RESUMEN
BACKGROUND AND PURPOSE: Inhibition of bradykinin metabolizing enzymes (BMEs) can cause acute angioedema, as demonstrated in a recent clinical trial in patients administered the antihypertensive, omapatrilat. However, the relative contribution of specific BMEs to this effect is unclear and confounded by the lack of a predictive pre-clinical model of angioedema. EXPERIMENTAL APPROACH: Rats were instrumented to record blood pressure and heart rate; inhibitors were infused for 35 min and bradykinin was infused during the last 5 min to elicit hypotension, as a functional marker of circulating bradykinin and relative angioedema risk. KEY RESULTS: In the presence of omapatrilat bradykinin produced dose-dependent hypotension, an effect abolished by B(2) blockade. In the presence of lisinopril (ACE inhibitor), but not candoxatril (NEP inhibitor) or apstatin (APP inhibitor), bradykinin also elicited hypotension. Lisinopril-mediated hypotension was unchanged with concomitant blockade of NEP or NEP/DPPIV (candoxatril+A-899301). However, hypotension was enhanced upon concomitant blockade of APP and further intensified in the presence of NEP inhibition to values not different from omapatrilat alone. CONCLUSIONS AND IMPLICATIONS: We demonstrated that bradykinin is degraded in vivo with an enzyme rank-efficacy of ACE>APP>>NEP or DPPIV. These results suggest the effects of omapatrilat are mediated by inhibition of three BMEs, ACE/APP/NEP. However, dual inhibition of ACE/NEP or ACE/NEP/DPPIV elicits no increased risk of angioedema compared to ACE inhibition alone. Thus, novel BME inhibitors must display no activity against APP to avoid angioedema risk due to high prevalence of ACE inhibitor therapy in patients with diabetes and cardiovascular disease.
Asunto(s)
Angioedema/etiología , Bradiquinina/metabolismo , Inhibidores Enzimáticos/farmacología , Hipotensión/etiología , Aminopeptidasas/antagonistas & inhibidores , Inhibidores de la Enzima Convertidora de Angiotensina/administración & dosificación , Inhibidores de la Enzima Convertidora de Angiotensina/farmacología , Animales , Bradiquinina/efectos de los fármacos , Relación Dosis-Respuesta a Droga , Evaluación Preclínica de Medicamentos/métodos , Indanos/farmacología , Lisinopril/farmacología , Masculino , Neprilisina/antagonistas & inhibidores , Péptidos/farmacología , Propionatos/farmacología , Piridinas/administración & dosificación , Piridinas/farmacología , Ratas , Ratas Sprague-Dawley , Tiazepinas/administración & dosificación , Tiazepinas/farmacologíaRESUMEN
Platelets are relatively short-lived, anucleated cells that are essential for proper hemostasis. The regulation of platelet survival in the circulation remains poorly understood. The process of platelet activation and senescence in vivo is associated with processes similar to those observed during apoptosis in nucleated cells, including loss of mitochondrial membrane potential, caspase activation, phosphatidylserine (PS) externalization, and cell shrinkage. ABT-737, a potent antagonist of Bcl-2, Bcl-X(L), and Bcl-w, induces apoptosis in nucleated cells dependent on these proteins for survival. In vivo, ABT-737 induces a reduction of circulating platelets that is maintained during drug therapy, followed by recovery to normal levels within several days after treatment cessation. Whole body scintography utilizing ([111])Indium-labeled platelets in dogs shows that ABT-737-induced platelet clearance is primarily mediated by the liver. In vitro, ABT-737 treatment leads to activation of key apoptotic processes including cytochrome c release, caspase-3 activation, and PS externalization in isolated platelets. Despite these changes, ABT-737 is ineffective in promoting platelet activation as measured by granule release markers and platelet aggregation. Taken together, these data suggest that ABT-737 induces an apoptosis-like response in platelets that is distinct from platelet activation and results in enhanced clearance in vivo by the reticuloendothelial system.
