Published online before print July 19, 2007, doi: 10.1161/CIRCRESAHA.107.157271
| |||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
© 2007 American Heart Association, Inc.
Cellular Biology |
Hypersensitivity of BKCa to Ca2+ Sparks Underlies Hyporeactivity of Arterial Smooth Muscle in Shock
From the Departments of Pathophysiology (G.Z., B.P., J.L., X.H., K.-s.Z) and Physiology (G.Z., Y.Z.), Southern Medical University, Guangzhou, China; and Institute of Molecular Medicine (X.Z., C.C., N.H., C.W., H.C.), Peking University, Beijing, China.
Correspondence to Dr Ke-seng Zhao, Southern Medical University, Department of Pathophysiology, Guangzhou 510515, China. E-mail zhaoks1937{at}jhyahoo.com
 |
Abstract |
|---|
 Top Abstract IntroductionMaterials and MethodsResultsDiscussionReferences |
|---|
Large conductance Ca2+-activated K+ channels (BKCa) play a critical role in blood pressure regulation by tuning the vascular smooth muscle tone, and hyposensitivity of BKCa to Ca2+ sparks resulting from its altered ß1 subunit stoichiometry underlies vasoconstriction in animal models of hypertension. Here we demonstrate hypersensitivity of BKCa to Ca2+ sparks that contributes to hypotension and blunted vasoreactivity in acute hemorrhagic shock. In arterial smooth muscle cells under voltage-clamp conditions (0 mV), the amplitude and duration, but not the frequency, of spontaneous transient outward currents of BKCa origin were markedly enhanced in hemorrhagic shock, resulting in a 265% greater hyperpolarizing current. Concomitantly, subsurface Ca2+ spark frequency was either unaltered (at 0 mV) or decreased in hyperpolarized resting cells. Examining the relationship between spark and spontaneous transient outward current amplitudes revealed a hypersensitive BKCa activity to Ca2+ spark in hemorrhagic shock, whereas the spark–spontaneous transient outward current coupling fidelity was near unity in both groups. Importantly, we found an acute upregulation of the ß1 subunit of the channel, and single-channel recording substantiated BKCa hypersensitivity at micromolar Ca2+, which promotes the
and ß1 subunit interaction. Treatment of shock animals with the BKCa inhibitors iberiotoxin and charybdotoxin partially restored vascular membrane potential and vasoreactivity to norepinephrine and blood reinfusion. Thus, the results underscore a dynamic regulation of the BKCa–Ca2+ spark coupling and its therapeutic potential in hemorrhagic shock–associated vascular disorders.
Key Words: Ca2+ sparks • Ca2+-activated K+ channels • vascular reactivity • hemorrhagic shock
|
Introduction |
|---|
|
TopAbstractIntroductionMaterials and MethodsResultsDiscussionReferences |
|---|
Ca2+ sparks are commonly thought to arise from the opening of one, or the coordinated opening of several, ryanodine receptor (RyR) Ca2+ release channels in the sarcoplasmic reticulum (SR) of cardiac,1 skeletal,2 and arterial myocytes.3 In arterial smooth muscle cells (ASMCs), Ca2+ sparks beneath the plasma membrane activate 10 to 100 large conductance Ca2+-activated K+ channels (BKCa) in the adjacent sarcolemma and elicit the discharge of spontaneous transient outward currents (STOCs), resulting in membrane hyperpolarization and, subsequently, dilation of vascular smooth muscle.3,4 Recent studies have revealed that the coupling between sparks and STOCs can continuously regulate, by negative feedback, the vascular tone in intact arteries.5–8 Furthermore, it has been recently shown that the spark–STOC coupling efficiency is altered during hypertension, because of modulation of the subunit stoichiometry of BKCa in vascular smooth muscle.5
Severe shock is well characterized by a sequence of events, including release of endogenous vasoconstrictors, delayed vascular decompensation, and refractory hypotension that is defined as a progressive vasodilatation, as well as a continuous decrease in peripheral vascular resistance resulting in death, despite augmentation of sympathetic nervous activity or treatment with vasoconstrictors.9 Various factors, such as -adrenoceptor desensitization, metabolic acidosis, cytokine release, transcription factor activation, endothelial cell (EC) damage, and overproduction of nitric oxide (NO) have been shown to be involved in this process.10–13 Nevertheless, the mechanisms underlying the low blood pressure in shock are not fully understood, and there are few effective clinical strategies to reverse the refractory hypotension in patients with severe shock.
