Erythropoietin (EPO), well known for its role in stimulation of erythropoiesis, has recently been shown to have a dramatic neuroprotective effect in animal models of cerebral ischemia, mechanical trauma of the nervous system, and excitotoxins, mainly by reducing apoptosis. We studied the effect of single systemic administration of recombinant human EPO (rhEPO) on left ventricular (LV) size and function in rats during 8 weeks after the induction of a myocardial infarction (MI) by permanent ligation of the left descending coronary artery. We found that an i.p. injection of 3,000 units͞kg of rhEPO immediately after the coronary artery ligation resulted, 24 h later, in a 50% reduction of apoptosis in the myocardial area at risk. Eight weeks after the induction of MI, rats treated with rhEPO had an infarct size 15-25% of the size of that in untreated animals. The reduction in myocardial damage was accompanied by reductions in LV size and functional decline as measured by repeated echocardiography. Thus, a single dose of rhEPO administered around the time of acute, sustained coronary insufficiency merits consideration with respect to its therapeutic potential to limit the extent of resultant MI and contractile dysfunction. E rythropoietin (EPO), a cytokine produced by the adult kidney, is a well known hematopoietic factor. EPO receptors (EPO-Rs) are expressed in adult bone marrow and spleen and are activated by hypoxia (1). Whether EPO-Rs are present in nonhematopoietic tissues is less certain. The predominant opinion is that the expression of EPO-Rs in nonhematopoietic tissues is limited to the fetal stage of development (2). Although some studies failed to detect EPO-R transcripts in the brain, kidney, liver, or heart of adult mice (3), others have reported an intensive immunoreactivity for EPO-Rs in many medium and large neurons of adult rat brain (4). Moreover, a weak EPO-R immunoreactivity of human brain was amplified by hypoxia (5). Recently, EPO-Rs have also been identified in the adult retina of mice (6). Although EPO-Rs have not been identified in adult hearts, their presence during embryogenesis is critical for cardiac development (7).Recombinant human EPO (rhEPO) is widely used for the treatment of anemia occurring in the context of surgery, cancer, HIV, kidney failure, etc. (8). Recently, rhEPO has been shown to have a dramatic neuroprotective effect in animal models of cerebral ischemia and mechanical trauma of the nervous system, and in response to excitotoxins. A single intracerebroventricular injection and, more importantly, systemic administration of rhEPO have resulted in a 50-75% reduction in brain injury induced by the focal ischemia (4). A reduction of apoptosis is a mechanism involved in this neuroprotective effect of rhEPO (9, 10).We hypothesized that the protective effect of systemic rhEPO administration that resulted in improvement of brain cell survival after cerebral ischemia would also occur in the ischemic heart model. Specifically, we studied the effect of a single systemic administration...

SummaryThe citric acid cycle (CAC) metabolite fumarate has been proposed to be cardioprotective; however, its mechanisms of action remain to be determined. To augment cardiac fumarate levels and to assess fumarate's cardioprotective properties, we generated fumarate hydratase (Fh1) cardiac knockout (KO) mice. These fumarate-replete hearts were robustly protected from ischemia-reperfusion injury (I/R). To compensate for the loss of Fh1 activity, KO hearts maintain ATP levels in part by channeling amino acids into the CAC. In addition, by stabilizing the transcriptional regulator Nrf2, Fh1 KO hearts upregulate protective antioxidant response element genes. Supporting the importance of the latter mechanism, clinically relevant doses of dimethylfumarate upregulated Nrf2 and its target genes, hence protecting control hearts, but failed to similarly protect Nrf2-KO hearts in an in vivo model of myocardial infarction. We propose that clinically established fumarate derivatives activate the Nrf2 pathway and are readily testable cytoprotective agents.

Background-Intermittent fasting (IF), a dietary regimen in which food is available only every other day, increases the life span and reduces the incidence of age-associated diseases in rodents. We have reported neuroprotective effects of IF against ischemic injury of the brain. In this study, we examined the effects of IF on ischemic injury of the heart in rats. Methods and Results-After 3 months of IF or regular every-day feeding (control) diets started in 2-month-old rats, myocardial infarction (MI) was induced by coronary artery ligation. Twenty-four hours after MI, its size in the IF group was 2-fold smaller, the number of apoptotic myocytes in the area at risk was 4-fold less, and the inflammatory response was significantly reduced compared with the control diet group. Serial echocardiography revealed that during 10 weeks after MI (with continuation of the IF regimen), the left ventricular (LV) remodeling and MI expansion that were observed in the control diet group were absent in the IF group. In a subgroup of animals with similar MI size at 1 week after MI, further observation revealed less remodeling, better LV function, and no MI expansion in the IF group compared with the control group. Conclusions-IF protects the heart from ischemic injury and attenuates post-MI cardiac remodeling, likely via antiapoptotic and antiinflammatory mechanisms.

