Cardiac ischemia and reperfusion in spontaneously diabetic rats with and without application of EGb 761: I. cardiomyocytes

Diabetic cardiomyopathy is known to result in increased mortality after ischemic events. Permanently increased oxidative stress with formation of oxygen-free radicals plays a key role in the development of specific heart muscle disease. Associated lesions include structural alterations to cardiomyocytes. Antioxidative treatment in addition to the usual insulin substitution would seem sensible in preventing or delaying long-term diabetic complications and protecting the myocardium against acute ischemic events. We investigated the effects of radical scavenger Ginkgo biloba extract EGb 761 against diabetes-induced damage to cardiomyocytes and additional ischemia/reperfusion injury in spontaneously diabetic BioBreeding/Ottawa Karlsburg (BB/OK) rats, as a model of diabetic myocardium infarction. Morphological and morphometric parameters of heart muscles were analyzed by light and electron-microscopic techniques. We used immunohistochemistry to evaluate parameters of oxidative stress (superoxide dismutase [SOD]) and inducible nitric oxide synthase (iNOS) protein expression. Our results indicated that A) Diabetic myocardium appears more vulnerable to ischemia/reperfusion damage concerning ultrastructure of cardiomyocytes (sarcomeres, vacuoles, mitochondria), expression of antioxidative enzymes (CuZnSOD, MnSOD), and iNOS than normal myocardium; B) Pre-treatment of diabetic myocardium with EGb and additional ischemia/reperfusion leads to a relative improvement in myocardial ultrastructure compared to unprotected myocardium. In summary, EGb appears to be promising as an adjuvant therapeutic drug in diabetics with respect to ischemic myocardium injury. It may contribute to the prevention of late diabetic complications in diabetic cardiomyopathy.     

Pupillary signs in diabetic autonomic neuropathy.

Pupillary function was investigated in 36 insulin-dependent diabetics and 36 controls matched for age and sex. About half of the diabetics had evidence of peripheral somatic or autonomic neuropathy, or both. The diabetic patients had abnormally small pupil diameters in the dark and less fluctuation in pupil size (hippus) during continuous illumination than the controls. They also had reduced reflex responses to light flashes of an intensity adjusted for individual retinal sensitivities. The pupillary findings were compared with results of five tests of cardiovascular function and five tests of peripheral sensory and motor nerve function. Almost all the patients with autonomic neuropathy had pupillary signs, which we therefore conclude are a common manifestation of diabetic autonomic neuropathy.     

EGb761对血管性痴呆大鼠海马突触可塑性的影响

目的：探讨银杏叶提取物（EGb761）对血管性痴呆（VD）模型大鼠海马突触可塑性的影响。方法：Morris水迷宫检测大鼠空间学习记忆能力;电生理学方法在体记录大鼠海马长时程增强。结果：各时间点模型组大鼠的逃逸潜伏期（EL）均较假手术组明显延长（P〈0.01）,药物组各亚组大鼠的EL均显著短于模型组（P〈0.01）,但仍长于假手术组（P〈0.01,P〈0.05）。模型组各亚组大鼠长时程增强（LTP）诱导率显著低于假手术组和药物组（P〈0.01）。模型组大鼠各时间点群发峰电位（PS）的相对幅值明显低于假手术组和药物组（P〈0.01,P〈0.05）。假手术组、模型组和药物组各时间点的PS潜伏期无显著差别。结论：VD模型大鼠长时间存在空间学习记忆障碍,EGb761能促进VD模型大鼠海马病理性突触可塑性的恢复,这可能是其促智作用的重要机制。     

