Rapid Solution Exchange and Ligand-Gated Channel Studies on the PatchXpress 7000A Automated Patch Clamp System
Poster Jan 09, 2006
Iris Yang, Cathy Smith-Maxwell, David Yamane and Naibo Yang
AbstractThe PatchXpress® 7000A patch clamp system is an automated electrophysiology workstation that allows users to increase throughput of research quality recordings and to measure ion channel activity from nearly any channel type. The unique SealChip™ planar electrodes by AVIVA Biosciences and the PatchXpress fluidics system support high quality patchclamp recordings of both voltage-gated and ligand-gated ion channels.
The system uses a robotic pipettor to deliver ligand and drugs directly to the patched cells in the whole-cell configuration. Washout of ligand in each well is achieved by employing a pair of dedicated perfusion tips in close proximity to the base of the recording well for rapid fluid exchange near the cell.
In order to quantify the speed of ligand and drug addition, we studied fluid exchange rates on the PatchXpress 7000A system using RBL cells endogenously-expressing inward-rectifier K channels and changed the concentration of external potassium [K+]. Going from low to high [K+] increased the current with an exponential time course and a single time constant of 19±11 ms. Washing out the high [K+] with the independent wash station has a time constant of 420±281 ms.
We further tested the system with cells expressing ligand-gated channels, including HEK cells endogenously expressing pH-sensitive ASIC channels as well as L-tk cells heterologously-expressing GABAA receptors. A number of ligands have been successfully tested on both cell/channel types. The interaction of ligand and specific agonists and antagonists with the channels was recorded.
Complete washout of ligand (GABA) is easily obtained with the independent wash station. Our results demonstrate that the properties of the fluid delivery system together with a newly added software feature for precise timing of ligand and drug delivery, make the PatchXpress 7000A platform suitable for a wide range of ligand-gated ion channel studies.
Early life stress (ELS) is highly associated with development of psychopathology
and mood disorders in adulthood. Genetic studies have identified variation in the gene calcium voltage-gated channel subunit alpha1C (CACNA1C) to increase risk for several psychiatric disorders. This poster assessed the expression of Cacna1c following prepubertal stress.
We found a distinct subpopulation of Tregs within BMSCs. Tregs and BMSCs in co-culture conferred neuroprotection that varied in a dose-dependent manner. Tregs minimized stem cell production of IL-6, a pro-inflammatory cytokine, and inhibited BMSC secretion of FGF-beta, a cytokine related to BMSC proliferation and differentiation. The ratio of Tregs found natively in BMSCs is optimally adapted to provide the maximum neuroprotective benefit of stem cell treatment after ischemic stroke.READ MORE
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