Angiotensin-converting enzyme inhibition augments the expression of rat elastase-2, an angiotensin II-forming enzyme

Becari C, Teixeira FR, Oliveira EB, Salgado MC. Angiotensin-converting enzyme inhibition augments the expression of rat elastase- 2, an angiotensin II-forming enzyme. Am J Physiol Heart Circ Physiol 301: H565-H570, 2011. First published May 20, 2011; doi:10.1152/ajpheart.00534.2010.-Mounting evidence suggest that tissue levels of angiotensin (ANG) II are maintained in animals submitted to chronic angiotensin-converting enzyme (ACE) inhibitor treatment. We examined the expression levels of transcripts for elastase-2, a chymostatin-sensitive serine protease identified as the alternative pathway for ANG II generation from ANG I in the rat vascular tissue and the relative role of ACE-dependent and -independent pathways in generating ANG II in the rat isolated carotid artery rings of spontaneously hypertensive rats (SHR) and Wistar normotensive rats (WNR) treated with enalapril for 7 days. Enalapril treatment decreased blood pressure of SHR only and resulted in significantly more elastase-2 mRNA expression in carotid artery of both enalapril-treated WNR and SHR. Captopril induced a comparable rightward shift of concentration-response curves to ANG I in vehicle and enalapril-treated rats, although this effect was of lesser magnitude in SHR group. Chymostatin induced a rightward shift of the dose response to ANG I in vehicle-treated and a decrease in maximal effect of 22% in enalapril-treated WNR group. Maximal response induced by ANG I was remarkably reduced by chymostatin in enalapril-treated SHR carotid artery (by 80%) compared with controls (by 23%). Our data show that chronic ACE inhibition was associated with augmented functional role of non-ACE pathway in generating ANG II and increased elastase-2 gene expression, suggesting that this protease may contribute as an alternative pathway for ANG II generation when ACE is inhibited in the rat vascular tissue.

Preoperative respiratory muscle dysfunction is a predictor of prolonged invasive mechanical ventilation in cardiorespiratory complications after heart valve surgery

Objective: To verify whether preoperative respiratory muscle strength and ventilometric parameters, among other clinically relevant factors, are associated with the need for prolonged invasive mechanical ventilation (PIMV) due to cardiorespiratory complications following heart valve surgery. Methods: Demographics, preoperative ventilometric and manometric data, and the hospital course of 171 patients, who had undergone heart valve surgery at Hospital das Clinicas da Faculdade de Medicina de Ribeirao Preto, were prospectively collected and subjected to univariate analysis for identifying the risk factors for PIMV. Results: The hospital mortality was 7%. About 6% of the patients, who had undergone heart valve surgery required PIMV because of postoperative cardiorespiratory dysfunction. Their hospital mortality was 60% (vs 4%, p < 0.001). Univariate analysis revealed that preoperative respiratory muscle dysfunction, characterized by maximal inspiratory and expiratory pressure below 70% of the predicted values combined with respiratory rate above 15 rpm during ventilometry, was associated with postoperative PIMV (p = 0.030, odds ratio: 50, 95% confidence interval (CI): 1.2-18). Postoperative PIMV was also associated with: (1) body mass index (BMI) < 18.5 (odds ratio: 7.2, 95% CI: 1.5-32), (2) body weight < 50 kg (odds ratio: 6.5, 95% CI: 1.6-25), (3) valve operation due to acute endocarditis (odds ratio: 5.5, 95% CI: 0.98-30), and (4) concomitant operation for mitral and tricuspid valve dysfunction (p = 0.047, odds ratio: 5.0, 95% CI: 1.1-22). Conclusion: Our results have demonstrated that respiratory muscle dysfunction, among other clinical factors, is associated with the need for PIMV due to cardiovascular or pulmonary dysfunction after heart valve surgery. (C) 2010 European Association for Cardio-Thoracic Surgery. Published by Elsevier B. V. All rights reserved.

