Volatile Anesthetic-induced Preconditioning Research Articles

Commentary on: Volatile anesthetic-induced preconditioning

Commentary on: Volatile anesthetic-induced preconditioning

Volatile anesthetic-induced preconditioning

The myocardium has an innate ability to protect itself from ischemic events. This protection occurs when the myocardium is exposed to a brief ischemic period prior to a more extreme ischemic event. This is termed ischemic preconditioning. Ischemic preconditioning induces a series of molecular pathways that protect the cardiac myocyte; first, for a period of 1-6 hours (early preconditioning) and, also, for a second period from 24-72 hours (delayed phase). The early preconditioning is mediated by the release of adenosine which induces a protective signal that is related to the mitochondrial KATP channel activation and activation of the δ-opioid and bradykinin receptors. The delayed phase is related to the induction of inducible nitric oxide synthase, superoxide dismutase and heat-shock proteins. Indirect evidence indicates that O2-derived free radicals are involved in the delayed phase, as noted in the early preconditioning phase. Applying ischemic preconditioning to clinical practice can be dangerous and difficult to implement in a controlled fashion. However, recent studies have shown that the use of volatile anesthetics, such as sevoflurane, isoflurane and desflurane, can mimic the early phase of ischemic preconditioning through a multi-pathway signaling of mitochondrial KATP channels. This important finding can easily be applied to clinical practice for patients undergoing surgery. It can also be significantly important for patients undergoing off-pump cardiac bypass surgery or cardiac bypass surgery where there is no cross-clamp or cardioplegia used where the probability of myocardial ischemia is greatly increased. This report will, therefore, discuss the mechanism, safety and efficacy of volatile anesthetics as inducers of cardiac preconditioning.

Abstract P094: Anesthetic Preconditioning in Human Cardiomyocytes Derived from Type II Diabetic Patient-Induced Pluripotent Stem Cells in a Varying Glucose Environment

Volatile anesthetic-induced preconditioning (APC) has been shown to pharmacologically precondition the myocardium, providing protection from an ischemia-reperfusion injury. However, APC is attenuated or even eliminated in diabetic individuals and in the presence of acute hyperglycemia with the underlying mechanism(s) being unknown. In this study, we used ventricular cardiomyocytes (CMs) differentiated from non-diabetic and type II diabetic-induced pluripotent stem cells (N-iPSCs and DM-iPSCs, respectively) to investigate the influence of a high glucose environment and genetic background on APC. Differentiated CMs were identified using cardiac-specific immunostaining and expression of green fluorescent protein under the transcriptional control of cardiac promoter myosin light chain-2v, a genetic construct delivered by lentiviral vector. N-iPSC- and DM-iPSC-derived CMs were exposed to varying glucose environments (11 mM and 25 mM). Confocal microscopy was utilized to measure mitochondrial membrane potential and mitochondrial permeability transition pore opening (mPTP) in CMs. The volatile anesthetic isoflurane depolarized mitochondria in N-iPSC-derived CMs via opening of the mitochondrial adenosine triphosphate-sensitive potassium channel; however, isoflurane depolarized mitochondria to a significantly lower level in diabetic-derived cardiomyocytes and in the presence of 25 mM glucose. APC delayed mPTP opening in both N-iPSC- and DM-iPSC-derived CMs in 11 mM glucose environment only. We have established an in vitro model based on directed differentiation of ventricular CMs from N-iPSCs and DM-iPSCs that allows us the unique opportunity to conduct comparative studies to address the inability of diabetic individuals to be preconditioned with anesthetics. Our preliminary results indicate for the first time that both a high glucose environment and a diabetic background have detrimental effects on the efficiency of APC to protect CMs from an ischemia-reperfusion injury.

Role of the O-linked β-N -acetylglucosamine in the Cardioprotection Induced by Isoflurane

