BTDAzo: A Photoswitchable TRPC5 Channel Activator

Photoswitchable reagents can be powerful tools for high-precision biological control. TRPC5 is a Ca2+-permeable cation channel with distinct tissue-specific roles, from synaptic function to hormone regulation. Reagents giving spatiotemporally-resolved control over TRPC5 activity may be key to understanding and harnessing its biology. Here we develop the first photoswitchable TRPC5-modulator, BTDAzo, to address this goal. BTDAzo can photocontrol TRPC5 currents in cell culture, as well as controlling endogenous TRPC5-based neuronal Ca2+ responses in mouse brain slices. BTDAzos are also the first reported azo-benzothiadiazines, an accessible and conveniently derivatised azoheteroarene with strong two-colour photoswitching. BTDAzo ' s ability to control TRPC5 across relevant channel biology settings makes it suitable for a range of dynamically reversible photoswitching studies in TRP channel biology, with the aim to decipher the various biological roles of this centrally important ion channel

An Ecological Study On Primary Productivity And Energy Dynamics Of Wetland Of Hafizpur , Saran

At the Site-1, net primary productivity ranged 3850 mg C/m2/day with an annual mean±S.D. of 2769.76±983.64mg C/m2/day . Maximum NPP (4150 mg C/m2/day) was recorded during February and minimum (400 mg C/m2/day) in May .Net primary productivity in the Site-2 ranged 5200 mg C/m2/day with an annual mean±S.D. of 2231.26±1202.1 mg C/m2/day .Maximum NPP (5200 mg C/m2/day) was observed in February and minimum (0 mg C/m2/day) in August. Community respiration at the Site-1 ranged 2575 mg C/m2/day with an annual mean ±S.D. of 6618.76±626.91 mg C/m2/day .Maximum community respiration (2650 mg C/m2/day) was recorded during October and minimum (76mg C/m2/day) in August . At the Site-2 community respiration ranged 3625 mg C/m2/day with an annual mean ±S.D. of 2331.26±887.21mg C/m2/day. Maximum community respiration (3775 mg C/m2/day) was observed in October and minimum (250 mg C/m2/day) in May

A diacylglycerol photoswitching protocol for studying TRPC channel functions in mammalian cells and tissue slices

Summary: Small molecular probes designed for photopharmacology and opto-chemogenetics are rapidly gaining widespread recognition for investigations of transient receptor potential canonical (TRPC) channels. This protocol describes the use of three photoswitchable diacylglycerol analogs—PhoDAG-1, PhoDAG-3, and OptoDArG—for ultrarapid activation and deactivation of native TRPC2 channels in mouse vomeronasal sensory neurons and olfactory type B cells, as well as heterologously expressed human TRPC6 channels. Photoconversion can be achieved in mammalian tissue slices and enables all-optical stimulation and shutoff of TRPC channels.For complete details on the use and execution of this protocol, please refer to Leinders-Zufall et al. (2018)

BTDAzo - a photoswitchable TRPC5 channel activator

Photoswitchable reagents to modulate protein activity are powerful tools for high-spatiotemporal-precision control over endogenous biological functions. TRPC5 is a Ca2+-permeable cation channel with distinct tissue-specific roles, ranging from synaptic function to hormone regulation. Achieving spatially-resolved control over TRPC5 activity in particular cells or tissues, and temporal regulation in targeted cells, are therefore crucial milestones towards understanding and harnessing the biology of TRPC5. Here we develop the first photoswitchable TRPC5-modulating reagent, BTDAzo, towards reaching this goal. BTDAzo can photocontrol TRPC5 currents in cell culture, as well as controlling endogenous TRPC5-based neuronal Ca2+ responses in mouse brain slices. BTDAzos are also the first reported azo-benzothiadiazines, an accessible and conveniently derivatised azoheteroarene that features excellent two-colour photoswitching. BTDAzo\u27s TRPC5 control across relevant channel biology settings makes it appropriate for a range of dynamically reversible photoswitching studies in TRP channel biology, aiming to decipher the various biological roles of this centrally important ion channel

Sodium channels as gateable non-photonic sensors for membrane-delimited reactive species

AbstractReactive oxygen species (ROS) and reactive oxygen intermediates (ROI) play crucial roles in physiological processes. While excessive ROS damages cells, small fluctuations in ROS levels represent physiological signals important for vital functions. Despite the physiological importance of ROS, many fundamental questions remain unanswered, such as which types of ROS occur in cells, how they distribute inside cells, and how long they remain in an active form. The current study presents a ratiometric sensor of intracellular ROS levels based on genetically engineered voltage-gated sodium channels (roNaV). roNaV can be used for detecting oxidative modification that occurs near the plasma membrane with a sensitivity similar to existing fluorescence-based ROS sensors. Moreover, roNaV has several advantages over traditional sensors because it does not need excitation light for sensing, and thus, can be used to detect phototoxic cellular modifications. In addition, the ROS dynamic range of roNaV is easily manipulated in real time by means of the endogenous channel inactivation mechanism. Measurements on ROS liberated from intracellular Lucifer Yellow and genetically encoded KillerRed have revealed an assessment of ROS lifetime in individual mammalian cells. Flashlight-induced ROS concentration decayed with two major time constants of about 10 and 1000ms

Impact of higher-order heme degradation products on hepatic function and hemodynamics

Background & Aims: Biliverdin and bilirubin were previously considered end products of heme catabolism; now, however, there is evidence for further degradation to diverse bioactive products. Z-BOX A and Z-BOX B arise upon oxidation with unknown implications for hepatocellular function and integrity. We studied the impact of Z-BOX A and B on hepatic functions and explored their alterations in health and cholestatic conditions. Methods: Functional implications and mechanisms were investigated in rats, hepatocytic HepG2 and HepaRG cells, human immortalized hepatocytes, and isolated perfused livers. Z-BOX A and B were determined by liquid chromatography-tandem mass spectrometry (LC-MS/MS) in acute and acute-on-chronic liver failure and hereditary unconjugated hyperbilirubinemia. Results: Z-BOX A and B are found in similar amounts in humans and rodents under physiological conditions. Serum concentrations increased similar to 20-fold during cholestatic liver failure in humans (p <0.001) and in hereditary deficiency of bilirubin glucuronidation in rats (p <0.001). Pharmacokinetic studies revealed shorter serum half-life of Z-BOX A compared to its regio-isomer Z-BOX B (p = 0.035). While both compounds were taken up by hepatocytes, Z-BOX A was enriched similar to 100-fold and excreted in bile. Despite their reported vasoconstrictive properties in the brain vasculature, BOXes did not affect portal hemodynamics. Both Z-BOX A and B showed dose-dependent cytotoxicity, affected the glutathione redox state, and differentially modulated activity of Rev-erba and Rev-erbb. Moreover, BOXes-triggered remodeling of the hepatocellular cytoskeleton. Conclusions: Our data provide evidence that higher-order heme degradation products, namely Z-BOX A and B, impair hepatocellular integrity and might mediate intra-and extrahepatic cytotoxic effects previously attributed to hyperbilirubinemia. Lay summary: Degradation of the blood pigment heme yields the bile pigment bilirubin and the oxidation products Z-BOX A and Z-BOX B. Serum concentrations of these bioactive molecules increase in jaundice and can impair liver function and integrity. Amounts of Z-BOX A and Z-BOX B that are observed during liver failure in humans have profound effects on hepatic function when added to cultured liver cells or infused into healthy rats. (C) 2017 European Association for the Study of the Liver. Published by Elsevier B.V