Abstract：
:We report a simple fluorometric method for the continuous monitoring of mitochondrial membrane potential and cell viability in suspensions of hepatocytes exposed in vitro to cytotoxic agents. Suspensions of freshly isolated hepatocytes (10(6) cells/mL) preloaded with rhodamine 123 (Rh 123, 100 mumol/L) are transferred to a thermostatically controlled mixed cuvette to which the desired cytotoxic agent is added. Rh 123 is a cationic fluorophore that is actively accumulated by cells in direct proportion to the mitochondrial membrane potential. Cell viability was estimated by monitoring propidium iodide (PI) fluorescence. Exposure of cell suspensions to the mitochondrial uncoupling agent FCCP caused an immediate and titratable increase in Rh 123 fluorescence. Subsequent treatment with digitonin did not change Rh 123 fluorescence, suggeseting that Rh 123 equilibrates rapidly across the intact cell membrane. Likewise, treatment of hepatocyte suspensions with inhibitors of mitochondrial respiration (rotenone, cyanide, or menadione) caused an immediate increase in Rh 123 fluorescence. This was accompanied by a progressive increase in PI fluorescence, suggesting a causal relationship between mitochondrial depolarization and cell injury. In contrast, 1,4-benzoquinone caused a time-dependent and linear increase in PI fluorescence that paralleled changes in Rh 123 fluorescence. Comparing the time courses for changes in PI and Rh 123 fluorescence suggests that for benzoquinone, the depolarization of the mitochondria is a consequence rather than a cause of the cell injury. This modified procedure provides a simple and specific technique for continuously monitoring mitochondrial membrane potential and cell viability in suspensions of freshly isolated hepatocytes. The advantage is that there is no need to separate cells from the incubation medium, making it possible to record real-time changes in mitochondrial membrane potential and cell viability throughout the in vitro exposure period.
journal_name
J Pharmacol Toxicol Methodsjournal_title
Journal of pharmacological and toxicological methodsauthors
Palmeira CM,Moreno AJ,Madeira VM,Wallace KBdoi
10.1016/1056-8719(95)00131-xsubject
Has Abstractpub_date
1996-02-01 00:00:00pages
35-43issue
1eissn
1056-8719issn
1873-488Xpii
1056-8719(95)00131-Xjournal_volume
35pub_type
杂志文章abstract：INTRODUCTION:Detection of drug effects on neuronal synapses is important for predicting their adverse effects. We have used drebrin as a marker to detect the synaptic changes in cultured neurons. High concentration of glutamate decreases the amount of drebrin in synapses. To increase the availability of this method for...
journal_title：Journal of pharmacological and toxicological methods
pub_type： 杂志文章
doi：10.1016/j.vascn.2019.106607
更新日期：2019-01-01 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
abstract：INTRODUCTION:Drug-induced QT interval prolongation may lead to ventricular arrhythmias. The aim of the study was to optimize QT interval data processing to quantify drug-induced QT interval prolongation in the telemetry instrumented conscious dog model. METHODS:The test substances cisapride, dofetilide, haloperidol, a...
journal_title：Journal of pharmacological and toxicological methods
pub_type： 杂志文章
doi：10.1016/j.vascn.2006.02.009
更新日期：2007-01-01 00:00:00
abstract：INTRODUCTION:The aim of this investigation was to evaluate methods for increasing Caco-2 cell throughput for assessing drug intestinal absorption. The use of 6-, 12-, and 24-well membranes and the effect of membrane size on permeability and the integrity of the Caco-2 cell monolayer were assessed. In an effort to optim...
journal_title：Journal of pharmacological and toxicological methods
pub_type： 杂志文章
doi：10.1016/s1056-8719(01)00161-7
更新日期：2001-07-01 00:00:00
abstract：INTRODUCTION:The standard Ficoll-Hypaque method used to isolate peripheral blood mononuclear cells (PBMC) gives very variable results when used with cynomolgus monkey blood. We have improved the method by using special cell processing tubes (CP-tubes), originally developed for clinical use. METHODS:Blood samples were ...
journal_title：Journal of pharmacological and toxicological methods
pub_type： 杂志文章
doi：10.1016/j.vascn.2004.04.003
更新日期：2004-09-01 00:00:00
abstract：:Induction of acute myocardial infarction in the rat is an established model for studying effects of therapeutic interventions. Images of sections of the rat left ventricle, stained with nitroblue tetrazolium, were digitized and several parameters estimated by dedicated software on an image analyzer (IBAS 2.0). The met...
journal_title：Journal of pharmacological and toxicological methods
pub_type： 杂志文章
doi：10.1016/1056-8719(95)00019-e
更新日期：1995-08-01 00:00:00
abstract：:The most commonly employed techniques for the in vivo measurement of arterial blood flow to individual organs involve the use of flow probes or sensors. Commercially available systems for the measurement of in vivo blood flow can be divided into two categories: ultrasonic and electromagnetic. Two types of ultrasonic p...
