异丙酚预先给药对抑郁大鼠电休克后海马Glu和GABA水平的影响

目的 探讨异丙酚预先给药对抑郁大鼠电休克后海马Glu和GABA水平的影响.方法 雄性SD大鼠30只,2～3月龄,体重200～250 g,随机分为5组(n=6):正常对照组(C组)、抑郁组(D组)、异丙酚组(P组)、电休克组(E组)、异丙酚+电休克组(PE组).采用孤养加慢性不可预见性应激法建立抑郁模型,建模后C组和D组腹腔注射生理盐水3 ml,P组腹腔注射异丙酚100 mg/kg,3组均不行电刺激;E组和PE组分别腹腔注射生理盐水3 ml或异丙酚100 mg/kg后行电休克治疗(频率50 Hz、电流50 mA、持续时间20 ms).各组处理隔日1次.共6次.于建模前1天(基础状态),建模后第1天(T1)及处理后第2天(T2)采用Open-field法行行为学评分.于T1.2时采用Morris水迷宫检测学习记忆功能,测定后处死,采用高效液相色谱法分析海马谷氨酸(Glu)、γ-氨基丁酸(GABA)含量,计算Gln/GABA比值.结果 与C组比较,其余4组行为学评分降低,学习记忆功能降低,D组和P组Glu含量升高,GABA含量降低,Glu/GABA比值升高,E组Glu含量降低,GABA含量升高,Glu/GABA比值降低(P<0.05);与D组和P组比较,E组行为学评分升高,学习记忆功能降低,Glu含量降低,GABA含量升高,Glu/GABA比值降低(P<0.05),PE组行为学评分升高,Glu含量降低,GABA含量升高,Glu/GABA比值降低(P<0.05),学习记忆功能差异无统计学意义(P>0.05);与E组比较,PE组学习记忆功能增强,Glu含量升高,GABA含量降低,Glu/GABA比值升高(P<0.05).结论 异丙酚预先给药改善电休克治疗后学习记忆功能的机制可能与异丙酚调节Glu和GABA功能状态的平衡有关.     

Deficient axonal transport of substance P in streptozocin-induced diabetic rats. Effects of sorbinil and insulin

This study measured the accumulation of substance P-like immunoreactivity (SPLI) proximal and distal to 12-h constricting ligatures applied to rat sciatic nerves. There were three separate experiments, and the baseline for each consisted of control and age-matched rats with 3 wk of untreated streptozocin-induced diabetes. We compared the effects of twice-daily insulin treatment, daily sorbinil (25 mg.kg-1.day-1 p.o.), and a combination of both treatments. In untreated diabetic rats the anterograde accumulation of SPLI was reduced by 30-40%. This deficit was unaffected by sorbinil alone but was attenuated by insulin and prevented completely by insulin and sorbinil combined. There were also indications that diabetes caused reductions in retrograde accumulation of SPLI and its content in unconstricted nerve and the L4 dorsal root ganglion. The fraction of SPLI undergoing net anterograde or retrograde movement and the velocities of accumulation were unaffected by diabetes or the treatment regimens. These findings indicate a reduction in the amount of substance P moved by axonal transport in diabetic rats that is related partly to aldose reductase activity and partly to some other insulin-correctable consequence of experimental diabetes.     

γ-氨基丁酸转运体-1在大鼠神经病理性痛形成中的作用

目的 探讨大鼠腰段脊髓γ-氨基丁酸(GABA)转运体-1(GAT-1)在神经病理性痛形成中的作用.方法 SD大鼠112只,随机分为2组(n=56):假手术组(s组)和慢性坐骨神经结扎损伤组(CCI组).结扎大鼠左侧坐骨神经建立CCI模型.于CCI前、CCI后1、3、5、7、10、14 d随机取8只大鼠,测定机械刺激缩足反应阈值(MWT)和热刺激缩足反应潜伏期(TWL).于CCI前、CCI后l、3、7、14d随机取6只大鼠,取腰段脊髓,采用免疫组织化学法检测大鼠脊髓背角GAT-1蛋白的表达.结果 与CCI前比较,2组CCI后各时点MWT和TWL均降低(P＜0.05);与S组比较,CCI组CCI后各时点MWT和TWL均降低,CCI后1、3 d GAT-1表达升高(P＜0.05或0.01).结论 大鼠神经病理性痛的形成与GAT-1表达增多有关.     

