Search

Background: Diabetic cardiomyopathy (DCM) increases the risk of heart failure. As yet, no effective therapeutic strategies exist. Recent evidence indicates that intracellular Na + concentration ([Na + ] i ) is augmented in the myocytes from diabetic hearts, where it causes oxidative stress, augments the sarcoplasmic reticulum Ca 2+ leak and contributes to electrical, structural and functional remodeling. Ranolazine (RAN), inhibiting persistent or late inward Na + current has been proposed to be a therapeutic choice for DCM. However, the role and mechanism of chronic RAN in DCM are unclear. We assessed the hypothesis that RAN improves myocyte function, [Ca 2+ ] i regulation, and β-adrenergic receptor (AR) signaling effectiveness, thus limiting DCM. Methods: We compared LV myocyte function, [Ca 2+ ] i transient ([Ca 2+ ] iT ) and responses to the stimulation of β-AR in 3 groups wild-type (WT) female mice over 10 weeks (W):1) DM (n=8), 10 W after receiving streptozotocin (STZ, 200 mg/kg, ip); 2) DM/RAN (n=6), 6 W after STZ, RAN (10 -5 M/kg/day, mini-pump) was initiated and was given for 4 W; and 3) Sham controls (C) (n=8). Results: Versus control, STZ-treated WT mice had DM with significantly elevated blood glucose levels (410 vs 128mg/dl) followed by LV myocyte dysfunction with decreases in myocyte contractility (dL/dt max ) (75.0 vs 140.1 μm/s), relengthening (dR/dt max ) (62.5 vs 116.6 μm/s) and [Ca 2+ ] iT (0.15 vs 0.22). In DM myocytes, the ability of β-AR agonist, isoproterenol (ISO, 10 -8 M) to increase cell contractility was blunted. Versus control, in DM myocytes, ISO-induced increases in dL/dt max (31% vs 60%), dR/dt max (23% vs 50%) and [Ca 2+ ] iT (15% vs 30%) were significantly reduced. By contrary, versus DM alone, DM/RAN myocytes showed normal basal cell contraction (137.8 μm/s), relaxation (117.2 μm/s) and [Ca 2+ ] iT (0.22) with preserved ISO-stimulated positive inotropic effect. Compared control, in DM/RAN, ISO caused similar increases in dL/dt max (62% vs 60%), dR/dt max (52% vs 50%) and [Ca 2+ ] iT (32% vs 30%). Conclusion: Chronic ranolazine leads to the preservation of myocyte function, [Ca 2+ ] iT and β-AR responsiveness in DCM. Thus, antagonizing myocyte [Na + ] i dysregulation might provide a new therapeutic strategy for DM-related decline in myocardial function.

Background: Previously we showed that rats expressing the human angiotensinogen (AGT) gene (hAogen) exhibit sustained hypertension and cardiac dysfunction, associated with increased cardiac angiotensin II (Ang II). Recent evidence indicates that Ang II increases CaMKII activation which plays a crucial role in ANG II-related pathological processes associated with hypertension, hypertrophy and cardiac dysfunction. Although CaMKII inhibitors are being developed as safe and effective therapeutic agents, their mechanism of action remains to be fully understood. We tested the hypothesis that in hAogen rats, CaMKII is upregulated. Increased CaMKII activation may produce direct depression in LV myocyte basal function and β-adrenergic reserve. Acute inactivation of CaMKII with KN93 would have beneficial actions in hAogen. Methods: We compared LV myocyte CaMKIIδ expression and activity and myocyte functional performance in 10 Sprague Dawley (SD) control and 10 hAogen male rats. In hAogen, we further assessed myocyte contractile and [Ca 2+ ] i transient ([Ca 2+ ] iT ) responses to β-AR stimulation by isoproterenol (ISO, 10 -8 M) with and without pretreatment of myocytes with KN93 (10 -6 M, 30 min). Results: Versus SD, in hAogen myocytes, the CaMKIIδ protein levels (19%, 0.50 vs 0.42) and CaMKIIδ phosphorylation (at Thr 287 ) (35%, 0.27 vs 0.20) were significantly increased. These changes were followed by significantly reduced cell contraction (dL/dtmax, 104.9 vs 138.4 μm/s), relaxation (dR/dtmax, 93.0 vs 104.2 μm/s) and [Ca 2+ ] iT (0.15 vs 0.20). ISO-stimulated increases in dL/dtmax (43% vs 62%), dR/dtmax (32% vs 49%) and [Ca 2+ ] iT (19% vs 29%) were also significantly reduced. Moreover, in hAogen myocytes, pretreatment with KN93 markedly improved myocyte basal contraction (134.4 μm/s), relaxation (107.4 μm/s) and [Ca 2+ ] iT (0.20). ISO-induced increases in dL/dtmax (60%), dR/dtmax (50%), and [Ca 2+ ] iT (31%) were also significantly augmented. Conclusions: In rats harboring the human AGT gene, upregulation of CaMKIIδ has adverse functional effects. Acute endogenous CaMKII activation exacerbates LV myocyte dysfunction and β-AR desensitization. Inhibition of CaMKII has a significant beneficial impact in this established humanized model of hypertension.

