Aging differentially modifies agonist-evoked mouse detrusor contraction and calcium signals

Although aging-induced changes in urinary bladder neurotransmission have been studied in some detail, information regarding alterations in detrusor muscle is scanty and addresses only partial aspects of the myogenic response of detrusor. Rodent bladder aging shows several features similar to those reported in humans. The aim of this study was to characterize in aged mouse the alterations of detrusor muscle contraction and the putative underlying changes in Ca²⁺ signals. We studied in vitro the myogenic contraction induced by agonists in detrusor strips from adult (3 months old) or aged (23–25 months old) mice. In addition, we determined the agonist-induced [Ca²⁺]_i signals by epifluorescence microscopy in fura-2 loaded isolated detrusor cells. Aging impaired the contractile response of bladder strips to cholinergic stimulation with bethanechol and to chemical depolarization with KCl-containing solutions. On the contrary, the response to purinergic stimulation (ATP) was enhanced. Aging also diminished the transient Ca²⁺ signal evoked by bethanechol and the Ca²⁺ influx induced by KCl in bladder strips. Treatments aimed to release calcium from intracellular stores (caffeine and a low level of ionomycin in Ca²⁺-free medium) showed that aging reduces the size of agonist-releasable stores. Similar to contraction, the mobilization of Ca²⁺ by ATP was increased in aged cells. Therefore, the differential effects of aging on detrusor contraction are associated to alterations of [Ca²⁺]_i signals: the cholinergic inhibition is due to inhibition of voltage-operated Ca²⁺ influx and reduction of the size of intracellular Ca²⁺ stores, while the age-induced ATP response is accompanied by an enhanced Ca²⁺ mobilization.     

Cardiac overexpression of metallothionein rescues cardiac contractile dysfunction and endoplasmic reticulum stress but not autophagy in sepsis

Sepsis is characterized by systematic inflammation where oxidative damage plays a key role in organ failure. This study was designed to examine the impact of the antioxidant metallothionein (MT) on lipopolysaccharide (LPS)-induced cardiac contractile and intracellular Ca²⁺ dysfunction, oxidative stress, endoplasmic reticulum (ER) stress and autophagy. Mechanical and intracellular Ca²⁺ properties were examined in hearts from FVB and cardiac-specific MT overexpression mice treated with LPS. Oxidative stress, activation of mitogen-activated protein kinase pathways (ERK, JNK and p38), ER stress, autophagy and inflammatory markers iNOS and TNFα were evaluated. Our data revealed enlarged end systolic diameter, decreased fractional shortening, myocyte peak shortening and maximal velocity of shortening/relengthening as well as prolonged duration of relengthening in LPS-treated FVB mice associated with reduced intracellular Ca²⁺ release and decay. LPS treatment promoted oxidative stress (reduced glutathione/glutathione disulfide ratio and ROS generation). Western blot analysis revealed greater iNOS and TNFα, activation of ERK, JNK and p38, upregulation of ER stress markers GRP78, Gadd153, PERK and IRE1α, as well as the autophagy markers Beclin-1, LCB3 and Atg7 in LPS-treated mouse hearts without any change in total ERK, JNK and p38. Interestingly, these LPS-induced changes in echocardiographic, cardiomyocyte mechanical and intracellular Ca²⁺ properties, ROS, stress signaling and ER stress (but not autophagy, iNOS and TNFα) were ablated by MT. Antioxidant N-acetylcysteine and the ER stress inhibitor tauroursodeoxycholic acid reversed LPS-elicited depression in cardiomyocyte contractile function. LPS activated AMPK and its downstream signaling ACC in conjunction with an elevated AMP/ATP ratio, which was unaffected by MT. Taken together, our data favor a beneficial effect of MT in the management of cardiac dysfunction in sepsis.     