Asunto(s)
Plaquetas/citología , Plaquetas/metabolismo , Proteínas Proto-Oncogénicas c-bcl-2/metabolismo , Animales , Apoptosis/efectos de los fármacos , Compuestos de Bifenilo/farmacología , Plaquetas/efectos de los fármacos , Separación Celular , Supervivencia Celular/efectos de los fármacos , Gránulos Citoplasmáticos/metabolismo , Perros , Relación Dosis-Respuesta a Droga , Exocitosis/efectos de los fármacos , Citometría de Flujo , Humanos , Hígado/efectos de los fármacos , Hígado/metabolismo , Masculino , Nitrofenoles/farmacología , Fosfatidilserinas/metabolismo , Piperazinas/farmacología , Agregación Plaquetaria/efectos de los fármacos , Recuento de Plaquetas , Sulfonamidas/farmacologíaRESUMEN
We investigated the possibility that opioids activate a tyrosine kinase (TK) that mediates cardioprotection in an in vivo rat model of myocardial infarction. All animals underwent 30 min of regional ischemia and 2 h of reperfusion. Infarct size was expressed as a percentage of the area at risk (IS/AAR). Control animals had an IS/AAR of 58.2 +/- 0.6. Cardioprotection was induced with the delta1- or delta1/delta2-selective opioid agonists, TAN-67, or D-Ala D-Leu enkephalin (DADLE). Both significantly reduced IS/AAR (28.8 +/- 3.6 and 34.8 +/- 3.8, respectively). The general TK inhibitor, genistein, abolished cardioprotection produced by TAN-67 or DADLE (59.1 +/- 3.2 and 61.5 +/- 3.4, respectively), whereas the structural analog, daidzein, lacking TK inhibitory activity, did not. Interestingly, the selective Src/epidermal growth factor (EGF) receptor TK inhibitor, lavendustin A, did not abolish TAN-67-induced cardioprotection (22.1 +/- 6.8). Similarly, the Src-selective TK antagonist, PP2, had no effect on DADLE-induced cardioprotection (31.1 +/- 7.3). These unexpected findings suggest that Src and EGF receptor TKs are not important in the genesis of cardioprotection produced by TAN-67. Finally, we demonstrate that genistein did not affect protein kinase C (PKC) translocation induced by TAN-67. These data suggest that a TK, but most likely not an Src/EGF receptor TK, is important in cardioprotection via opioid receptor stimulation and that the pathway for TK activation is downstream from or parallel to PKC activation in the in situ rat heart since genistein could not affect PKC translocation of selective isoforms induced by TAN-67 and assessed by immunohistochemistry.
Asunto(s)
Genes src/fisiología , Daño por Reperfusión Miocárdica/prevención & control , Proteínas Tirosina Quinasas/fisiología , Receptores Opioides delta/efectos de los fármacos , Transducción de Señal/fisiología , Animales , Técnica del Anticuerpo Fluorescente , Hemodinámica/efectos de los fármacos , Hemodinámica/fisiología , Inmunohistoquímica , Isoenzimas/metabolismo , Infarto del Miocardio/patología , Infarto del Miocardio/prevención & control , Daño por Reperfusión Miocárdica/patología , Proteína Quinasa C/metabolismo , Conejos , Ratas , Ratas Wistar , Translocación Genética/genéticaRESUMEN
Stress-activated protein kinases may be essential to cardioprotection. We assessed the role of p38 in an in vivo rat model of ischemia-reperfusion. Ischemic preconditioning (IPC) and the delta(1)-opioid receptor agonist 2-methyl-4aalpha-(3-hydroxyphenyl)-1,2,3,4,4a,5,12,12aalpha-octahydroquinolino [2,3,3-g]isoquinoline (TAN-67) significantly reduced infarct size (IS), expressed as a percentage of the area at risk (AAR), versus animals subjected only to 30 min of ischemia and 2 h of reperfusion (7.1 +/- 1.5 and 29.6 +/- 3.3 vs. 59.7 +/- 1.6%). The p38 antagonist SB-203580 attenuated IPC when it was administered before (34.0 +/- 6.9%) or after (25.0 +/- 3.8%) the IPC stimulus; however, it did not significantly attenuate TAN-67-induced cardioprotection (39.6 +/- 3.2). We also assessed the phosphorylation of p38 and c-jun NH(2)-terminal kinase (JNK) throughout ischemia-reperfusion in nuclear and cytosolic fractions. After either intervention, no increase was detected in the phosphorylation state of either enzyme in the nuclear fraction or for p38 in the cytosolic fraction versus control hearts. However, there was a robust increase in JNK activity in the cytosolic fraction immediately on reperfusion that was more pronounced in animals subjected to IPC or administered TAN-67. These data suggest that SB-203580 likely attenuates IPC via the inhibition of kinases other than p38, which may include JNK. The data also suggest that activation of JNK during early reperfusion may be an important component of cardioprotection.
