Qube Opto 384 – Optical modulation of ion channels
Sophion Bioscience Go to journalAuthor(s):
Kim Boddum, Sandra Wilson
Using the optical features of Qube Opto 384, it is possible to evaluate both light-activated ion channels and photoactivated ligands. Here we outline a few of the studies we have performed on Qube Opto 384.
Channelrhodopsin 2 (ChR2) could be activated and the channelrhodopsin 2 mediated current could be manipulated in both a light and voltage-dependent manner, with activation times as short as 4 ms. A light-induced chloride current could also be elicited, employing the chloride-conducting channelrhodopsin iC++.
Caged γ-aminobutyric acid (Rubi-GABA) could be activated by light and give rise to a GABAAR mediated current. The GABA response was both concentration and the light intensity dependent. The optical activation of GABA combined with microfluidic channels also enabled ultra-short ligand exposure times.
Optogenetics and Optical Tools in Automated Patch Clamping
Springer International Publishing Go to journalAuthor(s):
Boddum, K., Skafte-Pedersen, P., Rolland, J. F., & Wilson, S
Automated patch clamping (APC) has been used for almost two decades to increase the throughput of electrophysiological measurements, especially in preclinical safety screening of drug compounds. Typically, cells are suctioned onto holes in planar surfaces and a stronger subsequent suction allows access to a whole cell configuration for electrical measurement of ion channel activity. The development of optogenetic tools over a wide range of wavelengths (UV to IR) provides powerful tools for improving spatiotemporal control of in vivo and in vitro experiments and is emerging as a powerful means of investigating cell networks (neuronal), single-cell transduction, and subcellular pathways.
Combining APC and optogenetic tools paves the way for improved investigation and control of cell kinetics and provides the opportunity for collecting robust data for new and exciting applications and therapeutic areas. Here, we present an APC optogenetics capability on the Qube Opto 384 system including experiments on light-activated ion channels and photoactivated ligands.
Optical modulation of ion channels using Qube Opto
Mammalian Sensory Systems 2019 Go to journalAuthor(s):
Kim Boddum, Peder Skafte-Pedersen, Jens Henneke, Daniel Sauter, Jean Francois Rolland, Jiaye Zhang, Sandra Wilson
Optical modulation of ion channels is traditionally studied using a manual patch clamp system combined with a light source. This approach, however, is limited by very low throughput. In the present work, we show data recorded using a 384-well based automated patch clamp system equipped with 384 integrated light sources (Qube Opto 384).
In this study, we evaluated Channelrhodopsin 2 (ChR2), a light-sensitive non-selective cation channel permeable to Na+, K+ and Ca2+ opened upon illumination (Berndt et. al., 2012). Furthermore, we employed the chloride-conducting channelrhodopsin (iC++, Govorunova et Al. 2015), which was developed from a non-selective cation conducting channelrhodopsin through a mutational approach. Finally, we used photoactivatable adenylyl cyclase, bPAC, to modulate cellular cAMP levels and thereby alter biophysical properties of the HCN2 channel (Stierl et Al., 2011).
Compound activation by light enables the pharmacological manipulation of receptors, ion channels and other proteins with a high degree of temporal control. We used caged GABA (Rubi-GABA, Zayat et. al., 2003) to study the light activation of ligands, in combination with the microfluidic flow channel of the QChip 384, to give a higher degree of experimental control.
Ligand-activation of GABAA receptors on the automated patch clamp platforms QPatch and Qube 384 using conventional electrophysiology and optopharmacology
Society for Neuroscience 2018 Go to journalAuthor(s):
Melanie Schupp, Kim Boddum, Daniel R. Sauter, Peder Skafte-Pedersen, Linda Blomster, Hervør L. Olsen, Rasmus. B. Jacobsen
γ-Aminobutyric acid (GABA) is the major inhibitory neurotransmitter in the central nervous system (CNS) and the binding of GABA to ionotropic GABA receptors (GABAAR) is a crucial process in the healthy brain. An imbalance of GABA secretion or the malfunction of the receptor is associated with multiple disease areas like anxiety disorders, seizures and schizophrenia. Pharmacological manipulation of the receptor has, therefore, a large therapeutic potential, which is underscored by the amount of available treatment possibilities and the ongoing search for alternatives thereof1-4.
GABAA receptors are ligand-gated ion-channels that consist of 5 membrane-spanning subunits5,6 and are permeable to Cl– ions. So far, 16 different subunits have been identified in humans (ɑ1-6, ꞵ1-3, γ1-3, δ, ε, γ, θ, π) and their composition within the GABA receptor leads to different pharmacological responses7. Here, we show pharmacological modulation of the GABAAR using our high-throughput automated patch clamp (APC) systems QPatch and Qube 384. Our study includes a characterization of the heterogeneous GABAAR population of cultured primary hippocampal astrocytes and an evaluation of the GABAAR clone ɑ5ꞵ3γ2. In addition, we utilize the well-characterized GABAAR response to establish a novel method for ligand release, namely the light-stimulated release of ruthenium-bipyridine-triphenylphosphine(RuBi)-caged GABA using light stimulation on the Qube 384 platform.