有氧运动通过上调心肌细胞自噬改善衰老心肌收缩舒张功能

目的:探讨有氧运动训练能否通过调节心肌细胞自噬改善老年小鼠心肌收缩舒张功能。方法:采用老年C57小鼠(20月龄,10只),以成年小鼠(4月龄,10只)为对照,无负重游泳训练9周(每周5 d,每天1 h),建立有氧运动训练模型。模型建立后,急性分离各组小鼠心肌细胞,采用单心肌细胞动缘探测系统检测心肌收缩舒张能力。应用蛋白免疫印迹法(Western blot)检测自噬相关蛋白的表达。结果:有氧运动训练可有效地改善衰老小鼠心肌收缩舒张能力,表现为可显著增加老年组小鼠心肌细胞最大收缩幅度(PS)和最大收缩和舒张速率(±dL/dt),有效缩短舒张90%时程(TR90)(均P0.05)。有氧运动还可显著上调衰老心肌中单磷酸腺苷活化蛋白激酶(AMPK)磷酸化水平,进而抑制其下游雷帕霉素靶蛋白(mTOR)的活性(均P0.05)。检测心肌自噬标志物Beclin-1和Atg7发现,与成年组相比,老年组小鼠心肌自噬水平显著降低(P0.05),但有氧运动训练可显著改善衰老心肌细胞自噬,运动训练上调心肌细胞自噬的作用,在AMPK基因敲低小鼠(AMPK KD,10只)被显著抑制。采用自噬诱导剂雷帕霉素处理则可改善衰老小鼠心肌收缩舒张的能力(均P0.05)。结论:有氧运动训练可有效地上调衰老小鼠心肌细胞自噬的水平,进而改善衰老小鼠心肌收缩和舒张能力,其机制可能与激活心肌AMPK-mTOR通路有关。     

Age and hypertrophy related changes in contractile post-rest behavior and action potential properties in isolated rat myocytes

“Physiological” aging as well as early and progressive cardiac hypertrophy may affect action potential (AP) pattern, contractile function, and Ca²⁺ handling. We hypothesize that contractile function is disturbed in hypertrophy from early stages and is differently affected in aged myocardium. In vivo function, cardiomyocyte contractile behavior and APs were compared in Wistar-Kyoto (WIS) rats and spontaneously hypertensive rats (SHR) at different ages and degrees of hypertrophy (3–4, 9–11, 20–24 months). Post-rest (PR) behavior was used to investigate the relative contribution of the sarcoplasmic reticulum (SR) and the Na/Ca exchanger (NCX) to cytosolic Ca²⁺ removal. APs were recorded by whole-cell current-clamp and sarcomere shortening by video microscopy. Cyclopiazonic acid was used to suppress Ca²⁺ ATPase (SERCA) function. Heart weight/body weight ratio was increased in SHR versus WIS within all age groups. Myocyte steady state (SS) shortening amplitude was reduced in young SHR versus WIS. Aging led to a significant decay of SS contractile amplitude and relengthening velocity in WIS, but the PR potentiation was maintained. In contrast, aging in SHR led to a decrease of PR potentiation, while SS contraction and relengthening velocity increased. APD_50% was always prolonged in SHR versus WIS. With aging, APD_50% increased in both WIS and SHR, but was still shorter in WIS. However, in old WIS the late AP portion (APD_90%) was prolonged. Ca²⁺ handling and AP properties are disturbed progressively with aging and with increasing hypertrophy. Decreased amplitude of shortening and velocity of relengthening in aged WIS may be attributed to reduced SERCA function. In SHR, an increase in SR leak and shift towards transmembraneous Ca handling via NCX may be responsible for the changes in contractile function. A prolonged APD_90% in aged WIS may be an adaptive mechanism to preserve basal contractility. Therefore, the effects on contractile parameters and AP are different in hypertrophy and aging.     

