Epoxyeicosatrienoic acids (EET(11,12)) may partially restore endothelium-derived hyperpolarizing factor-mediated function in coronary microarteries.

Background. Endothelial cells derive nitric oxide, prostacyclin, and endothelium-derived hyperpolarizing factor (EDHF). The cytochrome P-450–monooxygenase metabolites of arachidonic acid (epoxyeicosatrienoic acids [EETs]) have been suggested to be EDHF. This study was designed to examine the effect of EET_11,12 with regard to the possibility of restoring EDHF function when added into hyperkalemic cardioplegic solution.

Methods. Porcine coronary microartery rings were studied in a myograph. In groups 1 and 2, paired arteries were incubated in either hyperkalemic solution (K⁺ 20 mmol/L) or Krebs’ solution (control). In group 3, the paired arteries were incubated in hyperkalemia plus EET_11,12 (1 × 10^−6.5 mol/L) or hyperkalemia alone (control) at 37°C for 1 hour, followed by Krebs’ washout and then precontracted with 1 × 10^−8.5 mol/L U46619. The EDHF-mediated relaxation to EET_11,12 (group 1) or bradykinin (groups 2 and 3) was studied in the presence of N^G-nitro-l-arginine, indomethacin, and oxyhemoglobin.

Results. After exposure to hyperkalemia, the EDHF-mediated maximal relaxation by bradykinin (72.5% ± 7.8% versus 41.6% ± 10.6%; p < 0.05), but not by EET_11,12 (18.4% ± 3.3% versus 25.1% ± 4.9%; p > 0.05) was significantly reduced. Incubation with EET_11,12 partially restored EDHF function (33.3% ± 9.5% versus 62.0% ± 8.5%; p < 0.05).

Conclusions. In coronary microarteries, hyperkalemia impairs EDHF-mediated relaxation, and EET_11,12 may partially mimic the EDHF function. Addition of EET_11,12 into cardioplegic solution may partially restore EDHF-mediated function reduced by exposure to hyperkalemia.     

Maintaining Ventilation During Cardiopulmonary Bypass Attenuates Polymorphonuclear Cell Activation and May Reduce Pulmonary Polymorphonuclear Cell Sequestration

Objectives: Endothelium‐derived hyperpolarizing factor (EDHF) plays a key role in vasorelaxation and the cytochrome P450‐monooxygenase metabolites of arachidonic acid epoxyeicosatrienoic acids (EETs), such as EET_11,12, have been suggested to be EDHF in various vasculatures. However, little is known about the role of EET_11,12 in the coronary and pulmonary circulation, especially in microcirculation. The present study was designed to examine the role of EET_11,12 in porcine coronary and pulmonary micro‐arteries. Methods: Porcine coronary and pulmonary micro‐arteries (diameter 200‐450 μm) were studied in a myograph (n = 8 in each group). The artery rings were set at the 90% of the circumference at 100 mm Hg for coronary or 30 mm Hg for pulmonary micro‐arteries, respectively. After precontraction with U₄₆₆₁₉ (?8.2 log M for coronary and ?7.5 log M for pulmonary micro‐arteries), EET_11,12 (?10 ～?6.5 log M) or bradykinin (BK, ?10 ～?6.5 log M)‐induced relaxation was established in the presence of inhibitors for cyclooxygenase (indomethacin, 7 μM), nitric oxide (NO) synthetase (N^G‐nitro‐_L‐arginine, 300 μM), and NO scavenger oxyhemoglobin (20 μM). Results: EET_11,12 induced a dose‐dependent relaxation in coronary micro‐arteries with the maximal relaxation of 18.3 ± 3.3% that was significantly less than the relaxation induced by BK (72.5 ± 7.8%; P < 0.001) . In contrast, in pulmonary micro‐arteries, BK induced a marked relaxation (69.6 ± 6.3%) whereas EET_11,12 did not have any effect. Conclusion: In porcine coronary micro‐arteries, EET_11,12 may partially mimic the action of EDHF whereas in pulmonary arteries, this substance is unlikely involved in the EDHF‐mediated relaxation.     

Effect of 11,12-epoxyeicosatrienoic acid as an additive to St. Thomas' cardioplegia and University of Wisconsin solutions on endothelium-derived hyperpolarizing factor-mediated function in coronary microarteries: influence of temperature and time