Background: The transgenic rats expressing the human AGT gene [TGR (hAGT) L1623] (hAogen) is an established humanized model of hypertension. Recently we further found that this model is associated with declines in cardiac contractile function and β-adrenergic receptor (AR) reserve. However, the molecular basis is unclear. It has been suggested that pivotal restructuring of β-AR system is a critical determinant of the dysfunctional β-AR regulation in several cardiovascular diseases, including hypertension. But the alterations of subtypes of β-ARs have not been systemically evaluated in this model. Furthermore, there is significant crosstalk between β-AR signaling and CaMKII activation presenting CaMKII as a possible downstream mediator of detrimental β-AR signaling in the abnormal contractile phenotype of cardiac hypertrophy. However, the changes in cardiac CaMKII in this model are unknown. We tested the hypothesis that in hAogen, CaMKII and β 3 -AR are upregulated in the myocardium, but β 1 -AR is down-regulated, which are important causes of cardiac dysfunction and β-AR desensitization. Methods: We compared LV myocyte CaMKIIδ expression and activity and β 1 - and β 3 -AR expression and determined myocyte functional performance and [Ca 2+ ] i transient ([Ca 2+ ] iT) response to β -AR stimulation in myocytes obtained from 6 Sprague Dawley (SD) control and 6 haogen male adult rats. Results: Compared with SD, in hAogen LV myocytes, β 3 -AR protein level significantly increased (26%, 0.29 vs 0.23), but β 1 -AR protein level (26%, 0.4 vs 0.54) significantly decreased. The CaMKIIδ protein levels (16%, 0.50 vs 0.43) and CaMKIIδ phosphorylation (at Thr287) (35%, 0.27 vs 0.20) were significantly increased. These changes were followed by significantly reduced cell contraction (dL/dt max , 102.1vs 138.8μm/s), relaxation (dR/dt max , 94.0 vs 104.7 μm/s) and [Ca 2+ ] iT (0.15 vs 0.21). ISO-stimulated increases in dL/dt max (42% vs 60%), dR/dt max (36% vs 51%) and [Ca 2+ ] iT (20% vs 31%) were also significantly reduced. Conclusions: The upregulation of LV myocyte CaMKIIδ and contrast changes in β 3 -AR and β 1 -AR may be the important cause of the abnormal contractile phenotype and β-AR desensitization in this humanized angiotensinogen model of hypertension of TGR (hAGT) L1623.

