Functional Coupling of TRPV4, IK, and SK Channels Contributes to Ca2+-Dependent Endothelial Injury in Rodent Lung

ABSTRACTGlutamatergic NMDA receptors (NMDAR) and small conductance Ca2+-activated K+ channels (SK) are critical synaptic and intrinsic mechanisms that regulate the activity of hypothalamic magnocellular neurosecretory neurons (MNNs) under physiological and pathological states, including lactation and heart failure (HF). Still, whether NMDARs and SK channels in MNNs are functionally coupled, and whether changes in this coupling contribute to exacerbated neuronal activity during HF is at present unknown. In the present study, we addressed these questions using patch-clamp electrophysiology and confocal Ca2+ imaging in a rat model of ischaemic HF. We found that in MNNs of sham rats, blockade of SK channels with apamin (200 nM) significantly increased the magnitude of an NMDAR-evoked current (INMDA). We also observed that blockade of SK channels potentiated NMDAR-evoked firing, and abolished spike frequency adaptation in MNNs from sham, but not HF rats. Importantly, a larger INMDA-ΔCa2+response was observed under basal conditions in HF compared to sham rats. Finally, we found that dialyzing recorded cells with the Ca2+ chelator BAPTA (10 mM) increased the magnitude of INMDA in MNNs from both sham and HF rats, and occluded the effects of apamin in the former. Together, our studies demonstrate that in MNNs, NMDARs and SK channels are functionally coupled, forming a local negative feedback loop that restrains the excitatory effect evoked by NMDAR activation. Moreover, our studies also support a blunted NMDAR-SK channel coupling in MNNs of HF rats, standing thus as a pathophysiological mechanism contributing to exacerbated hypothalamic neuronal activity during this prevalent neurogenic cardiovascular disease.

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Functional coupling between NMDA receptors and SK channels in rat hypothalamic magnocellular neurons: altered mechanisms during heart failure

The Journal of Physiology ◽

10.1113/jp278910 ◽

2019 ◽

Author(s):

Hildebrando C. Ferreira‐Neto ◽

Javier E. Stern

Keyword(s):

Heart Failure ◽

Nmda Receptors ◽

Functional Coupling ◽

Sk Channels ◽

Magnocellular Neurons

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Abstract #418: Effect of Testosterone (T) Replacement with a T-Gel or a New Oral T Formulation (Rextoro) on Selected Biomarkers of Endothelial Injury and Immune Response to Lipids

Endocrine Practice ◽

10.1016/s1530-891x(20)42633-3 ◽

2015 ◽

Vol 21 ◽

pp. 93

Author(s):

Merrell Magelli ◽

Ronald Swerdloff ◽

John Amory ◽

Gregory Flippo ◽

Wael Salameh ◽

...

Keyword(s):

Immune Response ◽

Endothelial Injury

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Differential Role of Components of the Fibrinolytic System in the Formation and Removal of Thrombus Induced by Endothelial Injury

Thrombosis and Haemostasis ◽

10.1055/s-0037-1614532 ◽

1999 ◽

Vol 81 (04) ◽

pp. 601-604 ◽

Cited By ~ 55

Author(s):

Hiroyuki Matsuno ◽

Osamu Kozawa ◽

Masayuki Niwa ◽

Shigeru Ueshima ◽

Osamu Matsuo ◽

...

Keyword(s):

Plasminogen Activator ◽

Thrombus Formation ◽

Endothelial Injury ◽

Fibrinolytic System ◽

Tissue Type ◽

Clot Lysis ◽

Wild Type ◽

Type Plasminogen Activator ◽

Pai 1

SummaryThe role of fibrinolytic system components in thrombus formation and removal in vivo was investigated in groups of six mice deficient in urokinase-type plasminogen activator (u-PA), tissue-type plasminogen activator (t-PA), or plasminogen activator inhibitor-1 (PAI-1) (u-PA-/-, t-PA-/- or PAI-1-/-, respectively) or of their wild type controls (u-PA+/+, t-PA+/+ or PAI-1+/+). Thrombus was induced in the murine carotid artery by endothelial injury using the photochemical reaction between rose bengal and green light (540 nm). Blood flow was continuously monitored for 90 min on day 0 and for 20 min on days 1, 2 and 3. The times to occlusion after the initiation of endothelial injury in u-PA+/+, t-PA+/+ or PAI-1+/+ mice were 9.4 ± 1.3, 9.8 ± 1.1 or 9.7 ± 1.6 min, respectively. u-PA-/- and t-PA-/- mice were indistinguishable from controls, whereas that of PAI-1-/- mice were significantly prolonged (18.4 ± 3.7 min). Occlusion persisted for the initial 90 min observation period in 10 of 18 wild type mice and was followed by cyclic reflow and reocclusion in the remaining 8 mice. At day 1, persistent occlusion was observed in 1 wild type mouse, 8 mice had cyclic reflow and reocclusion and 9 mice had persistent reflow. At day 2, all injured arteries had persistent reflow. Persistent occlusion for 90 min on day 0 was observed in 3 u-PA-/-, in all t-PA-/- mice at day 1 and in 2 of the t-PA-/-mice at day 2 (p <0.01 versus wild type mice). Persistent patency was observed in all PAI-1-/- mice at day 1 and in 5 of the 6 u-PA-/- mice at day 2 (both p <0.05 versus wild type mice). In conclusion, t-PA increases the rate of clot lysis after endothelial injury, PAI-1 reduces the time to occlusion and delays clot lysis, whereas u-PA has little effect on thrombus formation and spontaneous lysis.

