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Understanding the mechanisms of assembly and disassembly of macromolecular structures in cells relies on solving biomolecular interactions. However, those interactions often remain unclear because tools to track molecular dynamics are not sufficiently resolved in time or space. In this study, we present a straightforward method for resolving inter‐ and intra‐molecular interactions in cell adhesive machinery, using quantum dot (QD) based Förster resonance energy transfer (FRET) nanosensors. Using a mechanosensitive protein, talin, one of the major components of focal adhesions, we are investigating the mechanosensing ability of proteins to sense and respond to mechanical stimuli. First, we quantified the distances separating talin and a giant unilamellar vesicle membrane for three talin variants. These variants differ in molecular length. Second, we investigated the mechanosensing capabilities of talin, i.e., its conformational changes due to mechanical stretching initiated by cytoskeleton contraction. Our results suggest that in early focal adhesion, talin undergoes stretching, corresponding to a decrease in the talin‐membrane distance of 2.5 nm. We demonstrate that QD‐FRET nanosensors can be applied for the sensitive quantification of mechanosensing with a sub‐nanometer accuracy. [ABSTRACT FROM AUTHOR]

Neurovascular coupling (NVC), which mediates rapid increases in cerebral blood flow in response to neuronal activation, is commonly used to map brain activation or dysfunction. Here we tested the reemerging hypothesis that CO2 generated by neuronal metabolism contributes to NVC. We combined functional ultrasound and two-photon imaging in the mouse barrel cortex to specifically examine the onsets of local changes in vessel diameter, blood flow dynamics, vascular/perivascular/intracellular pH, and intracellular calcium signals along the vascular arbor in response to a short and strong CO2 challenge (10 s, 20%) and whisker stimulation. We report that the brief hypercapnia reversibly acidifies all cells of the arteriole wall and the periarteriolar space 3–4 s prior to the arteriole dilation. During this prolonged lag period, NVC triggered by whisker stimulation is not affected by the acidification of the entire neurovascular unit. As it also persists under condition of continuous inflow of CO2, we conclude that CO2 is not involved in NVC. A remerging hypothesis suggests that CO2 generated by neuronal metabolism contributes to neurovascular coupling (NVC). Here, the authors show that NVC is unaffected by the acidification of the entire arteriolar column during cerebrovascular reactivity to CO2, demonstrating that CO2 is not a mediator of NVC. [ABSTRACT FROM AUTHOR]

Fluorophores bearing cationic pendants, such as the pyridinium group, tend to preferentially accumulate in mitochondria, whereas those with pentafluorophenyl groups display a distinct affinity for the endoplasmic reticulum. In this study, we designed fluorophores incorporating pyridinium and pentafluorophenyl pendants and examined their impact on sub‐cellular localization. Remarkably, the fluorophores exhibited a notable propensity for the mitochondrial membrane. Furthermore, these fluorophores revealed dual functionality by facilitating the detection of viscosity changes within the sub‐cellular environment and serving as heavy‐atom‐free photosensitizers. With easy chemical tunability, wash‐free imaging, and a favorable signal‐to‐noise ratio, these fluorophores are valuable tools for imaging mitochondria and investigating their cellular processes. [ABSTRACT FROM AUTHOR]

We herein present a new concept to produce dual‐color photoconvertible probes based on a mechanism called Directed Photooxidation Induced Conversion (DPIC). As a support of this mechanism, styryl‐coumarins (SCs) bearing Aromatic Singlet Oxygen Reactive Moieties (ASORMs) like furan and pyrrole have been synthesized. SCs are bright fluorophores, which undergo a hypsochromic conversion upon visible light irradiation due to directed photooxidation of the ASORM that leads to the disruption of conjugation. SC‐P, a yellow emitting probe bearing a pyrrole moiety, converts to a stable blue emitting coumarin with a 68 nm shift allowing the photoconversion and tracking of lipid droplet in live cells. This new approach might pave the way to a new generation of photoconvertible dyes for advanced bioimaging applications. [ABSTRACT FROM AUTHOR]

Polymeric nanoparticles (NPs) continue to assert their high potential as efficient nanovectors for nanomedicine and imaging agents. Among them, biocompatible polyester block copolymers (BCP) possess appealing features as their physicochemical properties can be readily modified by the composition of their blocks. Although the fluorescent labeling of polymers is often used to track the formed NPs in bioimaging, it is rarely used to characterize their physicochemical properties. In this work, we assumed that the branching degree of the hydrophobic block copolymer might play an important role in the properties of the biocompatible NPs and thus could help in controlling their cargo encapsulation and release. To this end, clickable PEG-PCL block copolymers with various branching degrees were synthesized. Owing to their covalent fluorescent labeling using strain-promoted azide-alkyne cycloaddition, several parameters like the critical aggregation concentration of the BCPs as well as the colloidal stability, the core polarity, and stealth of the NPs have been studied. Taking advantage of the fluorescence labeling of the NPs' core, their ability to encapsulate and release a fluorescent cargo (Rhodamine C18) was assessed by Förster resonance energy transfer (FRET). The observed differences in the release profile were confirmed in cells where the fluorescent NPs and their cargo were tracked. Finally, the cargo release of endocytosed NPs was imaged and assessed by FRET ratiometric imaging. This study proved that the covalent fluorescent labeling of the BCPs is an efficient tool, offering various methods to characterize and assess the effects of polymers' modifications on the NPs' properties. [ABSTRACT FROM AUTHOR]