Phosphorylation of cardiac sodium channel at Ser571 anticipates manifestations of the aging myopathy.

Diastolic dysfunction and delayed ventricular repolarization are typically observed in the elderly, but whether these defects are intimately associated with the progressive manifestation of the aging myopathy remains to be determined. In this regard, aging in experimental animals is coupled with increased late Na+ current (INa,L) in cardiomyocytes, raising the possibility that INa,L conditions the modality of electrical recovery and myocardial relaxation of the aged heart. For this purpose, aging male and female wild-type (WT) C57Bl/6 mice were studied together with genetically engineered mice with phosphomimetic (gain of function, GoF) or ablated (loss of function, LoF) mutations of the sodium channel Nav1.5 at Ser571 associated with, respectively, increased and stabilized INa,L. At ∼18 mo of age, WT mice developed prolonged duration of the QT interval of the electrocardiogram and impaired diastolic left ventricular (LV) filling, defects that were reversed by INa,L inhibition. Prolonged repolarization and impaired LV filling occurred prematurely in adult (∼5 mo) GoF mutant mice, whereas these alterations were largely attenuated in aging LoF mutant animals. Ca2+ transient decay and kinetics of myocyte shortening/relengthening were delayed in aged (∼24 mo) WT myocytes, with respect to adult cells. In contrast, delayed Ca2+ transients and contractile dynamics occurred at adult stage in GoF myocytes and further deteriorated in old age. Conversely, myocyte mechanics were minimally affected in aging LoF cells. Collectively, these results document that Nav1.5 phosphorylation at Ser571 and the late Na+ current modulate the modality of myocyte relaxation, constituting the mechanism linking delayed ventricular repolarization and diastolic dysfunction.NEW & NOTEWORTHY We have investigated the impact of the late Na current (INa,L) on cardiac and myocyte function with aging by using genetically engineered animals with enhanced or stabilized INa,L, due to phosphomimetic or phosphoablated mutations of Nav1.5. Our findings support the notion that phosphorylation of Nav1.5 at Ser571 prolongs myocardial repolarization and impairs diastolic function, contributing to the manifestations of the aging myopathy.

Inhibition of soluble epoxide hydrolase increases coronary perfusion in mice.

Roles of soluble epoxide hydrolase (sEH), the enzyme responsible for hydrolysis of epoxyeicosatrienoic acids (EETs) to their diols (DHETs), in the coronary circulation and cardiac function remain unknown. We tested the hypothesis that compromising EET hydrolysis/degradation, via sEH deficiency, lowers the coronary resistance to promote cardiac perfusion and function. Hearts were isolated from wild type (WT), sEH knockout (KO) mice and WT mice chronically treated with t-TUCB (sEH inhibitor), and perfused with constant flow at different pre-loads. Compared to WT controls, sEH-deficient hearts required significantly greater basal coronary flow to maintain the perfusion pressure at 100 mmHg and exhibited a greater reduction in vascular resistance during tension-induced heart work, implying a better coronary perfusion during cardiac performance. Cardiac contractility, characterized by developed tension in response to changes in preload, was potentially increased in sEH-KO hearts, manifested by an enlarged magnitude at each step-wise increase in end-diastolic to peak-systolic tension. 14,15-EEZE (EET antagonist) prevented the adaptation of coronary circulation in sEH null hearts whereas responses in WT hearts were sensitive to the inhibition of NO. Cardiac expression of EET synthases (CYP2J2/2C29) was comparable in both genotypic mice whereas, levels of 14,15-, 11,12- and 8,9-EETs were significantly higher in sEH-KO hearts, accompanied with lower levels of DHETs. In conclusion, the elevation of cardiac EETs, as a function of sEH deficiency, plays key roles in the adaptation of coronary flow and cardiac function.

Elevated level of pro-inflammatory eicosanoids and EPC dysfunction in diabetic patients with cardiac ischemia.