Muscle fiber type-specific defects in insulin signal transduction to glucose transport in diabetic GK rats

To determine whether defects in the insulin signal transduction pathway to glucose transport occur in a muscle fiber type-specific manner, post-receptor insulin-signaling events were assessed in oxidative (soleus) and glycolytic (extensor digitorum longus [EDL]) skeletal muscle from Wistar or diabetic GK rats. In soleus muscle from GK rats, maximal insulin-stimulated (120 nmol/l) glucose transport was significantly decreased, compared with that of Wistar rats. In EDL muscle from GK rats, maximal insulin-stimulated glucose transport was normal, while the submaximal response was reduced compared with that of Wistar rats. We next treated diabetic GK rats with phlorizin for 4 weeks to determine whether restoration of glycemia would lead to improved insulin signal transduction. Phlorizin treatment of GK rats resulted in full restoration of insulin-stimulated glucose transport in soleus and EDL muscle. In soleus muscle from GK rats, submaximal and maximal insulin-stimulated insulin receptor substrate (IRS)-1 tyrosine phosphorylation and IRS-1-associated phosphatidylinositol (PI) 3-kinase activity were markedly reduced, compared with that of Wistar rats, but only submaximal insulin-stimulated PI 3-kinase was restored after phlorizin treatment. In EDL muscle, insulin-stimulated IRS-1 tyrosine phosphorylation and IRS-1-associated PI-3 kinase were not altered between GK and Wistar rats. Maximal insulin-stimulated Akt (protein kinase B) kinase activity is decreased in soleus muscle from GK rats and restored upon normalization of glycemia (Krook et al., Diabetes 46:2100-2114, 1997). Here, we show that in EDL muscle from GK rats, maximal insulin-stimulated Akt kinase activity is also impaired and restored to Wistar rat levels after phlorizin treatment. In conclusion, functional defects in IRS-1 and PI 3-kinase in skeletal muscle from diabetic GK rats are fiber-type-specific, with alterations observed in oxidative, but not glycolytic, muscle. Furthermore, regardless of muscle fiber type, downstream steps to PI 3-kinase (i.e., Akt and glucose transport) are sensitive to changes in the level of glycemia.     

Oxidative stress in arteriogenic erectile dysfunction: prophylactic role of antioxidants

PURPOSE: We searched for markers of oxidative stress in cavernous ischemia and examined the effect of long-term antioxidant intake on arteriogenic erectile dysfunction (ED) in the rabbit. MATERIALS AND METHODS: Antioxidant activity of known antioxidant beverages, such as pomegranate juice (PJ), red wine, blueberry juice, cranberry juice, orange juice and green tea, was examined spectrophotometrically. PJ demonstrated the highest free radical scavenging capacity. The effect of long-term PJ intake on intracavernous blood flow and penile erection was then examined in the rabbit model. Erectile tissues were processed to assess oxidative stress and smooth muscle relaxation, immunohistochemical staining of nitric oxide synthase (NOS) and histomorphometry. RESULTS: On spectrophotometric analysis PJ showed the highest capacity to decrease low density lipoprotein oxidation and inhibit cellular oxidative stress in macrophages. The rabbit model of arteriogenic ED demonstrated decreased intracavernous blood flow, erectile dysfunction, loss of smooth muscle relaxation, decreased endothelial NOS and neuronal NOS, increased inducible NOS expression, diffused cavernous fibrosis and increased cavernous levels of the oxidative product isoprostane 8-epi-prostaglandin F2alpha. Long-term PJ intake increased intracavernous blood flow, improved erectile response and smooth muscle relaxation in ED and control groups while having no significant effect on NOS expression. PJ intake prevented erectile tissue fibrosis in the ED group. CONCLUSIONS: Arteriogenic ED accumulates oxidative products in erectile tissue, possibly via an intrinsic mechanism. Oxidative stress may be of great importance in the pathophysiology of arteriogenic ED. Antioxidant therapy may be a useful prophylactic tool for preventing smooth muscle dysfunction and fibrosis in ED.     