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Protective Effect of Vitamin E on Immune Triggered, Granulocyte Mediated Endothelial Injury

Thrombosis and Haemostasis ◽

10.1055/s-0038-1661027 ◽

1984 ◽

Vol 51 (01) ◽

pp. 089-092 ◽

Cited By ~ 24

Author(s):

M A Boogaerts ◽

J Van de Broeck ◽

H Deckmyn ◽

C Roelant ◽

J Vermylen ◽

...

Keyword(s):

Vitamin E ◽

Protective Effect ◽

Umbilical Vein ◽

Endothelial Damage ◽

Endothelial Injury ◽

Mediated Effects ◽

Anti Oxidant ◽

Endothelial Cell Cultures ◽

Vitro Model

SummaryThe effect of alfa-tocopherol on the cell-cell interactions at the vessel wall were studied, using an in vitro model of human umbilical vein endothelial cell cultures (HUEC). Immune triggered granulocytes (PMN) will adhere to and damage HUEC and platelets enhance this PMN mediated endothelial injury. When HUEC are cultured in the presence of vitamin E, 51Cr-leakage induced by complement stimulated PMN is significantly decreased and the enhanced cytotoxicity by platelets is completely abolished (p <0.001).The inhibition of PMN induced endothelial injury is directly correlated to a diminished adherence of PMN to vitamin E- cultured HUEC (p <0.001), which may be mediated by an increase of both basal and stimulated endogenous prostacyclin (PGI2) from alfa-tocopherol-treated HUEC (p <0.025). The vitamin E-effect is abolished by incubation of HUEC with the irreversible cyclo-oxygenase inhibitor, acetylsalicylic acid, but the addition of exogenous PGI2 could not reproduce the vitamin E-mediated effects.We conclude that vitamin E exerts a protective effect on immune triggered endothelial damage, partly by increasing the endogenous anti-oxidant potential, partly by modulating intrinsic endothelial prostaglandin production. The failure to reproduce vitamin E-protection by exogenously added PGI2 may suggest additional, not yet elucidated vitamin E-effects on endothelial metabolism.

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Site-Specific Protein Covalent Attachment to Nanotubes and Its Electronic Impact on Single Molecule Function

10.26434/chemrxiv.7798973.v1 ◽

2019 ◽

Author(s):

Adam Beachey ◽

Harley Worthy ◽

William David Jamieson ◽

Suzanne Thomas ◽

Benjamin Bowen ◽

...

Keyword(s):

Single Molecule ◽

Fluorescent Protein ◽

Functional Integration ◽

Attachment Site ◽

Covalent Attachment ◽

Great Promise ◽

Functional Coupling ◽

Phenyl Azide ◽

Electronic Impact ◽

Protein Attachment

<p>Functional integration of proteins with carbon-based nanomaterials such as nanotubes holds great promise in emerging electronic and optoelectronic applications. Control over protein attachment poses a major challenge for consistent and useful device fabrication, especially when utilizing single/few molecule properties. Here, we exploit genetically encoded phenyl azide photochemistry to define the direct covalent attachment of three different proteins, including the fluorescent protein GFP, to carbon nanotube side walls. Single molecule fluorescence revealed that on attachment to SWCNTs GFP’s fluorescence changed in terms of intensity and improved resistance to photobleaching; essentially GFP is fluorescent for much longer on attachment. The site of attachment proved important in terms of electronic impact on GFP function, with the attachment site furthest from the functional center having the larger effect on fluorescence. Our approach provides a versatile and general method for generating intimate protein-CNT hybrid bioconjugates. It can be potentially applied easily to any protein of choice; attachment position and thus interface characteristics with the CNT can easily be changed by simply placing the phenyl azide chemistry at different residues by gene mutagenesis. Thus, our approach will allow consistent construction and modulate functional coupling through changing the protein attachment position.</p>

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