Ventilatory response to exercise in diabetic subjects with autonomic neuropathy

Tantucci, C., P. Bottini, M. L. Dottorini, E. Puxeddu, G. Casucci, L. Scionti, and C. A. Sorbini. Ventilatory response toexercise in diabetic subjects with autonomic neuropathy.J. Appl. Physiol. 81(5):1978-1986, 1996.We have used diabetic autonomic neuropathy as amodel of chronic pulmonary denervation to study the ventilatoryresponse to incremental exercise in 20 diabetic subjects, 10 with(Dan+) and 10 without (Dan) autonomic dysfunction, and in 10 normal control subjects. Although both Dan+ and Dan subjectsachieved lower O₂ consumption andCO₂ production(CO₂) thancontrol subjects at peak of exercise, they attained similar values ofeither minute ventilation(E) oradjusted ventilation (E/maximalvoluntary ventilation). The increment of respiratory rate withincreasing adjusted ventilation was much higher in Dan+ than inDan and control subjects (P < 0.05). The slope of the linearE/CO₂relationship was 0.032 ± 0.002, 0.027 ± 0.001 (P < 0.05), and 0.025 ± 0.001 (P < 0.001) ml/min inDan+, Dan, and control subjects, respectively. Bothneuromuscular and ventilatory outputs in relation to increasingCO₂ were progressivelyhigher in Dan+ than in Dan and control subjects. At peak ofexercise, end-tidal PCO₂ was muchlower in Dan+ (35.9 ± 1.6 Torr) than in Dan (42.1 ± 1.7 Torr; P < 0.02) and control (42.1 ± 0.9 Torr; P < 0.005) subjects.We conclude that pulmonary autonomic denervation affects ventilatoryresponse to stressful exercise by excessively increasing respiratoryrate and alveolar ventilation. Reduced neural inhibitory modulationfrom sympathetic pulmonary afferents and/or increasedchemosensitivity may be responsible for the higher inspiratoryoutput.

     

Chronic administration of EGb 761 modulates synaptic and mitochondrial plasticity in adult vitamin E-deficient rats.

A computer-assisted morphometric study has been carried out on synaptic junctions and synaptic mitochondria in the dentate gyrus supragranular layer of vitamin E-deficient rats undergone chronic administration of the extract EGb 761 from Ginkgo biloba leaves (100 mg/kg body weight, daily, from 4 to 7 months of age). Control animals were fed with the vitamin E-deficient diet from 1 to 7 months of age. Numeric density (Nv), surface density (Sv) and average size of the synaptic junctions (S), mitochondrial numeric density (Nvm), volume density (Vv) and average volume (V) were the measured parameters. In EGb 761-treated animals, Nv was significantly increased and S significantly decreased, while Sv was unchanged. EGb 761 administration resulted in an increased percentage of synapses of smaller size. In EGb 761-treated rats, Nvm significantly increased and V significantly decreased, while no significant difference of Vv was found. The population of synaptic mitochondria in EGb 761 -treated animals was composed of a higher number of smaller organelles. The measured parameters report on the structural dynamics of synapses and mitochondria, thus our findings support that EGb 761 administration is able to improve the physiological adaptive capacities of the investigated structures by a positive modulation of their morphofunctional features.     

Vagal impairment of gastric secretion in diabetic autonomic neuropathy.

Gastric acid output in response to insulin-induced hypoglycaemia and pentagastrin was measured in 18 diabetic patients with symptoms of autonomic neuropathy. Two patients had achlorhydria but the rest responded normally to pentagastrin. The acid output evoked by insulin-induced hypoglycaemia was low in 10 of the 16 patients who secreted acid in response to pentagastrin. These changes suggest that vagal impairment is common in diabetics with autonomic symptoms, which might explain the infrequency of duodenal ulcer in diabetics.     

Blood pressure regulation in diabetic patients with and without peripheral neuropathy

Cardiac and vascular dysfunctions resulting from autonomic neuropathy (AN) are complications of diabetes, often undiagnosed. Our objectives were to: 1) determine sympathetic and parasympathetic components of compromised blood pressure (BP) regulation in patients with peripheral neuropathy and 2) rank noninvasive indexes for their sensitivity in diagnosing AN. We continuously measured electrocardiogram, arterial BP, and respiration during supine rest and 70° head-up tilt in 12 able-bodied subjects, 7 diabetics without, 7 diabetics with possible, and 8 diabetics with definite, sensory, and/or motor neuropathy (D2). During the first 3 min of tilt, systolic BP (SBP) of D2 decreased [-10.9 ± 4.5 (SE) mmHg] but increased in able-bodied (+4.8 ± 5.4 mmHg). Compared with able-bodied, D2 had smaller low-frequency (0.04-0.15 Hz) spectral power of diastolic BP, lower baroreflex effectiveness index (BEI), and more SBP ramps. Except for low-frequency power of SBP, D2 had greater SBP and smaller RR interval harmonic and nonharmonic components at rest across the 0.003- to 0.45-Hz region. In addition, our results support previous findings of smaller HF RR interval power, smaller numbers of baroreflex sequences, and lower baroreflex sensitivity in D2. We conclude that diabetic peripheral neuropathy is accompanied by diminished parasympathetic and sympathetic control of heart rate and peripheral vasomotion and diminished baroreflex regulation. A novel finding of this study lies in the sensitivity of BEI to detect AN, presumably because of its combination of parameters that measure reductions in both sympathetic control of vasomotion and parasympathetic control of heart rate.     

