Linked Life Data

15671650

Source:http://linkedlifedata.com/resource/pubmed/id/15671650

Statements in which the resource exists as a subject.
PredicateObject
rdf:type
lifeskim:mentions
pubmed:issue
5
pubmed:dateCreated
2005-1-26
pubmed:abstractText
M-channels (M-current), encoded by KCNQ2/3 K(+) channel genes, have emerged as novel drug targets for a number of neurological disorders. The lack of direct high throughput assays combined with the low throughput of conventional electrophysiology (EP) has impeded rapid screening and evaluation of K(+)-channel modulators. Development of a sensitive and efficient assay for the direct measurement of M-current activity is critical for identifying novel M-channel modulators and subsequent investigation of their therapeutic potential. Using a stable CHO cell line expressing rat KCNQ2/3 K(+) channels confirmed by EP, we have developed and validated a nonradioactive rubidium (Rb(+)) efflux assay in a 96-well plate format. The Rb(+) efflux assay directly measures the activity of functional channels by atomic absorption spectroscopy using the automated Ion Channel Reader (ICR) 8000. The stimulated Rb(+) efflux from KCNQ2/3-expressing cells was blocked by the channel blockers XE991 and linopirdine with IC(50) values of 0.15 microM and 1.3 microM, respectively. Twelve compounds identified as KCNQ2/3 openers were further assessed in this assay, and their EC(50) values were compared with those obtained with EP. A higher positive correlation coefficient between these two assays (r = 0.60) was observed than that between FlexStation membrane potential and EP assays (r = 0.23). To simplify the assay and increase the throughput, we demonstrate that EC(50) values obtained by measuring Rb(+) levels in the supernatant are as robust and consistent as those obtained from the ratio of Rb(+) in supernatant/lysate. By measuring the supernatant only, the throughput of ICR8000 in an eight-point titration is estimated to be 40 compounds per day, which is suitable for a secondary confirmation assay.
pubmed:language
eng
pubmed:journal
pubmed:citationSubset
IM
pubmed:chemical
pubmed:status
MEDLINE
pubmed:month
Oct
pubmed:issn
1540-658X
pubmed:author
pubmed:issnType
Print
pubmed:volume
2
pubmed:owner
NLM
pubmed:authorsComplete
Y
pubmed:pagination
525-34
pubmed:dateRevised
2006-11-15
pubmed:meshHeading
pubmed:year
2004
pubmed:articleTitle
Validation of an atomic absorption rubidium ion efflux assay for KCNQ/M-channels using the ion Channel Reader 8000.
pubmed:affiliation
Discovery Neuroscience, Wyeth Research, CN-8000, Princeton, NJ 08543, USA. wangk@wyeth.com
pubmed:publicationType
Journal Article, Comparative Study, Validation Studies