Myocardial insulin signaling and glucose transport are up-regulated in Goto-Kakizaki type 2 diabetic rats after ileal transposition

Background  

Ileal transposition (IT) as one of the effective treatments for non-obese type 2 diabetes mellitus has been widely investigated. However, the mechanisms underlying profound improvements in glucose homeostasis are still uncertain. Our objective was to explore the myocardial insulin signal transduction and glucose disposal in non-obese type 2 diabetes mellitus rats after IT surgery.     

Oxidative stress in hypertension and chronic kidney disease: role of angiotensin II

Angiotensin II, via the type 1 (AT1) receptor, stimulates oxidative stress. The vasculature, interstitium, juxtaglomerular apparatus, and the distal nephron in the kidney express nicotinamide adenine dinucleotide phosphate (NADPH) oxidase that generates superoxide anion, which is an important component of angiotensin II-induced oxidative stress. The angiotensinogen gene is stimulated by NF-kappaB activation, which is sensitive to the redox ratio, providing a positive feedback loop that can upregulate angiotensin II production. Oxidative stress can accompany hypertension in many models, including the spontaneously hypertensive rat (SHR), angiotensin II-infused rats, renovascular hypertension, and the deoxycorticosterone acetate (DOCA) salt model of hypertension. AT1 receptor antagonists can abrogate the effects of angiotensin II on oxidative stress, thus providing an important mechanistic insight onto the renal protective effects of these agents in conditions associated with angiotensin II excess.     

Alterations in glomerular RNA in diabetic rats: roles of glucagon and insulin

Incorporation in vivo of labeled orotate into RNA and total nucleotides was measured in isolated glomeruli and whole renal cortex. In 2-day diabetic animals, glomerular RNA was increased, and there was greater incorporation of orotate into total nucleotides and RNA as compared with controls. Insulin reversed the exaggerated incorporation at infusion rates that corrected hyperglucagonemia without reducing plasma glucose and with only minimal changes in insulin concentrations. The addition of glucagon to insulin infusions reproduced the increased incorporation observed in untreated diabetics. Similar changes occurred in renal cortex, where differences in orotate incorporation into nucleotide precursors seemed to be the main cause for alterations in RNA labeling. Isotope incorporation in glomeruli correlated positively with plasma glucagon, but not with insulin or glucose concentrations. Although in 7-month diabetic animals orotate incorporation into RNA was less than in controls, probably as a consequence of renal disease, 24-hour insulin infusion decreased it further. Our results confirm that in the diabetic kidney, abnormal uracil nucleotide metabolism and increased cellular content of RNA are demonstrable in glomeruli as in the renal cortex. These changes appear to be related directly to hyperglucagonemia.     

大鼠骨骼肌细胞氧化应激损伤实验研究

目的构建体外压疮深部组织细胞氧化应激模型,为进一步在细胞层面探讨压疮深部组织损伤机制建立基础。方法将对数生长期大鼠骨骼肌细胞分为对照组及5个实验组,对照组行常规培养,5个实验组分别采用50、100、150、200、250μmol/L过氧化氢处理1~4h后分别检测四氮甲基唑蓝(MTT)、乳酸脱氧酶(LDH)指标并观察骨骼肌细胞Hoechst染色及形态学变化。结果对照组1~4h细胞存活100%,MTT、LDH指标及组织形态无明显变化。5个实验组2h时除50μmol/L组外,均呈现明显损伤变化;至4h,大鼠骨骼肌细胞存活率为20%~75%;显微镜下观察大鼠骨骼肌细胞随氧化应激程度加重而缩收,细胞间隙增大;Hoechst细胞荧光染色显示100μmol/L过氧化氢作用4h已有较多肌细胞趋向凋亡;细胞LDH释放率约为对照组的1.5倍。结论大鼠骨骼肌细胞随氧化应激程度加重,趋向凋亡;经100μmol/L过氧化氢作用4h的骨骼肌细胞可作为研究压疮深部组织细胞氧化应激的体外模型。     

