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A G_s-coupled purinergic receptor boosts Ca²⁺ influx and vascular contractility during diabetic hyperglycemia.

ABSTRACT: Elevated glucose increases vascular reactivity by promoting L-type Ca_V1.2 channel (LTCC) activity by protein kinase A (PKA). Yet, how glucose activates PKA is unknown. We hypothesized that a G_s-coupled P2Y receptor is an upstream activator of PKA mediating LTCC potentiation during diabetic hyperglycemia. Experiments in apyrase-treated cells suggested involvement of a P2Y receptor underlying the glucose effects on LTTCs. Using human tissue, expression for P2Y₁₁, the only G_s-coupled P2Y receptor, was detected in nanometer proximity to Ca_V1.2 and PKA. FRET-based experiments revealed that the selective P2Y₁₁ agonist NF546 and elevated glucose stimulate cAMP production resulting in enhanced PKA-dependent LTCC activity. These changes were blocked by the selective P2Y₁₁ inhibitor NF340. Comparable results were observed in mouse tissue, suggesting that a P2Y₁₁-like receptor is mediating the glucose response in these cells. These findings established a key role for P2Y₁₁ in regulating PKA-dependent LTCC function and vascular reactivity during diabetic hyperglycemia.

SUBMITTER: Prada MP

PROVIDER: S-EPMC6397001 | biostudies-literature | 2019 Mar

REPOSITORIES: biostudies-literature

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A Gs-coupled purinergic receptor boosts Ca2+ influx and vascular contractility during diabetic hyperglycemia.

Prada Maria Paz MP Syed Arsalan U AU Buonarati Olivia R OR Reddy Gopireddy R GR Nystoriak Matthew A MA Ghosh Debapriya D Simó Sergi S Sato Daisuke D Sasse Kent C KC Ward Sean M SM Santana Luis F LF Xiang Yang K YK Hell Johannes W JW Nieves-Cintrón Madeline M Navedo Manuel F MF

eLife 20190301

Elevated glucose increases vascular reactivity by promoting L-type CaV1.2 channel (LTCC) activity by protein kinase A (PKA). Yet, how glucose activates PKA is unknown. We hypothesized that a Gs-coupled P2Y receptor is an upstream activator of PKA mediating LTCC potentiation during diabetic hyperglycemia. Experiments in apyrase-treated cells suggested involvement of a P2Y receptor underlying the glucose effects on LTTCs. Using human tissue, expression for P2Y11, t ...[more]

PMID: 30821687

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Project description:In mesenteric arteries (MAs), aldosterone (ALDO) binds to the endogenous mineralocorticoid receptor (MR) and increases the expression of the voltage-gated L-type Cav1.2 channel, an essential ion channel for vascular contraction, sarcoplasmic reticulum (SR) Ca2+ store refilling, and Ca2+ spark generation. In mesenteric artery smooth muscle cells (MASMCs), Ca2+ influx through Cav1.2 is the indirect mechanism for triggering Ca2+ sparks. This process is facilitated by plasma membrane-sarcoplasmic reticulum (PM-SR) nanojunctions that drive Ca2+ from the extracellular space into the SR via Sarco/Endoplasmic Reticulum Ca2+ (SERCA) pump. Ca2+ sparks produced by clusters of Ryanodine receptors (RyRs) at PM-SR nanodomains, decrease contractility by activating large-conductance Ca2+-activated K+ channels (BKCa channels), which generate spontaneous transient outward currents (STOCs). Altogether, Cav1.2, SERCA pump, RyRs, and BKCa channels work as a functional unit at the PM-SR nanodomain, regulating intracellular Ca2+ and vascular function. However, the effect of the ALDO/MR signaling pathway on this functional unit has not been completely explored. Our results show that short-term exposure to ALDO (10 nM, 24 h) increased the expression of Cav1.2 in rat MAs. The depolarization-induced Ca2+ entry increased SR Ca2+ load, and the frequencies of both Ca2+ sparks and STOCs, while [Ca2+]cyt and vasoconstriction remained unaltered in Aldo-treated MAs. ALDO treatment significantly increased the mRNA and protein expression levels of the SERCA pump, which counterbalanced the augmented Cav1.2-mediated Ca2+ influx at the PM-SR nanodomain, increasing SR Ca2+ content, Ca2+ spark and STOC frequencies, and opposing to hyperpolarization-induced vasoconstriction while enhancing Acetylcholine-mediated vasorelaxation. This work provides novel evidence for short-term ALDO-induced upregulation of the functional unit comprising Cav1.2, SERCA2 pump, RyRs, and BKCa channels; in which the SERCA pump buffers ALDO-induced upregulation of Ca2+ entry at the superficial SR-PM nanodomain of MASMCs, preventing ALDO-triggered depolarization-induced vasoconstriction and enhancing vasodilation. Pathological conditions that lead to SERCA pump downregulation, for instance, chronic exposure to ALDO, might favor the development of ALDO/MR-mediated augmented vasoconstriction of mesenteric arteries.

Dataset Information

A G_s-coupled purinergic receptor boosts Ca²⁺ influx and vascular contractility during diabetic hyperglycemia.

Publications

A G<sub>s</sub>-coupled purinergic receptor boosts Ca<sup>2+</sup> influx and vascular contractility during diabetic hyperglycemia.

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