Influence of LH and high-density lipoprotein cholesterol (HDL-C) on metformin response in women with polycystic ovary syndrome

Objective: To search for predictors of metformin response in women with polycystic ovary syndrome (PCOS) through a detailed analysis of clinical and laboratory parameters. Study design: We designed a prospective study to investigate clinical and laboratory parameters to search for predictors of metformin response in women with PCOS. A total of 53 PCOS patients were given metformin 850 mg twice a day for 6 months, after which patients were classified as responders or non-responders. Parameters analyzed for comparison between the two groups were: plasma fasting insulin glucose/insulin ratio; oral glucose tolerance test (OGTT) with insulin (120 min); HOMA and QUICKI tests; total cholesterol and fractions, triglycerides; LH, FSH, estradiol, progesterone, testosterone, androstenedione, 17-OH progesterone, and DHEAS. Results: From all patients, 30(56.6%) were responders and 23(43.3%) were non-responders. Multinomial analysis showed that the positive response to metformin was associated with higher levels of basal LH (p = 0.038) and lower levels of high-density lipoprotein cholesterol (HDL-C) (p = 0.015). Conclusion: In weight-matched PCOS subjects, laboratory markers might predict the metformin response. Higher levels of basal LH and lower levels of HDL-C are correlated with a positive response to metformin treatment in PCOS subjects. (C) 2011 Elsevier Ireland Ltd. All rights reserved.

Relative Bioavailability of Two Oral Formulations of Risperidone 2 mg: A Single-Dose, Randomized-Sequence, Open-Label, Two-Period Crossover Comparison in Healthy Brazilian Volunteers

Background: Risperidone (RSP) is a benzisoxazole antipsychotic agent used to treat schizophrenia and other psychiatric illnesses in adults and children (including those with autism). After oral administration, RSP is completely absorbed from the gastrointestinal tract and undergoes hydroxylation to yield 9-hydroxyrisperidone (9-OH-RSP), an active metabolite that has a pharmacologic profile and potency similar to RSP. Objectives: The aims of this study were to compare the relative bioavailability of a pharmaceutical-equivalent (test) formulation with a reference formulation of oral RSP 2 mg, both available commercially on the Brazilian pharmaceutical market, and to generate data regarding the oral bioavailability of the tested drug in healthy Brazilian volunteers. Methods: This single-dose, randomized-sequence, open-label, 2-period crossover study was conducted in healthy Brazilian volunteers from August to December 2008. Subjects were randomly assigned to receive the test formulation followed by the reference formulation or vice versa, with a 30-day washout period between doses. Study drugs were administered after a 12-hour overnight fast. For pharmacokinetic analysis, blood samples were drawn at 0 (baseline), 0.25, 0.5, 1, 1.5, 3, 5, 8, 12, 24, 48, 72, 96, and 120 hours after administration. Plasma concentrations of RSP and 9-OH-RSP were determined using LC-MS/MS. The test and reference formulations were to be considered bioequivalent if the 90% CIs for the geometric mean test/reference ratios were within a predetermined range of 80% to 125%, in accordance with the policies of the Brazilian Sanitary Surveillance Agency and the US Food and Drug Administration. Tolerability was determined using clinical assessments, monitoring of vital signs, analysis of laboratory test results, and subject interviews regarding adverse events. Results: A total of 22 subjects were enrolled (11 men, 11 women; mean [SD] age, 32 [12] years [range, 18-58 years]; weight, 70.4 [11.9] kg [range, 50-103 kg]; height, 1.67 [0.08] m [range, 1.56-1.80 m]; and body mass index, 25 [4] kg/m(2) [range, 18-29 kg/m(2)]). For RSP, mean (SD) C(max) values were 12.6 (2.7) and 16.0 (2.3) ng/mL for the test and reference formulations, respectively. For 9-OH-RSP, mean C(max) values were 17.8 (1.3) and 21.0 (1.7) ng/mL for the test and reference formulations. The 90% CIs for the mean test/reference ratios for RSP C(max), AUC(0-120), and AUC(0-infinity) were 74% to 82%, 75% to 85%, and 76% to 85%, respectively, and 83% to 87%, 75% to 79%, and 75% to 78% for 9-OH-RSP. The related adverse events (headache, low back pain, drowsiness, standing hypotension, local postvenipuncture ecchymoses, insomnia, nausea, and vomiting) were transient and mild. Conclusions: This single-dose study found that the test and reference formulations of oral RSP 2 mg did not meet the Brazilian and US regulatory criteria for bioequivalence in these fasting, healthy volunteers. The study formulations appeared to be well tolerated. (Clin Ther 2010;32:2106-2115) (C) 2010 Elsevier HS Journals, Inc.

