Lipopolysaccharides directly decrease Ca2+ oscillations and the hyperpolarization-activated nonselective cation current If in immortalized HL-1 cardiomyocytes

Wondergem, Robert; Graves, Bridget M.; Ozment-Skelton, Tammy; Li, Chuanfu; Williams, David L.

doi:10.1152/ajpcell.00129.2010

Cited by 18 publications

(32 citation statements)

References 29 publications

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“…Prolongation and reduced rate of depolarization of the pacemaker potential by LPS is consistent with our report that LPS inhibits the hyperpolarization-activated cationic current, I f , (41) and HCN ion channels (19). This, however, does not account completely for the extended action potential duration.…”

Section: Resultssupporting

confidence: 91%

“…The magnitude of these inward currents is in keeping with that of I Na (43), and we have shown that they are eliminated by substituting NMDG ϩ for external Na ϩ (41). We also have reported that LPS does not inhibit HL-1 cell L-type Ca 2ϩ currents within this time course (41). Although it is apparent that voltage control of I Na is problematic in HL-1 cells (43) and other atrial myocytes (40), carefully controlled measurements demonstrated that LPS over 20 min reduced the magnitude of the largest and most rapid of the voltage-activated inward currents in HL-1 cells (Fig.…”

Section: Resultsmentioning

confidence: 55%

“…Action potentials occur spontaneously in ϳ40% of nonconfluent immortalized mouse Hl-1 cardiomyocytes, accompanied by transient increases in intracellular Ca 2ϩ (29,41). These action potentials display depolarizing, diastolic pacemaker potentials in phase 4 followed by rapid upstrokes and repolarizations, which are characteristic of sinoatrial node cardiomyocytes in situ, (Fig.…”

Section: Resultsmentioning

confidence: 99%

“…Based on telemetry studies in mice, this reduction in HRV is attributable to lipopolysaccharide (LPS; Ref. 5); however, it remains uncertain whether this action of LPS results from its direct effect on heart cells (41,47) or broader effects involving the central and autonomic nervous systems (6).…”

mentioning

confidence: 99%

“…We have reported that bacterial LPS added to an immortalized mouse myocardial cell line, HL-1 cells (4) (41). LPS also inhibited activation of the cardiac pacemaker, "funny" current, I f (41), which is attributable to direct inhibition of human HCN2 ion channel current (19).…”

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confidence: 99%

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Lipopolysaccharide prolongs action potential duration in HL-1 mouse cardiomyocytes

Wondergem

Graves

et al. 2012

American Journal of Physiology-Cell Physiology

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Wondergem R, Graves BM, Li C, Williams DL. Lipopolysaccharide prolongs action potential duration in HL-1 mouse cardiomyocytes. Am J Physiol Cell Physiol 303: C825-C833, 2012. First published August 15, 2012; doi:10.1152/ajpcell.00173.2012.-Sepsis has deleterious effects on cardiac function including reduced contractility. We have shown previously that lipopolysaccharides (LPS) directly affect HL-1 cardiac myocytes by inhibiting Ca 2ϩ regulation and by impairing pacemaker "funny" current, I f. We now explore further cellular mechanisms whereby LPS inhibits excitability in HL-1 cells. LPS (1 g/ml) derived from Salmonella enteritidis decreased rate of firing of spontaneous action potentials in HL-1 cells, and it increased their pacemaker potential durations and decreased their rates of depolarization, all measured by whole cell current clamp. LPS also increased action potential durations and decreased their amplitude in cells paced at 1 Hz with 0.1 nA, and 20 min were necessary for maximal effect. LPS decreased the amplitude of a rapidly inactivating inward current attributed to Na ϩ and of an outward current attributed to K ϩ ; both were measured by whole cell voltage clamp. The K ϩ currents displayed a resurgent outward tail current, which is characteristic of the rapid delayed-rectifier K ϩ current, IKr. LPS accordingly reduced outward currents measured with pipette Cs ϩ substituted for K ϩ to isolate IKr. E-4031 (1 M) markedly inhibited IKr in HL-1 cells and also increased action potential duration; however, the direct effects of E-4031 occurred minutes faster than the slow effects of LPS. We conclude that LPS increases action potential duration in HL-1 mouse cardiomyocytes by inhibition of IKr and decreases their rate of firing by inhibition of INa. This protracted time course points toward an intermediary metabolic event, which either decreases available mouse ether-a-go-go (mERG) and Na ϩ channels or potentiates their inactivation.delayed rectifier K ϩ current; patch-clamp electrophysiology; Salmonella enteritidis; E-4031 SEPSIS IS A COMPLEX CLINICAL problem that can progress into multiorgan dysfunction syndrome (MODS) and death. At least one-half of the high mortality rates of severe-sepsis, sepsis, and MODS result from cardiovascular failure (27). Approximately 40% of patients in septic shock also experience myocardial dysfunction (36, 44), which comprises reduced systolic contractility, impaired diastolic relaxation, and ventricular dilatation. The initial clinical pattern, however, ironically is termed "hyperdynamic," because it often presents with high cardiac output, elevated heart rate, and vasodilatation of the dermis (27). This is attributable in part to the volume loading that is frequently the initial treatment for septic shock patients to maintain cardiac output and organ perfusion (15, 28). As is often the case in the complexity of sepsis, the prevailing symptoms can mask organ and system failure occurring at a deeper level in the clinical spectrum.The most prevalent sign of myocardial depressio...

