Impaired cardiac mitochondrial membrane potential and respiration in copper-deficient rats

Cardiac mitochondrial respiration, ATP synthase activity, and membrane potential and intactness were evaluated in copper-deficient rats. In the presence of NADH, both copper-deficient and copper-adequate mitochondria had very low oxygen consumption rates, indicating membrane intactness. However copper-deficient mitochondria had significantly lower oxygen consumption rates with NADH than did copper-adequate mitochondria. Copper-deficient mitochondria had significantly lower membrane potential than did copper-adequate mitochondria using fluorescent dyes. Copper-deficient mitochondria had significantly lower state 3 oxygen consumption rates and were less sensitive to inhibition by oligomycin, an ATP synthase inhibitor. Copper-deficient and copper-adequate mitochondria responded similiarly to CCCP. No difference was observed in mitochondrial ATPase activity between copper-deficient and copper-adequate rats using submitochondrial particles. We conclude that cardiac mitochondrial respiration is compromised in copper-deficient rats, and may be related to an altered ATP synthase complex and/or a decreased mitochondrial membrane potential.     

Endothelin-1 in paraventricular nucleus modulates cardiac sympathetic afferent reflex and sympathetic activity in rats

Background

Cardiac sympathetic afferent reflex (CSAR) is a positive-feedback, sympathoexcitatory reflex. Paraventricular nucleus (PVN) is an important component of the central neurocircuitry of the CSAR. The present study is designed to determine whether endothelin-1 (ET-1) in the PVN modulates the CSAR and sympathetic activity, and whether superoxide anions are involved in modulating the effects of ET-1 in the PVN in rats.

Methodology/Principal Findings

In anaesthetized Sprague–Dawley rats with cervical vagotomy and sinoaortic denervation, renal sympathetic nerve activity (RSNA) and mean arterial pressure (MAP) were recorded. The CSAR was evaluated by the responses of the RSNA and MAP to epicardial application of capsaicin. Microinjection of ET-1 into the bilateral PVN dose-dependently enhanced the CSAR, increased the baseline RSNA and MAP. The effects of ET-1 were blocked by PVN pretreatment with the ET_A receptor antagonist BQ-123. However, BQ-123 alone had no significant effects on the CSAR, the baseline RSNA and MAP. Bilateral PVN pretreatment with either superoxide anion scavenger tempol or polyethylene glycol-superoxide dismutase (PEG-SOD) inhibited the effects of ET-1 on the CSAR, RSNA and MAP. Microinjection of ET-1 into the PVN increased the superoxide anion level in the PVN, which was abolished by PVN pretreatment with BQ-123. Epicardial application of capsaicin increased superoxide anion level in PVN which was further enhanced by PVN pretreatment with ET-1.

Conclusions

Exogenous activation of ET_A receptors with ET-1 in the PVN enhances the CSAR, increases RSNA and MAP. Superoxide anions in PVN are involved in the effects of ET-1 in the PVN.     

Presynaptic receptor systems on cardiac sympathetic nerves.

The presence of a number of presynaptic receptor mechanisms on postganglionic sympathetic nerve terminals has been described by various investigators. In the present review evidence is presented which supports the concept that activation of these presynaptic receptors results in either the inhibition or facilitation of transmitter release from sympathetic nerve endings. The role of these mechanisms in controlling sympathetic function to the myocardium in physiological as well as pathophysiological condition is discussed. The possibility that pharmacological actions of several agents may have a presynaptic component is also discussed.     

Cardiac sympathetic overactivity and decreased baroreflex sensitivity in L-NAME hypertensive rats

The present study evaluated the possible changes in the autonomic control of heart rate in the hypertensive model induced by the inhibition of nitric oxide synthase. Rats were treated with N(G)-nitro-L-arginine methyl ester (L-NAME group) in the drinking water during 7 days, whereas control groups were treated with tap water (control group) or with the N(G)-nitro-D-arginine methyl ester (D-NAME group), an inactive isomer of the L-NAME molecule. The L-NAME group developed hypertension and tachycardia. The sequential blockade of the autonomic influences with propranolol and methylatropine indicated that the intrinsic heart rate did not differ among groups and revealed a sympathetic overactivity in the control of heart rate in the L-NAME group. The spectral density power of heart rate, calculated using fast-Fourier transformation, indicated a reduced variability in the low-frequency band (0.20-0.60 Hz) for the L-NAME group. The baroreflex sensitivity was also attenuated in these animals when compared with the normotensive control or D-NAME group. Overall, these data indicate cardiac sympathetic overactivity associated with a decreased baroreflex sensitivity in L-NAME hypertensive rats.     