Cardiac protection by volatile anesthetic-induced preconditioning and ischemic preconditioning have similar signaling pathways. Recently, it was reported that augmentation of protein modified with O-linked β-N-acetylglucosamine (O-GlcNAc) contributes to cardiac protection. This study investigated the role of O-GlcNAc in cardiac protection induced by anesthetic-induced preconditioning. O-GlcNAc-modified proteins were visualized by immunoblotting. Tolerance against ischemia or reperfusion was tested in vivo (n = 8) and in vitro (n = 6). The opening of the mitochondrial permeability transition pore (mPTP) upon oxidative stress was examined in myocytes treated with calcein AM (n = 5). Coimmunoprecipitation and enzymatic labeling were performed to detect the mitochondrial protein responsible for the mPTP opening. Isoflurane treatment and the consequent augmentation of O-GlcNAc concentrations reduced the infarct size (26 ± 5% [mean ± SD], P < 0.001) compared with the control. The protective effect of O-GlcNAc was eliminated in the group pretreated with the O-GlcNAc transferase inhibitor alloxan (39 ± 5%, P < 0.001). Myocyte survival also showed the same result in vitro. Formation of the mPTP was abrogated in the isoflurane-treated cells (86 ± 4%, P < 0.001) compared with the control and alloxan-plus-isoflurane-treated cells (57 ± 7%, P < 0.001). Coimmunoprecipitation and enzymatic labeling studies revealed that the O-GlcNAc-modified, voltage-dependent anion channel restained the mPTP opening. Isoflurane induced O-GlcNAc modification of mitochondrial voltage-dependent anion channel. This modification inhibited the opening of the mPTP and conferred resistance to ischemia-reperfusion stress.

Anesthetic-induced Preconditioning Delays Opening of Mitochondrial Permeability Transition Pore via Protein Kinase C-ϵ–mediated Pathway

Cardioprotection by volatile anesthetic-induced preconditioning (APC) involves activation of protein kinase C (PKC). This study investigated the importance of APC-activated PKC in delaying mitochondrial permeability transition pore (mPTP) opening. Rat ventricular myocytes were exposed to isoflurane in the presence or absence of nonselective PKC inhibitor chelerythrine or isoform-specific inhibitors of PKC-delta (rottlerin) and PKC-epsilon (myristoylated PKC-epsilon V1-2 peptide), and the mPTP opening time was measured by using confocal microscopy. Ca-induced mPTP opening was measured in mitochondria isolated from rats exposed to isoflurane in the presence and absence of chelerythrine or in mitochondria directly treated with isoflurane after isolation. Translocation of PKC-epsilon was assessed in APC and control cardiomyocytes by Western blotting. In cardiomyocytes, APC prolonged time necessary to induce mPTP opening (261 +/- 26 s APC vs. 216 +/- 27 s control; P < 0.05), and chelerythrine abolished this delay to 213 +/- 22 s. The effect of isoflurane was also abolished when PKC-epsilon inhibitor was applied (210 +/- 22 s) but not in the presence of PKC-delta inhibitor (269 +/- 31 s). Western blotting revealed translocation of PKC-epsilon toward mitochondria in APC cells. The Ca concentration required for mPTP opening was significantly higher in mitochondria from APC rats (45 +/- 8 microM x mg control vs. 64 +/- 8 microM x mg APC), and APC effect was reversed with chelerythrine. In contrast, isoflurane did not protect directly treated mitochondria. APC induces delay of mPTP opening through PKC-epsilon mediated inhibition of mPTP opening, but not through PKC-delta. These results point to the connection between cytosolic and mitochondrial components of cardioprotection by isoflurane.

Time Course of Desflurane-induced Preconditioning in Rabbits

The authors tested the hypothesis that volatile anesthetic-induced preconditioning (APC) follows a similar time pattern as that described for ischemic preconditioning and that delayed APC is mediated by nitric oxide. A prospective randomized vehicle-controlled study. A university research laboratory. New Zealand white rabbits (n = 75). Rabbits were instrumented for the measurement of systemic hemodynamics and subjected to a 30-minute coronary artery occlusion (CAO) and 3 hours of reperfusion. Desflurane (1.0 minimum alveolar concentration) was administered for 30 minutes and was discontinued 0.5 hours, 2 hours, 3 hours, 12 hours, 24 hours, 48 hours, 72 hours, and 96 hours before CAO, respectively. In 2 separate experimental groups, the nitric oxide synthase inhibitor N-omega-nitro-L-arginine (L-NA) was administered 72 hours after the administration of 0.0 or 1.0 minimum alveolar concentration of desflurane. The infarct size was determined gravimetrically. Data are mean +/- standard deviation. Desflurane significantly (p < 0.05) reduced the infarct size compared with the control (63% +/- 12%, n = 7) when administered 0.5 hours (35% +/- 5%, n = 7), 2 hours (35% +/- 9%, n = 7), 24 hours (31% +/- 8%, n = 7), 48 hours (30% +/- 11%, n = 6), and 72 hours (39% +/- 5%, n = 6) before CAO. However, when desflurane was administered 3 hours (53% +/- 9%, n = 7), 12 hours (71% +/- 6%, n = 7), or 96 hours (66% +/- 5%, n = 7) before CAO, the myocardial infarct size was not reduced. The second window (72 hours) of preconditioning was abolished by the NOS inhibitor L-NA (52% +/- 16%, n = 7). L-NA alone had no effect on infarct size (64% +/- 11%, n = 7). Desflurane induces a first (0.5-2 hours) and second window of preconditioning (24-72 hours) in the rabbit model of acute myocardial infarction. The second window of APC is mediated by nitric oxide.