Vascular hyporeactivity is one of the major causes responsible for the progressive hypotension in severe shock, and membrane hyperpolarization of smooth muscle cells is considered to account for the low vasoreactivity during severe hemorrhagic as well as endotoxic shock.14–16 Given that the Ca2+ spark–STOC coupling constitutes a crucial component in determining ASMC membrane potential,3,4 and tuning the vascular tone,3,5,7 and is likely a contributing factor to the development of high blood pressure in hypertensive rats,5,6,8 we investigated whether BKCa, Ca2+ sparks, and their coupling in ASMCs are altered in acute hemorrhagic shock (HS) and, if so, whether such alterations contribute to the refractory hypotension and vascular hyporeactivity.
|
Materials and Methods |
|---|
|
TopAbstractIntroductionMaterials and MethodsResultsDiscussionReferences |
|---|
All experiments complied with the Chinese National Guidelines for the Use and Care of Experimental Animals and were approved by Southern Medical University. Wistar rats of either sex were anesthetized, and surgery was conducted as described previously.13,17,18 ASMCs from small mesenteric arteries (A2, A3) were isolated as previously described.19,20 Single-channel BKCa currents and membrane potential were measured in the inside-out or cell-attached patch-clamp configurations. ASMC membrane potential was evaluated according to a previously described procedure.21 STOCs were recorded in the perforated whole-cell patch-clamp configuration as described.22 Small mesenteric artery membrane potential was measured as previously described.23 For confocal Ca2+ imaging, isolated ASMCs were loaded with fluo-4 acetoxymethyl ester (5 µmol/L; Molecular Probes) for 15 minutes and then washed 3 times with Hepes-buffered whole-cell solution. A Zeiss LSM-510 laser-scanning confocal microscope was used in Ca2+ spark imaging. Each line-scan image consisted of 1000 scans obtained at 3-ms intervals, and each line comprised 512 pixels spaced at 0.056-µm intervals. In simultaneous recordings of STOCs and Ca2+ sparks, the scan line was set to cross the subsurface just beneath the patch site. For Ca2+ transient measurement, 3000 line scans were obtained at 15-ms intervals. For Western blotting, small mesenteric arteries were taken out and cleaned of connected tissues. Protein was extracted, estimated, separated, and transferred to the membrane using a routine method.24 Immunofluorescence labeling of isolated myocytes was performed using polyclonal antibodies for BKCa
or ß1 subunits5 (1:100; Affinity Bioreagents Inc). Data are presented as means±SEM of n observations. When appropriate, ANOVA, nonparametric test, and Student’s and paired t test were used to determine the significance of the differences among groups. A probability value of <0.05 was considered statistically significant.
An expanded Materials and Methods section is in the online data supplement at http://circres.ahajournals.org.
|
Results |
|---|
|
TopAbstractIntroductionMaterials and MethodsResultsDiscussionReferences |
|---|
ASMC Hyperpolarization and Vascular Hyporeactivity in HS
In the rat HS model, we determined changes of arteriole and ASMC reactivity to the vasoconstrictor and adrenoceptor agonist norepinephrine (NE). As shown in Table I in the online data supplement, the threshold concentrations of NE to induce arteriole and ASMC contraction were 0.16±0.06 (n=6) and 1.01±0.14 (n=8) µg/ml, respectively, for the sham group. However, thresholds were elevated to 2.50±0.13 (n=6, P<0.01) and 60.02±19.71 (n=8, P<0.01) µg/ml in the shock group, indicating a conspicuous decrease of arteriole as well as ASMC reactivity to NE. Because membrane potential (Em) is widely recognized as an important determinant of vascular tone and vasoreactivity,14,15 we hypothesized that a membrane hyperpolarization accompanies and underlies the arteriole and ASMC hyporeactivity during HS. Indeed, we found that ASMC Em was shifted from –41.9±4.3 mV (n=10) in the sham group to –66.2±2.9 mV (n=8, P<0.01) in the HS group, revealing that HS causes a hyperpolarization of ASMCs by as much as 24.3 mV.