Selective breeding of two lines of guinea pigs differing in bronchial sensitivity to acetylcholine and histamine exposure

We developed two lines of guinea pigs, one as model animals for bronchial asthma with bronchial hypersensitivity and the other with hyposensitivity as a control. In the last four years, the bronchial hypersensitive line (BHS) and hyposensitive line (BHR), both derived from Hartley strain guinea pigs, have been selected by using bronchial reactivity to acetylcholine and to histamine as parameters. Both lines have reached the F6 generation. The following results were obtained with the two lines: 1) Sib and cous in matings, and mating of selected consanguineous individuals were adopted in breeding BHS and BHR. The breeding started with six families, each, but in the F6 generation the number of families decreased to two in each line. 2) Appearance rates of hyper- or hyposensitivity to acetylcholine and histamine increased with successive generations in both lines, which had been completely separated by the F6 generation. 3) Coefficients of inbreeding in BHS and BHR in the F6 generation ranged from 42% to 45% in the former and 42% in the latter. 4) Heritabilities (h2) of BHS and BHR for the appearance rates of sensitivity to acetylcholine were presumed to be 0.54 in the former and 0.69 in the latter. 5) No difference in the body weight of 0, 20, and 40 day-old BHS was observed in any generation. On the other hand, the body weight of 20 and 40 day-old BHR tended to decrease with successive generations. 6) Mean litter sizes of BHS and BHR in each of the generations ranged from 2.24 to 3.47 animals in the former and from 2.63 to 3.38 animals in the latter.(ABSTRACT TRUNCATED AT 250 WORDS)     

Baroreflex sensitivity in obesity: relationship with cardiac autonomic nervous system activity

Objective: The aim of this study was to test the hypothesis that baroreflex sensitivity (BRS), assessed by indirect measurement of aortic pressure, is blunted in obesity. Additionally, the potential effect of cardiac autonomic nervous system (ANS) activity, aortic compliance, and metabolic parameters on BRS of obese subjects was investigated. Research Methods and Procedures: A group of 30 women with BMI >30 kg/m² and a group of 30 controls with BMI <25 kg/m² were examined. BRS was estimated by the sequence technique, cardiac ANS activity by short‐term spectral analysis of heart rate variability (HRV), and aortic compliance by the method of applanation tonometry. Results: BRS was lower in obese women (9.18 ± 3.77 vs. 19.63 ± 9.16 ms/mm Hg, p < 0.001). The median values (interquartile range) of the power of both the high‐frequency and low‐frequency components of the HRV were higher in the lean than in the obese participants [1079.2 (202.7 to 1716.9) vs. 224.1 (72.7 to 539.6) msec², p = 0.001 and 411.8 (199.3 to 798.0) vs. 235.8 (99.4 to 424.5) msec², p = 0.01 respectively]. Low‐to‐high‐frequency ratio values were higher in the obese subjects [0.82 (0.47 to 2.1) vs. 0.57 (0.28 to 0.89), p = 0.02]. Aortic augmentation values were not significantly different between lean and obese subjects. Multivariate analysis demonstrated a significant and independent association between BRS and age (p = 0.003), BMI (p < 0.001), and high‐frequency power of HRV (p < 0.001). These variables explained 72% of the variation of BRS values. Discussion: BRS is severely reduced in obese subjects. BMI, age, and the parasympathetic nervous system activity are the main determinants of BRS. Baroreflex behavior is of clinical relevance because an attenuated BRS represents a negative prognostic factor in cardiovascular diseases, which are common in obesity.     