Etiopathogenesis of Hepatic Osteodystrophy in Wistar Rats with Cholestatic Liver Disease

The pathophysiology of hepatic osteodystrophy (HO) remains poorly understood. Our aim was to evaluate bone histomorphometry, biomechanical properties, and the role of the growth hormone (GH)/insulin-like growth factor-I (IGF-I) system in the onset of this disorder. Forty-six male Wistar rats were divided into two groups: sham-operated (SO, n = 23) and bile duct-ligated (BDL, n = 23). Rats were killed on day 30 postoperatively. Immunohistochemical expression of IGF-I and GH receptor was determined in liver tissue and in the proximal growth plate cartilage of the left tibia. Histomorphometric analysis was performed in the right tibia, and the right femur was used for biomechanical analysis. The maximal force at fracture and the stiffness of the mid-shaft femur were, respectively, 53% and 24% lower in BDL compared to SO. Histomorphometric measurements showed low cancellous bone volume and decreased cancellous bone connectivity in BDL, compatible with osteoporosis. This group also showed increased mineralization lag time, indicating disturbance in bone mineralization. Serum levels of IGF-I were lower in BDL (basal 1,816 +/- A 336 vs. 30 days 1,062 +/- A 191 ng/ml, P < 0.0001). BDL also showed higher IGF-I expression in the liver tissue but lower IGF-I and GH receptor expression in growth plate cartilage than SO. Osteoporosis is the most important feature of HO; BDL rats show striking signs of reduced bone volume and decreased bone strength, as early as after 1 month of cholestasis. The endocrine and autocrine-paracrine IGF-I systems are deeply affected by cholestasis. Further studies will be necessary to establish their role in the pathogenesis of HO.

Effects of acute magnesium loading on pulmonary function of stable COPD patients

Background: Magnesium (Mg) use has the potential to promote bronchodilatation and to improve lung function in obstructive diseases. IV administration of Mg during exacerbations of chronic obstructive pulmonary disease (COPD) has led to improved peak flow. This study aimed to investigate the effects of acute IV Mg loading on respiratory parameters of stable COPD patients. Material/Methods: This was a randomized, double-blind, placebo-controlled crossover study. Twenty-two male COPD patients (64 +/- 6 years old, FEV1: 49 +/- 20%) received an IV infusion of 2 g of magnesium sulfate or placebo on two distinct occasions. Spirometry and mouth maximal respiratory pressures were obtained before and 45 minutes after the infusions. Results: Mg use led to significant changes in functional respiratory capacity (-0.48 1,95% CI: -0.96, -0.01), inspiratory capacity (0.21 1,95% CI: 0.04, 0.37). The treatment was also associated with a marginally significant decrease in residual volume (-0.47 1,95% CI: -0.96, 0.02, p=0.06). Conclusions: Acute IV Mg loading in stable COPD patients was associated with a reduction in lung hyperinflation and improvement of respiratory muscle strength. The clinical potential for chronic magnesium supplementation in COPD deserves further investigation.

Salicylates dilate blood vessels through inhibiting PYK2-mediated RhoA/Rho-kinase activation

Aims Compared with other non-steroid anti-inflammatory drugs (NSAIDs), aspirin is not correlated to hypertension. It has been shown that aspirin has unique vasodilator action in vivo, offering an explanation for the unique blood pressure effect of aspirin. In the present study, we investigate the mechanism whereby salicylates (aspirin and sodium salicylate) dilate blood vessels. Methods and results Rat aortic or mesenteric arterial rings were used to test the vascular effect of salicylates and other NSAIDs. RhoA translocation and the phosphorylation of MYPT1, the regulatory subunit of myosin light chain phosphatase, were measured by western blot, as evidenced for RhoA/Rho-kinase activation. Salicylates, but not other NSAIDs, relaxed contraction induced by most tested constrictors except for calyculin A, indicating that RhoA/Rho-kinase-mediated calcium sensitization is involved. The involvement of RhoA/Rho kinase in vasodilation by salicylates was confirmed by measurements of RhoA translocation and MYPT1 phosphorylation. The calculated half maximal inhibitory concentration (IC(50)) of vasodilation was apparently higher than that of cyclooxygenase inhibition, but comparable to that of proline-rich tyrosine kinase 2 (PYK2) inhibition. Over-expression of PYK2 induced RhoA translocation and MYPT1 phosphorylation, and these effects were markedly inhibited by sodium salicylate treatment. Consistent with the ex vitro vascular effects, sodium salicylate acutely decreased blood pressure in spontaneous hypertensive rats but not in Wistar Kyoto rats. Conclusion Salicylates dilate blood vessels through inhibiting PYK2-mediated RhoA/Rho-kinase activation and thus lower blood pressure.