HS Altered STOC Activity
Membrane permeability to K+ is among the crucial variables in controlling Em. Activation of BKCa results in membrane hyperpolarization in many types of smooth muscle cells,25–27 and increased K+ channel activity previously has been suggested to be associated with shock.14–16,28 Because of the relatively low sensitivity of BKCa channel to bulk cytosolic resting Ca2+ (
100 nmol/L), genesis of STOCs is usually coupled to dynamic high Ca2+ microdomains established during Ca2+ sparks.6,8,22,29,30 To determine possible roles of BKCa in HS, we first investigated shock-induced alterations of STOCs in ASMCs. Under perforated whole-cell patch-clamp conditions at 0 mV holding potential (Vm), STOCs were detected in sham control cells at a rate of 2.8±0.25 Hz (supplemental Table II). Individual STOCs displayed a peak current of 25.9±0.28 pA (n=9964 events from 88 cells), full duration at half maximum of 18.6±0.20 ms and an average outward charge transfer (Q) of 1112±21.8 fC (Figure 1A and 1B and supplemental Table II). Addition of 100 nmol/L charybdotoxin (ChTX), a BKCa inhibitor, in bath solution decreased STOC amplitude by 36% within 1 minute (supplemental Figure IAb). Iberiotoxin (IbTX) (200 nmol/L), a specific BKCa inhibitor, almost abolished STOCs within 2 minutes (supplemental Figure IAa and IB). STOCs were also partially inhibited by 10 µmol/L ryanodine, an inhibitor of the SR RyR Ca2+ release channel (supplemental Figure IAc). At a higher concentration (100 µmol/L), ryanodine nearly abolished STOCs (84% decrease of the amplitude) (supplemental Figure IAe and IB). Conversely, caffeine at 1 mmol/L augmented STOC activity, with rhythmic bursts of STOC hyperactivity (supplemental Figure IAd and IB). The caffeine effect was abolished in cells pretreated with high concentrations of ryanodine (supplemental Figure IAe and IB). These results indicate that STOCs in mesenteric ASMCs are both RyR and BKCa dependent, consistent with those described in other ASMCs.3
|
Although the frequency-dependent modulation of STOC activity accounts for changes in vasoactivity in many physiological and pathophysiological conditions,3,5–8,30,31 we failed to detect any significant change in the rate of STOC occurrence (2.6±0.18 Hz, n=110 cells, holding potential=0 mV, P>0.05 versus sham) in ASMCs from HS. Nevertheless, a close examination revealed prominent changes in unitary properties of STOCs between the sham and HS groups. As shown in Figure 1A and 1B, amplitude of STOCs in HS was increased by 57% (40.7±0.34 pA, n=18 684 events from 110 HS cells). Concomitantly, the rise time and the decay time of STOCs were prolonged by 53% and 73%, respectively. As a result, the amount of outward charge transfer in a single STOC was enhanced by 265%, reaching 2949±39.0 fC in HS (Figure 1B and supplemental Table II). These observations indicate that, in voltage-clamped cells at 0 mV, individual STOCs with greater amplitude and prolonged duration tend to hyperpolarize ASMCs nearly 3 times as effectively in HS cells as in sham controls.
Enhanced Ca2+ Spark–STOC Coupling Efficiency in Shock
Because STOCs in ASMCs are thought to be directly triggered by Ca2+ sparks,3,4,7 we next performed simultaneous confocal Ca2+ spark imaging and patch-clamp recording of STOCs, which allowed for determination of the coupling efficiency between Ca2+ sparks and STOCs. At a 0-mV holding potential with the scan line placed just beneath the surface of the fluo-4/acetoxymethyl ester–loaded ASMCs (Figure 2A, inset), we found that almost every subsurface spark triggered a detectable STOC in both sham and HS groups (Figure 2A). This indicates that the coupling fidelity between subsurface sparks and STOCs is near unity in mesenteric ASMCs regardless of the shock treatment. The rate of spark occurrence did not differ between the two groups (3.2±0.42 Hz/100 µm, n=31 sham cells versus 2.7±0.31 Hz/100 µm, n=38 HS cells; P>0.05), whereas the spark amplitude was significantly increased in HS (Figure 2B and supplemental Table III). The constancy of spark production suggests that HS does not alter the intrinsic propensity of spark production. As was the case in the indicator-unloaded ASMCs (see supplemental Table II), the STOC frequency was nearly identical in HS and sham control (3.5±0.32 Hz, n=61 sham cells versus 3.3±0.21 Hz, n=63 HS cells), whereas the STOC amplitude was significantly increased (28.1±0.74 pA in sham versus 43.8±1.43 pA in HS, P<0.001). More importantly, linear regression of STOC amplitude as a function of the corresponding spark amplitude uncovered a greater slope factor in HS (184±14 pA/Ca2+ unit in F/F0, n=191 spark–STOC pairs) than in sham control (97±8 pA/Ca2+ unit in F/F0, n=149 pairs, P<0.01) (Figure 2C). This observation indicates that HS confers a hypersensitivity to BKCa activation by Ca2+ sparks.
|
HS Decreased Spontaneous Ca2+ Spark Production in Resting ASMCs
Although the greater STOCs observed in voltage-clamped ASMCs tend to hyperpolarize the cell, membrane hyperpolarization exerts a negative-feedback regulation of the production of spontaneous Ca2+ sparks. As such, Ca2+ spark production in hyperpolarized resting cells from shock animals ought to reflect a new equilibrium between membrane hyperpolarization and intracellular Ca2+ homeostasis.