Macrophage populations and cardiac sympathetic denervation during L-NAME-induced hypertension in rats

The rat model of hypertension induced by prolonged treatment with Nomega-nitro-L-arginine methyl ester (L-NAME) has been extensively used. However, the effects on cardiac autonomic innervation are unknown. Here, the cardiac sympathetic innervation is analyzed in parallel with myocardial lesions and leukocyte infiltration during L-NAME (40 mg/Kg body weight/day, orally) treatment. The occurrence of cardiomyocyte hypertrophy, a controversial matter, is also addressed. Degenerating cardiomyocytes and focal inflammation occurred one day after treatment. Inflammatory lesions became gradually more frequent until day 7. At day 14 fibroblast-like cells were outstanding. Interstitial and perivascular connective tissue increased from day 28 on. In the left ventricle, cardiomyocyte hypertrophy occurred only around the damaged area during the first 14 days. After 28 days, it became more widespread. In the right ventricle, the hypertrophic cardiomyocytes were restricted to damaged areas. Significant reduction of the noradrenergic nerve terminals occurred from day 3 to 28. The area occupied by ED1+ (hematogenous) macrophages increased until day 7, and dropped to control levels by day 10. ED2+ (resident) macrophages increased from day 3 to 7 and remained higher than control values up to day 77. Animals receiving both L- NAME and aminoguanidine (AG), an inducible nitric oxide synthase (iNOS) inhibitor (65 mg/Kg body weight/day, orally), showed significant decrease in the nitrite serum levels, sympathetic denervation and macrophage infiltration at day 7. No denervation was detectable at day 14 of double treatment, using subcutaneous AG. Our findings favor a role for ED1+ macrophages and iNOS in the hypertension-induced denervation process.     

AT1 receptor mRNA antisense normalizes enhanced cardiac sympathetic afferent reflex in rats with chronic heart failure

Previous studies showed that the cardiac sympathetic afferent reflex (CSAR) is enhanced in dogs and rats with chronic heart failure (CHF) and that central ANG II type 1 receptors (AT(1)R) are involved in this augmented reflex. The aim of this study was to determine whether intracerebroventricular administration and microinjection of antisense oligodeoxynucleotides targeted to AT(1)R mRNA would attenuate the enhanced CSAR and decrease resting renal sympathetic nerve activity (RSNA) in rats with coronary ligation-induced CHF. The CSAR was elicited by application of bradykinin to the epicardial surface of the left ventricle. Reflex responses to epicardial administration of bradykinin were enhanced in rats with CHF. The response to bradykinin was determined every 50 min after intracerebroventricular administration (lateral ventricle) or microinjection (into paraventricular nucleus) of antisense or scrambled oligonucleotides to AT(1)R mRNA. AT(1)R mRNA and protein levels in the paraventricular nucleus were significantly reduced 5 h after administration of antisense. Antisense significantly decreased resting RSNA and normalized the enhanced CSAR responses to bradykinin in rats with CHF. Scrambled oligonucleotides did not alter resting RSNA or the enhanced responses to bradykinin in rats with CHF. No significant effects were found in sham-operated rats after administration of either antisense or scrambled oligonucleotides. These results strongly suggest that central AT(1)R mRNA antisense reduces expression of AT(1)R protein and normalizes the augmentation of this excitatory sympathetic reflex and that genetic manipulation of protein expression can be used to normalize the sympathetic enhancement in CHF.     

Physical properties of the purified cardiac muscarinic acetylcholine receptor

The physical properties of the cardiac muscarinic acetylcholine receptor (mAcChR) purified from porcine atria as recently described [Peterson, G.L., Herron, G.S., Yamaki, M., Fullerton, D.S., & Schimerlik, M.I. (1984) Proc. Natl. Acad. Sci. U.S.A. 81, 4993-4997] have been examined by D2O/H2O sucrose gradient sedimentation and Sephacryl S-300 gel filtration in Triton X-405 and by sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE). From the sedimentation experiments the partial specific volume and sedimentation constant for the mAcChR-Triton X-405 complex were determined to be 0.813 cm3/g and 5.30 S, respectively, which lead to an estimate of the molecular weight of the complex of 143 000. Gel filtration in Triton X-405 gave an estimate of the Stokes radius (4.29 nm) and an apparent molecular weight of 116 000. Combination of sedimentation and gel filtration gave an apparent molecular weight of 137 000 and a frictional ratio (f/f0) of 1.21 for the complex. The partial specific volume of the receptor calculated from composition was 0.717 cm3/g assuming 26.5% by weight carbohydrate. The amount of bound Triton X-405 was estimated at 1.011 g/g of mAcChR, which gave an apparent molecular weight of 70 900 (sedimentation) or 68 200 (sedimentation plus gel filtration) for the uncomplexed receptor. SDS-PAGE experiments at acrylamide concentrations ranging from 6% T [monomer plus bis(acrylamide)] to 17% T gave a linear range of apparent molecular weight from 67 600 (6% T) to 98 600 (17% T), and calibration against the retardation coefficient, Kr, determined from Ferguson plots gave an apparent molecular weight of 89 100 +/- 6700. From a newly developed, novel evaluation scheme the anomalous migration of the mAcChR in SDS-PAGE was found to be due to both an excess charge density and an abnormally large shape parameter (Kr), and the true molecular weight of the protein portion of the mAcChR ligand binding polypeptide was estimated to be between 50 000 and 60 000.     