Background: Diabetic cardiomyopathy (DCM) is the main cause of increased mortality in Diabetes mellitus (DM). There are no effective therapeutic strategies. Recently, we found that DM is associated with the upregulation of β 3 adrenergic receptor (AR) mediated cardiac inhibitory pathway. This suggests cardiac β 3 -AR activation may contribute to DCM progression and be a therapeutic target. We hypothesize that upregulation of β 3 -AR is maladaptive, and DM-caused progressive decline in cardiac function and β-adrenergic reserve will be prevented in β 3 -AR knockout (β 3 KO) mice. Methods: Studies were conducted in female mice of 2 Vehicle controls groups (n=8/group) of wild-type (CWT) and Cβ 3 KO, and 2 Type 2 DM (T2) of T2WT (n=8) and T2β 3 KO (n=6).T2 was induced by fed high-fat diet (HFD) for 14 weeks (W), but after HFD for 4 W receiving streptozotocin (STZ, 40 mg/kg/day, i.p. for 5 days). We compared LV myocyte contractile and [Ca 2+ ] iT responses to β-AR subtype stimulation by random exposure of myocytes to the superfusion of isoproterenol (ISO,10 -8 M) or a selective β 1 -, or β 3 -agonist, Norepinephrine (NE, 10 -7 M), and BRL-37,344 (BRL, 10 -8 M), respectively. Results: Mice received HFD plus STZ developed T2DM with elevated mean blood glucose from 128 mg/dl of control to 388 mg/dl and 382 mg/dl in T2WT and T2β 3 KO, respectively. However only T2WT mice developed DCM followed by significant decreases in myocyte contraction (dL/dt max , T2WT: 74.8 vs CWT:140.1 μm/s), relaxation (dR/dt max , 58.0 vs 117.9 μm/s) and [Ca 2+ ] iT (0.16 vs 0.22). ISO-stimulated increases in dL/dt max (37% vs 58%), dR/dt max (30% vs 53%), and [Ca 2+ ] iT (19% vs 30%) were attenuated accompanied by a diminished NE-caused increase in dL/dt max (28% vs 41%), but enhanced BRL-induced decrease in dL/dt max (29% vs 15%). By contrary, T2β 3 KO showed normal basal dL/dt max (137.6 μm/s), dR/dt max (111.4 μm/s) and [Ca 2+ ] iT (0.22). Importantly, T2β 3 KO myocytes showed preserved ISO-stimulated increases in dL/dt max (60%), dR/dt max (51%) and [Ca 2+ ] iT (32%) and restored normal dL/dt max responses to NE (41%). Conclusions: β 3 KO prevents T2DM-caused contrast changes in β 1 - and β 3 -AR-stimulated cardiac inotropic actions and leading to the preservation of normal myocyte function, [Ca 2+ ] iT , and β-AR responsiveness in DCM.

Background: Diabetic cardiomyopathy (DCM) leads to progressive decline in cardiac function, increasing the risk for heart failure. There are no known effective prevention approaches or therapeutic strategies. Recent evidence in diabetes mellitus (DM) human and animal hearts suggests that the up-regulation of β 3 -adrenergic receptor (AR)-mediated inhibitory pathway may be responsible for the progression of DCM. However, its precise role is still being debated. We hypothesize that β 3 -AR antagonists (β 3 -ANT) may rescue the detrimental effects of β 3 -AR activation, improve cardiomyocyte function, and preserve normal β-AR regulation, leading to the regression of DCM. Methods: We compared LV myocyte function, [Ca 2+ ] i transient ([Ca 2+ ] iT ) at baseline and responses to β-AR simulation with isoproterenol (ISO,10 -8 M) in 3 groups wild-type female mice over 14 weeks (W):1) Type 2 DM ( T2 ) (n=9), 14 W fed high-fat diet (HFD), but after HFD for 4 W receiving streptozotocin (STZ, 40 mg/kg/day, i.p. 5 days); 2) T2/β 3 -ANT (n=7), T2 mice at 10 W received L-748,337, a selective β 3 -ANT (10 -7 M/kg/day, mini-pump) for 4 W; and 3) Vehicle controls (C) (n=9). Results: Versus C, T2DM was induced in mice received HFD and low dose STZ with significantly elevated blood glucose levels (T2: 388, T2/β 3 -ANT: 369 vs C: 128 mg/dl). In T2, LV myocyte basal function and [Ca 2+ ] iT regulation were impaired measured as significantly decreased myocyte contractility (dL/dt max ) (76.8 vs 135.2 μm/s), relengthening (dR/dt max ) (62.1 vs 113.8 μ m/s) and [Ca 2+ ] iT (0.16 vs 0.21). Furthermore, versus C, in T2 myocytes, ISO-induced increases in dL/dt max (T2: 40% vs C: 58%), dR/dt max (35% vs 54%) and [Ca 2+ ] iT (19% vs 30%) were significantly reduced. By contrary, versus T2, T2/β 3 -ANT myocytes showed normal basal cell contraction (127.8 μm/s), relaxation (109.4 μm/s) and [Ca 2+ ] iT (0.21) with preserved ISO-stimulated positive inotropic effect. Versus C, in T2/β 3 -ANT, ISO caused similar increases in dL/dt max (57%), dR/dt max (52%) and [Ca 2+ ] iT (30%). Conclusion: Chronic β 3 -ANT leads to the preservation of LV myocyte function, [Ca 2+ ] iT , and β-AR responsiveness in a mouse model of type 2 diabetes. Thus, antagonizing β 3 -AR might provide a new therapeutic strategy for DM-related decline in myocardial function.

Background: Transgenic rats expressing the human angiotensinogen gene [TGR (hAogen) 1623] (hAogen) exhibit sustained hypertension and systolic dysfunction associated with altered cardiac renin-angiotensin system and β- adrenergic receptor (AR) signal transduction systems. We have shown previously, in contrast to β 1 - and β 2 -ARs, β 3 -ARs are linked to G i proteins, and stimulation of β 3 -ARs inhibits cardiac contraction and relaxation. Hypertension is associated with increased cardiac expression of G i proteins. This may alter β 3 -AR stimulation and play an important role in the cardiac functional impairment of this humanized model of hypertension. However, the alteration and functional effect of β 3 -AR activation in this model are unknown. We tested the hypothesis that high cardiac tissue Ang II content in hAogen may cause an up-regulation of cardiac β 3 -AR, which exacerbates myocyte dysfunction and impairs calcium regulation, thereby directly contributing to the cardiac dysfunction. Methods: We compared LV myocyte β 3 -AR expression and myocyte functional and [Ca 2+ ] i transient ([Ca 2+ ] iT ) responses to β- and β 3 -AR stimulation in myocytes obtained from 8 Sprague Dawley (SD) control and 8 hAogen male adult rats. Results: Versus SD, in hAGT rats, basal myocyte contraction (dL/dt max , 107.1 vs 138.8 μm/s), relaxation (dR/dt max , 94.0 vs 103.9 μm/s) and [Ca 2+ ] iT (0.15 vs 0.21) were depressed. A non-selective β-AR agonist, isoproterenol (ISO) (10 -8 M) produced significantly smaller increases in dL/dt max , (42% vs. 61%), dR / dtmax (35% vs. 51%), and [Ca 2+ ] iT (20% vs. 30%). Moreover, versus SD, in hAogen rats, LV myocyte β 3 -AR protein level increased by 32% (0.29 vs 0.22) with resulted significantly altered myocyte functional response to β 3 -AR stimulation. Compared with the changes in SD myocytes, in hAogen myocytes, BRL (10 -8 M) produced a significantly greater decreases in dL/dt max , (27% vs 12%), dR/dt max (28% vs 11%), and [Ca 2+ ] iT (17% vs 10%). Conclusions: TGR (hAogen) 1623, this humanized model of hypertension is associated with an up-regulation of LV myocyte β 3 -AR, which enhances β 3 -AR-caused inhibition of myocyte contractile, relaxation and [Ca 2+ ] iT and exacerbates β-AR desensitization, thereby directly contributing to the cardiac dysfunction.