Vitamin D status in a sunny country: Where has the sun gone?

Background & aims: Hypovitaminosis D [serum 25 vitamin D < 30 ng/ml] is related to the development of metabolic bone disease and greater risk of chronic illnesses. However, it is frequently under-diagnosed, mainly in countries where UV radiation is abundant. We prospectively determined the prevalence and the predictors of serum 25 vitamin D (s25(OH)D) in a healthy Brazilian population after the winter and after the summer. Methods: 603 (118M and 485F) healthy Brazilian volunteers aged 18-90 years from a universitary hospital were selected after the winter of 2006. From the initial sample, 209 volunteers (31M and 178F) accepted to participate in a second health check after the subsequent summer. Results: After the winter, median s25(OH)D was 21.4 ng/mL and 77.4% of the population presented hypovitaminosis D. s25(OH)D was significantly related to age, BMI, PTH and race. In multivariate linear regression analysis, s25(OH)D was significantly and independently dependent on age, glycemia and skin color. Significant increase in s25(OH)D was verified after summer [10.6 (3.7-19.3 ng/ml); p < 0.001] and this improvement was dependent on age. We also observed a significant decrease in hyperparathyroidism prevalence (20.8% vs. 4.9%; P < 0.0001). Conclusion: In Sao Paulo, at the end of winter, we observed a high prevalence of hypovitaminosis D and secondary hyperparathyroidism in healthy adults. s25(OH)D was dependent on age and skin color. After summer, we observed a decrease in the prevalence of hypovitaminosis D. This unexpected finding emphasizes the need for a strong recommendation to monitor s25(OH)D, even in a sunny country such as Brazil. (C) 2010 Elsevier Ltd and European Society for Clinical Nutrition and Metabolism. All rights reserved.

Laparoscopic resection of uncinate process of the pancreas

Solid pseudopapillary neoplasm of the pancreas is an uncommon but distinctive pancreatic neoplasm with low metastatic potential [1]. Therefore, whenever feasible, an organ-preserving operation should be performed. As previously reported, women with solid pseudopapillary neoplasm of the pancreas may be best treated by more conservative procedures [2]. Recently, laparoscopic pancreatic resections became more common and are being performed in highly specialized centers. There are only six cases of laparoscopic resection for solid pseudopapillary neoplasm of pancreas published in the English literature and, to our knowledge, laparoscopic resection of uncinate process of the pancreas has never been reported [3-6]. This video demonstrates the technical aspects of a totally laparoscopic resection of the uncinate process of the pancreas in a patient with solid pseudopapillary neoplasm. A 26-year-old woman with a 4-cm solid pseudopapillary pancreatic neoplasm was referred for surgical treatment. According to preoperative echoendoscopy, there was a safe margin between neoplasm and main pancreatic duct. The patient was placed in supine position with the surgeon standing between her legs. Four trocars, one 10-mm and three 5-mm, were used. At inspection, the inferior vena cava, transverse colon, duodenum, and pancreas are clearly identified. A Kocher maneuver was performed with complete exposure of pancreatic head and uncinate process. The uncinate process was dissected from the superior mesenteric vein and venous branches were divided between metallic clips or by use of laparoscopic coagulation shears (LCS; Ethicon Endo Surgery Industries, Cincinnati, OH, USA). Blood supply of the duodenum was preserved by ligature of small pancreatic branches from inferior pancreatoduodenal artery. Transection of pancreatic parenchyma was performed using laparoscopic coagulation shears, which is an effective tool for cutting the pancreas [7, 8]. Surgical specimen was removed through a suprapubic incision inside a retrieval bag. A hemostatic absorbable tissue (Surgicel; Ethicon Inc., Cincinnati, OH) was placed in the cutting pancreatic surface, and one round 19F Blake abdominal drain (Ethicon) was left in place. Operative time was 180 minutes and blood loss estimated in 40 ml with no blood transfusion. Hospital stay was 4 days. The patient did not have postoperative pancreatitis or pancreatic leakage, and the abdominal drain was removed on the tenth postoperative day. Final pathology confirmed the diagnosis of solid pseudopapillary neoplasm of pancreas with free surgical margins. The patient was well and asymptomatic 2 months after the procedure. Laparoscopic resection of uncinate process of the pancreas is safe and feasible and should be considered for patients suffering from pancreatic neoplasms.