In resting ASMCs under no voltage clamp, spontaneous Ca2+ sparks were observed in 83% (n=48 cells) and 85% (n=52 cells) of ASMCs during a 6-second period of line scan in the sham and HS groups, respectively. The rate of spark occurrence was 0.9±0.01 Hz/100 µm (n=39 cells) in the sham group (Figure 3A), suggesting that mesenteric ASMC spark activity is considerably higher than that reported in cerebral ASMCs.3 This spark frequency at rest is thus only a small fraction of that at 0 mV, consistent with a role of membrane potential in Ca2+ homeostasis. Interestingly, we detected a 55% decrease in subsurface spark frequency in HS (0.4±0.05, n=43 cells, P<0.001 versus sham) (Figure 3B and 3C). Morphometric measurement of sparks revealed a marginally enhanced spark amplitude (1.32±0.02 F/F0, n=324 events from 39 sham cells, 1.35±0.02, n=203 events from 43 HS cells, P<0.05) and small increases in full width at half maximum (2.0±0.05 µm in sham versus 2.3±0.076 µm in HS, P<0.01) and full duration at half maximum (46.5±3.52 ms in sham versus 50.3±3.79 ms in HS, P<0.01) (Figure 3C and supplemental Table III). Rapid application of 10 mmol/L caffeine elicited similar Ca2+ transients in both groups (Figure 3D), excluding a change in the SR Ca2+ store as the major cause of reduced spark frequency. Furthermore, because the intrinsic propensity of spark production was unaltered, as evidenced in voltage-clamped conditions (Figure 2), the decreased spark production in resting HS versus sham cells should be mainly accountable by the HS-associated membrane hyperpolarization.
|
Single-Channel Characterization of BKCa Ca2+ Sensitivity in HS
BKCa channels in smooth muscle cells consist of a pore-forming subunit and auxiliary ß1 subunit. The subunit possesses intrinsic sensitivity to activation by Ca2+, whereas the presence of ß1 subunit enhances its Ca2+ sensitivity.5,8,32,33 Because ß1 subunit downregulation and altered :ß1 stoichiometry underlie the BKCa hyposensitivity to Ca2+ sparks in a rat model of chronic hypertension,5,8 we sought to determine possible dynamic regulation of ß1 subunit expression in acute HS. To this end, immunocytochemistry data showed that - and ß1-subunit–associated immunofluorescence distributes mainly to the edge of the cell (Figure 4A); the expression of ß1 subunit was significantly elevated in HS (Figure 4B), whereas no significant difference was found in subunit expression between the HS and sham groups (Figure 4A and 4B). The protein expression in intact mesenteric arteries detected with Western blotting confirmed a significant augmentation of ß1 subunit expression without altering the subunit in HS (Figure 4C and 4D). The upregulation of the ß1 subunit of BKCa channel should explain, at least in part, the enhanced BKCa sensitivity to Ca2+ in vitro and in intact arteries.
|
It has been shown that interaction between the and ß1 subunits occurs at micromolar or higher Ca2+ concentrations, with little or no interaction at the resting level of Ca2+.32,33 To determine whether the upregulation of ß1 subunit expression accounts for the enhanced spark–STOC coupling in HS, we sought to examine BKCa activity at variable Ca2+ concentrations at the single channel level. For this purpose, the excised patch-clamp configuration was used to record activity of single BKCa from ASMCs, with symmetric 140 mmol/L K+ on both sides of the patch membrane (Figure 4E). To exclude the contamination of Ba2+, 50 µmol/L crown-ether, (+)-18-crown-6-tetra carboxylic acid (18C6TA) was added in the bath solution before the experiment.34 The identity of BKCa channel currents was confirmed by their ChTX (100 nmol/L) and tetraethylammonium sensitivity (200 µmol/L; data not shown) and by the high single-channel conductance typical of BKCa.21,33,35 Our results showed that, at 1 µmol/L [Ca2+]i, the slope conductance of BKCa was 216±7 pS in the shock group, which was nearly identical to that in sham (Figure 4G). At 1 µmol/L Ca2+, the channel open probability in HS was 1.5 times of that in sham (NPo=1.46±0.11, n=23 in HS versus 1.00±0.14, n=27 in sham, holding potential=20 mV, P<0.05) (Figure 4F), corroborating the aforementioned BKCa hypersensitivity. The fact that such hypersensitivity was absent at submicromolar Ca2+ concentrations (Figure 4E and 4F), when the subunit interaction is essentially negligible, suggests that altered ß1 subunit stoichiometry is likely the major mechanism underlying the HS-associated BKCa hypersensitivity to Ca2+ sparks.
Blocking BKCa Restored Membrane Potential and the Reactivity to NE in the HS Model
To further test the possibilities that activation of BKCa is responsible for HS-induced vascular hyperpolarization and hyporeactivity, mesenteric artery Em and reactivity were measured with either ChTX or IbTX as a BKCa blocker. Consistent with the aforementioned results, membrane was hyperpolarized from –37.5±1.4 mV in sham to –47.9±0.4 mV in HS and IbTX (100 nmol/L) depolarized Em by 7.9±0.7 mV in HS but only 3.3±0.7 mV in sham control (Figure 5A). This greater depolarization induced by IbTX in HS suggests higher HS-associated BKca activity. In whole-animal experiments, neither ChTX nor IbTX exerted any significant effect on mean arterial pressure (MAP) in the sham group (Figure 5B and 5C), demonstrating that basal activity of BKCa is not a major determinant of blood pressure in rats. When ChTX or IbTX was infused into the femoral vein 2 hours after shock, the blockade of BKCa significantly increased MAP (Figure 5D and 5E), consistent with the idea that an enhanced BKCa activity contributes to blood pressure dysregulation in HS. Furthermore, we found that ChTX or IbTX partially restored vasoreactivity both to the vasoconstrictor NE and the blood reinfusion (Figure 5D and 5E). As a control, infusion of physiological saline exerted no significant effects on either MAP or vasoreactivity (Figure 5D and 5E).