BACKGROUND: Circulating endothelial progenitor cells (EPCs) are recruited from the blood system to sites of ischemia and endothelial damage, where they contribute to the repair and development of blood vessels. Since numerous eicosanoids including leukotrienes (LTs) and hydroxyeicosatetraenoic acids (HETEs) have been shown to exert potent pro-inflammatory activities, we examined their levels in chronic diabetic patients with severe cardiac ischemia in conjunction with the level and function of EPCs. RESULTS: Lipidomic analysis revealed a diabetes-specific increase (p<0.05) in inflammatory and angiogenic eicosanoids including the 5-lipoxygenase-derived LTB (4.11±1.17 vs. 0.96±0.27 ng/ml), the lipoxygenase/CYP-derived 12-HETE (117.08±35.05 vs. 24.34±10.03 ng/ml), 12-HETrE (17.56±4.43 vs. 4.15±2.07 ng/ml), and the CYP-derived 20-HETE (0.32±0.04 vs. 0.06±0.05 ng/ml) the level of which correlated with BMI (p=0.0027). In contrast, levels of the CYP-derived EETs were not significantly (p=0.36) different between these two groups. EPC levels and their colony-forming units were lower (p<0.05) with a reduced viability in diabetic patients compared with non-diabetics. EPC function (colony-forming units (CFUs) and MTT assay) also negatively correlated with the circulating levels of HgA1C. CONCLUSION: This study demonstrates a close association between elevated levels of highly pro-inflammatory eicosonoids, diabetes and EPC dysfunction in patients with cardiac ischemia, indicating that chronic inflammation impact negatively on EPC function and angiogenic capacity in diabetes.

Effect of prenatal glucocorticoids on cerebral vasculature of the developing brain.

BACKGROUND AND PURPOSE: Prenatal glucocorticoids prevent germinal matrix hemorrhage in premature infants. The underlying mechanism, however, is elusive. Germinal matrix is enriched with angiogenic vessels exhibiting paucity of pericytes and glial fibrillary acidic protein-positive astrocyte end feet. Therefore, we asked whether glucocorticoid treatment would suppress angiogenesis and enhance periendothelial coverage by pericytes and glial fibrillary acidic protein-positive end feet in the germinal matrix microvasculature. METHODS: We treated pregnant rabbits with intramuscular betamethasone and delivered pups prematurely by cesarean section at E29 (term=32 days). Endothelial turnover, vascular density, pericyte coverage, glial fibrillary acidic protein-positive end feet, cell death, and growth factors orchestrating angiogenesis, including vascular endothelial growth factor, angiopoietins, transforming growth factor-beta, and platelet-derived growth factor-B, were compared between betamethasone-treated and untreated pups. Similar comparisons were done between autopsy materials from premature infants exposed and unexposed to prenatal glucocorticoids. RESULTS: Antenatal glucocorticoid treatment reduced endothelial proliferation, vascular density, and vascular endothelial growth factor expression in the germinal matrix of both rabbits and humans. The pericyte coverage was greater in glucocorticoid-treated rabbit pups and human infants than in controls, but not the glial fibrillary acidic protein-positive end feet coverage. Transforming growth factor-beta, but not angiopoietins and platelet-derived growth factor-B, were elevated in glucocorticoid-treated rabbit pups compared with controls. Betamethasone treatment induced apoptosis, neuronal degeneration, and gliosis in rabbit pups. However, there was no evidence of increased cell death in glucocorticoid-exposed human infants. CONCLUSIONS: Prenatal glucocorticoid suppresses vascular endothelial growth factor and elevates transforming growth factor-beta levels, which results in angiogenic inhibition, trimming of neovasculature, and enhanced pericyte coverage. These changes contribute to stabilizing the germinal matrix vasculature, thereby reducing its propensity to hemorrhage. Prenatal glucocorticoid exposure does not induce neural cell death in humans, unlike rabbits.

Cardiac structure and function in humans: a new cardiovascular physiology laboratory.

As the traditional cardiovascular control laboratory has disappeared from the first-year medical school curriculum, we have recognized the need to develop another "hands-on" experience as a vehicle for wide-ranging discussions of cardiovascular control mechanisms. Using an echocardiograph, an automatic blood pressure cuff, and a reclining bicycle, we developed protocols to illustrate the changes in cardiac and vascular function that occur with changes in posture, venous return, and graded exercise. We use medical student volunteers and a professional echocardiographer to generate and acquire data, respectively. In small-group sessions, we developed an interactive approach to discuss the data and to make a large number of calculations from a limited number of measurements. The sequence of cardiac events and cardiac structure in vivo were illustrated with the volunteers lying down, standing, and then with their legs raised passively above the heart to increase venous return. Volunteers were then asked to peddle the bicycle to achieve steady-state heart rates of 110 and 150 beats/min. Data were collected in all these states, and calculations were performed and used as the basis of a small-group discussion to illustrate physiological principles. Information related to a surprisingly large number of cardiovascular control mechanisms was derived, and its relevance to cardiovascular dysfunction was explored. This communication describes our experience in developing a new cardiovascular control laboratory to reinforce didactic material presented in lectures and small-group sessions.

