Deficiency of ER Ca2+ sensor STIM1 in AgRP neurons confers protection against dietary obesity.

Store-operated calcium entry (SOCE) is pivotal in maintaining intracellular Ca2+ level and cell function; however, its role in obesity development remains largely unknown. Here, we show that the stromal interaction molecule 1 (Stim1), an endoplasmic reticulum (ER) Ca2+ sensor for SOCE, is critically involved in obesity development. Pharmacological blockade of SOCE in the brain, or disruption of Stim1 in hypothalamic agouti-related peptide (AgRP)-producing neurons (ASKO), significantly ameliorates dietary obesity and its associated metabolic disorders. Conversely, constitutive activation of Stim1 in AgRP neurons leads to an obesity-like phenotype. We show that the blockade of SOCE suppresses general translation in neuronal cells via the 2',5'-oligoadenylate synthetase 3 (Oas3)-RNase L signaling. While Oas3 overexpression in AgRP neurons protects mice against dietary obesity, deactivation of RNase L in these neurons significantly abolishes the effect of ASKO. These findings highlight an important role of Stim1 and SOCE in the development of obesity.

Vasopressin-stimulated ORAI1 expression and store-operated Ca2+ entry in aortic smooth muscle cells.

Vascular calcification may result from stimulation of osteogenic signalling with upregulation of the transcription factors CBFA1, MSX2 and SOX9, as well as alkaline phosphatase (ALPL), which degrades and thus inactivates the calcification inhibitor pyrophosphate. Osteogenic signalling further involves upregulation of the Ca2+-channel ORAI1. The channel is activated by STIM1 and then accomplishes store-operated Ca2+ entry. ORAI1 and STIM1 are upregulated by the serum & glucocorticoid inducible kinase 1 (SGK1) which is critically important for osteogenic signalling. Stimulators of vascular calcification include vasopressin. The present study explored whether exposure of human aortic smooth muscle cells (HAoSMCs) to vasopressin upregulates ORAI1 and/or STIM1 expression, store-operated Ca2+ entry and osteogenic signalling. To this end, HAoSMCs were exposed to vasopressin (100 nM, 24 h) without or with additional exposure to ORAI1 blocker MRS1845 (10 µM) or SGK1 inhibitor GSK-650394 (1 µM). Transcript levels were measured using q-RT-PCR, cytosolic Ca2+-concentration ([Ca2+]i) by Fura-2-fluorescence, and store-operated Ca2+ entry from increase of [Ca2+]i following re-addition of extracellular Ca2+ after store depletion with thapsigargin (1 µM). As a result, vasopressin enhanced the transcript levels of ORAI1 and STIM1, store-operated Ca2+ entry, as well as the transcript levels of CBFA1, MSX2, SOX9 and ALPL. The effect of vasopressin on store-operated Ca2+ entry as well as on transcript levels of CBFA1, MSX2, SOX9 and ALPL was virtually abrogated by MRS1845 and GSK-650394. In conclusion, vasopressin stimulates expression of ORAI1/STIM1, thus augmenting store-operated Ca2+ entry and osteogenic signalling. In HAoSMCs, vasopressin (VP) upregulates Ca2+ channel ORAI1 and its activator STIM1. VP upregulates store-operated Ca2+ entry (SOCE) and osteogenic signalling (OS). VP-induced SOCE, OS and Ca2+-deposition are disrupted by ORAI1 inhibitor MRS1845. VP-induced SOCE, OS and Ca2+-deposition are disrupted by SGK1 blocker GSK-650394. KEY MESSAGES: • In HAoSMCs, vasopressin (VP) upregulates Ca2+ channel ORAI1 and its activator STIM1. • VP upregulates store-operated Ca2+ entry (SOCE) and osteogenic signalling (OS). • VP-induced SOCE, OS and Ca2+-deposition are disrupted by ORAI1 inhibitor MRS1845. • VP-induced SOCE, OS and Ca2+-deposition are disrupted by SGK1 blocker GSK-650394.

A Role for Astroglial Calcium in Mammalian Sleep and Sleep Regulation.

Mammalian sleep expression and regulation have historically been thought to reflect the activity of neurons. Changes in other brain cells (glia) across the sleep-wake cycle and their role in sleep regulation are comparatively unexplored. We show that sleep and wakefulness are accompanied by state-dependent changes in astroglial activity. Using a miniature microscope in freely behaving mice and a two-photon microscope in head-fixed, unanesthetized mice, we show that astroglial calcium signals are highest in wake and lowest in sleep and are most pronounced in astroglial processes. We also find that astroglial calcium signals during non-rapid eye movement sleep change in proportion to sleep need. In contrast to neurons, astrocytes become less synchronized during non-rapid eye movement sleep after sleep deprivation at the network and single-cell level. Finally, we show that conditionally reducing intracellular calcium in astrocytes impairs the homeostatic response to sleep deprivation. Thus, astroglial calcium activity changes dynamically across vigilance states, is proportional to sleep need, and is a component of the sleep homeostat.

Store-Operated Calcium Entry via STIM1 Contributes to MRGPRX2 Induced Mast Cell Functions.

Mast cells are inflammatory immune cells that play an essential role in mediating allergic reactions in humans. It is well-known that mast cell activation is critically regulated by intracellular calcium ion (Ca2+) concentrations. MAS-related G-protein coupled receptor-X2 (MRGPRX2) is a G-protein coupled receptor (GPCR) expressed on mast cells that is activated by various ligands, including several FDA approved drugs; consequently, this receptor has been implicated in causing pseudo-allergic reactions in humans. MRGPRX2 activation leads to an increase in intracellular Ca2+ levels; however, the Ca2+ mobilizing mechanisms utilized by this receptor are largely unknown. Previous reports showed that store-operated Ca2+ entry (SOCE) via the calcium sensor, stromal interaction molecule 1 (STIM1), regulates mast cell response induced by the high-affinity IgE receptor (FcεRI). In this study, using complementary pharmacologic and genetic ablation approaches we demonstrate that SOCE through STIM1 promotes MRGPRX2-induced human mast cell response in vitro. Importantly, SOCE also critically modulates MrgprB2 (mouse ortholog of human MRGPRX2) dependent inflammation in in vivo mouse models of pseudo-allergy. Collectively, our data suggests that MRGPRX2/MrgprB2 activation of mast cells is dependent on SOCE via STIM1, and further characterization of the MRGPRX2-SOCE-STIM1 pathway will lead to the identification of novel targets for the treatment of pseudo-allergic reactions in humans.