|
|
Discussion |
|---|
|
TopAbstractIntroductionMaterials and MethodsResultsDiscussionReferences |
|---|
The present study reports on a comprehensive characterization of possible roles of Ca2+ sparks, STOCs, and spark–STOC coupling in vascular responses to HS and has uncovered several novel and important findings. First, STOCs activity was enhanced in terms of amplitude, duration, and charge transfer, whereas the frequency of STOC occurrence was unchanged in voltage-clamped ASMCs from mesenteric small arteries and decreased in hyperpolarized resting cells in spite of unchanged store Ca2+ load. Second, simultaneous recordings of Ca2+ sparks and the triggered STOCs revealed a significantly enhanced spark–STOC coupling efficiency. Third, BKca ß1 subunit expression was elevated in HS, leading to BKCa hypersensitivity to Ca2+. By single-channel recording of BKCa in excised patches, we demonstrated that an increase of Ca2+ sensitivity occurs at micromolar Ca2+ concentration, which promotes interaction between the
and ß1 subunits of the channel but is absent at lower Ca2+ concentrations when little or no channel subunit interaction is expected. A significant role of BKCa in HS responses was further established by the observation that BKCa inhibition partially alleviates the blood pressure dysregulation and restores the vascular membrane potential and response to NE and the blood infusion. The present data, however, do not support a substantial change in the intrinsic propensity of Ca2+ spark production, and hence a frequency-dependent modulation of Ca2+ sparks in the rat HS model, because the spark frequency remained unchanged under controlled voltage. The reduction of spark frequency in resting cells from the shock group may stem from the fact that HS hyperpolarizes the cells, negatively regulating intracellular Ca2+ homeostasis. Based on these new findings, we propose a model for a mechanistic explanation of vasodilation and vascular hyporeactivity observed in HS (Figure 6). Owing to an HS-induced high expression of the BKCa ß1 subunits, BKCa becomes hypersensitive to high local Ca2+ produced by subsurface Ca2+ sparks, which leads to higher Ca2+ spark–STOC coupling efficiency by virtue of STOC amplitude, duration, and charge transfer. All else being equal, this tends to hyperpolarize the membrane potential, which, in turn, deactivates Ca2+ influx via voltage-operated Ca2+ influxes, leading to vasodilation and hypotension. The membrane hyperpolarization and deactivation of Ca2+ channels also tend to attenuate the ability of various vasoconstrictors such as NE to augment Ca2+ flux, contributing to the hyporeactivity in HS.
|
Amberg et al5 have demonstrated that an altered Ca2+ sensitivity of BKCa is associated with enhanced vascular tone in angiotensin II–induced hypertensive rats. The authors found a decrease in BKCa sensitivity to Ca2+ sparks caused by a decreased expression of the regulatory ß1 subunit of the channel, which seems to be in contrast to the acute HS reported here; both findings underscore an important role of the ß1 subunit in dynamic regulation of BKCa activity and hence the arterial smooth muscle tone. Furthermore, it has also been shown that the spark–STOC coupling can be reversibly enhanced in ASMCs in response to CO application.7 However, studies from Chang et al suggested that BKCa ß1 subunit is highly expressed in mesenteric artery from 12-week-old spontaneously hypertensive rats (SHR),24 whereas Cox et al36 reported that BKCa current is enhanced in SHR mesenteric artery and Wellman et al reported no change of BKCa current in cerebral artery from salt-sensitive hypertensive rat.37 Despite the controversy surrounding BKCa in chronic hypertension, our data implicate that targeting the spark–STOC signaling pathway might provide an effective therapeutic strategy to reverse the fatal prognosis of acute HS in the clinical setting.
Our results suggest that an acute upregulation of the regulatory ß1 subunit confers the BKCa channel hypersensitivity to spark Ca2+ in intact cells and to micromolar Ca2+ in excised patch membranes. Such a mechanism should activate in a time-dependent manner but retain a long memory that is essentially irreversible, so that overt effects are detectable in isolated ASMCs a few hours after the onset of the shock. Besides the alteration of ß1 subunit expression that we report here, previous studies have also shown that NO is abundantly released in sepsis and HS,13,28 and, among others, NO augments BKCa activation by increasing Ca2+ sparks in arterial myocytes.38–40 In this scenario, the NO and cGMP signaling pathway might be responsible for initial BKCa response in HS, whereas long-lasting effects could be retained via nitrosylation and phosphorylation of the channel. Nevertheless, the absence of BKCa hypersensitivity at submicromolar Ca2+ concentrations, as demonstrated in Figure 4F, argues against its contribution independently of the BKCa ß1 subunits (in the late stage of HS). Future studies are warranted to delineate their independent roles and possible synergistic interactions with the ß1 subunit upregulation in HS.
