Dissecting gating mechanisms of Orai calcium channel paralogs using constitutively active Orai mutants that mimic STIM1-gated state

In humans, there are three paralogs of the Orai Ca2+ channel, which lie at the heart of the store-operated calcium entry (SOCE) machinery. While the STIM-mediated gating mechanism of Orai channels is still being actively investigated, several artificial and natural variants are known to cause constitutive activity of the human Orai1 channel. Surprisingly, little is known about the conservation of the gating mechanism among the different human Orai paralogs and orthologs in other species. In our work, we show that the mutation corresponding to the activating mutation H134A in transmembrane helix 2 (TM2) of human Orai1 also activates Orai2 and Orai3, likely via a similar mechanism. However, this cross-paralog conservation does not apply to the “ANSGA” nexus mutations in TM4 of human Orai1 which mimic the STIM1-activated state of the channel. Investigating the mechanistic background of these differences, we identified two positions, H171 and F246 in human Orai1, which directly control the channel activation triggered by the “ANSGA” mutations in Orai1. Our results shed new light on these important gating checkpoints and show that the gating mechanism of the Orai channels is affected by multiple factors that are not necessarily evolutionarily conserved, such as the TM4-TM3 coupling

Sensitive multicolor indicators for monitoring norepinephrine in vivo

Genetically encoded indicators engineered from G-protein-coupled receptors are important tools that enable high-resolution in vivo neuromodulator imaging. Here, we introduce a family of sensitive multicolor norepinephrine (NE) indicators, which includes nLightG (green) and nLightR (red). These tools report endogenous NE release in vitro, ex vivo and in vivo with improved sensitivity, ligand selectivity and kinetics, as well as a distinct pharmacological profile compared with previous state-of-the-art GRAB$_{NE}$ indicators. Using in vivo multisite fiber photometry recordings of nLightG, we could simultaneously monitor optogenetically evoked NE release in the mouse locus coeruleus and hippocampus. Two-photon imaging of nLightG revealed locomotion and reward-related NE transients in the dorsal CA1 area of the hippocampus. Thus, the sensitive NE indicators introduced here represent an important addition to the current repertoire of indicators and provide the means for a thorough investigation of the NE system

The prevalence of abnormal glucose regulation in patients with coronary artery disease across Europe: The Euro Heart Survey on diabetes and the heart

Aim The objective behind the Euro Heart Survey on diabetes and the heart was to study the prevalence of abnormal glucose regulation in adult patients with coronary artery disease (CAD). Methods and results The survey engaged 110 centres in 25 countries recruiting 4196 patients referred to a cardiologist due to CAD out of whom 2107 were admitted on an acute basis and 2854 had an elective consultation. Patient data were collected via a web-based case record form. An oral glucose tolerance test (OGTT) was used for the characterisation of the glucose metabolism. Thirty-one per cent of the patients had diabetes. An OGTT was performed on the 1920 patients without known diabetes, of whom 923 had acute and 997 had a stable manifestation of CAD, respectively. In patients with acute CAD, 36% had impaired glucose regulation and 22% newly detected diabetes. In the stable group these proportions were 37% and 14%. Conclusion This survey demonstrates that normal glucose regulation is less common than abnormal glucose regulation in patients with CAD. OGTT easily discloses the glucometabolic state and should be a routine procedure. The knowledge of glucometabolic state among these patients should influence their future management because it has great potential to improve the outcome