Neri M. Cohen

Stretch-activated Channel Blockers Modulate Cell Volume in Cardiac Ventricular Myocytes

Stretch-activated channels (SAC) are postulated to regulate cell volume. While this hypothesis is appealing, direct evidence is lacking. Using digital video microscopy, we found that pharmacological blockade of SACs alters the cell volume of isolated rabbit ventricular myocytes during hypoosmotic stress. Under control conditions, relative cell volume increased from 1.0 to 1.311 +/- 0.019 after 10 min in 195 mosmol/l solution. The cation SAC blocker gadolinium (Gd3+; 10 microM) reduced the amount of swelling in hypoosmotic solution by 24% and induced a regulatory volume decrease otherwise not observed. In contrast, the anion SAC blocker 9-anthracene carboxylic acid (9-AC; 1 mM) increased swelling by 44% under the same conditions. Based on the direction of SAC currents, Gd3+ and 9-AC are expected to have opposite effects on cell volume. Furthermore, Gd3+ and 9-AC changed cell volume by only approximately 2% in isosmotic solutions when SACs are expected to be closed. This supports the idea that Gd3+ and 9-AC affect stretch-activated transport processes. In contrast, omitting bath Ca2+ did not alter cell volume under iso- or hypoosmotic conditions suggesting stretch-activated Ca2+ influx is not important in setting cell volume. Not all channels can affect cell volume. Opening ATP-sensitive K+ channels with aprikalim (100 microM) or blocking them with glibenclamide (1 microM) did not alter cell volume under isosmotic or hypoosmotic conditions. These data support the idea that SACs are involved in cardiac cell volume regulation.

Hyperpolarized cardiac arrest with a potassium-channel opener, aprikalim.

Cardioplegic solutions that arrest the heart at or near the resting membrane potential may provide better myocardial protection than standard depolarizing hyperkalemic cardioplegia by reducing both metabolic demand and harmful transmembrane ion fluxes. This hypothesis was investigated in an isolated, blood-perfused, rabbit heart Langendorff model during 30 minutes of normothermic global ischemia. Hyperpolarized cardiac arrest induced by aprikalim, an opener of adenosine triphosphate-dependent potassium channels, was compared with hyperkalemic depolarized arrest and with unprotected global ischemia. Left ventricular pressure was recorded over a wide range of balloon volumes before ischemia and 30 minutes after reperfusion. End-diastolic pressure versus balloon volume data were fitted to a two-coefficient exponential relationship. Changes in the diastolic compliance of the left ventricle were assessed by comparison of preischemic and postischemic coefficients within each cardioplegia group. Postischemic recovery of developed pressure was used to assess changes in left ventricular systolic function. The tissue water content of each heart was also determined. Myocardial protection with aprikalim resulted in better postischemic recovery of developed pressure (90% +/- 9%) than either protection with hyperkalemic cardioplegia (73% +/- 11%) or no protection (62% +/- 9%). Myocardial tissue water content in hearts protected with hyperkalemic cardioplegia (77.4% +/- 1.4%) was less than the tissue water content of either unprotected hearts (79.4% +/- 1.2%) or hearts protected with aprikalim (78.7% +/- 0.9%). Despite these differences, neither hyperkalemic cardioplegia (p = 0.15) nor aprikalim cardioplegia (p = 0.30) was associated with a significant postischemic decrease in ventricular compliance. By contrast, unprotected global ischemia was associated with a significant decrease in ventricular compliance (p < 0.001).

Is there an alternative to potassium arrest

BACKGROUND Mounting clinical and experimental evidence suggests that postoperative myocardial dysfunction is a frequent consequence of surgical global ischemia and reperfusion, despite our modern techniques of myocardial protection. The ubiquitous use of hyperkalemic depolarizing solutions in all forms of cardioplegia may be partly responsible for this phenomenon because of the known ongoing metabolic processes and damaging transmembrane ionic fluxes that occur at depolarized membrane potentials. Cardiac arrest at hyperpolarized potentials, the natural resting state of the heart, may avoid the shortcomings of depolarized arrest and provide an alternative means of myocardial protection. METHODS An adenosine triphosphate-sensitive potassium channel opener, aprikalim, was used to induce hyperpolarized arrest. Aprikalim was able to produce sustained and reproducible electromechanical arrest that was reversible by reperfusion. RESULTS In isolated heart models, when compared with depolarized hyperkalemic arrest, hyperpolarized arrest afforded better protection from global normothermic ischemia and resulted in better postischemic recovery of function upon reperfusion. Preliminary studies in a porcine cardiopulmonary bypass model also have revealed that hyperpolarized arrest can be achieved in a model more closely approximating the clinical setting, and can effectively protect the heart during normothermic surgical global ischemia. CONCLUSIONS Hyperpolarized cardiac arrest may offer an effective alternative to traditional potassium arrest.

