title: Β-adrenergic-stimulated L-type channel Ca²+ entry mediates hypoxic Ca²+ overload in intact heart.
abstract: Ca(2+) mishandling plays a key role in ischemia- and hypoxia-related cardiac dysfunction and injury. However, the cellular and molecular mechanisms underlying hypoxic intracellular Ca(2+) ([Ca(2+)]i) overload remain incompletely understood. This study aimed to investigate possible mechanisms of [Ca(2+)]i overload during hypoxia in the intact heart. In Langendorff-perfused heart expressing the Ca(2+) indicator GCaMP2, confocal microscopy was used to simultaneously visualize [Ca(2+)]i, mitochondrial membrane potential (ΔΨm, by tetramethylrhodamine methyl ester) and sarcolemmal integrity (by Evans blue). Upon hypoxia (pO2 ~20 mmHg in glucose-free perfusate), [Ca(2+)]i transients were initially enhanced and then became depressed, arrhythmic, and completely abolished within 12 min. At ~20 min, basal [Ca(2+)]i rose to its first peak at a supraphysiological level, coincident with loss of ΔΨm and onset of rigor. A greater [Ca(2+)]i rise occurred at ~2h and was linked to the loss of sarcolemmal integrity. Removal of extracellular Ca(2+) or blockade of the l-type Ca(2+) channel (LTCC) (10 μM diltiazem or nifedipine) prevented [Ca(2+)]i overload and markedly delayed the loss of ΔΨm; by contrast, depletion of the sarcoplasmic reticulum Ca(2+) store by thapsigargin did not have any significant effect. Importantly, β-adrenergic blockade or depletion of the sympathetic catecholamine store by reserpine slowed the Ca(2+) and mitochondrial responses to hypoxia in intact heart. This LTCC-mediated hypoxic [Ca(2+)]i overload was reproduced in isolated cardiomyocytes when β-adrenergic agonist was present. Taken together, we conclude that Ca(2+) entry through β-adrenergic-stimulated LTCC underlies hypoxia-induced [Ca(2+)]i overload and the ensuing loss of mitochondrial function in intact heart.
authors.affiliation: State Key Laboratory of Biomembrane and Membrane Biotechnology, Beijing Key Laboratory of Cardiometabolic Molecular Medicine, Institute of Molecular Medicine, Peking-Tsinghua Center for Life Sciences, Peking University, Beijing 100871, China.
authors.forename: Huiliang
authors.initials: H
authors.lastname: Zhang
authors.forename: Wei
authors.initials: W
authors.lastname: Shang
authors.forename: Xing
authors.initials: X
authors.lastname: Zhang
authors.forename: Jingli
authors.initials: J
authors.lastname: Gu
authors.forename: Xianhua
authors.initials: X
authors.lastname: Wang
authors.forename: Ming
authors.initials: M
authors.lastname: Zheng
authors.forename: Yanru
authors.initials: Y
authors.lastname: Wang
authors.forename: Zhuan
authors.initials: Z
authors.lastname: Zhou
authors.forename: Ji-Min
authors.initials: JM
authors.lastname: Cao
authors.forename: Guangju
authors.initials: G
authors.lastname: Ji
authors.forename: Rongli
authors.initials: R
authors.lastname: Zhang
authors.forename: Heping
authors.initials: H
authors.lastname: Cheng
mesh_terms: D000818:Animals; D002118:Calcium / Q000378:metabolism*; D020746:Calcium Channels, L-Type / Q000378:metabolism*; D020013:Calcium Signaling; D015687:Cell Hypoxia; D066298:In Vitro Techniques; D015640:Ion Channel Gating; D008297:Male; D053078:Membrane Potential, Mitochondrial; D051379:Mice; D008810:Mice, Inbred C57BL; D008822:Mice, Transgenic; D008928:Mitochondria / Q000378:metabolism; D008954:Models, Biological; D009206:Myocardium / Q000378:metabolism* / Q000473:pathology; D011943:Receptors, Adrenergic, beta / Q000378:metabolism*
chemical_list: D020746:Calcium Channels, L-Type; D011943:Receptors, Adrenergic, beta; D002118:Calcium
keywords: Ex vivo heart imaging; Hypoxia; Mitochondria; l-type Ca(2+) channel; β-Adrenergic stimulation