老年人心肌细胞自噬的变化及其调控机制研究进展

心肌细胞自噬对维持正常心脏结构和功能起重要作用。近年来的研究结果表明老年人心肌细胞自噬功能降低,老年人心肌自噬基因Atg5、Atg7和Beclin1表达降低;心肌细胞自噬下调与磷脂酰肌醇3-激酶/丝氨酸-苏氨酸激酶/雷帕霉素靶蛋白和单磷酸腺苷激活的蛋白激酶及SIRT1信号通路失调有关;此外,活性氧及一些神经内分泌因子也可介导老年人心肌细胞自噬下调。调控心肌细胞自噬将为老年人心肌病的预防和治疗提供新的途径。     

Caloric restriction attenuates aging-induced cardiac insulin resistance in male Wistar rats through activation of PI3K/Akt pathway

Background and Aim

Caloric restriction (CR) improves insulin sensitivity and is one of the dietetic strategies most commonly used to enlarge life and to prevent aging-induced cardiovascular alterations. The aim of this study was to analyze the possible beneficial effects of caloric restriction (CR) preventing the aging-induced insulin resistance in the heart of male Wistar rats.

Caloric restriction does not alter effects of aging in cardiac side population cells

The aged heart displays a loss of cardiomyocyte number and function, possibly due to the senescence and decreased regenerative potential that has been observed in some cardiac progenitor cells. An important cardiac progenitor that has not been studied in the context of aging is the cardiac side population (CSP) cell. To address this, flow cytometry-assisted cell sorting was used to isolate CSP cells from adult (6–10 months old) and aged (24–32 months old) C57Bl/6 mice that were fed either a control diet or an anti-aging diet (caloric restriction, CR). Aging caused a 2.3-fold increase in the total number of CSP cells and a 3.2-fold increase in the cardiomyogenic sca1⁺/CD31⁻ subpopulation. Aging did not affect markers of proliferation or senescence, including telomerase activity and expression of cell cycle genes, in sca1⁺/CD31⁻ CSP cells. In contrast, the aged cells had reduced expression of genes associated with differentiation, including smooth muscle actin and cardiac muscle actin (5.1- and 3.2-fold, respectively). None of these age effects were altered by CR diet. Therefore, it appears that the manner in which CSP cells age is distinct from the aging of post-mitotic tissue (and perhaps other progenitor cells) that can often be attenuated by CR.     

Cardiac-Specific Knockout of ET<Subscript>A</Subscript> Receptor Mitigates Paraquat-Induced Cardiac Contractile Dysfunction

Paraquat (1,1’-dimethyl-4-4’-bipyridinium dichloride), a highly toxic quaternary ammonium herbicide widely used in agriculture, exerts potent toxic prooxidant effects resulting in multi-organ failure including the lung and heart although the underlying mechanism remains elusive. Recent evidence suggests possible involvement of endothelin system in paraquat-induced acute lung injury. This study was designed to examine the role of endothelin receptor A (ET_A) in paraquat-induced cardiac contractile and mitochondrial injury. Wild-type (WT) and cardiac-specific ET_A receptor knockout mice were challenged to paraquat (45 mg/kg, i.p.) for 48 h prior to the assessment of echocardiographic, cardiomyocyte contractile and intracellular Ca²⁺ properties, as well as apoptosis and mitochondrial damage. Levels of the mitochondrial proteins for biogenesis and oxidative phosphorylation including UCP2, HSP90 and PGC1α were evaluated. Our results revealed that paraquat elicited cardiac enlargement, mechanical anomalies including compromised echocardiographic parameters (elevated left ventricular end-systolic and end-diastolic diameters as well as reduced factional shortening), suppressed cardiomyocyte contractile function, intracellular Ca²⁺ handling, overt apoptosis and mitochondrial damage. ET_A receptor knockout itself failed to affect myocardial function, apoptosis, mitochondrial integrity and mitochondrial protein expression. However, ET_A receptor knockout ablated or significantly attenuated paraquat-induced cardiac contractile and intracellular Ca²⁺ defect, apoptosis and mitochondrial damage. Taken together, these findings revealed that endothelin system in particular the ET_A receptor may be involved in paraquat-induced toxic myocardial contractile anomalies possibly related to apoptosis and mitochondrial damage.     