BACKGROUND: We examined the effect of 11,12-epoxyeicosatrienoic acid (EET(11,12)) added to St. Thomas' Hospital (ST) solution or University of Wisconsin (UW) solution on endothelium-derived hyperpolarizing factor (EDHF)-mediated relaxation under clinically relevant temperature and exposure time. METHODS: Porcine coronary microarteries (200 to 450 microm) were incubated with Krebs' solution (control), ST with or without EET(11,12) (300 nmol/L) at 22 degrees C for 1 hour as well as at 4 degrees C for 1 or 4 hours, and UW with or without EET(11,12) at 4 degrees C for 4 hours. The EDHF-mediated relaxation was induced by bradykinin (-10 to approximately -6.5 log M) in the precontraction evoked by U(46619) (10 nmol/L) or U(46619) (1 nmol/L) plus endothelin-1 (6 nmol/L). RESULTS: The EDHF-mediated relaxation was reduced after exposure to UW (79.7% +/- 4.6% versus 93.6% +/- 2.8%, p = 0.01) at 4 degrees C for 4 hours. One-hour exposure to ST under 22 degrees C or 4 degrees C decreased the relaxation (75.2% +/- 7.6% versus 96.7% +/- 1.6%, p < 0.05) or the sensitivity to bradykinin (-8.04 +/- 0.15 versus -8.50 +/- 0.20 log M, p < 0.05). The relaxation increased to 86.8% +/- 5.3% by addition of EET(11,12) to ST (1 hour at 22 degrees C, p < 0.05) but was unchanged when added to either ST or UW at 4 degrees C for 1 or 4 hours. CONCLUSIONS: As an additive to ST solution, EET(11,12) may partially restore EDHF-mediated endothelial function under moderate hypothermia but had no significant effect under profound hypothermia when added to either ST or UW solution. Further investigation is necessary to improve the effect.     

Impaired EDHF-Mediated Relaxation in Porcine Pulmonary Micro-Arteries by Cold Storage with University of Wisconsin and Euro-Collins Solutions

Background: Vascular endothelium plays a key role in regulation of vascular tone. Hyperkalemia has been demonstrated to impair the EDHF‐mediated endothelial function in coronary circulation. University of Wisconsin (UW) and Eruo‐collins (EC) solutions are used for organ preservation in transplantation surgery. The potassium concentration in UW or EC solutions is as high as 125 mmol/L or 115 mmol/L, respectively. This study was designed to examine whether hyperkalemia or storage with UW and EC solutions affects the relaxation mediated by EDHF in the porcine pulmonary micro‐arteries. Methods: Porcine pulmonary micro‐artery rings (diameter 200–450 μm) were studied in myograph (n = 8 in each group). After incubation with hyperkalemia (K⁺ 125 mmol/L, at 37° C), UW or EC solutions (at 4° C for 4 hours), EDHF‐mediated relaxation induced by bradykinin (BK, ?10 to ?6.5 log M) in the presence of inhibitors for cyclooxygenase (Indomethacin, 7 μM), nitric oxide synthase (N^G‐nitro‐_L‐arginine, 300 μM), and oxyhemoglobin (20 μM) was compared with control (Krebs' solution) in precontraction with U₄₆₆₁₉ (?7.5 log M). Results: The EDHF‐mediated relaxation to BK was 69.6 ± 6.3% compared with 97.1 ± 1.7% (p= 0.003) in control (no inhibitors). After incubation with hyperkalemia, the relaxation significantly decreased (38.6 ± 3.0% vs. 59.1 ± 7.4%, p= 0.03 ). Storage with UW or EC solutions also significantly decreased the relaxation (49.3 ± 7.3% vs. 65.2 ± 3.5%, p= 0.04 and 51.9 ± 8.4% vs. 60.3 ± 6.1%, p= 0.02 , respectively). Conclusions: In porcine pulmonary micro‐arteries, exposure to hyperkalemia or storage with UW or EC solutions at 4°C for 4 hours impairs the EDHF‐mediated endothelial function. The clinical significance of this effect should be further studied.     

Effect of potassium-channel openers on the release of endothelium-derived hyperpolarizing factor in porcine coronary arteries stored in cold hyperkalemic solution

Hyperkalemic solution is widely used to protect the myocardium during open-heart surgery or to preserve donor hearts during heart or heart/lung transplants. The inhibitory effects of hvperkalemic solution on the release of endothelium-derived hyperpolarizing factor (EDHF) of coronary arteries following deep hypothermic storage (4 degrees C) has been well studied. However, it has not been established whether potassium channel openers have protective effects on the coronary endothelial function after cold storage. This study was designed to examine this. Porcine coronary artery rings were studied in organ baths. Relaxation in response to the EDHF stimulus A23187 (nonreceptor-mediated stimulus calcium ionophore) in thromboxane A2 mimetic U46619 (30 nmol/L)-induced precontraction after incubation with hyperkalemic solution (20 mmol/L) with nicorandil (10 micromol/L) (either at 37 degrees C in the oxygenated organ chamber or at 4 degrees C in a refrigerator for 6 h) was compared with the control. There was significant difference between hyperkalemia group and hyperkalemia with nicorandil group under normothermia (p = .04). The difference was significant in the same solution between normothermia and hypothermia. After incubation in hyperkalemic solution without or with nicorandil, the A23187-induced relaxation was 32.8% +/- 9.1% and 72.6% +/- 16.9%, respectively (N = 8, p < .01). Potassium channel opener can attenuate the inhibitory effect of hyperkalemic solution on the release of EDHF after cold storage.     