Endothelin-1 Induces Contraction of Female Rat Internal Pudendal and Clitoral Arteries through ET(A) Receptor and Rho-Kinase Activation

Introduction. Endothelin-1 (ET-1), a potent vasoconstrictor peptide, acts mainly through the Gprotein-coupled ET(A) receptor (ET(A)R). Increased vascular ET-1 production and constrictor sensitivity have been observed in various cardiovascular diseases, including hypertension, as well as erectile dysfunction. The internal pudendal artery (IPA) supplies blood to the vagina and clitoris. Inadequate blood flow through the IPA may lead to insufficient vaginal engorgement and clitoral tumescence. Aim. Characterize the effects of ET-1 on the IPA and clitoral artery (CA). Methods. IPA and CA from female Sprague Dawley rats (225-250 g) were mounted in myograph chambers. Arterial segments were submitted to increasing concentrations of ET-1 (10-10-10-6 M). Segments were incubated with the ET(A)R antagonist, atrasentan (10-8 M) or the Rho-kinase inhibitor, Y-27632 (10-6 M) 30 minutes prior to agonist exposure. All E(max) values are expressed as % KCl-induced maximal contraction. ET(A)R, RhoA, and Rho-kinase expression from IPA was evaluated by Western blot. mRNA of preproET-1, ET(A)R, ET(B)R, RhoA, and Rho-kinase were measured by real time PCR. Main Outcome Measures. ET-1 constrictor sensitivity in IPA and CA, protein expression and messenger RNA levels of ET-1-mediated constriction components. Results. ET-1 concentration-dependently contracted IPA (% Contraction and pD2, respectively: 156 +/- 18, 8.2 +/- 0.1) and CA (163 +/- 12, 8.8 +/- 0.08), while ET(A)R antagonism reduced ET-1-mediated contraction (IPA: 104 +/- 23, 6.4 +/- 0.2; CA: 112 +/- 17, 6.6 +/- 0.08). Pretreatment with Y-27632 significantly shifted ET-1 pD2 in IPA (108 +/- 24, 7.9 +/- 0.1) and CA (147 +/- 58 and 8.0 +/- 0.25). Protein expression of ET(A)R, ET(B)R, RhoA, and Rho-kinase were detected in IPA. IPA and CA contained preproET-1, ET(A)R, ET(B)R, RhoA, and Rho-kinase message. Conclusion. We observed that the IPA and CA are sensitive to ET-1, signaling through the ET(A)R and Rho-kinase pathway. These data indicate that ET-1 may play a role in vaginal and clitoral blood flow and may be important in pathologies where ET-1 levels are elevated. Allahdadi KJ, Hannan JL, Tostes RC, and Webb RC. Endothelin-1 induces contraction of female rat internal pudendal and clitoral arteries through ETA receptor and Rho-kinase activation. J Sex Med 2010;7:2096-2103.

Pharmacological analysis of the neutrophil migration induced by D. rostrata lectin: Involvement of cytokines and nitric oxide

In the present study, we investigated the involvement of resident cell and inflammatory mediators in the neutrophil migration induced by chemotactic activity of a glucose/mannose-specific lectin isolated from Dioclea rostrata seeds (DrosL). Rats were injected i.p. with DrosL (125-1000 mu g/cavity), and at 2-96 h thereafter the leukocyte counts in peritoneal fluid were determined. DrosL-induced a dose-dependent neutrophil migration accumulation, which reached maximal response at 24 h after injection and declines thereafter. The carbohydrate ligand nearly abolished the neutrophil influx. Pre-treatment of peritoneal cavities with thioglycolate which increases peritoneal macrophage numbers, enhanced neutrophil migration induced by DrosL by 303%. However, the reduction of peritoneal mast cell numbers by treatment of the cavities with compound 48/80 did not modify DrosL-induced neutrophil migration. The injection into peritoneal cavities of supernatants from macrophage cultures stimulated with DrosL (125, 250 and 500 mu g/ml) induced neutrophil migration. In addition, DrosL treatment induced cytokines (TNF-alpha, IL-1 beta and CINC-1) and NO release into the peritoneal cavity of rats. Finally, neutrophil chemotaxis assay in vitro showed that the lectin (15 and 31 mu g/ml) induced neutrophil chemotaxis by even 180%. In conclusion, neutrophil migration induced by D. rostrata lectin occurs by way of the release of NO and cytokines such as IL-1 beta, TNF-alpha and CINC-1. (C) 2009 Elsevier Ltd. All rights reserved.