It should be noted that HS is a complex process and many other signaling mechanisms could be altered concurrently with the dysregulation of the Ca2+ spark–STOC coupling reported here. In this regard, it has been shown that ATP concentration decreases in the late stage of HS and that KATP channel activation ensues,41,42 which could also be contributing to membrane hyperpolarization.15,16,43–45 Previous work by us and others has also shown that KATP activation contributes to the large membrane hyperpolarization in ASMCs from shock animals14–16,44,45 and that KATP inhibition by glibenclamide restores the vascular reactivity to some extent.16
In summary, we have demonstrated an enhanced coupling efficiency between Ca2+ sparks and BKCa in ASMCs from shock animals. The increase in BKCa sensitivity to spark Ca2+ stems from an acute upregulation of the regulatory BKCa ß1 subunit. As a result, ASMCs in voltage-clamp conditions display markedly increased STOC amplitude, duration, and charge transfer, contributing to membrane hyperpolarization. Inhibition of BKCa partially reverses blood pressure dysregulation and restores Em and vascular reactivity. These findings in acute HS and hypotension may have important ramifications for the treatment of shock patients in emergency medicine.
|
Acknowledgments |
|---|
Sources of Funding
This work was supported by National Natural Science Foundation of China grants 30070735, 30271268, 30470407, and 30025022; Major State Basic Research Development Program, China, grant G2000057002; and Science Foundation of Guangdong Province, China, grants ZKM04702S and 001060.
Disclosures
None.
|
Footnotes |
|---|
*Both authors contributed equally to this work.
Original received September 14, 2004; first resubmission received November 16, 2006; second resubmission received June 5, 2007; revised second resubmission received June 19, 2007; accepted July 9, 2007.
|
References |
|---|
|
TopAbstractIntroductionMaterials and MethodsResultsDiscussionReferences |
|---|
1. Cheng H, Lederer WJ, Cannell MB. Calcium sparks: elementary events underlying excitation-contraction coupling in heart muscle. Science. 1993; 262: 740–744.
2. Tsugorka A, Rios E, Blatter LA. Imaging elementary events of calcium release in skeletal muscle cells. Science. 1995; 269: 1723–1726.
3. Nelson MT, Cheng H, Rubart M, Santana LF, Bonev AD, Knot HJ, Lederer WJ. Relaxation of arterial smooth muscle by calcium sparks. Science. 1995; 270: 633–637.
4. Jaggar JH, Porter VA, Lederer WJ, Nelson MT. Calcium sparks in smooth muscle. Am J Physiol. 2000; 278: C235–C256.
5. Amberg GC, Bonev AD, Rossow CF, Nelson MT, Santana LF. Modulation of the molecular composition of large conductance, Ca2+ activated K+ channels in vascular smooth muscle during hypertension. J Clin Invest. 2003; 112: 717–724.[CrossRef][Medline] [Order article via Infotrieve]
6. Brenner R, Perez GJ, Bonev AD, Eckman DM, Kosek JC, Wiler SW, Patterson AJ, Nelson MT, Aldrich RW. Vasoregulation by the b1 subunit of the calcium-activated potassium channel. Nature. 2000; 407: 870–876.[CrossRef][Medline] [Order article via Infotrieve]
7. Jaggar JH, Leffler CW, Cheranov SY, Tcheranova D, E S, Cheng X. Carbon monoxide dilates cerebral arterioles by enhancing the coupling of Ca2+ sparks to Ca2+-activated K+ channels. Circ Res. 2002; 91: 610–617.
8. Pluger S, Faulhaber J, Furstenau M, Lohn M, Waldschutz R, Gollasch M, Haller H, Luft FC, Ehmke H, Pongs O. Mice with disrupted BK channel b1 subunit gene feature abnormal Ca2+ spark/STOC coupling and elevated blood pressure. Circ Res. 2000; 87: e53–e60.[Medline] [Order article via Infotrieve]
9. Wurster SH, Wang P, Dean RE, Chaudry IH. Vascular smooth muscle contractile function is impaired during early and late stages of sepsis. J Surg Res. 1994; 56: 556–561.[CrossRef][Medline] [Order article via Infotrieve]
10. Altavilla D, Saitta A, Squadrito G, Galeano M, Venuti SF, Guarini S, Bazzani C, Bertolini A, Caputi AP, Squadrito F. Evidence for a role of nuclear factor-kappaB in acute hypovolemic hemorrhagic shock. Surgery. 2002; 131: 50–58.[CrossRef][Medline] [Order article via Infotrieve]
11. Bannerman DD, Goldblum SE. Direct effects of endotoxin on the endothelium: barrier function and injury. Lab Invest. 1999; 79: 1181–1199.[Medline] [Order article via Infotrieve]
12. Bond RF, Johnson G III. Vascular adrenergic interactions during hemorrhagic shock. Fed Proc. 1985; 44: 281–289.[Medline] [Order article via Infotrieve]
13. Thiemermann C, Szabo C, Mitchell JA, Vane JR. Vascular hyporeactivity to vasoconstrictor agents and hemodynamic decompensation in hemorrhagic shock is mediated by nitric oxide. Proc Natl Acad Sci U S A. 1993; 90: 267–271.