Cardiac overexpression of metallothionein attenuates chronic alcohol intake-induced cardiomyocyte contractile dysfunction

Chronic alcohol ingestion leads to alcoholic cardiomyopathy manifested by ventricular dysfunction and heart failure. Although accumulation of reactive oxygen species may play a role in alcohol-induced heart injury, direct impact of enhanced antioxidant defense on pathogenesis of alcoholic cardiomyopathy has not been elucidated. This study was designed to examine the effect of transgenic overexpression of the free radical scavengermetallothionein on alcohol-induced cardiac contractile dysfunction. Wild-type FVB and metallothionein mice were placed on a 4% alcohol or control diet for 12 wk. Cardiac contractile function was evaluated in cardiomyocytes including peak shortening (PS), time-to-peak shortening, time-to-90% relengthening (TR₉₀), maximal velocity of shortening/relengthening (±dL/dt), intracellular Ca²⁺ rise (change in fura-2 fluorescent intensity [ΔFF1]) and intracellular Ca²⁺ decay rate. Intracellular Ca²⁺ cycling proteins including sarco(endo)plasmic reticulum Ca²⁺-ATPase (SERCA2a), Na⁺−Ca²⁺ exchanger (NCX) and phospholamban were assessed using Western blot analysis. Alcohol intake depressed PS, ±dL/dt, and ΔFF1, increased baseline fura-2 fluorescence intensity (FF1), and prolonged intracellular Ca²⁺ decay and TR₉₀, all of which with the exception of ΔFF1 were abrogated by metallothionein. Enhanced stimulating frequency caused lessened PS decline at 1.0 Hz from FVB ethanol group, which was not affected by metallothionein. Immunoblotting data showed reduced SERCA2a, NCX and phospholamban expression in FVB group consuming alcohol. All of these alcohol-induced changes in cardiac proteins were nullified by the metallothionein transgene. In summary, our findings suggest a beneficial role of antioxidants in alcohol-induced cardiomyocyte dysfunction.     

Cardiac overexpression of catalase rescues cardiac contractile dysfunction induced by insulin resistance: role of oxidative stress, protein carbonyl formation and insulin sensitivity

Aims/hypothesis Insulin resistance leads to oxidative stress and cardiac dysfunction. This study examined the impact of catalase on insulin-resistance-induced cardiac dysfunction, oxidative damage and insulin sensitivity.Methods Insulin resistance was initiated in FVB and catalase-transgenic mice by 12 weeks of sucrose feeding. Contractile and intracellular Ca²⁺ properties were evaluated in cardiomyocytes including peak shortening (PS), time-to-PS (TPS), time-to-90% relengthening (TR₉₀), half-width duration (HWD), maximal velocity of shortening/relengthening (±dL/dt), fura-fluorescence intensity change (ΔFFI) and intracellular Ca²⁺ clearance rate (τ). Reactive oxygen species (ROS) and protein damage were evaluated with dichlorodihydrofluorescein and protein carbonyl formation.Results Sucrose-fed mice displayed hyperinsulinaemia, impaired glucose tolerance and normal body weight. Myocytes from FVB sucrose-fed mice exhibited depressed PS and ±dL/dt, prolonged TR₉₀ and τ, and reduced ΔFFI associated with normal TPS and HWD compared with those from starch-fed control mice. ROS and protein carbonyl formation were elevated in FVB sucrose-fed mice. Insulin sensitivity was reduced, evidenced by impaired insulin-stimulated 2-deoxy-d-[³H]glucose uptake. Western blot analysis indicated that sucrose feeding: (1) inhibited insulin-stimulated phosphorylation of insulin receptor and Akt; (2) enhanced protein-tyrosine phosphatase 1B (PTP1B) expression; and (3) suppressed endothelial nitric oxide synthase (eNOS) and Na⁺–Ca²⁺ exchanger expression without affecting peroxisome proliferator-activated receptor γ (PPARγ), sarco(endo)plasmic reticulum Ca²⁺-ATPase isozyme 2a and phospholamban. Catalase ablated insulin-resistance-induced mechanical dysfunction, ROS production and protein damage, and reduced eNOS, but not insulin insensitivity. Catalase itself decreased resting FFI and enhanced expression of PTP1B and PPARγ.Conclusions/interpretation These data indicate that catalase rescues insulin-resistance-induced cardiac dysfunction related to ROS production and protein oxidation but probably does not improve insulin sensitivity.     