Increased sympathetic outflow in juvenile rats submitted to chronic intermittent hypoxia correlates with enhanced expiratory activity

Chronic intermittent hypoxia (CIH) in rats produces changes in the central regulation of cardiovascular and respiratory systems by unknown mechanisms. We hypothesized that CIH (6% O(2) for 40 s, every 9 min, 8 h day(-1)) for 10 days alters the central respiratory modulation of sympathetic activity. After CIH, awake rats (n = 14) exhibited higher levels of mean arterial pressure than controls (101 +/- 3 versus 89 +/- 3 mmHg, n = 15, P < 0.01). Recordings of phrenic, thoracic sympathetic, cervical vagus and abdominal nerves were performed in the in situ working heart-brainstem preparations of control and CIH juvenile rats. The data obtained in CIH rats revealed that: (i) abdominal (Abd) nerves exhibited an additional burst discharge in late expiration; (ii) thoracic sympathetic nerve activity (tSNA) was greater during late expiration than in controls (52 +/- 5 versus 40 +/- 3%; n = 11, P < 0.05; values expressed according to the maximal activity observed during inspiration and the noise level recorded at the end of each experiment), which was not dependent on peripheral chemoreceptors; (iii) the additional late expiratory activity in the Abd nerve correlated with the increased tSNA; (iv) the enhanced late expiratory activity in the Abd nerve unique to CIH rats was accompanied by reduced post-inspiratory activity in cervical vagus nerve compared to controls. The data indicate that CIH rats present an altered pattern of central sympathetic-respiratory coupling, with increased tSNA that correlates with enhanced late expiratory discharge in the Abd nerve. Thus, CIH alters the coupling between the central respiratory generator and sympathetic networks that may contribute to the induced hypertension in this experimental model.

Ageing and surface EMG activity patterns of masticatory muscles

P>The purpose of this study was to evaluate the influence of age on the electromyographic activity of masticatory muscles. All volunteers were Brazilian, fully dentate (except for Group I - mixed dentition), Caucasian, aged 7-80, and divided into five groups: I (7-12 years), II (13-20 years), III (21-40 years), IV (41-60 years) and V (61-80 years). Except for Group V, which comprised nine women and eight men, all groups were equally divided with respect to gender (20 M/20 F). Surface electromyographic records of masticatory muscles were obtained at rest and during maximal voluntary contraction, right and left laterality, maximal jaw protrusion and maximal clenching in the intercuspal position. Statistically significant differences (P < 0 center dot 05) were found in all clinical conditions among the different age groups. Considerably different patterns of muscle activation were found across ages, with greater electromyographic activity in children and youth, and decreasing from adults to aged people.

Evaluation of molar and incisor bite force in indigenous compared with white population in Brazil

The purpose of this study was to deter-mine maximum bite force in molar and incisor regions in young Brazilian indigenous individuals, who have had a natural diet since birth, and compare the sample with white Brazilian individuals. To do this, individuals were paired one-to-one (same weight, height, and Class I facial pattern). A secondary purpose was to elucidate the relation between bite force and gender in both populations. Eighty-two Brazilians took part in this study. Participants were aged between 18 and 28 years and were divided into two groups: 41 Xingu indigenous individuals and 41 white Brazilian individuals, with 28 men and 13 women in each group. The inclusion criteria were: having complete dentition; normal occlusion; no neurological, psychiatric or movement disorders.; no reports of toothaches; having satisfactory periodontal health; absence of large facial skeletal alterations (typical Class II and Class III individuals); and no previous treatments using occlusal splints. To measure maximum bite force, a digital dynamometer model IDDK (Kratos-Equipamentos Industriais Ltda, Cotia, Sao Paulo, Brazil) was used, with a capacity of 1000 N, adapted for oral conditions. Assessments were made in the first molar (right and left) and central incisive regions. Results reveal that mean maximum bite forces in indigenous individuals of the right molar is 421 N, left molar 429 N and incisor region is 194 14 and for white individuals of the right molar is 410 N, left molar 422 N and incisor region is 117 N. Comparing indigenous with white individuals, maximal bite force showed a tendency of being greater in the indigenous group. It was observed that the incisor region showed statistical significance (p < 0.0005) but no significance was observed in the molar region. Moreover, indigenous men showed the highest bite force values. (C) 2007 Elsevier Ltd. All rights reserved.