Asunto(s)
Citoprotección , Precondicionamiento Isquémico Miocárdico , Proteínas Quinasas Activadas por Mitógenos/metabolismo , Isquemia Miocárdica/fisiopatología , Miocardio/metabolismo , Animales , Vasos Coronarios/fisiopatología , Vasos Coronarios/cirugía , Citoprotección/efectos de los fármacos , Inhibidores Enzimáticos/farmacología , Hemodinámica/efectos de los fármacos , Imidazoles/farmacología , Proteínas Quinasas JNK Activadas por Mitógenos , Masculino , Proteínas Quinasas Activadas por Mitógenos/antagonistas & inhibidores , Isquemia Miocárdica/patología , Miocardio/patología , Fosforilación/efectos de los fármacos , Piridinas/farmacología , Quinolinas/farmacología , Ratas , Ratas Wistar , Receptores Opioides delta/agonistas , Daño por Reperfusión/patología , Daño por Reperfusión/prevención & control , Proteínas Quinasas p38 Activadas por MitógenosRESUMEN
Ceramide has been shown to be a key signaling molecule involved in the apoptotic effect of tumor necrosis factor alpha (TNF-alpha) and other cytokines. Given the importance of cytokines such as TNF-alpha in myocardial ischemia-reperfusion injury, we hypothesize that ceramide is increased during ischemia or reperfusion, and that the activity of enzymes responsible for its production or breakdown should be increased and/or decreased, respectively. Therefore, in the present study, we characterized the enzymatic activities responsible for ceramide production and metabolism in the myocardium of rats, and determined the contribution of these enzymes to altered ceramide levels during myocardial ischemia and reperfusion. The basal ceramide concentration in the myocardium of rats was 34.0 pmol/mg tissue. As determined by the conversion of 14C-sphingomyelin into ceramide and 14C-choline phosphate, both neutral (N-) and acidic (A-) SMase were detected in the myocardium, with a conversion rate of 0.09 +/- 0.008 and 0.32 +/- 0.05 nmol/min per mg protein, respectively. The activity of A-SMase (78 % of total cellular activity) was significantly higher in microsomes than in cytosol, while the activity of N-SMase was similar in both fractions. Ceramidase, a ceramide-metabolizing enzyme, was also detected in the myocardium of rats. It metabolized ceramide into sphingosine at a rate of 9.94 +/- 0.42 pmol/min per mg protein. In anesthetized rats, 30 min of ischemia had no apparent effect on ceramide concentrations in the myocardium, while 30 min of ischemia followed by 3 h of reperfusion resulted in a significant increase in ceramide by 48 %. The activities of both N- and A-SMase were reduced by 44 % and 32 %, respectively, in the myocardium subjected to ischemia followed by reperfusion, but unaltered in the ischemic myocardium. It was also found that myocardial ischemia followed by reperfusion produced a marked inhibition of ceramidase (by 29 %). These results demonstrate that the myocardium of rats expresses N- and A-SMase and ceramidase, which contribute to the production and metabolism of ceramide, respectively. Tissue ceramide concentrations increased in reperfused myocardium. These increases in ceramide were not associated with enhanced SMase activity, but rather with reduced ceramidase activity.
Asunto(s)
Ceramidas/metabolismo , Daño por Reperfusión Miocárdica/metabolismo , Miocardio/metabolismo , Animales , Citosol/metabolismo , Modelos Animales de Enfermedad , Masculino , Microsomas/metabolismo , Ratas , Ratas Wistar , Esfingomielina Fosfodiesterasa/metabolismoRESUMEN
Opioids have been previously shown to confer acute and delayed cardioprotection against a prolonged ischemic insult. We have extensively characterized the signal transduction pathway mediating acute cardioprotection and have suggested a role for extracellular signal regulated kinase (ERK) in this cardioprotection. Therefore, we attempted to determine a role for ERK and the stress activated MAP kinase, p38, in opioid-induced delayed cardioprotection by using selective inhibitors of these pathways. All rats were subjected to 30 min of ischemia and 2 h of reperfusion (I/R). Control animals, injected with saline 48 h prior to I/R, had an infarct size/area at risk (IS/AAR) of 61.6 +/- 1.6.48-h pretreatment with TAN-67 (30 mg/kg), a delta1-opioid receptor agonist, maximally reduced IS/AAR (31.2 +/- 6.5). The involvement of ERK was examined with PD 098059, a selective pharmacological antagonist which inhibits the upstream kinase, MEK-1, that phosphorylates and activates ERK. PD 098059 (0.3 mg/kg) did not alter IS/AAR when administered alone (60.7 +/- 4.9). However, PD 098059 (0.3 mg/kg) administration 30 min prior to TAN-67 (30 mg/kg) completely abolished cardioprotection (61.0 +/- 7.6). The selective p38 inhibitor, SB 203580 (1.0 mg/kg), had no effect on IS/AAR in the absence of TAN-67 (53.1 +/- 2.3). Additionally, SB 203580 (1.0 mg/kg) when administered prior to TAN-67 (30 mg/kg) partially abolished cardioprotection (51.3 +/- 6.4). These results suggest that both ERK and p38 are integral components of opioid-induced delayed cardioprotection and may act via parallel pathways.