Mutations affecting agonist sensitivity of the nicotinic acetylcholine receptor.

The nicotinic acetylcholine receptor (AChR) is a pentameric transmembrane protein (alpha 2 beta gamma delta) that binds the neurotransmitter acetylcholine (ACh) and transduces this binding into the opening of a cation selective channel. The agonist, competitive antagonist, and snake toxin binding functions of the AChR are associated with the alpha subunit (Kao et al., 1984; Tzartos and Changeux, 1984; Wilson et al., 1985; Kao and Karlin, 1986; Pederson et al., 1986). We used site-directed mutagenesis and expression of AChR in Xenopus oocytes to identify amino acid residues critical for ligand binding and channel activation. Several mutations in the alpha subunit sequence were constructed based on information from sequence homology and from previous biochemical (Barkas et al., 1987; Dennis et al., 1988; Middleton and Cohen, 1990) and spectroscopic (Pearce and Hawrot, 1990; Pearce et al., 1990) studies. We have identified one mutation, Tyr190 to Phe (Y190F), that had a dramatic effect on ligand binding and channel activation. These mutant channels required more than 50-fold higher concentrations of ACh for channel activation than did wild type channels. This functional change is largely accounted for by a comparable shift in the agonist binding affinity, as assessed by the ability of ACh to compete with alpha-bungarotoxin binding. Other mutations at nearby conserved positions of the alpha subunit (H186F, P194S, Y198F) produce less dramatic changes in channel properties. Our results demonstrate that ligand binding and channel gating are separable properties of the receptor protein, and that Tyr190 appears to play a specific role in the receptor site for acetylcholine.     

Neural control of embryonic acetylcholine receptor and skeletal muscle

The manner by which motor neurons exert control over the distribution and number of acetylcholine receptors, and muscle development was investigated in the superior oblique muscle of white Peking duck embryos. Clusters of receptors in the normally developing muscle first appeared on day 10 of incubation as determined with I125 alpha-bungarotoxin autoradiography. The initial appearance of receptor clusters coincided with the arrival of motor nerve fibers in the muscle. Clusters of receptors also appeared in normal fashion in muscles made aneural by destruction of motor neurons on day 7. But after day 14 these clusters had disappeared and no new clusters were seen thereafter in the aneural muscle. Receptor clusters persisted throughout development in muscle in which neuromuscular transmission was blocked with either curare or botulinum toxin and in muscles denervated on day 10.5, i.e., shortly after the initial nerve-muscle contact but prior to the onset of muscle activity. A progressive increase in the total number of receptors and in the total amount of protein occurred during the course of normal development. However, the specific activity of the receptor protein declined sharply following innervation on day 10. The total number of receptors and the specific activity of the receptor was affected depending on whether the motor neurons were destroyed before or after innervation and following chronic blockade of neuromuscular transmission. The half-life of the receptor protein was similar in normal, aneural, and paralyzed muscles (26, 25, 26 h, respectively). Measurements of total protein indicated that essentially no muscle growth occurred in the complete absence of innervation. Paralyzed muscles continued to develop but at a slower pace.     

Specific baroreceptor control of vertebral and cardiac sympathetic activity.

The effect of bilateral carotid occlusion (BCO) on the activity of the vertebral and cardiac sympathetic efferent nerves was studied in gallamine-immobilized and artificially ventilated cats under chloralose-urethane anaesthesia. Electrical activity of the vertebral and cardiac nerves (VNA and CNA), their integram, arterial blood pressure and respiration were recorded. BCO led to an increase in VNA persisting throughout the occlusion period, while merely a transient increase took place in CNA. When blood pressure was kept at a constant level or the depressor nerves was transected, CNA responded to BCO with a lasting increase. Electrical stimulation of the central stump of the left depressor nerve inhibited CNA much more than VNA. It is assumed that the selective inhibition of CNA, after a transient increase, arises as a consequence of a rise in blood pressure, i.e. of consecutive aortic baroreceptor excitation.     