Background: The negative inotrope and up-regulation of β 3 -adrenergic receptors (AR) in human and animal failing hearts suggest a direct and contributing role of cardiac β 3 -AR activation on the progression of congestive heart failure (CHF). However, its precise role is still being debated. We hypothesize that up-regulation of cardiac β 3 -AR is detrimental and chronic β 3 -AR blockade may prevent CHF-caused intrinsic defects of left ventricular (LV) myocyte force-generating capacity and relaxation and improve β-AR regulation, thereby limiting the progression of CHF. Methods: We compared the alterations of LV and myocyte functional responses and [Ca 2+ ] i transient ([Ca 2+ ] iT ) in SD rats divided into 3 groups (8/group): 1) CHF 3 months after isoproterenol (ISO) (170 mg/kg, sq, for 2 days); 2) ISO/β 3 -ANT , 2 months after receiving ISO, a selective β 3 -AR antagonist (ANT), L-748,337, was initiated (10 -7 M/kg/day, sq. by mini-pump) and was given for 1 month; and 3) Sham controls . Results: Compared with controls, the animals that received ISO treatment had CHF onset at 1 month and progressed to severe HF at 3 months after ISO. Plasma norepinephrine (NE) (1295 vs 259 pg/ml) increased 5-fold; whereas, stroke volume (SV) (39 vs 91 μl) and ejection fraction (EF) (39 vs 62%) significantly decreased, and LV end-diastolic pressure (P ED ) (13.9 vs 6.0 mmHg) was doubled. These changes were paralleled with about 50% reductions in cell contraction (dL/dt max , 93 vs 186 μm/s) and relaxation (dR/dt max , 96 vs 159 μm/s) associated with a significant decrease in the peak systolic [Ca 2+ ] iT , (0.17 vs 0.26). In addition, superfusion of ISO (10 -8 M) caused much less increases in dL/dt max (39 vs 68%), dR/dt max (23 vs 54%), and [Ca 2+ ] IT (14 vs 28%). Treatment with β 3 -ANT increased SV (89 μl) and EF (60%), decreased P ED more than 90% from ISO-treated values, and corrected the elevation of plasma NE (301 pg/ml), dL/dt max (184 μm/s), dR/dt max (152 μm/s), and [Ca 2+ ] iT (0.24). ISO-induced increase in dL/dt max and [Ca 2+ ] iT also returned close to control levels. Conclusion: Chronic β 3 -ANT treatment after CHF significantly improves LV and myocyte contractile function and [Ca 2+ ] i regulation and limits the development of CHF. Thus, β 3 -AR blocker may provide a new therapeutic strategy for the treatment of CHF.

Background: Angiotensin-(1-12) [Ang-(1-12)] is a chymase-dependent source for Angiotensin II (Ang II) inotropic activity that may be impaired in a model of sustained hypertension with high cardiac Ang II content due to insertion of the human angiotensinogen (AGT) gene in the rat genome. Accordingly, we evaluated the effects of Ang-(1-12) and Ang II on myocyte contractility and [Ca 2+ ] i regulation in 9 adult male transgenic rats expressing the human AGT gene [TGR(hAGT)L1623)] and 9 SD controls. Methods: We compared LV myocyte contraction, relaxation and [Ca 2+ ] i transient ([Ca 2+ ] iT ) responses to Ang-(1-12) (4x10 -6 M) and Ang II (10 -6 M) in freshly isolated LV myocytes. Results: Myocyte contraction (dL/dt max , 109.6 vs 127.9 μm/s), relaxation (dR/dt max , 95.3 vs 107.5 μm/s) and [Ca 2+ ] iT (0.15 vs 0.24) were depressed in TGR(hAGT)L1623 rats compared to SD controls. Moreover, cell contractile and [Ca 2+ ] iT responses following exposure to Ang-(1-12) or Ang II were markedly blunted. In SD myocytes, versus baseline, Ang II or Ang-(1-12) superfusion produced significant increases in dL/dt max [Ang II: 44% vs Ang-(1-12): 34%], dR/dt max (33% vs 26%) and [Ca 2+ ] iT (31% vs 25%). Importantly, the magnitude of the responses to the two agents in TGR(hAGT)L1623 myocytes was significantly reduced. Versus the changes in SD myocytes, Ang-(1-12) caused significantly less increases in dL/dt max (22%), dR/dt max (16%) and [Ca 2+ ] iT (15%) in TGR(hAGT)L1623 myocytes . Ang II also caused similar significantly attenuated increases in dL/dt max (27%), dR/dt max (25%) and [Ca 2+ ] iT (23%). The Ang-(1-12)-induced inotropic effects were completely prevented in the presence of the inhibitory cAMP analog, Rp-cAMPS (10 –4 M, 2 hours) in both SD and TGR(hAGT)L1623 myocytes, but were further augmented only intransgenic rats after incubation of myocytes with the G i inhibitor, pertussis toxin (PTX, 2 μg/ml, 36°C, 5 hours). Conclusions: Ang-(1-12) stimulates LV myocyte contractile function and [Ca 2+ ] iT in both SD and TGR(hAGT)L1623 rats. Furthermore, we now show that the blunted inotropic responses to Ang-(1-12) and Ang II in rats expressing the human AGT gene is mediated through a cAMP-dependent mechanism that is coupled to both stimulatory G and inhibitory PTX-sensitive G proteins.