Endoplasmic reticulum chaperon tauroursodeoxycholic acid alleviates obesity-induced myocardial contractile dysfunction

ER stress is involved in the pathophysiology of obesity although little is known about the role of ER stress on obesity-associated cardiac dysfunction. This study was designed to examine the effect of ER chaperone tauroursodeoxycholic acid (TUDCA) on obesity-induced myocardial dysfunction. Adult lean and ob/ob obese mice were treated with TUDCA (50 mg/kg/day, p.o.) or vehicle for 5 weeks. Oral glucose tolerance test (OGTT) was performed. Echocardiography, cardiomyocyte contractile and intracellular Ca²⁺ properties were assessed. Sarco(endo)plasmic reticulum Ca²⁺-ATPase (SERCA) activity and protein expression of intracellular Ca²⁺ regulatory proteins were measured using ⁴⁵Ca²⁺ uptake and Western blot analysis, respectively. Insulin signaling, ER stress markers and HSP90 were evaluated. Our results revealed that chronic TUDCA treatment lowered systolic blood pressure and lessened glucose intolerance in obese mice. Obesity led to increased diastolic diameter, cardiac hypertrophy, compromised fractional shortening, cardiomyocyte contractile (peak shortening, maximal velocity of shortening/relengthening, and duration of contraction/relaxation) and intracellular Ca²⁺ properties, all of which were significantly attenuated by TUDCA. TUDCA reconciled obesity-associated decrease in SERCA activity and expression, and increase in serine phosphorylation of IRS, total and phosphorylated cJun, ER stress markers Bip, peIF2α and pPERK. Obesity-induced changes in phospholamban and HSP90 were unaffected by TUDCA. In vitro finding revealed that TUDCA ablated palmitic acid-induced cardiomyocyte contractile dysfunction. In summary, these data depicted a pivotal role of ER stress in obesity-associated cardiac contractile dysfunction, suggesting the therapeutic potential of ER stress as a target in the management of cardiac dysfunction in obesity.     

Ageing-related cardiomyocyte functional decline is sex and angiotensin II dependent

Clinically, heart failure is an age-dependent pathological phenomenon and displays sex-specific characteristics. The renin-angiotensin system mediates cardiac pathology in heart failure. This study investigated the sexually dimorphic functional effects of ageing combined with angiotensin II (AngII) on cardiac muscle cell function, twitch and Ca²⁺-handling characteristics of isolated cardiomyocytes from young (~13 weeks) and aged (~87 weeks) adult wild type (WT) and AngII-transgenic (TG) mice. We hypothesised that AngII-induced contractile impairment would be exacerbated in aged female cardiomyocytes and linked to Ca²⁺-handling disturbances. AngII-induced cardiomyocyte hypertrophy was evident in young adult mice of both sexes and accentuated by age (aged adult ~21–23 % increases in cell length relative to WT). In female AngII-TG mice, ageing was associated with suppressed cardiomyocyte contractility (% shortening, maximum rate of shortening, maximum rate of relaxation). This was associated with delayed cytosolic Ca²⁺ removal during twitch relaxation (Tau ~20 % increase relative to young adult female WT), and myofilament responsiveness to Ca²⁺ was maintained. In contrast, aged AngII-TG male cardiomyocytes exhibited peak shortening equivalent to young TG; yet, myofilament Ca²⁺ responsiveness was profoundly reduced with ageing. Increased pro-arrhythmogenic spontaneous activity was evident with age and cardiac AngII overexpression in male mice (42–55 % of myocytes) but relatively suppressed in female aged transgenic mice. Female myocytes with elevated AngII appear more susceptible to an age-related contractile deficit, whereas male AngII-TG myocytes preserve contractile function with age but exhibit desensitisation of myofilaments to Ca²⁺ and a heightened vulnerability to arrhythmic activity. These findings support the contention that sex-specific therapies are required for the treatment of age-progressive heart failure.     

