ABSTRACT
Ionic conductivity and membrane capacitance are two foundational parameters that govern neuron excitability. Conventional optogenetics has emerged as a powerful tool to temporarily manipulate membrane ionic conductivity in intact biological systems. However, no analogous method exists for precisely manipulating cell membrane capacitance to enable long-lasting modulation of neuronal excitability. Genetically targetable chemical assembly of conductive and insulating polymers can modulate cell membrane capacitance, but further development of this technique has been hindered by poor spatiotemporal control of the polymer deposition and cytotoxicity from the widely diffused peroxide. We address these issues by harnessing genetically targetable photosensitizer proteins to assemble electrically functional polymers in neurons with precise spatiotemporal control. Using whole-cell patch-clamp recordings, we demonstrate that this optogenetic polymerization can achieve stepwise modulation of both neuron membrane capacitance and intrinsic excitability. Furthermore, cytotoxicity can be limited by controlling light exposure, demonstrating a promising new method for precisely modulating cell excitability.
ABSTRACT
Fluorescent sensors for mobile zinc are valuable for studying complex biological systems. Because these sensors typically bind zinc rapidly and tightly, there has been little temporal control over the activity of the probe after its application to a sample. The ability to control the activity of a zinc sensor in vivo during imaging experiments would greatly improve the time resolution of the measurement. Here, we describe photoactivatable zinc sensors that can be triggered with short pulses of UV light. These probes are prepared by functionalizing a zinc sensor with protecting groups that render the probe insensitive to metal ions. Photoinduced removal of the protecting groups restores the binding site, allowing for zinc-responsive changes in fluorescence that can be observed in live cells and tissues.
Subject(s)
Fluorescent Dyes/chemistry , Zinc/analysis , Brain Chemistry , Fluorescence , HeLa Cells , Humans , Microscopy, Fluorescence/methods , Spectrometry, Fluorescence/methods , Ultraviolet RaysABSTRACT
The CF2H group, a potential surrogate for the OH group, can act as an unusual hydrogen bond donor, as confirmed by crystallographic, spectroscopic, and computational methods. Here, we demonstrate the bioisosterism of the OH and CF2H groups and the important roles of CF2-H···O hydrogen bonds in influencing intermolecular interactions and conformational preferences. Experimental evidence, corroborated by theory, reveals the distinctive nature of CF2H hydrogen bonding interactions relative to their normal OH hydrogen bonding counterparts.