Procaine in Cardioplegia: The Effect on EDHF‐Mediated Function in Porcine Coronary Arteries

Abstract Objectives: Hyperkalemia in cardioplegia impairs the endothelium‐derived hyperpolarizing factor (EDHF)‐mediated function. This study examined the effect of procaine in cardioplegia on the EDHF‐mediated response in porcine coronary arteries. Methods: An isometric force study was performed in a myograph. Two rings taken from the same artery (diameter 200–450 μm) were incubated with Krebs solution (group I) or 20 mM K⁺ (group II) with/without procaine (1 mM) at 37°C for 1 hour. The EDHF‐mediated relaxation was induced by bradykinin (BK,‐10 ?‐6.5 log M) after U₄₆₆₁₉ (‐8 log M, in group I) or K⁺‐precontraction (in group II) in the presence of indomethacin (7 μM), N^G‐nitro‐L‐arginine (300 μM), and hemoglobin (20 μM). The membrane potential of a single smooth muscle cell was measured by a microelectrode after superfusion with Krebs solution with/without procaine for 1 hour. Results: The EDHF‐mediated relaxation was increased by the treatment with procaine with the EC₅₀ shifted leftward (97.3 ± 0.6% vs. 83.0 ± 5.1% at‐7 log M and 99.4 ± 0.6% vs. 96.7 ± 1.6% at‐6.5 log M, p < 0.05; EC₅₀.‐8.57 ± 0.24 vs.‐7.92 ± 0.23 log M, p < 0.05). Procaine decreased the BK‐induced hyperpolarization from‐72.3 ± 0.7 mV to‐68.8 ± 0.8 mV (‐6.5 log M, p < 0.01). The EDHF‐mediated relaxation in arteries exposed to 20 mM K⁺ was not altered by procaine (49.9 ± 7.4% vs. 55.8 ± 7.6%, p > 0.05). Conclusions: In the coronary arteries, procaine has a depolarizing effect but it enhances EDHF‐mediated relaxation. Addition of procaine in cardioplegia did not change the EDHF‐mediated endothelial function.     

Effects of l-arginine administration before cardioplegic arrest on ischemia-reperfusion injury

Background. Administration of l-arginine during reperfusion or its addition to cardioplegic solution has been shown to protect myocardium against ischemia-reperfusion injury. This study aimed at evaluating the role of l-arginine in ischemia-reperfusion injury when administered intraperitoneally 24 hours before cardioplegic arrest.Methods. Two groups of Sprague-Dawley rats (control, n = 10; and l-arginine, n = 10) were studied in an isolated buffer-perfused heart model. Both groups were injected intraperitoneally 24 hours before ischemia. Before experimentation blood samples were collected for cardiac troponin I and cGMP analysis. In the coronary effluents, cardiac troponin I, adenosine, cyclic guanosine monophosphate, and nitric oxide metabolites were assayed.Results. Before heart excision, serum cardiac troponin I concentrations were higher in the l-arginine than in the control group (0.037 ± 0.01 versus 0.02 ± 0.05 μg · L⁻¹; p < 0.05). During reperfusion, cardiac troponin I release was lower in the l-arginine than in the control group (0.04 ± 0.01 versus 0.19 ± 0.03 ng · min⁻¹; p < 0.05). The coronary flow as well as the left ventricular developed pressure were higher in the l-arginine than in the control group before ischemia and remained so throughout the experimentation.Conclusions. These results indicate that l-arginine administered intraperitoneally 24 hours before cardioplegic arrest reduced myocardial cell injury and seems to protect myocardium against ischemia-reperfusion injury.     

Comparison of University of Wisconsin and St Thomas' Hospital solutions on endothelium-derived hyperpolarizing factor-mediated function in coronary micro-arteries