Hyperpolarized Arrest Attenuates Myocardial Stunning Following Global Surgical Ischemia: An Alternative to Traditional Hyperkalemic Cardioplegia?

There is clinical evidence that myocardial stunning is a frequent sequela of surgical global ischemia, despite our modern techniques of myocardial protection. The ubiquitous usage of hyperkalemic depolarizing solutions in all forms of cardloplegia may be partly responsible for this phenomenon because of the known ongoing metabolic requirements and damaging transmembrane ionic fluxes that occur at depolarized membrane potentials. Cardiac arrest at hyperpolarized potentials, the natural resting state of the heart, may avoid the shortcomings of depolarized arrest and provide an alternative means of myocardial protection. To test this hypothesis, a potassium channel opener, aprikalim, was used to induce hyperpolarized arrest in an isolated rabbit heart model. Aprikalim was able to produce sustained and reproducible electromechanical arrest that was reversible by reperfusion. When compared with depolarized hyperkalemic arrest, hyperpolarized arrest afforded better protection after short 20‐minute periods of global ischemia and resulted in less myocardial stunning. Moreover, aprikalim was able to significantly prolong the time to ischemic contracture and improve functional recovery after the onset of ischemic contracture when compared with either traditional hyperkalemic cardioplegia or no cardloplegia at all. There was a dose dependence to the protective effect of aprikalim. Preliminary studies in the intact porcine cardiopulmonary bypass model also have revealed that hyperpolarized arrest can effectively protect the heart during surgical global ischemia.

Electrophysiologic consequences of hyperkalemic cardioplegia during surgical ischemia

Myocardial protection strategies use cardioplegic solutions to reduce the injury induced by surgical ischemia and reperfusion. However, there is a high incidence of electrophysiologic abnormalities after cardioplegic arrest. A computerized epicardial mapping system in a porcine cardiopulmonary bypass model was used to measure the electrophysiologic consequences of different myocardial protection techniques. Both warm and cold, crystalloid and blood cardioplegic solutions were compared. The effects of hypothermia and prolonged cardiopulmonary bypass were examined in a control group that underwent a 2-hour period of hypothermia without cardioplegia or aortic cross-clamping, followed by 2 hours of normothermic reperfusion. Isochronous activation maps, unipolar electrograms, ventricular refractory periods, and pacing thresholds were measured before cardioplegic arrest and during reperfusion. Compared with the control group, crystalloid cardioplegia, but not blood cardioplegia, was accompanied by large changes in the pattern of ventricular activation and by persistent (> 2 hours) and significant slowing of the time required for complete ventricular activation. This was not the result of hypoxia. Moreover, the effective refractory period and the pacing threshold were unchanged by any cardioplegia. Our data suggest that crystalloid cardioplegia increases myocardial resistance to current flow leading to a derangement of electrical impulse propagation that may underlie arrhythmogenesis.

Electrophysiological Consequences of Hypothermic Hyperkalemic Elective Cardiac Arrest

Abstract While the development of pharmacological cardioplegic solutions for myocardial protection during cardiopulmonary bypass (CPB) have significantly lengthened the safe operating time for cardiac surgical procedures, the introduction of hypothermic hyperkalemic cardioplegia (CPG) has markedly increased the incidence of postoperative arrhythmias and conduction abnormalities. Using a customized modification of a computerized mapping system, we have developed a large animal porcine model of CPB that is exquisitely sensitive to the electrophysiological (EP) derangements imposed by ischemia and cardiac arrest. This model is able to measure spatial and temporal parameters of ventricular activation with high resolution, using an array of up to 84 epicardial electrodes that can be reproducibly placed on the surface of the heart utilizing known epicardial anatomical markers (e.g., coronary arteries). With this system we have measured the spectrum of clinically observed EP disturbances caused by CPG, from slowed intraventricular conduction to complete heart block. Compared to the control group of hypothermia alone, 2 hours of crystalloid CPG arrest had a significant slowing effect on ventricular activation (p < 0.05). CPG was accompanied, in each animal, by profound changes in the spatial distribution of ventricular activation and persistent slowing of ventricular activation. Traditional EP parameters of effective refractory period and pacing threshold were unchanged by CPG. Smaller temporal and spatial changes were observed in the control group, but were always reversed by 90 minutes of warm reperfusion. We conclude that CPG induces injury of the specialized conducting system and, to a lesser degree, the myocardium. This model will afford us the opportunity to test new methods of CPG to further improve myocardial preservation during CPB.

Warm ischemia lung protection with pinacidil: an ATP regulated potassium channel opener.

