013.04.010
更新日期:2013-07-01 00:00:00
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journal_title:Journal of pharmacological and toxicological methods
pub_type: 杂志文章
doi:10.1016/j.vascn.2015.04.006
更新日期:2015-07-01 00:00:00
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journal_title:Journal of pharmacological and toxicological methods
pub_type: 杂志文章,评审
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更新日期:2017-01-01 00:00:00
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journal_title:Journal of pharmacological and toxicological methods
pub_type: 杂志文章
doi:10.1016/j.vascn.2018.02.007
更新日期:2018-01-01 00:00:00
abstract::The acetylcholine-evoked relaxation of rabbit isolated thoracic aorta precontracted by phenylephrine was studied. Phenylephrine caused a steady contraction that was maintained for 6 h. In the presence of calcium disodium ethylenediaminetetraacetate (EDTA) and ascorbic acid the contraction decreased with time. N(G)-Nit...
journal_title:Journal of pharmacological and toxicological methods
pub_type: 杂志文章
doi:10.1016/s1056-8719(99)00035-0
更新日期:1999-08-01 00:00:00
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journal_title:Journal of pharmacological and toxicological methods
pub_type: 杂志文章
doi:10.1016/j.vascn.2007.03.002
更新日期:2007-07-01 00:00:00
abstract:INTRODUCTION:Gastrointestinal (GI) motility disorders include many clinical manifestations associated with various pathologies. They are widespread and can be considered a primary symptom or can be associated to other diseases, such as Parkinson's disease. Understanding the type and site of GI dysmotility is crucial to...
journal_title:Journal of pharmacological and toxicological methods
pub_type: 杂志文章
doi:10.1016/j.vascn.2014.08.003
更新日期:2014-09-01 00:00:00
abstract::In many circumstances, rapid information is required about the effects of drugs on neurotransmitter release in brain, and a common method used is measurement of radiolabelled release from superfused brain slices or synaptosomes in vitro. However, the method requires expensive equipment and is not readily adapted to th...
journal_title:Journal of pharmacological and toxicological methods
pub_type: 杂志文章
doi:10.1016/s1056-8719(97)00010-5
更新日期:1997-04-01 00:00:00
abstract::Undesired immune responses against protein therapeutics may adversely affect the pharmacokinetics, efficacy, and safety of the product. The presence of anti-drug-antibodies (ADA) has been the key determinant of immunogenic responses. Here we describe the use of a capillary electrophoresis platform for the identificati...
journal_title:Journal of pharmacological and toxicological methods
pub_type: 杂志文章
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更新日期:2020-01-01 00:00:00
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journal_title:Journal of pharmacological and toxicological methods
pub_type: 杂志文章
doi:10.1016/s1056-8719(02)00219-8
更新日期:2002-03-01 00:00:00
abstract::Evaluation of the myocardial relaxation has become important in the last years. An impaired relaxation may precede contractile dysfunctions and even cause heart failure. To treat this impaired lusitropism it is necessary to properly assess the lusitropic state of the heart and understand how drugs affect the cellular ...
journal_title:Journal of pharmacological and toxicological methods
pub_type: 杂志文章,评审
doi:10.1016/1056-8719(94)90011-6
更新日期:1994-09-01 00:00:00
abstract::Evaluation of angiotensin II (AII) receptor binding often necessitates freezing of the tissue of interest prior to assay of radioligand binding. This study evaluated the effects of freezing of various rat tissues at different rates on 125I-sarcosine1, isoleucine8 angiotensin II (125I-SI AII) binding to AII receptor su...
journal_title:Journal of pharmacological and toxicological methods
pub_type: 杂志文章
doi:10.1016/1056-8719(94)00061-8
更新日期:1995-04-01 00:00:00