BACKGROUND: It is controversial whether coronary endothelial function is impaired after cold exposure to University of Wisconsin (UW) or St. Thomas' Hospital (ST) solution during heart transplantation. We therefore examined the effects of cold storage of coronary micro-arteries with UW or ST solution on endothelium-derived hyperpolarizing factor (EDHF)-mediated function. METHODS: Porcine and human coronary arteries were immersed in either UW or ST solution at 4 degrees C for 4 hr and then normalized in a wire myograph. RESULTS: In the rings (normalized diameter: 200-500 microM) precontracted by U46619, EDHF-mediated relaxation and hyperpolarization were initiated by bradykinin (BK) or A23187 in the presence of indomethacin and NG-nitro-L-arginine. In the human coronary arteries, the EDHF-mediated relaxation to BK was reduced by UW solution from 53.2+/-5.6% to 24.0+/-2.7% (P=0.006). The reduced EDHF-mediated relaxation occurred concurrently with the decreased hyperpolarization to BK (17.0+/-1.5 vs. 10.5+/-1.1 mV, n=10, P=0.004) or A23187 in porcine coronary arteries. In the control arteries, K+ channel blockers, either glybenclamide or tetraethylammonium reduced the EDHF-mediated relaxation. After exposure to UW solution, the EDHF-mediated relaxation was further significantly inhibited. In contrast, ST solution did not affect these responses. CONCLUSIONS: These results show that in coronary micro-arteries, UW, but not ST, solution impairs the EDHF-mediated function and inhibits the Ca2+-activated and ATP-sensitive K+ channels. Our comparative study suggests that ST solution may be superior to UW solution in preserving the EDHF-related endothelial function of coronary micro-arteries.     

Adenosine instead of supranormal potassium in cardioplegic solution preserves endothelium-derived hyperpolarization factor-dependent vasodilation.

OBJECTIVE: We have recently shown that adenosine instead of supranormal potassium in cold crystalloid cardioplegia improves cardioprotection. Studies indicate that hyperkalemia has unfavorable effects on vascular endothelial function. Three pathways have been identified as major vasodilatory pathways: the nitric oxide (NO) pathway, the cyclooxygenase (COX) pathway, and the endothelium-derived hyperpolarization (EDHF) pathway, where the EDHF pathway, in particular, seems susceptible to hyperkalemia. We hypothesized that adenosine cardioplegia improves postcardioplegic endothelial function. METHODS: Sixteen pigs were randomized to receive either cold (6 degrees C) hyperkalemic cardioplegia (n=8) or cardioplegia where hyperkalemia was substituted with 1.2 mM adenosine (n=8). After 1h of cold ischemic arrest, coronary blood flow was monitored for the following 2h. The LAD artery was then explanted, and cylindrical rings were mounted for isometric tension recordings in organ chambers. Vessels were preconstricted with U46610 (Thromboxane A(2) analog) and then bradykinin-mediated relaxation was investigated. To differentiate between the vasodilatory pathways the relaxation was assessed in the absence and presence of inhibitors of the COX (indomethacin), NO (L-NAME+carboxy-PTIO), and EDHF (apamin+charybdotoxin) pathways. RESULTS: Invivo: The adenosine group had, as distinct from the hyperkalemic group, a significantly increased coronary blood flow index 1h after cross-clamp release (from (ml/min/100 g, mean+/-SD) 50.9+/-13.9 to 72.8+/-21.9, p=0.010). The difference was, however, not statistically significant between groups. Invitro: Maximal relaxation without blockers was 27.4+/-10.1% of maximal tension in the adenosine group and 22.2+/-7.5% in the hyperkalemic group. To investigate EDHF-dependent vasodilation the vessel rings were simultaneously treated with indomethacin, L-NAME, and carboxy-PTIO. Maximal relaxation in the hyperkalemic group was then reduced to 47.4+/-17.4% of maximal tension, which was a significant reduction compared to the adenosine group with a maximal relaxation of 20.6+/-8.7% (p=0.028). CONCLUSION: Adenosine instead of supranormal potassium in cold crystalloid cardioplegia increases postcardioplegic myocardial blood flow and preserves EDHF-dependent vasodilation.     

Hyperkalemia Complicating Cardiopulmonary Bypass: Analysis of Risk Factors

This study was undertaken to assess the hyperkalemic effect of several factors, including cardioplegia containing 25 mEq K⁺ per liter. We measured potassium balance at termination of cardiopulmonary bypass (CPB) in each of 20 patients with hyperkalemia (minimum K⁺, 6.0 mEq/L) and 20 patients with normal potassium levels (K⁺ up to 5.5 mEq/L) by subtracting urinary excretion of potassium from the sum of contributions from cardioplegia, hemolysis, and transfusion. The effects of potassium balance, diabetes, blood glucose, catecholamines, and propranolol on the degree of potassium change during CPB were assessed by multiple linear regression.We found no effect of potassium load on potassium change in either the hyperkalemic or normal group. This suggested that derangement of potassium homeostasis was more important than exogenous potassium loading as a hyperkalemic stimulus. Use of catecholamines reduced the potassium change in the hyperkalemic group (p < 0.048), but any effect of propranolol on potassium change was not significant.A striking finding was the frequency of diabetics in the hyperkalemic group (12 of 20 patients) compared with the control group (2 of 20) (p < 0.001). In addition, the preoperative serum glucose level in the hyperkalemic group (mean, 154 ± 75 mg/dl) exceeded that of the normal group (mean, 103 ± 13 mg/dl) (p < 0.001).It is concluded that dangerous hyperkalemia is related to derangements of potassium homeostasis rather than excessive potassium loads, and that the use of cardioplegic solutions containing 25 mEq/L of potassium is safe. Known diabetics or patients noted preoperatively to have abnormal elevations of serum glucose deserve special attention during CPB, as many hyperkalemic patients require pacing or prolongation of bypass to manage the resulting electromechanical disturbance.     