BACKGROUND Ischemia/reperfusion injury remains a limiting factor in lung transplantation. Traditional hyperkalemic preservation solutions are associated with a host of metabolic derangements. ATP-regulated potassium channel openers (PCOs) may provide an attractive alternative to traditional solutions by utilizing inherent mechanisms of ischemic preconditioning. The purpose of this study was to assess warm ischemia graft protection with pinacidil, a nonspecific PCO. METHODS An isolated recirculating blood perfused ventilated rabbit lung model was used (n = 15). No ischemia control lungs underwent immediate reperfusion (n = 5). Warm ischemia control lungs were flushed with lactated Ringers (LR), stored at 37 degrees C for 2.5 hours and then reperfused for 2 hours (n = 5). PCO protected lungs were flushed with LR + 100 micromol/L pinacidil, stored, and then reperfused (n = 5). Intermittent blood gases were taken from the pulmonary artery and left atria. Every 30 minutes, graft function was assessed with a 10-minute 100% fractional inspired oxygen concentration challenge to measure maximal gas exchange. Lung samples were graded for histologic injury and assayed for myeloperoxidase activity. RESULTS A mixed-models repeated measures ANOVA demonstrated a significant difference between groups. Tukeys honestly significant difference multiple comparison test demonstrated significantly improved graft function and reduced histologic injury with pinacidil protection compared with the warm ischemia controls. There was no significant difference in graft function or pathology grade between the pinacidil protected lungs and the no ischemia controls. A similar trend, although not significant, was seen in myeloperoxdiase activity. CONCLUSIONS Potassium channel openers with pinacidil can provide pulmonary protection against warm ischemia reperfusion injury.

The Effect of Left Ventricular Intracavitary Volume on the Unipolar Electrogram

In order to examine the effects of ventricular distention on the unipolar electrogram (UEG), an isolated rabbit heart modified Langendorff preparation was utilized. Left ventricular (LV) volume was adjusted using ionically permeable (PB = 9 hearts) or ionically impermeable balloons (IB = 4 hearts). LV UEGs, LV end‐diastolic pressure (EDP), and LV minor axis dimension (MAD), as measured by ultrasonic transducers, were recorded. Three hundred twenty‐five eiectrograms were digitized and analyzed with customdesigned software, In the PB group, a significant inverse linear relationship was found between UEG amplitude and changes in MAD (P < 0.0001). For each animal, this relationship had an R value > 0.8 and a P value < 0.0001. There was also a significant inverse linear relationship between UEG slope and changes in MAD (P < 0.01). UEG amplitude and slope also exhibited a significant inverse relationship to changes in LV EDP, which were best described by a third order polynomial function. In the IB group, no significont relationship was found between either UEG amplitude or slope and MAD or EDP. In this study, intracavitary volume exerted a profound and significant influence on UEG amplitude and slope. This effect was due to increases in conductive intraventricular volume and not to myocardial stretch

Novel protection strategy for pulmonary transplantation.

BACKGROUND Ischemia-reperfusion injury continues to represent a significant challenge to successful lung transplantation. Traditional pulmonary ischemic protection is performed using hypothermic hyperkalemic depolarizing solutions to reduce the metabolic demands of the ischemic organ. Measures to further reduce the effects of ischemic injury have focused on the reperfusion period. We tested the hypothesis that novel physiologic hyperpolarizing solutions-using ATP-dependent potassium channel (K(ATP)) openers-given at the induction of ischemia, will reduce cellular injury and provide superior graft function even after prolonged periods of ischemia. METHODS An isolated blood-perfused ventilated rabbit lung model was used to study lung injury. Airway, left atrial, and pulmonary artery pressures were measured continuously during the 2-h reperfusion period. Oxygenation, as a surrogate of graft function, was measured using intermittent blood gas analysis of paired left atrial and pulmonary artery blood samples. Graft function was measured by oxygen challenge technique (F(i)O(2) = 1.0). Wet-to-dry ratio was measured at the conclusion of the 2-h reperfusion period. Control (Group I) lungs were perfused with modified Euro-Collins solution (depolarizing) and reperfused immediately (no ischemia). Traditional protection lungs were perfused with modified Euro-Collins flush solution and stored for 4 h (Group II) or 18 h (Group III) at 4 degrees C before reperfusion. Novel protection (Group IV) lungs were protected with a hyperpolarizing solution containing 100 nM Aprikalim, a specific K(ATP) channel opener, added to the modified Euro-Collins flush solution and underwent 18 h of ischemic storage at 4 degrees C before reperfusion. RESULTS Profound graft failure was measured after 18 h of ischemic storage with traditional protection strategies (Group III). Graft function was preserved by protection with hyperpolarizing solutions even for prolonged ischemic periods (Group IV). Wet-to-dry weight ratio, airway, left atrial, and pulmonary artery pressures were not significantly different between the groups. CONCLUSIONS We have created a model of predictable lung injury. Membrane hyperpolarization with a K(ATP) channel opener (PCO) provides superior prolonged protection from ischemia-reperfusion injury in an in vitro model of pulmonary transplantation.