Protein kinase C-epsilon primes the cardiac sarcolemmal adenosine triphosphate-sensitive potassium channel to modulation by isoflurane.

Cardioprotection by volatile anesthetic-induced preconditioning is known to involve intracellular signaling pathways. Recent studies have shown that protein kinase C (PKC) plays an important role in anesthetic-induced preconditioning. In this study, the effects of the activation of specific isozymes of PKC, specifically PKC-epsilon and -delta, on the modulation of the sarcolemmal adenosine triphosphate-sensitive potassium (sarcKATP) channel by isoflurane were investigated. The sarcKATP current was measured in ventricular myocytes isolated from guinea pig hearts using the whole cell configuration of the patch clamp technique. Peptides that induced the translocation of specific PKC isozymes were used to activate PKC-epsilon and PKC-delta. Under whole cell conditions, isoflurane alone was unable to elicit the opening of the sarcKATP channel. Pretreatment with the specific PKC-epsilon activator, PP106, primed the sarcKATP channel to open in the presence of isoflurane. The resulting sarcKATP current densities in the presence of 0.88 mm isoflurane were 6.5 +/- 6.0 pA/pF (n = 7) and 40.4 +/- 18.2 pA/pF (n = 7) after pretreatment with 100 and 200 nm PP106, respectively. The PKC-epsilon antagonist PP93 abolished this effect. A scrambled peptide of the PKC-epsilon activator PP105 did not prime the sarcKATP channel. The PKC-delta activator PP114 was significantly less effective in priming the sarcKATP channel. 5-Hydroxydecanoate significantly attenuated the effect of the PKC-epdsilon activator on the sarcKATP channel. In addition, immunohistochemical analysis showed that the PKC-epsilon isoform translocated to both the mitochondria and sarcolemma after anesthetic-induced preconditioning, whereas the PKC-delta isoform translocated to the mitochondria. The PKC-epsilon isozyme primed the sarcKATP channel to open in the presence of isoflurane. The PKC-delta isozyme was significantly less effective in modulating the isoflurane effect on this channel.

Colchicine inhibits isoflurane-induced preconditioning.

When administered before prolonged myocardial ischemia and reperfusion, isoflurane exerts potent cardioprotective effects similar to those inferred by ischemic preconditioning. To determine whether an intact cytoskeleton is critically important in isoflurane-induced preconditioning, the authors used a rabbit model in which isoflurane-induced myocardial preconditioning decreases myocardial infarct size (IS) substantially. In this model, the authors tested whether the microtubule depolymerizing agent, colchicine, would inhibit isoflurane-induced myocardial preconditioning. Myocardial IS was measured in four groups of propofol-anesthetized rabbits, each subjected to 30 min of anterolateral coronary occlusion followed by 3 h of reperfusion. Groups differed only in the pretreatments given, and only the control group received no pretreatment. An isoflurane-preconditioned group was pretreated with 15 min of end-tidal isoflurane, 1.1%, and then 15 min of washout. An isoflurane-plus-colchicine group was administered 2 mg/kg colchicine intravenously before isoflurane pretreatment. A colchicine-control group was administered 2 mg/kg colchicine but no isoflurane pretreatment. Myocardial IS and area at risk (AR) were defined by staining. Data were analyzed by analysis of variance or covariance. Infarct size, expressed as a percentage of AR (IS:AR) was 33.6%+/-8.8% (SD) in the control group. Isoflurane preexposure reduced myocardial IS:AR significantly, to 11.8%+/-9.1%. Colchicine pretreatment eliminated the preconditioning-like effect of isoflurane (IS:AR = 32.6%+/-8.7%). Colchicine alone did not alter IS (IS:AR = 27.6%+/-7.1%; P = not significant). Colchicine abolished the preconditioning effect of isoflurane but did not increase IS when administered alone. An intact microtubular cytoskeleton is critically important in the process of volatile anesthetic-induced preconditioning.