Background-Studies in isolated cardiac myocytes have demonstrated that signaling via specific ␤ 1 -adrenergic receptor subtypes (␤ 1 ARs) promotes but that signaling via ␤ 2 ARs protects from cell death. We hypothesized that prolonged ␤ 2 AR stimulation or ␤ 1 AR blockade would each protect myocytes from death and thereby ameliorate cardiac remodeling in chronic heart failure. Methods and Results-A large myocardial infarction (MI) induced in rats by coronary artery ligation resulted in a dilated cardiomyopathy (DCM) characterized by infarct expansion and a progressive increase in left ventricular (LV) end-diastolic volume, accompanied by a reduction in ejection fraction (EF), as assessed by repeated echocardiography. Pressure-volume analysis at 8 weeks after ligation showed that diastolic stiffness (Eed) and arterial elastance (Ea) were increased, end-systolic elastance (Ees) was decreased, and arterioventricular (AV) coupling (Ea/Ees) had deteriorated. Apoptosis was present in both peri-infarct and remote myocardium. Chronic (6-week) administration of the ␤ 2 AR agonists fenoterol or zinterol, starting at 2 weeks after MI, reduced the extent of LV dilation, infarct expansion, and EF decline. The ␤ 1 AR blocker metoprolol did not affect the former and preserved EF to a lesser extent than did the ␤ 2 AR agonists. At 8 weeks after ligation, apoptosis was reduced by all drugs but to a greater extent by ␤ 2 AR agonists than by the ␤ 1 AR blocker. Both ␤ 2 AR agonists and the ␤ 1 AR blocker improved AV coupling, the former mainly by reducing Ea and the latter mainly by increasing Ees. Only the ␤ 2 AR agonists reduced the Eed and the MI size by reducing infarct expansion. Conclusions-These results provide proof of concept for the efficacy of chronic ␤ 2 AR stimulation in this DCM model.

Heart rate (HR) and HR variability (HRV), predictors of over-all organism health, are widely believed to be driven by autonomic input to the sinoatrial node (SAN), with sympathetic input increasing HR and reducing HRV. However, variability in spontaneous beating intervals in isolated SAN tissue and single SAN cells, devoid of autonomic neural input, suggests that clocks intrinsic to SAN cells may also contribute to HR and HRV in vivo . We assessed contributions of both intrinsic and autonomic neuronal input mechanisms of SAN cell function on HR and HRV via in vivo , telemetric EKG recordings. This was done in both wild type (WT) mice, and those in which adenylyl cyclase type 8 (ADCY8), a main driver of intrinsic cAMP-PKA-Ca 2+ mediated pacemaker function, was overexpressed exclusively in the heart (TG AC8 ). We hypothesized that TG AC8 mice would: (1) manifest a more coherent pattern of HRV in vivo , i.e., a reduced HRV driven by mechanisms intrinsic to SAN cells, and less so to modulation by autonomic input and (2) utilize unique adaptations to limit sympathetic input to a heart with high levels of intrinsic cAMP-Ca 2+ signaling. Increased adenylyl cyclase (AC) activity in TG AC8 SAN tissue was accompanied by a marked increase in HR and a concurrent marked reduction in HRV, both in the absence or presence of dual autonomic blockade. The marked increase in intrinsic HR and coherence of HRV in TG AC8 mice occurred in the context of: (1) reduced HR and HRV responses to β-adrenergic receptor (β-AR) stimulation; (2) increased transcription of genes and expression of proteins [β-Arrestin, G Protein-Coupled Receptor Kinase 5 (GRK5) and Clathrin Adaptor Protein (Dab2)] that desensitize β-AR signaling within SAN tissue, (3) reduced transcripts or protein levels of enzymes [dopamine beta-hydorxylase (DBH) and phenylethanolamine N -methyltransferase (PNMT)] required for catecholamine production in intrinsic cardiac adrenergic cells, and (4) substantially reduced plasma catecholamine levels. Thus, mechanisms driven by cAMP-PKA-Ca 2+ signaling intrinsic to SAN cells underlie the marked coherence of TG AC8 mice HRV. Adaptations to limit additional activation of AC signaling, via decreased neuronal sympathetic input, are utilized to ensure the hearts survival and prevent Ca 2+ overload.

