A fast microfluidic based 30-barrel application system to functionally characterize ligand-gated ion channels

in: Acta Physiologica (2016)
Schmauder, R.; Schwabe, T.; Kusche, J.; Nache, V.; Herbst, E.; Mayer, Günter; Benndorf, K.
The interaction of ligands with the binding site in ligand-gat-ed ion channels is usually characterized by measuring the steady-state concentration-activation relationships. In addi-tion to this, the kinetics of the gating process is investigated with fast concentration jumps at selected concentrations.Here we present a fast multichannel-flow system enabling measurement of fast kinetics and detailed concentration-ac-tivation relationships over a wide concentration range in one experiment. The system consists of a 30-channel microflu-idic application device, combined with a step-motor and a fast piezo-actuator to evoke concentration jumps for all 30barrels. The microfluidic chip also ensures a minute re-agent consumption.Proof-of concept data on cyclic-nucleotide gated (CNG) channels in inside-out patches and purinergic P2X channels in whole cells and outside-out patches are presented. For whole cells and outside patches solution exchange rates well below 10 ms are routinely achieved.The system enables fast screening of ligands and receptors using minute reagent quantities and recording of more or less smooth concentration-activation relationships. Further-more the system allows for complex concentration-step pro-tocols, similar to conventional voltage-protocols. With such specialized protocols certain transitions in Markov-models, describing complex, highly cooperative ligand gated ion channels, can be preferentially populated and studied in greater detail.

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