Hyperkalemia in hospitalized patients

Objective: Evaluate the prevalence of hyperkalemia(potassium < 5.5 mmol/l) in hospitalized patientsnot on dialysis, as well as the association ofmedications, impaired renal function and comorbidconditions with hyperkalemia.Design: A retrospective case-control method.Setting: A tertiary care teaching hospital.Patients: Hyperkalemic adults not on dialysis withage and sex matched controls.Interventions: None.Main outcome measures: The use of medicationsassociated with hyperkalemia and renal function usinga calculated creatinine clearance were compared in thehyperkalemic and control groups.Results: 35 adult patients with hyperkalemia who werenot receiving dialysis were identified, with aprevalence in the hospitalized population of 3.3%.The hyperkalemic patients were older than the generalhospital population (p < 0.05). Compared withcontrols, hyperkalemic patients: had a lowercreatinine clearance (p < 0.05), were more likely tobe taking angiotensin-converting enzyme inhibitors (p < 0.05), and had an increased frequency of diabetesmellitus (p < 0.001). All of the control patientssurvived their hospitalization, but the mortality rate in the hyperkalemic group was 17% (p < 0.0001). Noneof the deaths were directly attributable to hyperkalemia.Conclusions: Hyperkalemia is more frequent in olderpatients and is usually mild. Hyperkalemia isassociated with diabetes mellitus, diminished renalfunction and the use of angiotensin-converting enzymeinhibitors. An elevated serum potassium level in ahospitalized patient may be a marker for asignificantly increased risk of death, which is due tounderlying medical problems and is not a consequenceof the hyperkalemia. This revised version was published online in August 2006 with corrections to the Cover Date.     

Synergistic interaction of 3 M KCl-extracted donor antigens (e-HAg) with cyclosporine or cyclosporine/sirolimus for prolongation of rat heart allograft survival

Extracted donor histocompatibility antigens (e-HAg) may potentiate the effects of drugs to protect organ allografts from rejection. We examined the capacity of e-HAg when combined with cyclosporine (CsA) alone, sirolimus (rapamycin, RAPA) alone, or CsA/RAPA combinations to prolong heart allograft survival in rats. Wistar-Furth (WF: RT1^u) rats that received CsA (10 mg/kg/day) by oral gavage for 3 (days 0, 1 and 2) or 7 (days 0, 1, 2, 3, 4, 5 and 6) consecutive days displayed modest prolongation of Brown Norway (BN; RT1ⁿ) heart allograft survival from a mean survival time of 7.2 ± 0.8 days in untreated controls to 12.2 ± 1.1 days and 18.6 ± 2.7 days, respectively (p < 0.01). Although administration on the day of transplantation (day 0) of a single intravenous (i.v.) dose of BN e-HAg (5 mg/kg) failed to affect allograft survival, both three (days 0, 1 and 2) and five (days 0, 1, 2, 3 and 4) injections significantly potentiated the effect of a 3-day course of oral CsA (18.6 ± 1.3 days (p < 0.01) and 20.0 ± 1.4 days (p < 0.01), respectively) and of a 7-day course of oral CsA (25.3 ± 4.4 days (p < 0.05) and 33.5 ± 9.3 days (p < 0.01), respectively). Median-effect analysis confirmed a synergistic interaction between CsA (0.5 mg/kg × 7 days, i.v.) and e-HAg with combination index (CI) values less than 0.7 (CI = 1 shows additive interactions, CI < 1 synergistic, and CI> 1 antagonistic, interactions). In contrast, e-HAg failed to affect the immunosuppressive effect of RAPA. However, e-HAg (5.0 mg/kg × 3 days) significantly potentiated the effects of a 7-day or 14-day course of RAPA (0.01 mg/kg)/CsA (0.5 mg/kg) combination therapy, namely from 26.0 ± 4.8 days with a 7-day treatment of CsA/RAPA alone to 32.6 ± 3.6 days (p < 0.01) and from 28.2 ± 2.7 days with a 14-day course of CsA/RAPA alone to 42.0 ± 4.9 days (p < 0.05), respectively (CI = 0.2–0.5). Thus, e-HAg potentiates the immunosuppressive effects of CsA alone and of the CsA/RAPA combination, but not of sirolimus alone.     