Background A reduction of complexity of heart-beat interval variability (BIV) that is associated with an increased morbidity and mortality in cardiovascular disease states is thought to derive from the balance of sympathetic and parasympathetic neural impulses to the heart. But rhythmic clock-like behavior intrinsic to pacemaker cells within the sinoatrial node (SAN) drives their beating, even in the absence of autonomic neural input. Objective To test how this rhythmic clock-like behavior intrinsic to pacemaker cells interacts with autonomic impulses to the heart-beat interval variability in vivo. Methods We analyzed BIV in the time and frequency domains and by fractal and entropy analyses: i) in vivo, when the brain input to the SAN is intact; ii) during autonomic denervation in vivo; iii) in isolated SAN tissue (i.e., in which the autonomic-neural input is completely absent); iv) in single pacemaker cells isolated from the SAN; and v) following autonomic receptor stimulation of these cells. Results Spontaneous-beating intervals of pacemaker cells residing within the isolated SAN tissue exhibit fractal-like behavior and have lower approximate entropy than in the intact heart. Isolation of pacemaker cells from SAN tissue, however, leads to a loss in the beating-interval order and fractal-like behavior. β adrenergic receptor stimulation of isolated pacemaker cells increases intrinsic clock synchronization, decreases their action potential period and increases system complexity. Conclusions Both the average-beating interval in vivo and beating interval complexity are conferred by the combined effects of clock periodicity intrinsic to pacemaker cells and their response to autonomic-neural input.

We have reported therapeutic effectiveness of pharmacological stimulation of ␤ 2 adrenoreceptors (ARs) to attenuate the cardiac remodeling and myocardial infarction (MI) expansion in a rat model of dilated cardiomyopathy (DCM) post-MI. Furthermore, the combination of ␤ 2 AR stimulation with ␤ 1

Multiple health benefits of calorie restriction (CR) and alternate day fasting (ADF) regimens are widely recognized. Experimental data concerning the effects of calorie restriction on cardiac health are more controversial, ranging from evidence that ADF protects heart from ischemic damage but results in developing of diastolic dysfunction, to reports that CR ameliorates the age-associated diastolic dysfunction. Here we investigated the effects of chronic CR on morphology and function of the cardiovascular system of aged rats and cardioprotective effect of CR against ischemic damage in the experimental rat model of MI. Cardiovascular fitness of 24-mo old Fisher 344 rats maintained through life on ad libitum (AL) or CR diets was extensively evaluated via echocardiography, dobutamine stress test, pressure-volume loop analyses, pulse wave velocity measurements, and histology. Groups of 2-mo old AL and 29-mo old CR rats were studied for comparison. Myocardial infarction (MI) was induced by a permanent ligation of the anterior descending coronary artery in 5-mo old rats maintained for 3 months on CR or AL. MI size was evaluated histologically 24 hrs following coronary ligation. Cardiac remodeling was followed-up via echocardiography. Age-associated changes in 24-mo old rats consisted of 33% increase of fibrosis in the myocardium and more than 2 fold increase of the collagen in the tunica media of the aorta. There was a significant decrease in the density and total number of cardiomyocytes, while their size was increased. These morphological changes were manifested in a decline of systolic and diastolic cardiac function, increase of left ventricular and aortic stiffness, and arterio-ventricular uncoupling. Tachycardic response to dobutamine challenge was absent in the old rats. Compared to AL rats, 24-mo old CR rats had reduced levels of cardiac and aortic fibrosis, increased density of cardiomyocytes that were smaller in size, attenuated diastolic dysfunction, normal systolic function and arterio-ventricular coupling. Tachycardic response to dobutamine was also intact in CR 24-mo old rats and aortic stiffness was reduced. Adjustment for body weight differences through ratiometric or allometric scaling did not affect the overall pattern of differences between AL and CR rats. Attenuation of morphological and functional age-associated changes in 24-mo old CR rats either was not observed at all or was smaller in 29-mo old CR rats. Size of MI induced by a permanent coronary ligation as well as post-MI cardiac remodeling and function were similar in CR and AL rats. CR does not increase tolerance of myocardium to ischemic damage, but attenuates the age-associated changes in the heart and major vessels. The attenuation of age-associated changes by CR cannot be explained by the effect of lower body weight but are attributable to more intimate cellular mechanisms of CR itself. Attenuation of age-associated changes by CR waned with advancing age, and is consistent with the idea that CR postponed senescence.