Delayed insulin transport across endothelium in insulin-resistant JCR:LA-cp rats

Capillary endothelial cells are thought to limit the transport of insulin across the endothelium, resulting in attenuated insulin action at target sites. Whether endothelial insulin transport is altered in dysglycemic insulin-resistant states is not clear and was therefore investigated in the JCR:LA-cp corpulent male rat, which exhibits the metabolic syndrome of obesity, insulin resistance, hyperlipidemia, and hyperinsulinemia. Lean littermates that did not develop these alterations served as controls. Animals of both groups were normotensive (mean arterial pressure 136+/-2 mmHg). Hearts from obese and lean rats aged 7 (n = 6) or 18 (n = 8) weeks were perfused in vitro at 10 ml/min per gram wet wt over 51 min with Krebs-Henseleit buffer containing 0.1 or 0.5 U human insulin/l (equivalent to 0.6 and 3 nmol/l). Interstitial fluid was collected using a validated method, and interstitial insulin was determined with a radioimmunoassay. At 0.1 U/l, insulin transfer velocity was similar in both experimental groups (half-times of transfer: 11+/-0.2 min in obese and 18+/-4 min in lean rats; NS), but at 0.5 U/l, the respective half-times were 7+/-1 min in lean and 13+/-2 min in obese rats (P < 0.05). The steady-state level of insulin in the interstitium was 34+/-1% of the vascular level at 0.1 U/l and reached the vascular level (102+/-2%) at 0.5 U/l in both lean and obese rats. In rats aged 18 weeks, the half-times of insulin transfer were 31+/-2 and 14+/-l min in obese rats and 10+/-0.3 and 7+/-0.3 min in lean rats (P < 0.05). Again, interstitial steady-state levels were similar in both groups. Finally, postprandial insulin dynamics were simulated over a period of 120 min with a peak concentration of 0.8 U/l in rats aged 27 weeks (n = 4). The maximal interstitial level was 0.38+/-0.02 U/l in lean rats and 0.24+/-0.02 U/l in obese rats (P < 0.05), and a similar difference was noted throughout insulin infusion (areas under the transudate concentration-time curves: 17 and 11 U/min per 1, respectively). These data show, for the first time in a genetic animal model of insulin resistance, that transfer of insulin across the endothelium is substantially delayed in obese insulin-resistant rats and that it likely contributes to the postprandial alterations of glucose metabolism observed in the metabolic syndrome.     

Normalization of insulin sensitivity with lithium in diabetic rats

Lithium salts are commonly used in psychiatric patients and have been shown to have an insulinlike action in vitro. To define the impact of lithium ion on in vivo glucose metabolism, the effect of 2 wk of lithium treatment on plasma glucose and insulin concentrations, insulin-mediated glucose disposal, and skeletal muscle glycogen synthesis in normal and diabetic rats was examined. Our results demonstrated the ability of lithium ions to completely restore insulin sensitivity to normal in diabetic rats. The insulin-mimetic activity of the cation seems to be highly specific for the glycogenic pathway in skeletal muscle. These results raise the possibility that lithium ion may prove effective in reversing the defect in glycogen storage that characterizes non-insulin-dependent diabetes mellitus in humans.     

Oxidative stress in the liver and the brain of rats in fulminant hepatic failure

The etiological mechanisms of brain edema in fulminant hepatic failure are incompletely understood. In a surgical model of fulminant hepatic failure in the rat, we tested whether oxidative stress may be involved in the early steps of brain edema. Moreover, we took advantage of this model to determine if oxidative stress may be involved in the hepatocyte dysfunction observed in the setting of fulminant hepatic failure. Oxidative stress was evaluated by measurement of tissue ascorbic acid in the brain and liver of rats at 6 hours after induction of fulminant hepatic failure versus in control or partially hepatectomized rats. After 6 hours, the level of ascorbic acid was not different in the brain tissue of the various groups, indicating no oxidative stress. The liver showed a significant decrease in ascorbic acid levels, both in ischemic and nonischemic liver tissue, suggesting that oxidative stress might be involved in the failure of liver regeneration in fulminant hepatic failure. In this rat model no oxidative stress was demonstrated in the brain during the early phase of fulminant liver failure.     