Cellular electrophysiological and mechanical effects of celsior solution on endothelial function in resistance coronary arteries

BACKGROUND: We investigated a relatively new organ preservation (Celsior) solution regarding its effect on the endothelium-derived hyperpolarizing factor (EDHF)-mediated function with comparison to St. Thomas Hospital (ST) solution. METHODS: The EDHF-mediated relaxation was induced by bradykinin (BK, -10 to -6.5 logM) in the presence of inhibitors of nitric oxide and prostacyclin in porcine small resistance coronary arteries, before and after incubation in ST (Group Ia, n=11), Celsior (Group Ib, n=13), or Krebs (Group Ic, control, n=12) at 4 degrees C for 4 hr. The EDHF-mediated hyperpolarization of the membrane potential of smooth muscle cells was measured by microelectrode with simultaneous relaxation after cold storage in ST (IIa, n=7), Celsior (IIb, n=6), or Krebs (IIc, control, n=6), or followed by washout with Krebs (ST: IIIa, n=6, Celsior: IIIb, n=6). RESULTS: The EDHF-mediated relaxation was significantly decreased in Group Ia (56.4+/-7.2% vs. 71.2+/-5.3%, P<0.05) and Ib (44.8+/-4.9% vs. 74.7+/-3.3%, P<0.05) but not in Ic. The sensitivity to BK was also significantly decreased (Ia: -7.51+/-0.14 vs. -7.76+/-0.12 log M, P<0.05; Ib: -7.36+/-0.09 vs. -7.60+/-0.09 logM, P<0.05). The resting membrane potential was depolarized in IIa (-44.3+/-1.9 mV, n=7, P<0.05) and IIb (-33.0+/-2.2 mV, n=6, P<0.05) compared with IIc (-57.1+/-1.5 mV, n=6). The EDHF-mediated hyperpolarization decreased significantly in IIa and IIb (3.4+/-0.3 and 3.0+/-0.2 vs. 6.3+/-0.5 mV, P<0.05) and partially restored in IIIa (5.0+/-0.2 vs. 3.4+/-0.3 mV, P<0.05) and IIIb (4.1+/-0.3 vs. 3.0+/-0.2 mV, P<0.05). CONCLUSIONS: Storage with Celsior and ST solutions reduces the EDHF-mediated endothelial function (hyperpolarization and associated relaxation) in porcine small resistance coronary arteries.     

Hyperkalemia in chronic peritoneal dialysis patients

Background. Chronic peritoneal dialysis (PD) patients often develop hypokalemia but less commonly hyperkalemia.Methods. We explored incidence and mechanisms of hyperkalemia in 779 serum samples from 33 patients on PD for 1 − 59 months. Normal serum potassium concentration was defined as 3.5 − 5.1 meq/l.Results. Mean monthly serum potassium concentrations were normal (except for 1 month), but we observed hypokalemia (<3.5 meq/l) in 5% and hyperkalemia (>5.1 meq/l) in 14% of 779 serum samples. Incidence of hyperkalemia did not change over time on PD: Year 1 (15%), Year 2 (11%), Year 3 (19%), Years 4–5 (22%). Hyperkalemia was mostly modest but occasionally extreme [5.2–5.4 meq/l (55%), 5.5–5.7 meq/l (21%), 5.8–6.0 meq/l (10%), >6.0 meq/l (14%)]. Of 31 patients (2 excluded due to brief PD time), 39% displayed hyperkalemia only, 23% displayed hypokalemia only, and the remainder (38%) displayed both or neither. Comparing hypokalemia-only with hyperkalemia-only patients, we found no difference in potassium chloride therapy, medications interrupting the renin-angiotensin system, small-molecule transport status, and renal urea clearance. We compared biochemical parameters from the hypokalemic and hyperkalemic serum samples and observed lower bicarbonate concentrations, higher creatinine concentrations, and higher urea nitrogen concentrations in the hyperkalemic samples (p < 0.001 for each), without difference in glucose concentrations.Conclusion. We observed hyperkalemia 3 times as frequently as hypokalemia in our PD population. High-potassium diet, PD noncompliance, increased muscle mass, potassium shifts, and/or the daytime period without PD might contribute to hyperkalemia.     

Histidine-tryptophan-ketoglutarate solution maximally preserves endothelium-derived hyperpolarizing factor-mediated function during heart preservation: comparison with University of Wisconsin solution.