Background: We have shown that diabetes (DM)-induced cardiac dysfunction and β-adrenergic desensitization were prevented in β 3 -adrenergic receptor (AR) knockout (β 3 KO) mice. However, the molecular mechanism is unclear. We hypothesize that reversal of type 2 (T2) DM-induced alterations of cardiac SR Ca 2+ -ATPase (SERCA2a) and inducible nitric oxide synthase (iNOS) by β 3 KO may play a key role for the protective effect. Methods: LV myocyte protein levels of β 1 - and β 3 -AR, SERCA2a and iNOS, myocyte contractile and [Ca 2+ ] i transient ([Ca 2+ ] iT ) responses to isoproterenol (ISO, 10 -8 M) were compared in 4 groups (n=6) female mice: 2 Vehicle controls wild-type (CWT) and Cβ 3 KO; and 2 T2DM of T2WT and T2β 3 KO.T2 was induced by fed high-fat diet (HFD) for 14 weeks (W), but after HFD for 4 W receiving streptozotocin (STZ, 40 mg/kg/day, i.p. for 5 days). Results: Mice-treated with HFD plus STZ had high blood glucose (~350 to 380 mg/dl). Versus CWT, T2WT myocytes had significantly decreased protein levels of SERCA2a (0.18 vs 0.29), and β 1 -AR (0.35 vs 0.49), but increased iNOS (0.25 vs 0.15) and β 3 -AR (0.25 vs 0.14) with reduced cell contraction (dL/dt max ) (72.7 vs 140.9 μm/s), relaxation (dR/dt max ) (50.9 vs 117.3 μm/s) and [Ca 2+ ] iT (0.16 vs 0.22). Moreover, in T2WT myocytes, ISO caused significantly less increases in dL/dt max (37% vs 59%), dR/dt max (30% vs 53%) and [Ca 2+ ] iT (19% vs 30%). Versus CWT, T2β 3 KO did not alter basal myocyte contraction and relaxation and response to ISO stimulation, but had significantly increased protein levels of SERCA2a (0.38 vs 0.29). T2β 3 KO prevented DM-caused increase in iNOS (T2β 3 KO: 0.16 vs CWT: 0.15) and decrease in β 1 -AR (0.46 vs 0.49). Versus T2β 3 KO, Cβ 3 KO mice had similar alterations. Importantly, in contrast to T2WT, in T2β 3 KO myocytes, the increased SERCA2a and reduced iNOS correlated with normal basal cell contraction and relaxation with preserved ISO-stimulated inotropic response. ISO caused similar increases in dL/dt max (60% vs 59%), dR/dt max (51% vs 53%) and [Ca 2+ ] iT (32% vs 30%). Conclusions: β 3 KO prevents T2DM-caused downregulation of cardiac β 1 -ARs and reverses increased iNOS and decreased SERCA2a, leading to the preservation of myocyte function, [Ca 2+ ] iT , and β-adrenergic reserve. Thus, β 3 -AR may provide a new target for DCM.