Emotional fever after habituation to the temperature-recording procedure.

To examine whether habituation to having temperatures taken might reduce the emotional fever induced by other stressors, 20 rats were habituated by having three colonic temperatures taken within 6 min twice a week for 8 weeks. Two novel stressors were added during Week 9; rats were given an intraperitoneal saline injection before taking their temperatures on Day 1, and on a second day, they were housed with a group of unfamiliar animals for 5 min before taking their temperatures. Temperatures during Weeks 10-11 were taken as during Weeks 1-8. Results showed that, overall, the third daily temperature averaged 1 degrees C higher than the first, indicating that taking temperatures was stressful. Between weeks, temperatures rose between Weeks 1 and 2, suggesting a conditioned fever. Temperatures fell 1.5 degrees C from Weeks 2-8, indicating habituation. During Week 9, temperatures after the intraperitoneal injection were no higher than during Week 8; however, 5 min of group housing raised temperatures 1.5 degrees C for males and 2.5 degrees C for females. Temperatures during Weeks 10 and 11 were below those of Week 8, suggesting that the fever induced by group housing in Week 9 was situation specific. These results indicate that rats habituated to having their temperatures taken exhibit emotional fever only to selected new stimuli. Body temperatures do not rise after an intraperitoneal injection given by a familiar handler, but briefly placing rats with unfamiliar animals induces a substantial febrile response.

Plasma constituents and mortality in rat pups given chronic insulin via injection, pellet, or osmotic minipump.

Two studies compared the glucose responses of 9-day-old rats given subcutaneous insulin, either continuously or via daily injection, for 10 days. In Experiment 1, implanted pellets released a total of 0, 1.9, or 5.7 U insulin/kg the first 24 h. Injected doses were larger, 0 or 8 U/kg. Injections caused no deaths, but insulin-releasing pellets caused high mortality within 24 h. Pups surviving the pellets were normoglycemic by treatment day 8. In Experiment 2, pups received 0.184 U of insulin daily, approximately 8 U/kg at 9 days, via either injection or osmotic minipump. All pups survived. Injected pups were hypoglycemic 2 h postinjection through treatment day 10, whereas pups with insulin minipumps were normoglycemic by day 5. Insulin injections, but not minipumps, lowered plasma triglycerides on day 10. To examine age differences in response to insulin, additional pups and adults received daily injections of 0 or 8 U/kg for 10 days. All survived. Insulin lowered plasma glucose more in pups than in adults and reduced triglycerides in pups but not in adults. The rapid development of normoglycemia in pups with insulin minipumps, compared with pups injected daily with the same dose, suggests that continuous early insulin may produce insulin resistance.

Regulation of renal A(1) adenosine receptors in vivo.

We have compared renal A(1) adenosine receptor (AR) regulations in rats after chronic agonist and antagonist treatments. In one group, R-phenylisopropyladenosine (R-PIA), a selective A(1) AR agonist, was infused subcutaneously for 7 days. Another group was fed theophylline, a non-selective AR antagonist, for 2 weeks. Renal cortex membrane A(1) AR binding with 1,3-[(3)H]-dipropyl-8-cyclopentylxanthine demonstrated approximately 40% reduction in the B(max ) for the R-PIA group without any changes in the K(d) values. Neither the B(max) nor the K(d) changed following chronic theophylline treatment. Renal cortex G(i)alpha-proteins from the R-PIA treated rats decreased by approximately 30%. Renal G(i)alpha levels did not change in theophylline-treated rats. Consistent with the A(1) AR desensitization, R-PIA-treated rats had significantly higher basal renin release and showed attenuated A(1) AR-mediated inhibition of renin release. These data suggest that prolonged A(1) AR stimulation results in downregulation of renal A(1) ARs and G(i)alpha, accompanied by desensitization of A(1) AR-mediated inhibitory effects on renin release. Unlike cardiac and brain A(1) ARs, renal A(1) receptors are not subject to up-regulation following chronic antagonist treatment.