14. Chen SJ, Wu CC, Yang SN, Lin CI, Yen MH. Abnormal activation of K+ channels in aortic smooth muscle of rats with endotoxic shock: electrophysiological and functional evidence. Br J Pharmacol. 2000; 131: 213–222.[CrossRef][Medline] [Order article via Infotrieve]
15. Liu J, Zhao K. The ATP-sensitive K+ channel and membrane potential in the pathogenesis of vascular hyporeactivity in severe hemorrhagic shock. Chin J Traumatol. 2000; 3: 39–44.[Medline] [Order article via Infotrieve]
16. Zhao KS, Huang X, Liu J, Huang Q, Jin C, Jiang Y, Jin J, Zhao G. New approach to treatment of shock-restitution of vasoreactivity. Shock. 2002; 18: 189–192.[CrossRef][Medline] [Order article via Infotrieve]
17. Gray SD. Rat spinotrapezius muscle preparation for microscopic observation of the terminal vascular bed. Microvasc Res. 1973; 5: 395–400.[CrossRef][Medline] [Order article via Infotrieve]
18. Zhao KS, Junker D, Delano FA, Zweifach BW. Microvascular adjustments during irreversible hemorrhagic shock in rat skeletal muscle. Microvasc Res. 1985; 30: 143–153.[CrossRef][Medline] [Order article via Infotrieve]
19. Pan BX, Zhao GL, Huang XL, Zhao KS. Calcium mobilization is required for peroxynitrite-mediated enhancement of spontaneous transient outward currents in arteriolar smooth muscle cells. Free Radic Biol Med. 2004; 37: 823–838.[CrossRef][Medline] [Order article via Infotrieve]
20. Zhao Y, Wu ZH, Zhao GL. [Isolation and physiological characterization of mesenteric arterial smooth muscle cells]. Nan Fang Yi Ke Da Xue Xue Bao. 2006; 26: 954–958.[Medline] [Order article via Infotrieve]
21. Maruyama Y, Petersen OH, Flanagan P, Pearson GT. Quantification of Ca2+-activated K+ channels under hormonal control in pig pancreas acinar cells. Nature. 1983; 305: 228–232.[CrossRef][Medline] [Order article via Infotrieve]
22. Perez GJ, Bonev AD, Patlak JB, Nelson MT. Functional coupling of ryanodine receptors to KCa channels in smooth muscle cells from rat cerebral arteries. J Gen Physiol. 1999; 113: 229–238.
23. Pan BX, Zhao GL, Huang XL, Jin JQ, Zhao KS. Peroxynitrite induces arteriolar smooth muscle cells membrane hyperpolarization with arteriolar hyporeactivity in rats. Life Sci. 2004; 74: 1199–1210.[CrossRef][Medline] [Order article via Infotrieve]
24. Chang T, Wu L, Wang R. Altered expression of BK channel b 1 subunit in vascular tissues from spontaneously hypertensive rats. Am J Hypertens. 2006; 19: 678–685.[CrossRef][Medline] [Order article via Infotrieve]
25. Brayden JE, Nelson MT. Regulation of arterial tone by activation of calcium-dependent potassium channels. Science. 1992; 256: 532–535.
26. Jackson WF. Potassium channels and regulation of the microcirculation. Microcirculation. 1998; 5: 85–90.[CrossRef][Medline] [Order article via Infotrieve]
27. Nelson MT, Quayle JM. Physiological roles and properties of potassium channels in arterial smooth muscle. Am J Physiol. 1995; 268: C799–C822.[Medline] [Order article via Infotrieve]
28. Landry DW, Oliver JA. The ATP-sensitive K+ channel mediates hypotension in endotoxemia and hypoxic lactic acidosis in dog. J Clin Invest. 1992; 89: 2071–2074.[Medline] [Order article via Infotrieve]
29. Wang SQ, Wei C, Zhao G, Brochet DX, Shen J, Song LS, Wang W, Yang D, Cheng H. Imaging microdomain Ca2+ in muscle cells. Circ Res. 2004; 94: 1011–1022.