The interaction between gamma-aminobutyric acid agonists and diltiazem in visceral antinociception in rats

To examine whether the gamma-aminobutyric acid (GABA) receptor agonists and L-type voltage-dependent calcium channel blockers potentiate each other on the visceral antinociceptive effects at the spinal cord, we assessed visceral nociception with colorectal distension (CD) test in rats with an intrathecal catheter. The measurements were performed after intrathecal administration of a GABA agonist (muscimol or baclofen), a calcium channel blocker (diltiazem), or the combination of the two. CD threshold did not change after muscimol 0.1 microg, baclofen 0.01 microg, or diltiazem 100 microg, but increased slightly after muscimol 1 microg and baclofen 0.1 microg. When muscimol 0.1 microg or 1 microg was administered with diltiazem, the increase in CD threshold was significantly larger than muscimol alone (at 5 min, 26.2% versus 0.6% MPE (maximum possible effect) or 84.5% versus 19.5%MPE, respectively; P < 0.01). The CD threshold after the combination of baclofen 0.1 microg and diltiazem also showed a significantly larger increase than that seen after baclofen alone (at 5 min, 48.0% versus 14.3% MPE; P < 0.01). Motor paralysis observed with muscimol 1 microg did not increase when muscimol was coadministered with diltiazem. In conclusion, intrathecal diltiazem in combination with a GABA agonist, muscimol or baclofen, potentiated the GABA agonists-induced visceral antinociception without increasing motor paralysis. IMPLICATIONS: Intrathecal administration of diltiazem in combination with a gamma-aminobutyric acid (GABA) agonist, muscimol or baclofen, potentiated the GABA agonists-induced visceral antinociception but did not affect motor paralysis. The present results indicate that the coadministration of the two types of drugs may be clinically useful.     

Oxidative stress and haemodialysis: role of inflammation and duration of dialysis treatment.

BACKGROUND: Oxidative stress has long been demonstrated in haemodialysis patients. However, the factors influencing their oxidative status have not been characterized extensively in these patients. Therefore, the present study was designed to investigate the influence of a large number of factors known to be associated with oxidative stress. METHODS: In the present cross-sectional study, we determined the plasma levels of lipid and protein oxidation markers in 31 non-smoking haemodialysis patients and 18 non-smoking healthy subjects, together with various components of the antioxidant system at the plasma and erythrocyte level. RESULTS: No influence of age, diabetes or iron overload on oxidative markers and plasma and erythrocyte antioxidant systems was detected in these haemodialysis patients. The lack of an association between iron overload and oxidative status may be related to the lower level of plasma ascorbate in haemodialysis patients, since ascorbate favours the generation of free iron from ferritin-bound iron. Interestingly, plasma C reactive protein (CRP) levels measured by highly sensitive CRP assay were correlated positively with plasma levels of thiobarbituric acid reactive substances (r=0.38, P<0.04) and negatively with plasma alpha-tocopherol levels (r=-0.46, P<0.01). Moreover, significant inverse correlations were observed between duration of dialysis treatment and plasma levels of alpha-tocopherol (r=-0.49, P<0.02) and ubiquinol (r=-0.40, P<0.05). CONCLUSIONS: Our results suggest that inflammatory status and duration of dialysis treatment are the most important factors relating to oxidative stress in haemodialysis patients.     

Oxidative stress and nitric oxide synthase in rat diabetic nephropathy: effects of ACEI and ARB.

BACKGROUND: Angiotensin II (Ang II) can up-regulate nicotinamide adenine dinucleotide phosphate [NAD(P)H] oxidase, whose product superoxide anion (O2-) can interact with nitric oxide (NO) to form peroxynitrite (ONOO-). We tested the hypothesis that Ang II subtype 1 (AT1) receptor activation enhances oxidative stress and nitrotyrosine deposition in the kidneys of rats with diabetes mellitus (DM). METHODS: After two weeks of streptozotocin-induced DM, rats received either no treatment, an angiotensin-converting enzyme inhibitor (ACEI) or an angiotensin receptor blocker (ARB) for two weeks. At four weeks, renal expression of the p47phox component of NAD(P)H oxidase, endothelial nitric oxide synthase (eNOS), neuronal nitric oxide synthase (nNOS), and nitrotyrosine were evaluated by Western blot and immunohistochemistry and related to plasma lipid peroxidation products (LPO), hydrogen peroxide production in the kidney and 24-hour protein excretion. RESULTS: Immunoreactive expression of p47phox and eNOS were increased in DM with an increase in plasma LPO, renal hydrogen peroxide production and nitrotyrosine deposition. Expression of nNOS was unaltered. Treatment with either ACEI or ARB prevented all these findings and also prevented significant microalbuminuria. The treatments did not affect the elevated blood sugar, nor did DM or its treatment affect the blood pressure or the creatinine clearance. CONCLUSION: Early proteinuric diabetic nephropathy increases renal expression of the p47phox component of NAD(P)H oxidase and eNOS with increased indices of systemic and renal oxidative/nitrosative stress. An ACEI or an ARB prevents these changes and prevents the development of proteinuria, independent of blood pressure or blood sugar. This finding indicates a pathogenic role for AT1 receptors in the development of oxidative damage in the kidneys during early DM.     