Asunto(s)
Precondicionamiento Isquémico Miocárdico , Proteínas Quinasas Activadas por Mitógenos/metabolismo , Isquemia Miocárdica/metabolismo , Receptores Opioides/metabolismo , Analgésicos/farmacología , Animales , Presión Sanguínea , Activación Enzimática/fisiología , Inhibidores Enzimáticos/farmacología , Flavonoides/farmacología , Frecuencia Cardíaca , Imidazoles/farmacología , Sistema de Señalización de MAP Quinasas/efectos de los fármacos , Sistema de Señalización de MAP Quinasas/fisiología , Masculino , Isquemia Miocárdica/tratamiento farmacológico , Miocardio/enzimología , Piridinas/farmacología , Quinolinas/farmacología , Ratas , Ratas Wistar , Proteínas Quinasas p38 Activadas por MitógenosRESUMEN
We have previously demonstrated that K(ATP)channel openers administered just prior to and throughout reperfusion induce cardioprotection in the blood-perfused canine heart. However, a recent report suggests that the mitochondrial K(ATP)channel is only a trigger of ischemic preconditioning (IPC). These recent data are, however, in contrast to most previous investigations that suggested that activation of the mitochondrial K(ATP)channel is an important downstream mediator of cardioprotection. Therefore, we examined the role of the mitochondrial K(ATP)channel as a downstream mediator of IPC in a rat model by administering the selective mitochondrial K(ATP)channel antagonist, 5-hydroxydecanoate (5-HD), at several points during IPC. Infarct size (IS) was determined by tetrazolium chloride staining and expressed as a percent of the area at risk (AAR). Control animals had an IS/AAR of 58.4+/-0.6 and IS/AAR was reduced to 6.2+/-1.7 following IPC. 5-HD (10 mg/kg), attenuated cardioprotection when administered either 5 min prior to the IPC stimulus (40.4+/-1.4), during the reperfusion phase of the IPC stimulus (39.7+/-5.9), or 5 min prior to reperfusion during prolonged ischemia (34.3+/-6.9). Additionally, when 5-HD was administered at 5 mg/kg during the reperfusion phase of index ischemia plus 5 min prior to IPC or plus during the reperfusion phase of IPC, cardioprotection was also attenuated (36.3+/-5.5 and 43.8+/-6.9, respectively). These data suggest that activation of the mitochondrial K(ATP) channel is an important downstream regulator of myocardial protection with effects lasting into the reperfusion period following prolonged ischemia.
Asunto(s)
Adenosina Trifosfato/metabolismo , Activación del Canal Iónico/fisiología , Precondicionamiento Isquémico Miocárdico , Mitocondrias/metabolismo , Isquemia Miocárdica/fisiopatología , Reperfusión Miocárdica , Canales de Potasio/fisiología , Animales , Corazón , Hemodinámica , Masculino , Infarto del Miocardio/patología , Infarto del Miocardio/fisiopatología , Canales de Potasio/metabolismo , Ratas , Ratas WistarRESUMEN
Stimulation of the delta(1)-opioid receptor confers cardioprotection to the ischemic myocardium. We examined the role of protein kinase C (PKC) after delta-opioid receptor stimulation with TAN-67 or D-Ala(2)-D-Leu(5)-enkephalin (DADLE) in a rat model of myocardial infarction induced by a 30-min coronary artery occlusion and 2-h reperfusion. Infarct size (IS) was determined by tetrazolium staining and expressed as a percentage of the area at risk (IS/AAR). Control animals, subjected to ischemia and reperfusion, had an IS/AAR of 59.9 +/- 1.8. DADLE and TAN-67 administered before ischemia significantly reduced IS/AAR (36.9 +/- 3.9 and 36.7 +/- 4.7, respectively). The delta(1)-selective opioid antagonist 7-benzylidenenaltrexone (BNTX) abolished TAN-67-induced cardioprotection (54.4 +/- 1.3). Treatment with the PKC antagonist chelerythrine completely abolished DADLE- (61.8 +/- 3.2) and TAN-67-induced cardioprotection (55.4 +/- 4.0). Similarly, the PKC antagonist GF 109203X completely abolished TAN-67-induced cardioprotection (54.6 +/- 6.6). Immunofluorescent staining with antibodies directed against specific PKC isoforms was performed in myocardial biopsies obtained after 15 min of treatment with saline, chelerythrine, BNTX, or TAN-67 and chelerythrine or BNTX in the presence of TAN-67. TAN-67 induced the translocation of PKC-alpha to the sarcolemma, PKC-beta(1) to the nucleus, PKC-delta to the mitochondria, and PKC-epsilon to the intercalated disk and mitochondria. PKC translocation was abolished by chelerythrine and BNTX in TAN-67-treated rats. To more closely examine the role of these isoforms in cardioprotection, we utilized the PKC-delta selective antagonist rottlerin. Rottlerin abolished opioid-induced cardioprotection (48.9 +/- 4.8) and PKC-delta translocation without affecting the translocation of PKC-alpha, -beta(1), or -epsilon. These results suggest that PKC-delta is a key second messenger in the cardioprotective effects of delta(1)-opioid receptor stimulation in rats.