Abnormalities in plasma catecholamine response and tissue catecholamine accumulation in streptozotocin diabetic rats: a possible role for diabetic autonomic neuropathy

Plasma catecholamine levels, determined by high performance liquid chromatography, were elevated in response to blood withdrawal in normal rats. Such a response was also observed in streptozotocin diabetic rats 2 and 6 weeks after disease onset, but was no longer seen at 13 weeks. Tissue (adrenal, heart, skin, kidney) catecholamine levels in diabetic rats were increased at 6 weeks as well as at 13 weeks. These abnormalities were corrected by insulin treatment in at least a part of diabetic rats. The present data suggest that there might be a catecholamine accumulation, which is later accompanied with an impairment of catecholamine secretion, in diabetic rats, and they gave a basis for an inference that similar changes might play some role in the pathogenesis of diabetic autonomic neuropathy in man.     

Cerebrovascular reactivity and hypercapnic respiratory drive in diabetic autonomic neuropathy.

Because abnormalities in cerebrovascular reactivity (CVR) in subjects with long-term diabetes could partly be ascribed to autonomic neuropathy and related to central chemosensitivity, CVR and the respiratory drive output during progressive hypercapnia were studied in 15 diabetic patients without (DAN-) and 30 with autonomic neuropathy (DAN+), of whom 15 had postural hypotension (PH) (DAN+PH+) and 15 did not (DAN+PH-), and in 15 control (C) subjects. During CO(2) rebreathing, changes in occlusion pressure and minute ventilation were assessed, and seven subjects in each group had simultaneous measurements of the middle cerebral artery mean blood velocity (MCAV) by transcranial Doppler. The respiratory output to CO(2) was greater in DAN+PH+ than in DAN+PH- and DAN- (P < 0.01), whereas a reduced chemosensitivity was found in DAN+PH- (P < 0.05 vs. C). MCAV increased linearly with the end-tidal PCO(2) (PET(CO(2))) in DAN+PH- but less than in C and DAN- (P < 0.01). In contrast, DAN+PH+ showed an exponential increment in MCAV with PET(CO(2)) mainly >55 Torr. Thus CVR was lower in DAN+ than in C at PET(CO(2)) <55 Torr (P < 0.01), whereas it was greater in DAN+PH+ than in DAN+PH- (P < 0.01) and DAN- (P < 0.05) at PET(CO(2)) >55 Torr. CVR and occlusion pressure during hypercapnia were correlated only in DAN+ (r = 0.91, P < 0.001). We conclude that, in diabetic patients with autonomic neuropathy, CVR to CO(2) is reduced or increased according to the severity of dysautonomy and intensity of stimulus and appears to modulate the hypercapnic respiratory drive.     

Improvement of cardiovascular autonomic reflexes after amelioration of metabolic control in insulin-dependent diabetic subjects with severe autonomic neuropathy

To evaluate the influence of good metabolic equilibrium on Diabetic Autonomic Neuropathy (DAN), cardiovascular autonomic reflexes were monitored in 9 male insulin-dependent diabetic patients with DAN, treated with Continuous Subcutaneous Insulin Infusion (CSII) by pump: 9 for 10 days, 4 for 1 year and 2 for 20 months. Autonomic neuropathy was assessed evaluating 5 cardiovascular autonomic tests: Valsalva Manoeuvre (VR), Deep Breathing (DB), Lying-to-Standing (L-S), Sustained Handgrip (SHG), and Postural Hypotension (PH). Metabolic control was assessed evaluating the mean daily plasma glucose, glucosuria and glycosylated hemoglobin. Ten days of CSII treatment induced a normalization of glucose balance and a slight but significant improvement in some parasympathetic cardiovascular tests (VR: from 1.09 +/- 0.01 to 1.13 +/- 0.02; P less than 0.05). After 4-8 months of CSII treatment a significant improvement in VR (P less than 0.05); DB (P less than 0.01) and L-S (P less than 0.05) was recorded. The long-term treatment with CSII did not seem to induce a further amelioration in cardiovascular autonomic reflexes. These results show that the slight improvement induced by good metabolic balance in the cardiovascular autonomic response could be related to functional-metabolic rather than structural changes in the nerves.     