30. Perez GJ, Bonev AD, Nelson MT. Micromolar Ca2+ from sparks activates Ca2+-sensitive K+ channels in rat cerebral artery smooth muscle. Am J Physiol. 2001; 281: C1769–C1775.
31. Zhuge R, Fogarty KE, Tuft RA, Walsh JV Jr. Spontaneous transient outward currents arise from microdomains where BK channels are exposed to a mean Ca2+ concentration on the order of 10 microM during a Ca2+ spark. J Gen Physiol. 2002; 120: 15–27.
32. Meera P, Wallner M, Jiang Z, Toro L. A calcium switch for the functional coupling between a (hslo) and b subunits (Kv,Ca b) of maxi K channels. FEBS Lett. 1996; 385: 127–128.[CrossRef][Medline] [Order article via Infotrieve]
33. Toro L, Wallner M, Meera P, Tanaka Y. Maxi-KCa, a unique member of the voltage-gated K channel superfamily. News Physiol Sci. 1998; 13: 112–117.
34. Neyton J. A Ba2+ chelator suppresses long shut events in fully activated high-conductance Ca2+-dependent K+ channels. Biophys J. 1996; 71: 220–226.[Medline] [Order article via Infotrieve]
35. Magleby KL, Pallotta BS. Calcium dependence of open and shut interval distributions from calcium-activated potassium channels in cultured rat muscle. J Physiol. 1983; 344: 585–604.
36. Cox RH, Lozinskaya I, Dietz NJ. Calcium exerts a larger regulatory effect on potassium+ channels in small mesenteric artery myocytes from spontaneously hypertensive rats compared to Wistar-Kyoto rats. Am J Hypertens. 2003; 16: 21–27.[CrossRef][Medline] [Order article via Infotrieve]
37. Wellman GC, Cartin L, Eckman DM, Stevenson AS, Saundry CM, Lederer WJ, Nelson MT. Membrane depolarization, elevated Ca2+ entry, and gene expression in cerebral arteries of hypertensive rats. Am J Physiol Heart Circ Physiol. 2001; 281: H2559–H2567.
38. Bolotina VM, Najibi S, Palacino JJ, Pagano PJ, Cohen RA. Nitric oxide directly activates calcium-dependent potassium channels in vascular smooth muscle. Nature. 1994; 368: 850–853.[CrossRef][Medline] [Order article via Infotrieve]
39. Pucovsky V, Gordienko DV, Bolton TB. Effect of nitric oxide donors and noradrenaline on Ca2+ release sites and global intracellular Ca2+ in myocytes from guinea-pig small mesenteric arteries. J Physiol. 2002; 539: 25–39.
40. Tare M, Parkington HC, Coleman HA, Neild TO, Dusting GJ. Hyperpolarization and relaxation of arterial smooth muscle caused by nitric oxide derived from the endothelium. Nature. 1990; 346: 69–71.[CrossRef][Medline] [Order article via Infotrieve]
41. Takano M, Noma A. The ATP-sensitive K+ channel. Prog Neurobiol. 1993; 41: 21–30.[CrossRef][Medline] [Order article via Infotrieve]
42. Wattanasirichaigoon S, Menconi MJ, Delude RL, Fink MP. Effect of mesenteric ischemia and reperfusion or hemorrhagic shock on intestinal mucosal permeability and ATP content in rats. Shock. 1999; 12: 127–133.[Medline] [Order article via Infotrieve]
43. Landry DW, Oliver JA. The pathogenesis of vasodilatory shock. N Engl J Med. 2001; 345: 588–595.
44. Musser JB, Bentley TB, Griffith S, Sharma P, Karaian JE, Mongan PD. Hemorrhagic shock in swine: nitric oxide and potassium sensitive adenosine triphosphate channel activation. Anesthesiology. 2004; 101: 399–408.[CrossRef][Medline] [Order article via Infotrieve]
45. Szabo C, Salzman AL. Inhibition of ATP-activated potassium channels exerts pressor effects and improves survival in a rat model of severe hemorrhagic shock. Shock. 1996; 5: 391–394.[Medline] [Order article via Infotrieve]
Related Article:
Circ. Res. 2007 101: 436-438.
This article has been cited by other articles:
 |
|
 |
|
  R. Tiniakov and K. E. Scrogin The spleen is required for 5-HT1A receptor agonist-mediated increases in mean circulatory filling pressure during hemorrhagic shock in the rat Am J Physiol Regulatory Integrative Comp Physiol, May 1, 2009; 296(5): R1392 - R1401. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 H. Cheng and W. J. Lederer Calcium Sparks Physiol Rev, October 1, 2008; 88(4): 1491 - 1545. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 L. H. Clapp and N. N. Orie Stoking Up BKCa Channels in Hemorrhagic Shock: Which Channel Subunit Is Really Fueling the Fire? Circ. Res., August 31, 2007; 101(5): 436 - 438. [Full Text] [PDF]
|
|
| |||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||