journal_title：Journal of pharmacological and toxicological methods
pub_type： 杂志文章,评审
doi：10.1016/s1056-8719(00)00123-4
更新日期：2000-09-01 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: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：:High-content screening (HCS) systems can be used for high-throughput screening of drugs in human embryonic stem cells (hESCs). However, hESCs require immunofluorescence staining with stemness markers (e.g., Oct-4) prior to HCS, which can be time consuming and labor intensive. In this study, we employed transgenic hESC...
journal_title：Journal of pharmacological and toxicological methods
pub_type： 杂志文章
doi：10.1016/j.vascn.2020.106913
更新日期：2020-01-01 00:00:00
abstract：INTRODUCTION:Implantable telemetry enables continuous monitoring of physiological functions in freely moving animals and can greatly complement pharmacological research. Despite its miniaturization, a sensor/transmitter constitutes 5% or more of a mouse's bodyweight. The aim of the present study was to evaluate whether...
journal_title：Journal of pharmacological and toxicological methods
pub_type： 杂志文章
doi：10.1016/j.vascn.2017.02.002
更新日期：2017-01-01 00:00:00
abstract：:For the rapid assay of norepinephrine (NE) and its major metabolite, 3-methoxy-4-hydroxyphenylglycol (MHPG) in the mouse brain, we developed a simple method using isocratic HPLC with coulometric detection. This method permits NE and MHPG assay within 5 min in one chromatographic run. Within-run coefficients of variati...
journal_title：Journal of pharmacological and toxicological methods
pub_type： 杂志文章
doi：10.1016/1056-8719(94)90060-4
更新日期：1994-10-01 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
pub_type： 杂志文章
doi：10.1016/j.vascn.2010.04.004
更新日期：2010-07-01 00:00:00
abstract：INTRODUCTION:The ICH S7B guidelines recommend that all new chemical entities should be subjected to hERG repolarization screening due to its association with life-threatening "Torsades de Pointes" (TdP) arrhythmia. However, it has become evident that not all hERG channel inhibitors result in TdP and not all compounds t...
journal_title：Journal of pharmacological and toxicological methods
pub_type： 杂志文章
doi：10.1016/j.vascn.2016.06.004
更新日期：2016-09-01 00:00:00
abstract：:The majority of articles published in J Pharmacol Tox Methods describe new approaches, or modifications to those established (Curtis, 2006). However, even established methods have their limitations. It is part of the practice of research to troubleshoot: to identify methodological weaknesses and correct them. This pro...
journal_title：Journal of pharmacological and toxicological methods
pub_type： 社论
doi：10.1016/j.vascn.2010.02.013
更新日期：2010-03-01 00:00:00
abstract：INTRODUCTION:In response to the ongoing shift of the regulatory cardiac safety paradigm, a recent White Paper proposed general principles for developing and implementing proarrhythmia risk prediction models. These principles included development strategies to validate models, and implementation strategies to ensure a m...
journal_title：Journal of pharmacological and toxicological methods
pub_type： 杂志文章
doi：10.1016/j.vascn.2020.106890
更新日期：2020-09-01 00:00:00
abstract：BACKGROUND:Animal models of allergy and bronchial hyperresponsiveness (BHR) are useful in researching pulmonary diseases and evaluating drug effects on the airways. Neonatally immunised rabbits exhibit several features of asthma as adults, including early and late airway responses following antigen challenge, oedema an...
journal_title：Journal of pharmacological and toxicological methods
pub_type： 杂志文章
doi：10.1016/j.vascn.2011.08.003
更新日期：2011-09-01 00:00:00
abstract：INTRODUCTION:In vitro metabolism models such as liver microsomes represent an important tool for the development of novel radioligands. Comparability and physiological relevance of in vitro metabolism data critically depend on the careful evaluation and optimization of assay protocols. We therefore investigated the inf...
journal_title：Journal of pharmacological and toxicological methods
pub_type： 杂志文章
doi：10.1016/j.vascn.2018.11.005
更新日期：2019-01-01 00:00:00
abstract：INTRODUCTION:The ability to quantify systemic concentrations of protein therapeutics is complicated by the presence of endogenous analyte, specific binding proteins, and nonspecific matrix components in biological matrices (Lee & Ma, 2007). Further complications can be introduced following pegylation whereby polyethyle...
journal_title：Journal of pharmacological and toxicological methods
pub_type： 杂志文章
doi：10.1016/j.vascn.2009.12.004
更新日期：2010-03-01 00:00:00
abstract：:A sensitive and precise gas chromatography-mass spectrometry method with selected ion monitoring has been developed for determination of ketamine in the brain using chlorpheniramine as an internal standard. The assay is based on the acid extraction of brain homogenate with hexane and ethyl ether with subsequent alkali...