Diminished bronchial reactivity to cold air in diabetic patients with autonomic neuropathy.

To investigate the role of neural pathways in the nonasthmatic response to eucapnic hyperventilation with below freezing air five diabetic patients with severe symptomatic autonomic neuropathy were studied. Their responses were compared with those shown by five diabetic patients without autonomic neuropathy and five non-diabetic controls. After bronchial provocation testing with cold air the diabetic patients with autonomic neuropathy did not show a significant fall in specific airways conductance (mean (SE) maximum percentage fall 2.0 (3)%), whereas conductance fell in the diabetic patients without neuropathy by 30.8 (2.0)% (p less than 0.001) and in the non-diabetic controls by 22.7 (4.6)% (p less than 0.02). In subjects who do not have asthma the bronchial response to cold air is mediated largely via neural mechanisms.     

Glomerular function in spontaneously diabetic rats.

This study was undertaken to determine whether hyperfiltration exists at the single nephron level and whether albumin excretion is increased early in the course of diabetes in Biobreeding rats. Diabetic rats were studied at 8-12 weeks after the onset of diabetes. Control animals were age-matched, diabetes-resistant rats. Urinary and tubular fluid albumin concentrations were measured by polyacrylamide gel electrophoresis. Clearance and micropuncture techniques were used to determine whole kidney and single nephron glomerular filtration rate, renal blood flow, and glomerular capillary pressure. The urinary albumin excretion rate (1.3 +/- 0.1 mg/24 hr) and the tubular fluid albumin concentration (4.7 +/- 0.7 mg/dl) in the diabetic group were significantly elevated when compared with urinary albumin excretion (0.9 +/- 0.1 mg/24 hr) and tubular fluid albumin concentration (2.5 +/- 0.5 mg/dl) in the control group. There were no significant differences in glomerular hemodynamics (whole kidney or single nephron glomerular filtration rate or glomerular capillary pressure) between diabetic and control rats. The kidney weight and kidney weight to body weight ratio were significantly higher in diabetic rats when compared with control rats. Early diabetes in Biobreeding rats is characterized by mild albuminuria and increased kidney size, but not glomerular hyperfiltration.     

糖尿病植物神经病变病人心率变异的非线性定量分析

目的：研究糖尿病人植物神经病变与心率变异的关系。对象：正常对照组和根据临床有无糖尿病神经病变（ＤＡＮ）分组的糖尿病病人,方法：应用２４小时动态心电图对正常和糖尿病人进行心率变异的线性,非线性散点图和非线性定量参数分析,结果：单纯糖尿病组ＳＤＮＮ,ＳＤＡＮＮ和ＰＮＮ５０低于正常组（Ｐ〈０．０５）;糖尿病＋ＤＡＮ组各项线性时域分析指标均低于正常和单纯糖尿病组（Ｐ〈０．０１－０．００１）,散点图分析结果     

Chronotropic function in spontaneously diabetic BB rats

In the present study, isolated atrial function in spontaneously diabetic BB Wistar rats maintained for 12 weeks on a low (BB-LI) and a high (BB-HI) dosage of insulin was examined. Basal atrial rates were unchanged in the diabetic animals, relative to nondiabetic littermates (ND-BB) or Wistar controls. The BB-HI animals were euglycemic and responded to isoproterenol in a similar manner to the ND-BB and Wistar control animals. In contrast, BB-LI animals remained hyperglycemic and exhibited lower responses to the maximum chronotropic effects of isoproterenol. Plasma thyroid hormone levels were unaltered in the BB-diabetic animals. These results therefore reveal an absence of bradycardia and hypothyroidism in spontaneously diabetic BB rats, in contrast to previous observations in streptozotocin diabetic rats. However, a decrease in chronotropic response to isoproterenol was still noted in the BB-LI animals. These findings suggest that decreased positive chronotropic effect of isoproterenol in diabetes may not be a direct consequence of altered thyroid status.     

Identifying diabetic patients with cardiac autonomic neuropathy by heart rate complexity analysis

Background  

Cardiac autonomic neuropathy (CAN) in diabetes has been called a "silent killer", because so few patients realize that they suffer from it, and yet its effect can be lethal. Early sub clinical detection of CAN and intervention are of prime importance for risk stratification in preventing sudden death due to silent myocardial infarction. This study presents the usefulness of heart rate variability (HRV) and complexity analyses from short term ECG recordings as a screening tool for CAN.