BACKGROUND: The University of Wisconsin (UW) solution is used widely in heart preservation but has been demonstrated to be detrimental to the endothelial function. The present study compares the effect of histidine-tryptophan-ketoglutarate (HTK) and UW solutions on endothelium-derived hyperpolarizing factor (EDHF)-mediated function in porcine small coronary arteries. METHODS: An isometric force study was performed in a myograph and the membrane potential of a single smooth muscle cell was measured electrophysiologically. Small coronary arteries (diameter 457 +/- 15 microm) were incubated with UW (n = 8), HTK (n = 7) or Krebs solution (n = 15) at 4 degrees C for 4 hours. After washout, in the presence of indomethacin (Indo; 7 micromol/liter), N(G)-nitro-l-arginine (l-NNA; 300 micromol/liter) and oxyhemoglobin (HbO; 20 micromol/liter), bradykinin (BK; -10 to -6.5 log M)-induced relaxation was compared in U46619 (-8 log M) pre-contraction. EDHF-mediated hyperpolarization was elicited by BK (-6.5 log M) in the presence of Indo, l-NNA and HbO. RESULTS: BK-induced, EDHF-mediated relaxation was reduced from 93.6 +/- 2.8% to 79.7 +/- 4.6% after UW preservation (p = 0.01 by unpaired t-test and p = 0.005 by 2-way analysis of variance [ANOVA]), whereas HTK incubation did not decrease EDHF-mediated relaxation (87.0 +/- 6.5%, p = 0.3 by unpaired t-test and p = 0.6 by 2-way ANOVA, compared with control, and p = 0.001 by 2-way ANOVA, compared with UW). EDHF-mediated hyperpolarization (10.3 +/- 1.6 mV) was attenuated by UW exposure (3.4 +/- 0.6 mV, [p = 0.002] vs control), but not by HTK exposure (8.3 +/- 1.1 mV, [p = 0.3] vs control). CONCLUSIONS: HTK is superior to UW solution in protecting EDHF-mediated endothelial function in porcine small coronary arteries. The present findings supports the use of HTK solution in heart preservation.     

Procaine in cardioplegia: the effect on EDHF-mediated function in porcine coronary arteries

OBJECTIVES: Hyperkalemia in cardioplegia impairs the endothelium-derived hyperpolarizing factor (EDHF)-mediated function. This study examined the effect of procaine in cardioplegia on the EDHF-mediated response in porcine coronary arteries. METHODS: An isometric force study was performed in a myograph. Two rings taken from the same artery (diameter 200-450 microm) were incubated with Krebs solution (group I) or 20 mM K+ (group II) with/without procaine (1 mM) at 37 degrees C for 1 hour. The EDHF-mediated relaxation was induced by bradykinin (BK, -10 approximately -6.5 log M) after U46619 (-8 log M, in group I) or K+-precontraction (in group II) in the presence of indomethacin (7 microM), NG-nitro-L-arginine (300 microM), and hemoglobin (20 microM). The membrane potential of a single smooth muscle cell was measured by a microelectrode after superfusion with Krebs solution with/without procaine for 1 hour. RESULTS: The EDHF-mediated relaxation was increased by the treatment with procaine with the EC50 shifted leftward (97.3 +/- 0.6% vs. 83.0 +/- 5.1% at -7 log M and 99.4 +/- 0.6% vs. 96.7 +/- 1.6% at -6.5 log M, p < 0.05; EC50: -8.57 +/- 0.24 vs. -7.92 +/- 0.23 log M, p < 0.05). Procaine decreased the BK-induced hyperpolarization from -72.3 +/- 0.7 mV to -68.8 +/- 0.8 mV (-6.5 log M, p < 0.01). The EDHF-mediated relaxation in arteries exposed to 20 mM K+ was not altered by procaine (49.9 +/- 7.4% vs. 55.8 +/- 7.6%, p > 0.05). CONCLUSIONS: In the coronary arteries, procaine has a depolarizing effect but it enhances EDHF-mediated relaxation. Addition of procaine in cardioplegia did not change the EDHF-mediated endothelial function.     

The dependency of solute diffusion on molecular weight and shape in intact bone

Solute transport through the bone lacunar–canalicular system (LCS) is essential for osteocyte survival and function, but quantitative data on the diffusivity of various biological molecules in the LCS are scarce. Using our recently developed approach based on fluorescence recovery after photobleaching (FRAP), diffusion coefficients of five exogenous fluorescent tracers (sodium fluorescein, dextran-3k, dextran-10k, parvalbumin, and ovalbumin) were measured in murine tibiae in situ. These tracers were chosen to test the dependency of solute diffusion on molecular weight (376–43,000 Da) and shape (linear vs. globular). Among the five tracers, no fluorescence recovery (and thus mobility) was detected for dextran-10k and the diffusion coefficients (D_LCS) of the other four tracers were 295 ± 46, 128 ± 32, 157 ± 88, 65 ± 21 μm² s^− 1 in the LCS, respectively. Overall, the rate of solute diffusion in the bone LCS showed strong dependency on molecular size and shape. Diffusivity decreased with increasing molecular weight for both linear and globular molecules, with the linear molecules decreasing at a faster rate. Compared with free diffusion (D_free) in aqueous solutions, the relative diffusivities (D_LCS / D_free) of the four tracers were not significantly different for sodium fluorescein, dextran-3k, parvalbumin, and ovalbumin (55.0 ± 8.6%, 68.1 ± 17.0%, 79.7 ± 44.7%, 61.0 ± 19.6%, respectively). This result did not agree with the homogenous molecular sieve model proposed for the osteocytic pericellular matrix structure. Instead, a heterogeneous porous model of the pericellular matrix may account for the observed solute transport in the LCS. In summary, the present study provides quantitative data on diffusion of various nutrients and signaling molecules in the LCS that are important for bone metabolism and mechanotransduction.     