Background: Cardiac aging (CA) leads to the progressive decline in cardiac function- increasing risk for heart failure (HF) and cardiac mortality. Both down-regulation of β 1 -adrenergic receptors (AR) and up-regulation of a newly-described inhibitory pathway mediated by β 3 -AR may play a crucial role in the development of CA. β-AR blockers are a mainstay in the therapy of several cardiovascular diseases, including HF. However the therapeutic effects of different β blockers in CA are unknown. We tested the hypothesis that up-regulation of cardiac β 3 -AR with enhanced negative inotropic actions, rather than the desensitization of β 1 -AR, is the critical determinant of the dysfunctional β-AR regulation that occurs in CA. Thus, β 3 -AR antagonists (β 3 ANT), but not β 1 Blockade, may reverse the progression of age-associated deficits in LV myocyte functional performance. Methods: LV myocyte basal and isoproterenol (ISO)-stimulated functional and [Ca 2+ ] i transient ([Ca 2+ ] iT ) responses were compared in 4 groups (7/group) of wild-type (WT) mice: 1) YWT , young (Y, 6 mo); 2) Aβ 3 ANT , aged (A, 26 mo) treated with β 3 ANT (L-748,337, 10 -7 M/kg/day, mini-pump) for 12 weeks (W);3) Aβ 1 Blockade , metoprolol XL (1.5 mg/kg/day, po), a selective β 1 blocker was given for 12 W and 4) AWT , aged sham control. Results: Compared with YWT, AWT myocytes had about 30% decreases in basal cell contraction (dL/dt max , 85.9 vs 122.3 μm/s) and relaxation (dR/dt max , 69.4 vs 99.2 μm/s) with reduced the peak [Ca 2+ ] iT (0.17 vs 0.21). ISO (10 -8 M)-stimulated increases in dL/dt max (33% vs 69%), dR/dt max (31% vs 62%) and [Ca 2+ ] iT (15% vs 34%) also significantly reduced. In Aβ 1 Blockade group, metoprolol therapy failed to improve basal dL/dt max (86.3 μm/s), dR/dt max (68.9 μm/s) and [Ca 2+ ] iT (0.18). ISO-stimulated increases in dL/dt max (34%), dR/dt max (35%) and [Ca 2+ ] iT (17%) remained attenuated. By contrary, Aβ 3 ANT myocytes showed normal basal cell contraction (131.0 μm/s), relaxation (101.9 μm/s) and [Ca 2+ ] iT (0.21) with preserved ISO-stimulated increases in dL/dt max (68%), dR/dt max (59%) and [Ca 2+ ] iT (32%). Conclusions: Chronic β 3 -AR antagonist, but not β 1 -AR blocker leads to the preservation of myocyte function, [Ca 2+ ] iT , and β-adrenergic responsiveness in a murine model of cardiac aging.

