Kinetics of xylem loading, membrane potential maintenance, and sensitivity of K+-permeable channels to reactive oxygen species: physiological traits that differentiate salinity tolerance between pea and barley

Salt sensitive (pea) and salt tolerant (barley) species were used to understand the physiological basis of differential salinity tolerance in crops. Pea plants were much more efficient in restoring otherwise depolarized membrane potential thereby effectively decreasing K+ efflux through depolarization-activated outward rectifying potassium channels. At the same time, pea root apex was 10-fold more sensitive to physiologically relevant H2O2 concentration and accumulated larger amounts of H2O2 under saline conditions. This resulted in a rapid loss of cell viability in the pea root apex. Barley plants rapidly loaded Na+ into the xylem; this increase was only transient, and xylem and leaf Na+ concentration remained at a steady level for weeks. On the contrary, pea plants restricted xylem Na+ loading during the first few days of treatment but failed to prevent shoot Na+ elevation in the long term. It is concluded that superior salinity tolerance of barley plants compared with pea is conferred by at least three different mechanisms: (1) efficient control of xylem Na+ loading; (2) efficient control of H2O2 accumulation and reduced sensitivity of non-selective cation channels to H2O2 in the root apex; and (3) higher energy saving efficiency, with less ATP spent to maintain membrane potential under saline conditions.

GABA-mediated tonic inhibition differentially modulates gain in functional subtypes of cortical interneurons

Sean Lewis Hill, Steven Petrou, Christian Andreas Rössert, Bas-Jan Zandt

The binding of GABA (gamma-aminobutyric acid) to extrasynaptic GABA(A) receptors generates tonic inhibition that acts as a powerful modulator of cortical network activity. Despite GABA being present throughout the extracellular space of the brain, previous work has shown that GABA may differentially modulate the excitability of neuron subtypes according to variation in chloride gradient. Here, using biophysically detailed neuron models, we predict that tonic inhibition can differentially modulate the excitability of neuron subtypes according to variation in electrophysiological properties. Surprisingly, tonic inhibition increased the responsiveness (or gain) in models with features typical for somatostatin interneurons but decreased gain in models with features typical for parvalbumin interneurons. Patch-clamp recordings from cortical interneurons supported these predictions, and further in silico analysis was then performed to seek a putative mechanism underlying gain modulation. We found that gain modulation in models was dependent upon the magnitude of tonic current generated at depolarized membrane potential-a property associated with outward rectifying GABA(A) receptors. Furthermore, tonic inhibition produced two biophysical changes in models of relevance to neuronal excitability: 1) enhanced action potential repolarization via increased current flow into the dendritic compartment, and 2) reduced activation of voltage-dependent potassium channels. Finally, we show theoretically that reduced potassium channel activation selectively increases gain in models possessing action potential dynamics typical for somatostatin interneurons. Potassium channels in parvalbumin-type models deactivate rapidly and are unavailable for further modulation. These findings show that GABA can differentially modulate interneuron excitability and suggest a mechanism through which this occurs in silico via differences of intrinsic electrophysiological properties.

NATL ACAD SCIENCES2020

Water as a contrast agent for imaging interfacial structure and ion transport in giant vesicles

David Roesel

Hydrated lipid bilayer membranes and their asymmetry play a fundamental role in living cells by maintaining and regulating concentration gradients between cells, their environment, and their compartments. They achieve this not only through various channels and transporters, but also by being inherently semi-permeable to various molecules, including water and ions. However, molecular information about passive ion transport, as well as the complex structure of membrane hydration remain elusive, mainly due to a lack of experimental methods that can access this information, and relate it to micro- and macroscopic membrane properties. In this work, we study the molecular structure of lipid bilayer membranes and ion transport across them with high throughput wide-field second harmonic (SH) microscopy by utilizing water as the contrast agent. We show that by detecting small amounts of asymmetry in the structure of interfacial water, it is possible to measure the distribution of charged species across the membrane with high sensitivity. We apply this surface-selective technique to measure ion-lipid-water interactions, track ion transport, and quantify electro-chemical surface properties at the interfaces of lipid bilayer membranes in the form of giant unilamellar vesicles (GUVs). We start by improving the throughput of wide field SH microscopy in order to probe low-asymmetry interfaces with high contrast in a label-free manner. To do this, we design a custom optical parametric amplifier (OPA) with a tunable output in the 670-1000 nm range, up to a 1 MHz repetition rate, and an ultra-short 23 fs pulse duration. We then experimentally demonstrate the achieved throughput improvement. Next, we establish a way to probe interfacial hydration of GUVs. We quantify the surface properties of vesicles composed of different ratios of zwitterionic and anionic lipids, and show that only a few percent of anionic headgroups are ionized. We also observe spatial and temporal fluctuations in surface properties, and demonstrate that these fluctuations are universally found in lipid bilayer systems.Following that, we demonstrate a direct link between membrane potential fluctuations and divalent ion transport. Molecular dynamics simulations reveal that these fluctuations reduce the free energy cost of transient pore formation and increase the ion flux across an open pore. These transient pores can act as conduits for ion transport, which we SH image for a series of divalent cations (Cu2+, Ca2+, Ba2+, Mg2+) passing through GUV membranes. Combining the experimental and computational results, we show that permeation through pores formed via an ion-induced electrostatic field is a viable mechanism for unassisted ion transport.Then, we focus on unassisted Ca2+ translocation in more detail. We vary the hydrophobic core of bilayer membranes and observe different types of behavior in high throughput wide-field SH images. Ca2+ translocation is observed through mono-unsaturated membranes, significantly reduced upon adding cholesterol, and completely inhibited for branched and poly-unsaturated membranes. We propose, using molecular dynamics simulations, that ion transport occurs through ion induced transient pores, which require non-equilibrium membrane restructuring. This results in different transport rates at different locations and suggests that the hydrophobic structure of lipids plays a much more sophisticated regulating role than previously thought.

EPFL2022

GABA-mediated tonic inhibition differentially modulates gain in functional subtypes of cortical interneurons

Sean Lewis Hill, Steven Petrou, Christian Andreas Rössert, Bas-Jan Zandt

NATL ACAD SCIENCES2020

Water as a contrast agent for imaging interfacial structure and ion transport in giant vesicles

David Roesel

EPFL2022