Oxidative stress in testicular tissues of rats exposed to cigarette smoke and protective effects of caffeic acid phenethyl ester

Aim： To show the oxidative stress after cigarette smoke exposure in rat testis and to evaluate the effects of caffeic acid phenethyl ester （CAPE）. Methods： Twenty-one rats were divided into three groups of seven. Animals in Group Ⅰ were used as control. Rats in Group Ⅱ were exposed to cigarette smoke only （4 × 30 min/d） and rats in Group Ⅲ were exposed to cigarette smoke and received daily intraperitoneal injections of CAPE （10 μmol/kg.d）. After 60 days all the rats were killed and the levels of nitric oxide （NO） and anti-oxidant enzymes such as superoxide-dismutase, catalase and glutathione peroxidase （GSH-Px） and the level of malondialdehyde were studied in the testicular tissues of rats with spectrophotometric analysis. Results： There was a significant increase in catalase and superoxide-dismutase activities in Group Ⅱ when compared to the controls, but the levels of both decreased after CAPE administration in Group Ⅲ. GSH-Px activity was decreased in Group Ⅱ but CAPE caused an elevation in GSH-Px activity in Group Ⅲ. The difference between the levels of GSH-Px in Group Ⅰ and Group Ⅱ was significant, but the difference between groups Ⅱ and Ⅲ was not significant. Elevation of malondialdehyde after smoke exposure was significant and CAPE caused a decrease to a level which was not statistically different to the control group. A significantly increased level of NO after exposure to smoke was reversed by CAPE administration and the difference between NO levels in groups Ⅰ and Ⅲ was statistically insignificant. Conclusion： Exposure to cigarette smoke causes changes in the oxidative enzyme levels in rat testis, but CAPE can reverse these harmful effects. （Asian J Andro12006 Mar; 8： 189-193）     

Histomorphometric analysis of diabetic osteopenia in streptozotocin-induced diabetic mice: a possible role of oxidative stress

Diabetic osteopenia causes an increase in bone fracture and a delay in healing of fractures, and affects the quality of life. However, the mechanisms responsible for the disease have not been clearly identified. Oxidative stress may be a potential candidate for the pathogenesis, since it is increased under diabetic conditions and is known to induce cellular dysfunction in a wide variety of cell types. Although in vitro studies have shown that oxidative stress inhibits osteoblastic differentiation and induces osteoblast insults and apoptosis, the relationship between diabetic osteopenia and oxidative stress remains unclear. To explore these issues, analysis of a mouse model that represents the diabetic osteopenia as seen in patients with diabetes is necessary. However, there are few reports of such a model. Therefore, we focused on the streptozotocin (STZ)-induced diabetic mouse, one of the most common animal models of type 1 diabetes. Eight-week-old male C57BL/6 mice were randomly assigned to the following three groups: 1) control group, 2) diabetic group, and 3) insulin-treated diabetic group. After 12 weeks of STZ treatment, the physical properties of the femora, and the static and dynamic parameters of bone histomorphometry of the tibiae from STZ-induced diabetic mice (STZ-mice) were assessed, and oxidative stress in the whole body and bone of the mice was evaluated. Renal function was comparable in all three groups at the end of the experimental period. In addition, no significant difference in serum PTH, Ca, and P was found among the three groups. In contrast, radiological analysis demonstrated a significant decrease in trabecular bone volume, and histomorphometric analyses confirmed that parameters for both bone formation (OV/BV, OS/BS, and BFR/BS) and bone resorption (ES/BS and Oc.S/BS) were also significantly lower in STZ-mice. In addition, urinary excretion of 8-hydroxydeoxyguanosine, a marker of oxidative DNA damage, was elevated in STZ-mice. Further immunohistological studies showed intensified immunostaining of an oxidative stress marker in bone tissue including the osteoblasts of diabetic mice. Here, we demonstrated that STZ-mice exhibit low-turnover osteopenia associated with increased oxidative stress.