Asunto(s)
Leucina Encefalina-2-Alanina/farmacología , Precondicionamiento Isquémico Miocárdico , Isoenzimas/metabolismo , Isquemia Miocárdica/tratamiento farmacológico , Isquemia Miocárdica/metabolismo , Naltrexona/análogos & derivados , Proteína Quinasa C/metabolismo , Acetofenonas/farmacología , Alcaloides , Analgésicos/farmacología , Animales , Benzofenantridinas , Benzopiranos/farmacología , Compuestos de Bencilideno/farmacología , Activación Enzimática/efectos de los fármacos , Inhibidores Enzimáticos/farmacología , Frecuencia Cardíaca , Indoles/farmacología , Masculino , Maleimidas/farmacología , Infarto del Miocardio/tratamiento farmacológico , Infarto del Miocardio/metabolismo , Daño por Reperfusión Miocárdica/tratamiento farmacológico , Daño por Reperfusión Miocárdica/metabolismo , Miocardio/enzimología , Naltrexona/farmacología , Antagonistas de Narcóticos/farmacología , Fenantridinas/farmacología , Proteína Quinasa C-delta , Quinolinas/farmacología , Ratas , Ratas Wistar , Receptores Opioides delta/agonistas , Receptores Opioides delta/antagonistas & inhibidoresRESUMEN
Stimulation of the delta(1)-opioid receptor has been shown to trigger ischemic preconditioning (IPC). Additionally, myocardial ischemia/reperfusion induces the activation of extracellular signal-regulated kinase (ERK). Therefore, we examined the role of ERK in acute cardioprotection induced by delta(1)-opioid receptor stimulation or IPC. Infarct size (IS) was expressed as a percentage of the area at risk (AAR). Control animals had an IS/AAR of 60.6 +/- 1.8. IPC and delta(1)-opioid receptor stimulation with TAN-67 reduced IS/AAR (8.2 +/- 1.3 and 30.2 +/- 2.4). Inhibition of ERK with the selective MEK-1 antagonist, PD 098059 during IPC or TAN-67 administration significantly reduced cardioprotection (41.5 +/- 6.4 and 63.0 +/- 4.8). Western Blot analysis and subsequent densitometry corroborated these observations. Control, TAN-67-, or IPC-treated hearts were harvested after 0, 5, 15, and 30 min of ischemia or 5, 30, and 60 min of reperfusion and separated into cytosolic and nuclear fractions. Both isoforms of ERK (p44 and p42) rapidly increased to greater levels throughout reperfusion in the nuclear fraction of IPC- and opioid-treated versus control rats, however, this increase was not attenuated by PD 098059. Conversely, the rapid activation of the 44-kDa isoform of ERK after 5 min of reperfusion in the cytosolic fraction was significantly increased in IPC- and opioid-treated hearts versus control, and this increase was abolished by pretreatment with PD 098059. Additionally, p42 was activated in the cytosolic fraction of IPC-treated animals. These results suggest a key role for the 44-kDa isoform of ERK in the cytoplasm during cardioprotection induced by either IPC or stimulation of the delta(1)-opioid receptor.
