Hyperpolarization contributes to vascular hyporeactivity in rats with lipopolysaccharide-induced endotoxic shock

doi:10.1016/S0024-3205(00)00978-4

Life Sciences

Volume 68, Issue 6, 29 December 2000, Pages 659-668

https://doi.org/10.1016/S0024-3205(00)00978-4 Get rights and content

Abstract

We have examined the role of membrane hyperpolarization in mediating vascular hyporeactivity induced by bacterial lipopolysaccharide (LPS) in endothelial-denuded strips of rat thoracic aorta ex vivo. The injection of rats with LPS caused a significant fall of blood pressure and a severe vascular hyporeactivity to norepinephrine. The membrane potential recording showed that endotoxemia caused a hyperpolarization when compared to the control. This hyperpolarization was fully restored by methylene blue (MB; 10 μM) and partially reversed by N^ω-nitro-L-arginine methyl ester (L-NAME; 0.3 mM), 1H-[1,2,4]oxadiazolo[4,3-a]quinoxalin-1-one (ODQ; 1 μM), tetraethylammonium (TEA; 10 mM), charybdotoxin (CTX; 0.1 μM), or glibenclamide (GB; 10 μM), however, this hyperpolarization was not significantly affected by apamin (0.1 μM), 4-aminopyridine (4-AP; 1 mM), or Ba²+ (50 μM). In addition, the basal tension of the tissues obtained from endotoxemic rats was enhanced by the following order: MB ≥ ODQ > TEA ≥ L-NAME ≥ CTX > GB; whereas apamin, 4-AP or Ba²+ had no significant effects on these tissues. In contrast, none of these inhibitors had significant effects on the membrane potential or the basal tension in control tissues. Our electrophysiological results further confirmed previous studies showing that in addition to nitric oxide, the large conductance Ca²+-activated K⁺-channels and ATP-sensitive K⁺-channels are, most likely, responsible for endotoxin-mediated hyporeactivity to vasoconstrictor agents in vascular smooth muscle.