Warm-up phenomenon and cardiac autonomic control in patients with coronary artery disease

Decreased heart rate variability (HRV) and heart rate turbulence (HRT) are independent predictors of mortality after acute myocardial infarction (AMI). There are no previous studies on the relationship between warm-up phenomenon and cardiac autonomic control in stable coronary artery disease (CAD). We investigated the responses in HRV to repeated exercise induced ischemia and differences in global HRV and HRT in patients with and without adaptation to ischemia (warm-up phenomenon). Fifty male patients with CAD underwent two successive exercise tests with ambulatory electrocardiogram (ECG) recordings. HRV was evaluated using time and frequency domain measures and HRT was determined among patients with ventricular premature complexes (VPCs). The patients were divided in two groups on the basis of either positive (warm-up+) or negative (warm-up-) ischemia adaptation. Total power, ULF and VLF power and pNN50 calculated from the entire ECG recording were higher in the group demonstrating warm-up phenomenon (P<0.05 for all). In the assessment of the four short-term stationary phases (pre-and post-test 1 and 2) total power, VLF power and pNN50 were significantly higher in the warm-up positive group already at the baseline (P<0.05 for all). Furthermore, in the entire recordings total power, ULF and VLF power and SDNN correlated positively with the decrease in ischemic burden in the recovery phase (P相似文献   

Altered autonomic control in conscious transgenic rabbits with overexpressed cardiac Gsalpha

Both enhanced sympathetic drive and altered autonomic control are involved in the pathogenesis of heart failure. The goal of the present study was to determine the extent to which chronically enhanced sympathetic drive, in the absence of heart failure, alters reflex autonomic control in conscious, transgenic (TG) rabbits with overexpressed cardiac Gsalpha. Nine TG rabbits and seven wild-type (WT) littermates were instrumented with a left ventricular (LV) pressure micromanometer and arterial catheters and studied in the conscious state. Compared with WT rabbits, LV function was enhanced in TG rabbits, as reflected by increased levels of LV dP/dt (5,600 +/- 413 vs. 3,933 +/- 161 mmHg/s). Baseline heart rate was also higher (P < 0.05) in conscious TG (247 +/- 10 beats/min) than in WT (207 +/- 10 beats/min) rabbits and was higher in TG after muscarinic blockade (281 +/- 9 vs. 259 +/- 8 beats/min) or combined beta-adrenergic receptor and muscarinic blockade (251 +/- 6 vs. 225 +/- 9 beats/min). Bradycardia was blunted (P < 0.05), whether induced by intravenous phenylephrine (arterial baroreflex), by cigarette smoke inhalation (nasopharyngeal reflex), or by veratrine administration (Bezold-Jarisch reflex). With veratrine administration, the bradycardia was enhanced in TG for any given decrease in arterial pressure. Thus the chronically enhanced sympathetic drive in TG rabbits with overexpressed cardiac Gsalpha resulted in enhanced LV function and heart rate and impaired reflex autonomic control. The impaired reflex control was generalized, not only affecting the high-pressure arterial baroreflex but also the low-pressure Bezold-Jarisch reflex and the nasopharyngeal reflex.     

Impaired estrogen sensitivity in bone by inhibiting both estrogen receptor alpha and beta pathways

Although it is well established that estrogen deficiency causes osteoporosis among the postmenopausal women, the involvement of estrogen receptor (ER) in its pathogenesis still remains uncertain. In the present study, we have generated rats harboring a dominant negative ERalpha, which inhibits the actions of not only ERalpha but also recently identified ERbeta. Contrary to our expectation, the bone mineral density (BMD) of the resulting transgenic female rats was maintained at the same level with that of the wild-type littermates when sham-operated. In addition, ovariectomy-induced bone loss was observed almost equally in both groups. Strikingly, however, the BMD of the transgenic female rats, after ovariectomized, remained decreased even if 17beta-estradiol (E(2)) was administrated, whereas, in contrast, the decrease of littermate BMD was completely prevented by E(2). Moreover, bone histomorphometrical analysis of ovariectomized transgenic rats revealed that the higher rates of bone turnover still remained after treatment with E(2). These results demonstrate that the prevention from the ovariectomy-induced bone loss by estrogen is mediated by ER pathways and that the maintenance of BMD before ovariectomy might be compensated by other mechanisms distinct from ERalpha and ERbeta pathways.