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Section: Resultssupporting

confidence: 91%

Section: Resultsmentioning

confidence: 55%

Section: Resultsmentioning

confidence: 99%

mentioning

confidence: 99%

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confidence: 99%

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Lipopolysaccharide prolongs action potential duration in HL-1 mouse cardiomyocytes

Wondergem

Graves

et al. 2012

American Journal of Physiology-Cell Physiology

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Long-term culture of HL-1 cardiomyocytes in modular poly(ethylene glycol) microsphere-based scaffolds crosslinked in the phase-separated state

Smith¹,

Segar²,

Nguyen³

et al. 2012

Acta Biomaterialia

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Poly(ethylene glycol) (PEG) microspheres were assembled around HL-1 cardiomyocytes to produce highly porous modular scaffolds. In this study, we took advantage of the immiscibility of PEG and dextran to improve upon our previously described modular scaffold fabrication methods. Phase separating the PEG microspheres in dextran solutions caused them to deswell and crosslink together rapidly, eliminating the need for serum protein-based crosslinking. This also led to a dramatic increase in the stiffness of the scaffolds and greatly improved the handling characteristics. HL-1 cardiomyocytes were present during the microsphere crosslinking in the cytocompatible dextran solution, exhibiting high cell viability following scaffold formation. Over the course of 2 weeks, a 9-fold expansion in cell number was observed. The cardiac functional markers sarcomeric α-actinin and connexin 43 were expressed at 13 and 24 days after scaffold formation. HL-1 cells were spontaneously depolarizing 38 days after scaffold formation, which was visualized by confocal microscopy using a calcium-sensitive dye. Electrical stimulation resulted in synchronization of activation peaks throughout the scaffolds. These findings demonstrate that PEG microsphere scaffolds fabricated in the presence of dextran can support the long-term three-dimensional culture of cells, suggesting applications in cardiovascular tissue engineering.

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9-Phenanthrol and flufenamic acid inhibit calcium oscillations in HL-1 mouse cardiomyocytes

Burt¹,

Graves²,

Gao³

et al. 2013

Cell Calcium

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It is well established that intracellular calcium ([Ca2+]i) controls the inotropic state of the myocardium, and evidence mounts that a “Ca2+ clock” controls the chronotropic state of the heart. Recent findings describe a calcium-activated nonselective cation channel (NSCCa) in various cardiac preparations sharing hallmark characteristics of the Transient Receptor Potential Melastatin 4, (TRPM4). TRPM4 is functionally expressed throughout the heart and has been implicated as a NSCCa that mediates membrane depolarization. However, the functional significance of TRPM4 in regards to Ca2+ signaling and its effects on cellular excitability and pacemaker function remains inconclusive. Here, we show by Fura2 Ca-imaging that pharmacological inhibition of TRPM4 in HL-1 mouse cardiac myocytes by 9-phenanthrol (10 μM) and flufenamic acid (10 & 100 μM) decreases Ca2+ oscillations followed by an overall increase in [Ca2+]i. The latter occurs also in HL-1 cells in Ca2+-free solution and after depletion of sarcoplasmic reticulum Ca2+ with thapsigargin (10 μM). These pharmacologic agents also depolarize HL-1 cell mitochondrial membrane potential. Furthermore, by on-cell voltage clamp we show that 9-phenanthrol reversibly inhibits membrane current; by fluorescence immunohistochemistry we demonstrate that HL-1 cells display punctate surface labeling with TRPM4 antibody; and by immunoblotting using this antibody we show these cells express a 130-150 kD protein, as expected for TRPM4. We conclude that 9-phenanthrol inhibits TRPM4 ion channels in HL-1 cells, which in turn decreases Ca2+ oscillations followed by a compensatory increase in [Ca2+]i from an intracellular store other than the sarcoplasmic reticulum. We speculate that the most likely source is the mitochondrion.

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Lipopolysaccharides directly decrease Ca²⁺ oscillations and the hyperpolarization-activated nonselective cation current I_f in immortalized HL-1 cardiomyocytes

Cited by 18 publications

References 29 publications

Lipopolysaccharide prolongs action potential duration in HL-1 mouse cardiomyocytes

Lipopolysaccharide prolongs action potential duration in HL-1 mouse cardiomyocytes

Long-term culture of HL-1 cardiomyocytes in modular poly(ethylene glycol) microsphere-based scaffolds crosslinked in the phase-separated state

9-Phenanthrol and flufenamic acid inhibit calcium oscillations in HL-1 mouse cardiomyocytes

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