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2015, Life Sciences
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Sepsis is a life threatening condition that is characterized by the loss of vascular reactivity. The factor(s) responsible for the diminished vascular function seen in sepsis are not well understood. The purpose of this study was to characterize the vascular dysfunction from the rat cecal inoculum (CI) sepsis model using cecal ligation and puncture (CLP), and lipopolysaccharide (LPS) sepsis as reference models.
Experiments were performed on isolated aorta from CI, CLP and LPS treated rats using a combination of pharmacological approaches.
Phenylephrine (PE)-induced aortic contraction was significantly decreased in each model (p < 0.05) and not normalized by L-NAME or indomethacin. The vascular response elicited in the CI model for acetylcholine (Ach) was more similar to that seen in the CLP than the LPS model. The removal of the endothelial layer increased sensitivity to L-NAME (p < 0.05) in aortae from CI group. Inhibition of the large conductance Ca^2 +/voltage sensitive K⁺ (BK_Ca) channel did not normalize PE hyporesponsiveness but did abolish sepsis-induced contractile oscillation. Inhibition of the voltage dependent Kv_1.5 channel was not able to reverse the vascular hyporesponsiveness, however, inhibition of the ATP dependent (K_ATP) channel inhibition partially restored the contractile response (p < 0.05). Elevation of VCAM expression and aortic structural alternation were observed in each model.
These results suggest that the CI model may be an additional tool that could be used to investigate the mechanisms of vascular hyporesponsiveness in sepsis.
Role of non-MLC20 phosphorylation pathway in the regulation of vascular reactivity during shock
2014, Journal of Surgical Research
Citation Excerpt :
Elucidating the mechanism for vascular hyporesponsiveness has appreciable implications for the treatment of these critical conditions. Previous studies have demonstrated that the mechanisms for the vascular hyporeactivity after shock are related to receptor desensitization, membrane hyperpolarization of vascular smooth muscle cells (VSMCs), and calcium desensitization [4–6]. These mechanisms are all involved in the inhibition of 20-kDa myosin light chain (MLC20) phosphorylation.
Studies have shown that shock-induced vascular hyporeactivity is associated with the decrease in 20-kDa myosin light chain (MLC20) phosphorylation. Whether and how a non-MLC20 phosphorylation pathway participates in the regulation of vascular reactivity after shock is not known.
With superior mesentery artery (SMA) obtained from rats in hemorrhagic shock and hypoxia-treated SMA, the regulatory effect of platelet-derived growth factor (PDGF) on vascular reactivity and the roles of caldesmon, 27-kDa heat shock protein (HSP27), extracellular signal–regulated protein kinase (Erk), and p38 mitogen–activated protein kinase (MAPK), the main molecules that are involved in the non-MLC20 phosphorylation pathway of the regulation of smooth-muscle contraction, were investigated.
PDGF (40–100 ng/mL) increased the vascular reactivity after shock in a dose-dependent manner, whereas it did not increase the MLC20 phosphorylation in a dose-dependent manner. PDGF with concentration more than 60 ng/mL did not further increase the MLC20 phosphorylation, whereas upregulated the phosphorylation of HSP27, Erk, and p38MAPK, and the activity of myosin adenosine triphosphatase in SMAs, and downregulated the phosphorylation of caldesmon. p38MAPK antagonist, SB203580, not only antagonized PDGF-induced increase in the phosphorylation of HSP27, but also antagonized PDGF-induced decrease in the phosphorylation of caldesmon, whereas Erk antagonist, PD98059, only antagonized PDGF-induced decrease in the phosphorylation of caldesmon.
These findings suggested that a non-MLC20 phosphorylation pathway participated in the regulation of vascular reactivity after shock. Caldesmon- and HSP27-mediated change in myosin adenosine triphosphatase activity and Erk and p38MAPK played an important role in this process. These findings may provide some potential targets for the treatment of vascular hyporeactivity after shock.
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Vasopressin has myriad end-organ effects that are mediated through several different receptors. Endotoxins diminish the contractile response of arteries to norepinephrine in vitro.40 Vasopressin is capable of reversing this diminished responsiveness through the V1 receptor.41
Sepsis brings about neuroendocrine dysfunction in children that differs significantly from that of adults and can thus be difficult to interpret and manage. Aggressive treatment of sepsis with appropriate and judicious use of antibiotics remains a top priority. Strict glycemic control in children has been associated with significant risk of hypoglycemia, which may independently contribute to morbidity and mortality. Timely initiation of hydrocortisone in persistently hypotensive children with fluid-refractory, catecholamine-resistant shock is controversial, but its use in children with suspected or proven adrenal insufficiency is suggested. Fluid and electrolyte abnormalities must be corrected. Treatment of thyroid dysfunction has been shown to be beneficial in certain specific populations but cannot be extrapolated to all septic patients with the current available data.
Role of adenosine A2A receptor in organ-specific vascular reactivity following hemorrhagic shock in rats
2013, Journal of Surgical Research
Citation Excerpt :
Research has shown that vascular reactivity is significantly decreased following severe trauma, shock, or sepsis. This vascular hyporeactivity interferes with the development, therapy, and outcome of these critical illnesses, especially with the application of vasoactive agents [1–7]. Many studies have focused on single blood vessels such as the aorta or the pulmonary, renal, or mesenteric artery [5–10], and a few studies have identified different responses to vasoactive agents among different vessels.
Previous studies have demonstrated differences among organs in terms of shock-induced vascular reactivity and a role for adenosine A2A receptors (A2ARs) in protection against ischemia/reperfusion injury. However, the contributions of A2ARs to organ-specific vascular reactivity and the protection of vascular responsiveness following shock are currently unknown.