Adenosine A3 pretreatment before cardioplegic arrest attenuates postischemic cardiac dysfunction

Background. The cardioprotective effects of the adenosine A₃ receptor in a cardioplegia model have not been described. We tested the hypothesis that infusion of the A₃ receptor agonist, Cl-IB-MECA (100 nM), as a pretreatment (PTx) and/or as a cardioplegic (CP) additive reduces postischemic myocardial injury.Methods. Isolated perfused rat hearts underwent 30 minutes of normothermic ischemia, 60 minutes of intermittent hypothermic cardioplegia (10°C), followed by 2 hours of reperfusion. Hearts were divided into four groups: (1) no pretreatment (PTx) and unsupplemented cardioplegia (CP) (control), (2) Cl-IB-MECA PTx and unsupplemented CP (A₃-PTx), (3) no PTx and Cl-IB-MECA CP (A₃-CP), or (4) Cl-IB-MECA PTx and Cl-IB-MECA CP (A₃-[PTx+CP]).Results. Coronary flow was not increased after A₃ pretreatment when compared to baseline values. After 2 hours of reperfusion, left ventricular developed pressure in control and A₃-CP groups was depressed to 43% ± 3% and 47% ± 2% of baseline; while A₃-PTx and A₃-[PTx+CP] significantly increased left ventricular developed pressure (65% ± 3% and 61% ± 5%) from baseline relative to control and A₃-CP. Effluent creatine kinase activity was significantly decreased by A₃-PTx (1520 ± 32 IU/L), A₃-[PTx+CP] (1481 ± 41 IU/L) from control (1734 ± 54 IU/L) and A₃-CP (1750 ± 43 IU/L). Myocardial edema (% tissue water) was significantly less in A₃-PTx (78 ± 0.6%) and A₃-[PTx+CP] (76% ± 2%) compared with control (85% ± 0.4%) and A₃-CP (83% ± 2%).Conclusions. Adenosine A₃ receptor stimulation as a pretreatment attenuates postischemic cardiodynamic dysfunction and creatine kinase release but has no cardioprotection as an adjunct to cold cardioplegia.     

Aprikalim reduces the Na-Ca exchange outward current enhanced by hyperkalemia in rat ventricular myocytes

Bacground. Aprikalim, an adenosine triphosphate (ATP) sensitive K⁺ (K_ATP) channel opener, attenuates the elevation of intracellular Ca²⁺ concentration ([Ca²⁺]_i) and improves the contractile functions after hyperkalemic and hypothermic cardioplegia. There is evidence that cardioplegia increases the Na⁺-Ca²⁺ exchange activity without affecting Ca²⁺ influx through L-type Ca²⁺ channels or Ca²⁺ content in the sarcoplasmic reticulum, the intracellular Ca²⁺ store.Methods. We measured the Na⁺-Ca²⁺ exchange outward current with the patch-clamp technique in single rat ventricular myocytes exposed to hyperkalemia and hypothermia in the presence of aprikalim. The intracellular calcium concentration ([Ca²⁺]_i) during cardioplegia, and the contractile function and [Ca²⁺]_i transients induced by electrical stimulation or caffeine during rewarming and reperfusion in single ventricular myocytes were also determined. Contraction and [Ca²⁺]_i were determined with video tracking and spectrofluorometry, respectively.Results. Aprikalim, 100 μmol/L, the effect of which was blocked by glibamclamide, a K_ATP inhibitor, significantly attenuated the hyperkalemia-elevated Na⁺-Ca²⁺ exchange current by 26% and 11% at 22°C and 4°C, respectively. Aprikalim also attenuated significantly the [Ca²⁺]_i elevated during cardioplegia. Furthermore aprikalim significantly attenuated the reduction in amplitude and prolongation in duration of contraction of myocytes after cardioplegia. The effects of aprikalim mimicked those of nickle (Ni²⁺), a Na⁺-Ca²⁺ exchange blocker. The electrically or caffeine-induced [Ca²⁺]_i transients were unaltered by cardioplegia or aprikalim.Conclusions. Aprikalim attenuates the Na⁺-Ca²⁺ exchange outward current elevated by hyperkalemia, which may attenuate the [Ca²⁺]_i elevation during hyperkalemia and improve the contractile function after cardioplegia in the ventricular myocyte. The study provides further support that addition of a K_ATP channel opener to the cardioplegic solution may produce beneficial effects in open heart surgery.