Asunto(s)
Analgésicos Opioides/uso terapéutico , Cardiopatías/prevención & control , Precondicionamiento Isquémico Miocárdico , Proteínas Quinasas Activadas por Mitógenos/metabolismo , Animales , Presión Sanguínea/efectos de los fármacos , Western Blotting , Células COS , Núcleo Celular/efectos de los fármacos , Núcleo Celular/enzimología , Citosol/efectos de los fármacos , Citosol/enzimología , Activación Enzimática/efectos de los fármacos , Inhibidores Enzimáticos/uso terapéutico , Frecuencia Cardíaca/efectos de los fármacos , Isoenzimas/efectos de los fármacos , Isoenzimas/metabolismo , Masculino , Proteína Quinasa 1 Activada por Mitógenos/metabolismo , Proteínas Quinasas Activadas por Mitógenos/antagonistas & inhibidores , Infarto del Miocardio/patología , Infarto del Miocardio/prevención & control , Daño por Reperfusión Miocárdica/patología , Daño por Reperfusión Miocárdica/prevención & control , Ratas , Ratas Wistar , Receptores Opioides delta/agonistas , Receptores Opioides mu/agonistasRESUMEN
We examined the role of the sarcolemmal and mitochondrial ATP-sensitive potassium (K(ATP)) channel in a rat model of myocardial infarction after stimulation with the selective delta(1)-opioid receptor agonist TAN-67. Hearts were subjected to 30 min of regional ischemia and 2 h of reperfusion. Infarct size was expressed as a percentage of the area at risk. TAN-67 significantly reduced infarct size/area at risk (29.6 +/- 3.3) versus control (63. 1 +/- 2.3). The sarcolemmal-selective K(ATP) channel antagonist HMR 1098, administered 10 min before TAN-67, did not significantly attenuate cardioprotection (26.0 +/- 7.3) at a dose (3 mg/kg) that had no effect in the absence of TAN-67 (56.3 +/- 4.3). Pretreatment with the mitochondrial selective antagonist 5-hydroxydecanoic acid (5-HD) 5 min before the 30-min occlusion completely abolished TAN-67-induced cardioprotection (54.3 +/- 2.7), but had no effect in the absence of TAN-67 (62.6 +/- 4.1), suggesting the involvement of the mitochondrial K(ATP) channel. Additionally, we examined the antiarrhythmic effects of TAN-67 in the presence or absence of 5-HD and HMR 1098 during 30 min of ischemia. Control animals had an average arrhythmia score of 10.40 +/- 2.41. TAN-67 significantly reduced the arrhythmia score during 30 min of ischemia (2.38 +/- 0. 85). 5-HD and HMR 1098 in the absence of TAN-67 produced an insignificant decrease in the arrhythmia score (8.80 +/- 2.56 and 4. 20 +/- 1.07, respectively). 5-HD administration before TAN-67 treatment abolished its antiarrhythmic effect (4.71 +/- 1.11). However, HMR 1098 did not abolish TAN-67-induced protection against arrhythmias (1.67 +/- 0.80). These data suggest that delta(1)-opioid receptor stimulation is cardioprotective against myocardial ischemia and sublethal arrhythmias and suggest a role for the mitochondrial K(ATP) channel in mediating these cardioprotective effects.
Asunto(s)
Analgésicos Opioides/uso terapéutico , Antiarrítmicos/farmacología , Arritmias Cardíacas/prevención & control , Mitocondrias Cardíacas/efectos de los fármacos , Infarto del Miocardio/prevención & control , Bloqueadores de los Canales de Potasio , Quinolinas/uso terapéutico , Sarcolema/efectos de los fármacos , Adenosina Trifosfato/fisiología , Analgésicos Opioides/antagonistas & inhibidores , Animales , Antiarrítmicos/antagonistas & inhibidores , Benzamidas/farmacología , Presión Sanguínea/efectos de los fármacos , Ácidos Decanoicos/farmacología , Interacciones Farmacológicas , Frecuencia Cardíaca/efectos de los fármacos , Hidroxiácidos/farmacología , Masculino , Mitocondrias Cardíacas/fisiología , Isquemia Miocárdica/complicaciones , Daño por Reperfusión Miocárdica/prevención & control , Canales de Potasio/efectos de los fármacos , Canales de Potasio/fisiología , Quinolinas/antagonistas & inhibidores , Ratas , Ratas Wistar , Receptores Opioides delta/agonistas , Sarcolema/fisiología , Fibrilación Ventricular/prevención & controlRESUMEN
We examined the role of the sarcolemmal and mitochondrial K(ATP) channels in a rat model of ischemic preconditioning (IPC). Infarct size was expressed as a percentage of the area at risk (IS/AAR). IPC significantly reduced infarct size (7 +/- 1%) versus control (56 +/- 1%). The sarcolemmal K(ATP) channel-selective antagonist HMR-1098 administered before IPC did not significantly attenuate cardioprotection. However, pretreatment with the mitochondrial K(ATP) channel-selective antagonist 5-hydroxydecanoic acid (5-HD) 5 min before IPC partially abolished cardioprotection (40 +/- 1%). Diazoxide (10 mg/kg iv) also reduced IS/AAR (36.2 +/- 4.8%), but this effect was abolished by 5-HD. As an index of mitochondrial bioenergetic function, the rate of ATP synthesis in the AAR was examined. Untreated animals synthesized ATP at 2.12 +/- 0.30 micromol x min(-1) x mg mitochondrial protein(-1). Rats subjected to ischemia-reperfusion synthesized ATP at 0.67 +/- 0.06 micromol x min(-1) x mg mitochondrial protein(-1). IPC significantly increased ATP synthesis to 1.86 +/- 0.23 micromol x min(-1) x mg mitochondrial protein(-1). However, when 5-HD was administered before IPC, the preservation of ATP synthesis was attenuated (1.18 +/- 0.15 micromol x min(-1) x mg mitochondrial protein(-1)). These data are consistent with the notion that inhibition of mitochondrial K(ATP) channels attenuates IPC by reducing IPC-induced protection of mitochondrial function.