We investigated the role of A2ARs in different arteries, including the left femoral artery (LFA), thoracic aorta (TA), superior mesenteric artery (SMA), right renal artery (RRA), pulmonary artery (PA), and middle cerebral artery (MCA), in hemorrhagic-shock rats.
The vascular reactivities of the LFA, SMA, RRA, and MCA increased slightly during early shock and then gradually decreased, whereas those of the PA and TA decreased from the start of shock. Different blood vessels lost vascular reactivity at different rates compared with controls; the LFA had the highest rate of loss (64.51%), followed by the SMA (44.69%), TA (36.06%), PA (37.83%), and RRA (32.33%), whereas the MCA had the lowest rate (18.45%). The rate of loss of vascular reactivity in the different vessels was negatively correlated with A2AR expression levels in normal and shock conditions. The highly selective A2AR agonist CGS 21680 significantly improved vascular reactivity, hemodynamic parameters, and animal survival, whereas the specific antagonist SCH58261 further decreased the shock-induced reduction in vascular reactivity and hemodynamic parameters.
A2ARs are involved in the regulation and protection of vascular reactivity following shock. A2AR activation may have a beneficial effect on hemorrhagic shock by improving vascular reactivity and hemodynamic parameters.
Glibenclamide reduces proinflammatory cytokines in an ex vivo model of human endotoxinaemia under hypoxaemic conditions
2011, Life Sciences
In vivo application of the K_ATP-channel blocker glibenclamide can reverse endotoxin-induced hypotension, vascular hyporeactivity and shock in experimental animals. The hypothesis of the present study is, that the drug effects might not only be based on direct inhibition of K_ATP-channels of vascular smooth muscle cells, but might also reflect reduction of shock-induced excess proinflammatory cytokines and procoagulatory molecules produced in the blood monocytes.
Human whole blood (normoxaemic or hypoxaemic) supplemented ex vivo with 100 ng/ml LPS was used to assess glibenclamide (3–100 μM) effects on IL-1beta, IL-6, TNF-alpha, tissue factor, and plasminogen-activator-inhibitor-2 (PAI-2). Co-incubations with monocytes and erythrocytes and cytosolic calcium measurements were performed to reveal their purinergic intercellular interaction.
In heparinized blood, glibenclamide reduced LPS-induced release of IL-1beta and TNF-alpha, tissue factor and PAI-2 mRNA in a concentration-dependent manner. When samples were subjected to strong hypoxemia using 95% N₂/5% CO₂, these parameters became even more sensitive to the drug. No drug effect was observable in citrated blood or in isolated monocytes. IL-1beta mRNA inhibition by glibenclamide appeared to be dependent on P2X7-receptor activation of monocytes by ATP-releasing erythrocytes during hypoxia. Cytosolic calcium values as well as the duration of calcium transients elicited by P2X7-receptor stimulation in isolated monocytes were strongly increased during hypoxia, both of which could be abolished by glibenclamide.
We conclude that the anti-inflammatory effect of glibenclamide is mainly based on the reduction of calcium entry by drug-induced depolarization of hypoxic monocytes. Thus, glibenclamide possesses a potentially beneficial shock-specific anti-inflammatory action.
Role of V<inf>1a</inf> Receptor in AVP-Induced Restoration of Vascular Hyporeactivity and Its Relationship to MLCP-MLC<inf>20</inf> Phosphorylation Pathway
2010, Journal of Surgical Research
Citation Excerpt :
Studies have demonstrated that vascular hyporeactivity plays an important role in the incidence, development, and the outcome of shock and interferes with the application of vasoactive agents in the therapy of shock. Vascular hyporeactivity may be related to the functional disorder of the multiple receptors and ion channels in VSMC, the hyperpolarization of cell membrane [24], or calcium desensitization of VSMC [13]. Our previous study showed that AVP significantly restored the decreased vascular reactivity and calcium sensitivity of VSMC following hemorrhagic shock [5].
Our previous study showed that small dosage of arginine vasopressin (AVP) had beneficial effect on shock by improving the shock-induced vascular hyporeactivity. But the mechanism is not clear. The objective of the present study is to investigate the role of V_1a and V₂ receptors in AVP-mediated restoration of hemorrhage-induced vascular hyporesponsiveness and its relationship to protein kinase C (PKC), myosin light chain phosphatase (MLCP), and myosin light chain (MLC₂₀) phosphorylation signal transduction pathway.
We used isolated superior mesenteric artery (SMA) from hemorrhagic shock rats (40 mmHg for 2hours) and 90minutes hypoxia-treated vascular smooth muscle cell (VSMC) to study the effects of V_1a and V₂ receptor inhibitors on AVP-mediated regulation of vascular reactivity and calcium sensitivity. Meanwhile the effects of AVP and V_1a and V₂ receptor antagonists on the activity of MLCP and myosin light chain kinase (MLCK), and the phosphorylation of MLC₂₀, and the expression of PKC-α, δ, and ε isoforms were also studied.
AVP significantly improved the reactivity of SMA to norepinephrine (NE) and Ca²⁺ following hemorrhagic shock and increased the hypoxia-induced decrease of contractile response of VSMC to NE. The PKC-α and ε expression in particulate fractions of VSMC following hemorrhagic shock and hypoxia was also increased by treatment with AVP. V_1a receptor inhibitor significantly antagonized these effects of AVP. AVP treatment resulted in a significant increase of MLC₂₀ phosphorylation in SMA and a significant decrease of MLCP activity in VSMC, which was also inhibited by V_1a receptor inhibitor.
V_1a receptor, not V₂ receptor, played an important role in AVP-mediated regulation of vascular reactivity and calcium sensitivity following hemorrhagic shock. The mechanism is mainly through PKC-MLCP-MLC₂₀ phosphorylation calcium sensitivity pathway.

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Original articlesHyperpolarization contributes to vascular hyporeactivity in rats with lipopolysaccharide-induced endotoxic shock

Abstract

Original articles
Hyperpolarization contributes to vascular hyporeactivity in rats with lipopolysaccharide-induced endotoxic shock