Aging impairs Ca2+ sensitization pathways in gallbladder smooth muscle

Calcium sensitization is an important physiological process in agonist-induced contraction of smooth muscle. In brief, calcium sensitization is a pathway that leads to smooth muscle contraction independently of changes in [Ca²⁺]_i by mean of inhibition of myosin light chain phosphatase. Aging has negative impacts on gallbladder contractile response due to partial impairment in calcium signaling and alterations in the contractile machinery. However, information regarding aging-induced alterations in calcium sensitization is scanty. We hypothesized that the calcium sensitization system is negatively affected by age. To investigate this, gallbladders were collected from adult (4 months old) and aged (22–24 months old) guinea pigs. To evaluate the contribution of calcium sensitization pathways we assayed the effect of the specific inhibitors Y-27632 and GF109203X on the “in vitro” isometric gallbladder contractions induced by agonist challenges. In addition, expression and phosphorylation (as activation index) of proteins participating in the calcium sensitization pathways were quantified by Western blotting. Aging reduced bethanechol- and cholecystokinin-evoked contractions, an effect associated with a reduction in MLC20 phosphorylation and in the effects of both Y-27632 and GF109203X. In addition, there was a drop in ROCK I, ROCK II, MYPT-1 and PKC expression and in the activation/phosphorylation of MYPT-1, PKC and CPI-17 in response to agonists. Interestingly, melatonin treatment for 4 weeks restored gallbladder contractile responses due to re-establishment of calcium sensitization pathways. These results demonstrate that age-related gallbladder hypocontractility is associated to alterations of calcium sensitization pathways and that melatonin treatment exerts beneficial effects in the recovery of gallbladder contractility.     

Impaired autophagic function in rat islets with aging

Type 2 diabetes is characterized by a deficit in β-cell function and mass, and its incidence increases with age. Autophagy is a highly regulated intracellular process for degrading cytoplasmic components, particularly protein aggregates and damaged organelles. Impaired or deficient autophagy is believed to cause or contribute to aging and age-related disease. Autophagy may be necessary to maintain structure, mass, and function of pancreatic β-cells. In this study, we investigated the effects of age on β-cell function and autophagy in pancreatic islets of 4-month-old (young), 14-month-old (adult), and 24-month-old (old) male Wistar rats. We found that islet β-cell function decreased gradually with age. Protein expression of the autophagy markers LC3/Atg8 and Atg7 exhibited a marked decline in aged islets. The expression of Lamp-2, a good indicator of autophagic degradation rate, was significantly reduced in the islets of old rats, suggesting that autophagic degradation is decreased in the islets of aged rats. However, protein expression of beclin-1/Atg6, which plays an important role in the induction and formation of the pre-autophagosome structure by associating with a multimeric complex of autophagy regulatory proteins (Atg14, Vps34/class 3 PI3 kinase, and Vps15), was most prominent in the islets of adult rats, and was higher in 24-month-old islets than in 4-month-old islets. The levels of p62/SQSTM1 and polyubiquitin aggregates, representing the functions of autophagy and proteasomal degradation, were increased in aging islets. 8-Hydroxydeoxyguanosine, a marker of mitochondrial and nuclear DNA oxidative damage, exhibited strong immunostaining in old islets. Analysis by electron microscopy demonstrated swelling and disintegration of cristae in the mitochondria of aged islets. These results suggest that β-cell and autophagic function in islets decline simultaneously with increasing age in Wistar rats, and that impaired autophagy in the islets of older rats may cause accumulation of misfolded and aggregated proteins and reduce the removal of abnormal mitochondria in β-cells, leading to reduced β-cell function. Dysfunctional autophagy in islets during the aging process may be an important mechanism leading to the development of type 2 diabetes.     