Asunto(s)
Precondicionamiento Isquémico Miocárdico , Proteínas de la Membrana/fisiología , Mitocondrias Cardíacas/fisiología , Adenosina Trifosfato/biosíntesis , Adenosina Trifosfato/metabolismo , Animales , Ácidos Decanoicos/farmacología , Diazóxido/farmacología , Hemodinámica/efectos de los fármacos , Hidroxiácidos/farmacología , Masculino , Proteínas de la Membrana/antagonistas & inhibidores , Mitocondrias Cardíacas/metabolismo , Infarto del Miocardio/patología , Miocardio/metabolismo , Concentración Osmolar , Canales de Potasio , Ratas , Ratas WistarRESUMEN
Ischemic preconditioning (IPC) is a phenomenon in which single or multiple brief periods of ischemia have been shown to protect the heart against a more prolonged ischemic insult, the result of which is a marked reduction in myocardial infarct size, severity of stunning, or incidence of cardiac arrhythmias. Although a number of substances and signaling pathways have been proposed to be involved in mediating the cardioprotective effect of IPC, the overwhelming majority of evidence suggests that the ATP-sensitive potassium channel (KATP channel) is an important component of this phenomenon and may serve as the end effector in this process. Initially, it was hypothesized that the surface or sarcolemmal KATP (sarc KATP) channel mediated protection observed after IPC; however, subsequent evidence suggested that the recently identified mitochondrial KATP channel (mito KATP) may be the potassium channel mediating IPC-induced cardioprotection. In this review, evidence will be presented supporting a role for either the sarc KATP or the mito KATP in IPC and potential mechanisms by which opening these channels may produce cardioprotection; additionally, we will address important questions that still need to be investigated to define the role of the sarc or mito KATP channel, or both, in cardiac pathophysiology.
Asunto(s)
Adenosina Trifosfato/fisiología , Precondicionamiento Isquémico Miocárdico , Mitocondrias Cardíacas/metabolismo , Canales de Potasio/metabolismo , Sarcolema/metabolismo , Animales , HumanosRESUMEN
Both tyrosine kinase (TK) and protein kinase C (PKC) inhibitors have been shown individually to completely abolish the cardioprotective effects of ischemic preconditioning (IPC) in rabbits; however, blockade of both enzymes is necessary to totally abolish IPC in pigs. Recently, we have shown that TK inhibition partially attenuates the cardioprotective effect of IPC in intact rat hearts. Therefore, the present study was designed to test the hypothesis that inhibition of both TK and PKC is necessary to completely abolish IPC in the intact rat and that this effect is dependent on the intensity of the preconditioning stimulus. All animals were subjected to 30 min of coronary artery occlusion and 2 h of reperfusion. In series 1, multiple-cycle-induced IPC was produced via three 5-min occlusions interspersed with 5 min of reperfusion (3 x 5 IPC). Genistein (5 mg/kg), a TK inhibitor infused 30 min before IPC, and chelerythrine chloride (5 mg/kg), a PKC inhibitor infused 5 min before the prolonged ischemic insult, were administered alone or in combination in the absence or presence of 3 x 5 IPC. 3 x 5 IPC produced a marked reduction in infarct size as a percentage of area at risk compared with control (8.0 +/- 0.8 vs. 56.1 +/- 0.8%). The effects of 3 x 5 IPC were partially blocked by pretreatment with genistein (34.0 +/- 2.0%) or chelerythrine (26.4 +/- 2.8%) alone; however, combined administration of genistein and chelerythrine completely abolished the effects of 3 x 5 IPC (50.7 +/- 3.6%). In series 2, single-cycle IPC was elicited by one 5-min occlusion followed by 10 min of reperfusion (1 x 5 IPC). Compared with control, 1 x 5 IPC also significantly reduced infarct size (15.4 +/- 3.0%). Genistein or chelerythrine administered alone completely abolished 1 x 5 IPC-induced cardioprotection. These results suggest that the efficacy of TK and PKC inhibition to block IPC depends on the intensity of the preconditioning stimulus and that these kinases may work through parallel pathways.
