Cell Membrane and Ion Channel Studies


Chemical permeation measured with droplet bilayers

02_five_droplets_array_BF_FITC


Aqueous droplets dispensed within a lipid/oil solution in a microfluidic device acquire self-assembled lipid monolayers on their surfaces. Contact of these droplets results in the formation of lipid bilayers. Molecules within these droplets may permeate through the lipid bilayers into the neighboring droplets. This chemical permeation from donor droplets to acceptor droplets can be monitored using fluorescence microscopy or UV-Vis microspectroscopy. Analysis of the permeation as a function of time allows the determination of permeability coefficients in under 20 minutes, much faster than conventional PAMPA measurements, a result of the 5 nm thick lipid bilayer and the ~nL volume droplets.

TRPM8DrugDose


Ion channels reconstituted into lipid bilayers may be characterized electrically by applying a trans-membrane potential and measuring the resultant current. The conductance of these ion channels may be altered by pharmaceutical compounds in the surrounding solution. We have used lipid bilayers to measure the interactions of hERG and TRPM8 ion channels with several drugs. hERG is an important cardiac ion channel implicated in many off-target dug interactions. TRPM8 is a temperature sensitive channel activated by cold temperatures and "cool" compounds such as menthol and has been implicated in several cancers. We reconstituted purified rTRPM8 ion channels into lipid bilayers and measured the dose dependent reduction in TRPM8 conductance with increasing concentration of Menthol (a TRPM8 activator) and 2-APB (an inhibitor) in the surrounding solution. A Hill equation fit of this reduction in measured current due to the drugs allows us to quantify the drug potency.





References
  1. “Hydrogel-Stabilized Droplet Bilayers for High Speed Solution Exchange,” Shiv A. Acharya, Alexander Portman, Carl S. Salazar, Jacob J. Schmidt, Scientific Reports 3 : 3139, DOI: 10.1038/srep03139
  2. “Microfluidic Passive Permeability Assay using Nanoliter Droplet Interface Lipid Bilayers,” Takasi Nisisako, Shiva Portonovo, Jacob Schmidt, Analyst 138(22), 6793-6800 (2013)
  3. “hERG Drug Response Measured in Droplet Bilayers,” Shiva A. Portonovo, Carl S. Salazar, and Jacob J. Schmidt, Biomedical Microdevices 2013, 15 (2), 255-259 (2012)
  4. “Ion channel drug potency assay with an artificial bilayer chip,” Ahmad M. El-Arabi, Carl J Salazar, and Jacob Schmidt, Lab Chip 2012, 12, 2409-2413