Characteristics of cardiac aging in C57BL/6 mice

The specific processes that cause aging of the cardiac tissue remain elusive. C57BL/6 (B6) mice are commonly used for investigating age-related diseases in mammals. We thus sought to evaluate the cardiac aging process in B6 mice. Cardiac tissues from the newborn (B6 NB), 2 month-old (B6 2M) and 21–27 month-old B6 mice (B6 aged) were used for the investigation. Several age-related cellular processes were evaluated, including telomere shortening, changes in p53 and p16 expression, changes in mitochondria DNA expression and DNA deletion, and alteration of mitochondria. We found that the aging of the B6 mice cardiac tissue is associated with the maintenance of telomere length, increased expression of p53 and p16, mild changes in mitochondrial DNA expression but widespread DNA deletion, and significant alterations of the mitochondrial ultrastructure within the cardiac tissue. The results of our studies suggest that mitochondrial DNA deletions, which affect the mitochondrial ultrastructure, cytochrome C oxidase activity, and p53 expression, are significantly associated with cardiac aging and may be a source of age-related heart failure.     

Cardiac hypertrophy induced by sustained β-adrenoreceptor activation: pathophysiological aspects

Cardiac hypertrophy is promoted by adrenergic over-activation and represents an independent risk factor for cardiovascular morbidity and mortality. The basic knowledge about mechanisms by which sustained adrenergic activation promotes myocardial growth, as well as understanding how structural changes in hypertrophied myocardium could affect myocardial function has been acquired from studies using an animal model of chronic systemic β-adrenoreceptor agonist administration. Sustained β-adrenoreceptor activation was shown to enhance the synthesis of myocardial proteins, an effect mediated via stimulation of myocardial growth factors, up-regulation of nuclear proto-oncogenes, induction of cardiac oxidative stress, as well as activation of mitogen-activated protein kinases and phosphatidylinositol 3-kinase. Sustained β-adrenoreceptor activation contributes to impaired cardiac autonomic regulation as evidenced by blunted parasympathetically-mediated cardiovascular reflexes as well as abnormal storage of myocardial catecholamines. Catecholamine-induced cardiac hypertrophy is associated with reduced contractile responses to adrenergic agonists, an effect attributed to downregulation of myocardial β-adrenoreceptors, uncoupling of β-adrenoreceptors and adenylate cyclase, as well as modifications of downstream cAMP-mediated signaling. In compensated cardiac hypertrophy, these changes are associated with preserved or even enhanced basal ventricular systolic function due to increased sarcoplasmic reticulum Ca²⁺ content and Ca²⁺-induced sarcoplasmic reticulum Ca²⁺ release. The increased availability of Ca²⁺ to maintain cardiomyocyte contraction is attributed to prolongation of the action potential due to inhibition of the transient outward potassium current as well as stimulation of the reverse mode of the Na⁺–Ca²⁺ exchange. Further progression of cardiac hypertrophy towards heart failure is due to abnormalities in Ca²⁺ handling, necrotic myocardial injury, and increased myocardial stiffness due to interstitial fibrosis.     