Asunto(s)
Precondicionamiento Isquémico Miocárdico , Proteína Quinasa C/fisiología , Proteínas Tirosina Quinasas/fisiología , Alcaloides , Animales , Benzofenantridinas , Combinación de Medicamentos , Inhibidores Enzimáticos/farmacología , Genisteína/farmacología , Corazón/efectos de los fármacos , Hemodinámica/fisiología , Masculino , Infarto del Miocardio/patología , Fenantridinas/farmacología , Proteína Quinasa C/antagonistas & inhibidores , Proteínas Tirosina Quinasas/antagonistas & inhibidores , Ratas , Ratas WistarRESUMEN
Opioids have been previously shown to confer short-term cardioprotection against a prolonged ischemic insult. Therefore, the present study was designed to determine whether opioids can induce a delayed or "second window" of cardioprotection and to assess the potential involvement of the mitochondrial KATP channel. All rats were subjected to 30 minutes of ischemia and 2 hours of reperfusion (I/R). Control animals, injected with saline 24 hours before I/R, elicited an infarct size/area at risk (IS/AAR) of 62.9+/-3.4. TAN-67, a delta1-opioid receptor agonist, was administered 10 or 30 mg/kg IP 12, 24, 48, or 72 hours before I/R. TAN-67 (10 mg/kg) 12- or 24-hour pretreatment did not significantly reduce IS/AAR (62.1+/-6.3 and 43.3+/-7.3, respectively). Similarly, 12-hour pretreatment with TAN-67 (30 mg/kg) did not reduce IS/AAR (60.0+/-5.6); however, 24-hour pretreatment significantly reduced IS/AAR (34.5+/-5.9). Forty-eight-hour pretreatment with TAN-67 maximally reduced IS/AAR (29.2+/-7.0), and opioid-induced cardioprotection was lost after 72-hour pretreatment (61.7+/-3.8). TAN-67-induced cardioprotection could be abolished by pretreatment with the selective delta1-opioid receptor antagonist 7-benzylidenenaltrexone, BNTX, administered either 30 minutes before TAN-67 given 48 hours before I/R or 10 minutes before I/R in rats previously treated for 48 hours with TAN-67 (59.6+/-3.1 and 58.7+/-3.5, respectively). The involvement of the KATP channel was investigated with 2 inhibitors: glibenclamide, a nonselective KATP channel inhibitor, and 5-hydroxydecanoic acid, selective for the mitochondrial KATP channel in rabbits. Glibenclamide, administered 30 minutes before I/R in 48-hour TAN-67-pretreated rats, completely abolished cardioprotection (60. 4+/-3.2). Similarly, 5-hydroxydecanoic acid, administered 5 minutes before I/R in rats pretreated 48 hours previously with TAN-67, completely abolished cardioprotection (57.8+/-2.5). These results suggest that delta1-opioid receptor stimulation, 24 to 48 hours before an ischemic insult, produces a delayed cardioprotective effect that is possibly the result of mitochondrial KATP channel activation.
Asunto(s)
Mitocondrias/química , Isquemia Miocárdica/fisiopatología , Daño por Reperfusión Miocárdica/fisiopatología , Canales de Potasio/fisiología , Receptores Opioides delta/fisiología , Analgésicos/farmacología , Animales , Compuestos de Bencilideno/farmacología , Presión Sanguínea , Circulación Coronaria , Gliburida/farmacología , Hipoglucemiantes/farmacología , Precondicionamiento Isquémico Miocárdico , Masculino , Infarto del Miocardio/tratamiento farmacológico , Infarto del Miocardio/fisiopatología , Isquemia Miocárdica/tratamiento farmacológico , Daño por Reperfusión Miocárdica/tratamiento farmacológico , Miocardio/química , Naltrexona/análogos & derivados , Naltrexona/farmacología , Antagonistas de Narcóticos/farmacología , Quinolinas/farmacología , Ratas , Ratas WistarRESUMEN
Ischemic preconditioning (IPC) confers cardioprotection against a prolonged ischemic insult. Tyrosine kinase (TK) inhibitors have been shown to attenuate IPC; however, it is unclear whether TK is involved in the initiation of and/or the maintenance of this phenomenon. Thus the hypothesis that TK acts primarily during the initiation of IPC was examined in a rat model of myocardial infarction. Hearts were subjected to 30 min of coronary artery occlusion and 2 h of reperfusion. IPC was elicited by three 5-min occlusions interspersed with 5 min of reperfusion before the prolonged occlusion period. Genistein, a nonspecific TK inhibitor, was administered before or during the final 2 min of the first or third occlusion period of IPC. Daidzein, an inactive structural analog of genistein, and lavendustin A, a more specific TK inhibitor, were also tested in this model. IPC markedly reduced infarct size expressed as a percentage of the area at risk compared with control (56.3 +/- 2.8 to 7.1 +/- 2.0%). This cardioprotection was attenuated by genistein pretreatment (5 mg/kg: 34.7 +/- 2.2%, 10 mg/kg: 33.5 +/- 5.9%). However, genistein administered during the first or third occlusion period of IPC did not significantly attenuate cardioprotection (10.3 +/- 2.9% and 6.4 +/- 2.0%). Lavendustin A (1. 0 mg/kg) pretreatment also attenuated IPC (30.1 +/- 2.2%), whereas daidzein (5 mg/kg) had no effect (7.9 +/- 2.4%). These results suggest that activation of a TK is involved in the initiation but not the maintenance of IPC in the rat myocardium.