Masticatory muscle activity in children with a skeletal or dentoalveolar open bite

Forty-five children (31 boys and 14 girls), aged 6-11 years, were included in the study, 15 with a skeletal anterior open bite (SAOB), 15 with a dentoalveolar anterior open bite (DAOB), and 15 with a normal occlusion (CG), defined by clinical evaluation and lateral cephalograms. EMG recordings of the temporal and masseter muscles were performed under maximal voluntary clenching and during chewing. Analysis of variance was used for inter-group analysis, followed by the Tukey post hoc test. A Student`s t-test for paired data was used for intra-group analysis. There were statistically significant differences among the three groups (P < 0.05), with the mean EMG being highest in the CG and lowest in children with a SAOB. The percentage EMG activity during chewing in relation to that during maximal voluntary clenching was more than 100 per cent in the SAOB group. The CG and DAOB groups presented higher EMG activity during clenching compared with chewing (P < 0.001), as well as a greater difference between tasks. In the SAOB group, the neuromuscular system appeared to have a lower capacity to produce EMG activity according to the task, while that in the DAOB group suggests that their functional capacity during growth should also be carefully observed.

Mineralocorticoid Receptor Mutations Differentially Affect Individual Gene Expression Profiles in Pseudohypoaldosteronism Type 1

Context: Type 1 pseudohypoaldosteronism (PHA1), a primary form of mineralocorticoid resistance, isdueto inactivating mutations of the NR3C2 gene, coding for the mineralocorticoid receptor (MR). Objective: The objective of the study was to assess whether different NR3C2 mutations have distinct effects on the pattern of MR-dependent transcriptional regulation of aldosterone-regulated genes. Design and Methods: Four MR mutations affecting residues in the ligand binding domain, identified in families with PHA1, were tested. MR proteins generated by site-directed mutagenesis were analyzed for their binding to aldosterone and were transiently transfected into renal cells to explore the functional effects on the transcriptional activity of the receptors by cis-trans-cotrans-activation assays and by measuring the induction of endogenous gene transcription. Results: Binding assays showed very low or absent aldosterone binding for mutants MR(877Pro), MR(848Pro), and MR(947stop) and decreased affinity for aldosterone of MR(843Pro). Compared with wildtype MR, the mutations p.Leu843Pro and p.Leu877Pro displayed half-maximal aldosterone-dependent transactivation of reporter genes driven by mouse mammary tumor virus or glucocorticoid response element-2 dependent promoters, whereas MR(848Pro) and MR(947stop) nearly or completely lost transcriptional activity. Although MR(848Pro) and MR(947stop) were also incapable of inducing aldosterone-dependent gene expression ofendogenoussgk1, GILZ, NDRG2, and SCNN1A, MR(843Pro) retained complete transcriptional activity on sgk1 and GILZ gene expression, and MR(877Pro) negatively affected the expression of sgk1, NDRG2, and SCNN1A. Conclusions: Our data demonstrate that MR mutations differentially affect individual gene expression in a promoter-dependent manner. Investigation of differential gene expression profiles in PHA1 may allow a better understanding of the molecular substrate of phenotypic variability and to elucidate pathogenic mechanisms underlying the disease. (J Clin Endocrinol Metab 96: E519-E527, 2011)

Obesity due to Melanocortin 4 Receptor (MC4R) Deficiency Is Associated with Increased Linear Growth and Final Height, Fasting Hyperinsulinemia, and Incompletely Suppressed Growth Hormone Secretion