Background: Recent evidence links impaired nitric oxide (NO) signaling pathway in the pathogenesis of diabetes-induced cardiac dysfunction. However, the different nitric oxide synthases (NOS) isoforms involved in this pathology are controversial. Recent reports have shown that in failing myocardium, increased inducible NOS (iNOS) contributes to the attenuation of β-adrenergic receptor (AR)-mediated inotropic effect. The alteration and functional significance of cardiac iNOS in diabetic cardiomyopathy (DCM) are unclear. We assessed the hypothesis that increased cardiomyocyte iNOS expression and stimulation may inhibit myocyte contraction, relaxation, [Ca 2+ ] i transient ([Ca 2+ ] iT ), and depress its response to β-AR stimulation, thereby directly contributing to the functional impairment in DCM. Methods: Left ventricular (LV) myocyte protein levels of 3 NOS and myocyte functional responses were evaluated in 2 groups of wild-type female mice (11/group): control and DCM-induced by streptozotocin (STZ) (at 10 weeks after receiving 200 mg/kg STZ, ip). In DCM, we further assessed myocyte contractile and ([Ca 2+ ] iT ) responses to β-AR stimulation by isoproterenol (ISO, 10 -8 M) with and without pretreatment of myocytes with a selective iNOS inhibitor, 1400W (10 -5 M). Results: Compared with controls, DCM myocytes had significantly increased iNOS (1.76 vs 1.03) with decreased eNOS (0.29 vs 0.34), but relatively unchanged nNOS (0.17 vs 0.18). These changes were followed by significantly reduced basal cell contraction (dL/dt max , 73.8 vs 140.7 μm/s), relaxation (dR/dt max , 61.5 vs 121.1 μm/s) and [Ca 2+ ] iT (0.17 vs 0.20). ISO-stimulated increases in dL/dt max (32% vs 59%), dR/dt max (30% vs 55%) and [Ca 2+ ] iT (17% vs 31%) were also significantly reduced. Moreover, in DCM myocytes, pretreatment with 1400W markedly improved myocyte basal contraction (128.0 μm/s) and relaxation (111.7 μm/s). The ISO-induced increases in dL/dt max (55%), dR/dt max (52%), and [Ca 2+ ] iT (29%) were also significantly augmented. Conclusions: Our findings indicate that myocardial iNOS is activated in diabetic mice and suggest that increased iNOS expression contributes to depressed myocardial contractility, impaired [Ca 2+ ] i regulation and β-adrenergic hyporesponsiveness.

Background: Natriuretic peptides (NPs) play a crucial role in maintaining cardiovascular homeostasis. NPs stimulate the production and release of cGMP, leading to the vasodilating and natriuretic actions. In heart failure (HF), circulating and cardiac ANP, BNP, and CNP are increased and exhibit a range of actions. However, although they serve as therapeutic agents, their direct cardiac effects in HF are uncertain due to the confounding influence of NPs-produced changes in loading condition on conventional measures of LV function. We test the hypothesis that equal hypotensive 3 NPs may have different inotropic effects on LV contractility and relaxation in HF. Methods: We assessed the cardiac effects of intravenous infusion (20 min) of ANP (2 μg/kg plus 0.5 μg/kg/min), BNP (2 μg/kg plus 0.04 μg/kg/min) and CNP (2 μg/kg plus 0.4 μg/kg/min) on different days in 6 instrumented conscious dogs with pacing-induced HF by using pressure (P)-volume (V) analysis, a load-independent measure of LV contractility. Results: Versus baselines, 3 NPs produced arterial vasodilation with similar and significant decreases in LV end-systolic pressure (10 to 12 mmHg) with relatively unchanged heart rate. ANP caused significant reductions (13%) of E ES (4.2 vs 4.8 mmHg/ml) and M SW (54.6 vs 62.8 mmHg). The time constant of LV relaxation (τ, 45.1 vs 37.6 ms) was lengthened. The LV-arterial coupling, E ES / E A (arterial elastance) (0.57 vs 0.58) was unaltered. The peak mitral flow, dV/dt max was only increased by 7% (178 vs 166 ml/s). With BNP, there were no significant changes in E ES (5.1 vs 4.9 mmHg/ml) and M SW, but E ES /E A was improved 30% (0.74 vs 0.57) due to decreased E A . τ (33.4 vs 37.9 ms) was significantly shortened and dV/dt max increased 15% (189 vs 165 ml/s). In contrast, CNP produced significant increases (~30%) in E ES (6.3 vs 4.8 mmHg/ml) and M SW (80.5 vs 62.4 mmHg) with enhanced increase in E ES /E A (50%, 0.87vs 0.58), but decrease in τ (25%, 28.4 vs 38.1 ms) and significantly greater augmented dV/dt max (25%, 205 vs 164 ml/s). Similar observations of NPs were made at constant heart rate, or after autonomic blockade. Conclusion: In conscious dogs with HF, equal hypotensive ANP, BNP and CNP have negative, no effect, and positive inotropic effects on LV contractility and relaxation, respectively.