journal_title：Journal of pharmacological and toxicological methods
pub_type： 杂志文章
doi：10.1016/s1056-8719(99)00023-4
更新日期：1999-04-01 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
abstract：:The origins of safety pharmacology are grounded upon observations that organ functions (like organ structures) can be toxicological targets in humans exposed to novel therapeutic agents, and that drug effects on organ functions (unlike organ structures) are not readily detected by standard toxicological testing. Safet...
journal_title：Journal of pharmacological and toxicological methods
pub_type： 杂志文章,评审
doi：10.1016/j.vascn.2004.02.007
更新日期：2004-05-01 00:00:00
abstract：INTRODUCTION:The purpose of this study was to optimize carbon tetrachloride-induced hepatotoxicity in the rat with respect to dose, route of injection, and time course. METHODS:Male Wistar albino rats, 4 to 6 weeks old and weighing 130-180 g were used. Hepatotoxicity was evaluated by measuring the activity of serum en...
journal_title：Journal of pharmacological and toxicological methods
pub_type： 杂志文章
doi：10.1016/S1056-8719(03)00019-4
更新日期：2002-07-01 00:00:00
abstract：INTRODUCTION:Oxidative stress plays an important role in drug-induced toxicity. Oxidative stress-mediated toxicities can be detected using conventional animal models but their sensitivity is insufficient, and novel models to improve susceptibility to oxidative stress have been researched. In recent years, gene targetin...
journal_title：Journal of pharmacological and toxicological methods
pub_type： 杂志文章
doi：10.1016/j.vascn.2018.12.006
更新日期：2019-01-01 00:00:00
abstract：:In a typical flowchart for discovery of novel analgesic (or other) agents, a critical path often involves maximization of the separation of the therapeutic endpoint from known adverse-effect (AE) endpoint(s). Although strategies can easily be designed for in vitro paradigms such as high-throughput screening, extension...
journal_title：Journal of pharmacological and toxicological methods
pub_type： 杂志文章
doi：10.1016/s1056-8719(00)00041-1
更新日期：1999-11-01 00:00:00
abstract：:This article describes the application of a Microplate Filtration System (MFS) to a binding assay, with the results being compared to those obtained with a conventional 24-Well Filtration Manifold (24WFM). The data reported here characterize the PAF receptor on rabbit platelet membranes using [3H]apafant. The results ...
journal_title：Journal of pharmacological and toxicological methods
pub_type： 杂志文章
doi：10.1016/1056-8719(96)00074-3
更新日期：1996-09-01 00:00:00
abstract：INTRODUCTION:Inhalation of antigen in atopic asthma induces early (EAR) and late asthmatic responses (LARs), inflammatory cell infiltration and airways hyperresponsiveness (AHR). Previously, we have established a protocol of sensitisation and subsequent ovalbumin (Ova) inhalation challenge in guinea-pigs which induced ...
journal_title：Journal of pharmacological and toxicological methods
pub_type： 杂志文章
doi：10.1016/j.vascn.2014.10.007
更新日期：2015-03-01 00:00:00
abstract：:Fluorescent labeling of amine functional groups using dansyl chloride (DNS-Cl), and in sodium carbonate buffer, allowed the detection of 1 microg amounts of analytes. The methodology presented allows dansylation of primary, secondary, and tertiary amine groups at a temperature of 25 degrees C. The dansylation of terti...
journal_title：Journal of pharmacological and toxicological methods
pub_type： 杂志文章
doi：10.1016/s1056-8719(01)00157-5
更新日期：2001-05-01 00:00:00
abstract：INTRODUCTION:Fosmidomycin is a phosphonic acid derivative originally isolated as a natural antibiotic from Streptomyces lavendulae. It was originally used as an antibiotic, and later on as an antimalarial drug. A simple, sensitive, selective and reproducible bioassay based on colorimetric method was developed for the d...
journal_title：Journal of pharmacological and toxicological methods
pub_type： 杂志文章
doi：10.1016/j.vascn.2010.02.012
更新日期：2010-05-01 00:00:00
abstract：:A method for determination of cis-diaminedichloroplatinum (II) (cisplatin) in ultrafiltered plasma, cell, and tumour samples is described. Cisplatin separation was carried out on a reversed-phase column using methanol-acetonitrile-water as the mobile phase. The flow rate was maintained constant at 1.6 mL/min and analy...
journal_title：Journal of pharmacological and toxicological methods
pub_type： 杂志文章
doi：10.1016/j.vascn.2005.06.005
更新日期：2005-11-01 00:00:00