Context: Melanocortin receptor 4 (MC4R) deficiency is characterized by increased linear growth greater than expected for the degree of obesity. Objective: The objective of the investigation was to study the somatotroph axis in obese MC4R-deficient patients and equally obese controls. Patients and Methods: We obtained anthropometric measurements and insulin concentrations in 153 MC4R-deficient subjects and 1392 controls matched for age and severity of obesity. We measured fasting IGF-I, IGF-II, IGF binding protein (IGFBP)-1, IGFBP-3, and acid-labile subunit levels in a subset of 33 MC4R-deficient patients and 36 control subjects. We examined pulsatile GH secretion in six adult MC4R-deficient subjects and six obese controls. Results: Height so score was significantly greater in MC4R-deficient children under 5 yr of age compared with controls (mean +/- SEM: 2.3 +/- 0.06 vs. 1.8 +/- 0.04, P < 0.001), an effect that persisted throughout childhood. Final height (cm) was greater in MC4R-deficient men (mean +/- SEM 173 +/- 2.5 vs. 168 +/- 2.1, P < 0.001) and women (mean 165 +/- 2.1 vs. 158 +/- 1.9, P < 0.001). Fasting IGF-I, IGF-II, acid-labile subunit, and IGFBP-3 concentrations were similar in the two groups. GH levels were markedly suppressed in obese controls, but pulsatile GH secretion was retained in MC4R deficiency. The mean maximal GH secretion rate per burst (P < 0.05) and mass per burst (P < 0.05) were increased in MC4R deficiency, consistent with increased pulsatile and total GH secretion. Fasting insulin levels were markedly elevated in MC4R-deficient children. Conclusions: In MC4R deficiency, increased linear growth in childhood leads to increased adult final height, greater than predicted by obesity alone. GH pulsatility is maintained in MC4R deficiency, a finding consistent with animal studies, suggesting a role for MC4R in controlling hypothalamic somatostatinergic tone. Fasting insulin levels are significantly higher in children carrying MC4R mutations. Both of these factors may contribute to the accelerated growth phenotype characteristic of MC4R deficiency. (J Clin Endocrinol Metab 96: E181-E188, 2011)

Assessment of two central auditory tests in elderly patients without hearing complaints

Speech understanding disorders in the elderly may be due to peripheral or central auditory dysfunctions. Asymmetry of results in dichotic testing increases with age, and may reflect on a lack of inter-hemisphere transmission and cognitive decline. Aim: To investigate auditory processing of aged people with no hearing complaints. Study design: clinical prospective. Materials and Methods: Twenty-two voluntary individuals, aged between 55 and 75 years, were evaluated. They reported no hearing complaints and had maximal auditory thresholds of 40 dB HL until 4 KHz, 80% of minimal speech recognition scores and peripheral symmetry between the ears. We used two kinds of tests: speech in noise and dichotic alternated dissyllables (SSW). Results were compared between males and females, right and left ears and between age groups. Results: There were no significant differences between genders, in both tests. Their Left ears showed worse results, in the competitive condition of SSW. Individuals aged 65 or older had poorer performances than those aged 55 to 64. Conclusion: Central auditory tests showed worse performance with aging. The employment of a dichotic test in the auditory evaluation setting in the elderly may help in the early identification of degenerative processes, which are common among these patients.

Chronic alcoholism associated with diabetes impairs erectile function in rats

OBJECTIVE To investigate the effects of chronic ethanol consumption and diabetes on nitric oxide (NO)-mediated relaxation of cavernosal smooth muscle (CSM). MATERIAL AND METHODS Male Wistar rats were divided into four groups: control, isocaloric, diabetic and ethanol-diabetic. The CSMs were mounted in organ chambers for measurement of isometric tension. Contraction of the strips was induced by electrical field stimulation (EFS, 1-32 Hz) and phenylephrine. We also evaluated the effect of ethanol consumption on the relaxation induced by acetylcholine (ACh; 0.01-1000 mu mol/L), sodium nitroprusside (SNP, 0.01-1000 mu mol/L) or EFS (1-32 Hz) in strips pre-contracted with phenylephrine (10 mu mol/L). Immunoexpression of endothelial NO synthase (eNOS) and inducible NOS (iNOS) was also accessed. RESULTS The endothelium-dependent relaxation induced by ACh was decreased in CSM from ethanol-diabetic rats when compared with the controls, with a mean (sem) of 21 (4) vs 37 (2)%. Similarly, the potency and maximal responses induced by SNP were reduced in the ethanol-diabetic [3.97 (0.38) and 85 (1)%, respectively] and diabetic groups [3.78 (0.56) and 81 (2)%, respectively] when compared with the controls [5.3 (0.22) and 90 (3)%, respectively] and isocaloric [5.3 (0.19) and 92 (1)%, respectively] groups. Noradrenergic nerve-mediated contractions of CSM in response to EFS were increased in rats from ethanol-diabetic and diabetic groups when compared with the control and isocaloric groups. Conversely, there were no differences in EFS-induced relaxation among the groups. The immunostaining assays showed overexpression of eNOS and iNOS in the CSM from diabetic and ethanol-diabetic rats when compared with the control and isocaloric rats. CONCLUSION There was an impairment of relaxation of CSM from ethanol-diabetic and diabetic rats that involved a decrease in the NO-cyclic guanosine monophosphate signalling pathway by endothelium-dependent mechanisms accompanied by a change in the CSM contractile sensitivity.