Increased susceptibility to Ca2+‐induced permeability transition and to cytochrome c release in rat heart mitochondria with aging: effect of melatonin

Abstract: Aging is associated with a decline of cardiac function. The mitochondrial permeability transition (MPT) may be a factor in cardiac dysfunction associated with aging. We investigated the effect of aging and long‐term treatment with melatonin (approximately 10 mg/kg b.w./day for 2 months), a known natural antioxidant, on the susceptibility to Ca²⁺‐induced MPT opening and cytochrome c release in rat heart mitochondria. The mitochondrial content of normal and oxidized cardiolipin as a function of aging and melatonin treatment was also analyzed. Mitochondria from aged rats (24 month old) displayed an increased susceptibility to Ca²⁺‐induced MPT opening, associated with an elevated release of cytochrome c, when compared with young control animals (5 month old). Melatonin treatment counteracted both these processes. Aging was also associated with an oxidation/depletion of cardiolipin which could be counteracted as well by melatonin. It is proposed that the increased level of oxidized cardiolipin could be responsible, at least in part, for the increased susceptibility to Ca²⁺‐induced MPT opening and cytochrome c release in rat heart mitochondria with aging. Melatonin treatment counteracts both these processes, most likely, by preventing the oxidation/depletion of cardiolipin. Our results might have implications in the necrotic and apoptotic myocytes cell death in aged myocardium, particularly in ischemia/reperfusion injury.     

Deficiency in AMP-activated protein kinase exaggerates high fat diet-induced cardiac hypertrophy and contractile dysfunction

AMPK, a metabolic sensor, protects against ischemic injury and cardiac hypertrophy although its role in obesity is unclear. This study was designed to examine the impact of AMPK deficiency on cardiac dysfunction following high fat feeding. Adult WT and transgenic mice overexpressing a kinase dead (KD) α2 isoform (K45R mutation) of AMPK were fed a low or high fat diet for 20 weeks. DEXA was used to confirm adiposity. Wheat germ agglutinin immunostaining was used to evaluate myocardial histology. Myocardial function was evaluated using echocardiography and edge-detection. AMPK activity was analyzed using fluorescence polarization assays. [1-¹⁴C] oleate was used to determine fatty acid oxidation. Expression of AMPK, α1, α2, ACC, Akt, the Glut-4 translocation mediator Akt substrate of 160KD (AS160), mTOR, total and membrane Glut-4 was evaluated using Western blot. AMPK activity was decreased in KD mice regardless of diet regimen. High fat diet led to obesity, glucose intolerance and cardiac hypertrophy with accentuated glucose intolerance, dampened fatty acid oxidation and cardiac hypertrophy in KD mice. High fat feeding triggered lower fractional shortening, increased LV mass, left ventricular end diastolic/systolic diameter, decreased PS, ± dL/dt, prolonged TR₉₀ and intracellular Ca²⁺ mishandling with a more pronounced effect in KD mice. High fat diet and AMPK KD lessened AMPKα2 isoform activity and ACC phosphorylation. AMPK deficiency unveiled or accentuated high fat diet-induced decrease in phosphorylation of Akt and AS160, membrane fraction of Glut-4 and mTOR expression (a greater mTOR phosphorylation). Taken together, these data suggest that AMPK deficiency exacerbates obesity-induced cardiac hypertrophy and contractile dysfunction, possibly associated with AS160 and mTOR signaling.     

Abnormalities of calcium metabolism and myocardial contractility depression in the failing heart

Heart failure (HF) is characterized by molecular and cellular defects which jointly contribute to decreased cardiac pump function. During the development of the initial cardiac damage which leads to HF, adaptive responses activate physiological countermeasures to overcome depressed cardiac function and to maintain blood supply to vital organs in demand of nutrients. However, during the chronic course of most HF syndromes, these compensatory mechanisms are sustained beyond months and contribute to progressive maladaptive remodeling of the heart which is associated with a worse outcome. Of pathophysiological significance are mechanisms which directly control cardiac contractile function including ion- and receptor-mediated intracellular signaling pathways. Importantly, signaling cascades of stress adaptation such as intracellular calcium (Ca²⁺) and 3′-5′-cyclic adenosine monophosphate (cAMP) become dysregulated in HF directly contributing to adverse cardiac remodeling and depression of systolic and diastolic function. Here, we provide an update about Ca²⁺ and cAMP dependent signaling changes in HF, how these changes affect cardiac function, and novel therapeutic strategies which directly address the signaling defects.