Stimulation of 5-HT1A or 5-HT2A receptors in the ventrolateral periaqueductal gray causes anxiolytic-, but not panicolytic-like effect in rats

Evidences from studies using electrical or chemical stimulation of the midbrain periaqueductal gray (PAG) suggest that whereas the dorsal PAG is critical for the regulation of panic-related defensive behaviors, the ventrolateral PAG (vlPAG) modulates generalized anxiety-related responses. In the present study we evaluated whether the activation of 5-HT1A and 5-HT2A/2C receptors in the ventrolateral column of the periaqueductal gray (vlPAG) causes differential effects on an anxiety- and a panic-related defensive behavior, respectively, inhibitory avoidance and escape, in male Wistar rats submitted to the elevated T-maze. Our results showed that intra-vlPAG injection of the endogenous agonist serotonin, the 5-HT1A/7 agonist 8-OH-DPAT or 5-HT2A/2C agonist DOI impaired the acquisition of inhibitory avoidance, without interfering with escape performance. The same selective anxiolytic effect was also observed after local administration of the benzodiazepine receptor agonist midazolam. Moreover, as shown by the results of antagonism studies, 5-HT2A receptors are recruited for the anxiolysis caused by serotonin and DOI. while 5-HT1A receptors account for the effect of 8-OH-DPAT. In conclusion, our data show that the activation of 5-HT1A and 5-HT2A receptors in the vlPAG affects defensive responses related to generalized anxiety, but not panic disorder. (C) 2008 Elsevier B.V. All rights reserved.

Involvement of median raphe nucleus 5-HT1A receptors in the regulation of generalized anxiety-related defensive behaviours in rats

Changes in 5-HT1A receptor-mediated neurotransmission at the level of the median raphe nucleus (MRN) are reported to affect the expression of defensive responses that are associated with generalized anxiety disorder (e.g. inhibitory avoidance) but not with panic (e.g. escape). The objective of this study was to further explore the involvement of MRN 5-HT1A receptors in the regulation of generalized anxiety-related behaviours. Results of experiment 1 showed that intra-MRN injection of the 5-HT1A/7 receptor agonist 8-OH-DPAT (0.6 nmol) in male Wistar rats impaired the acquisition of inhibitory avoidance, without interfering with the performance of escape in the elevated T-maze test of anxiety. Pre-treatment with the 5-HT1A receptor antagonist WAY-100635 (0.18 nmol) fully blocked this anxiolytic-like effect. As revealed by experiment 2, intra-MRN injection of 8-OH-DPAT (0.6, 3 or 15 nmol) also caused anxiolytic effect in rats submitted to the light-dark transition test, another animal model that has been associated with generalized anxiety. In the same test, intra-MRN injection of WAY-100635 (0.18, 0.37 or 0.74 nmol) caused the opposite effect. Overall, the current findings support the view that MRN 5-HT neurons, through the regulation of 5-HT1A somatodendritic autoreceptors, are implicated in the regulation of generalized anxiety-associated behaviours. (C) 2008 Elsevier Ireland Ltd. All rights reserved.

5-HT1A receptors in the dorsal hippocampus mediate the anxiogenic effect induced by the stimulation of 5-HT neurons in the median raphe nucleus

We evaluated the involvement of dorsal hippocampus (DH) 5-HT1A receptors in the mediation of the behavioral effects caused by the pharmacological manipulation of 5-HT neurons in the median raphe nucleus (MRN). To this end, we used the rat elevated T-maze test of anxiety. The results showed that intra-DH injection of the 5-HT1A/7 agonist 8-OH-DPAT facilitated inhibitory avoidance, an anxiogenic effect, without affecting escape. Microinjection of the 5-HT1A antagonist WAY-100635 was ineffective. In the elevated T-maze, inhibitory avoidance and escape have been related to generalized anxiety and panic disorders, respectively. Intra-MRN administration of the excitatory aminoacid kainic acid, which non-selectively stimulates 5-HT neurons in this brain area facilitated inhibitory avoidance and impaired escape performance, but also affected locomotion. Intra-MRN injection of WAY-100635, which has a disinhibitory effect on the activity of 5-HT neurons in this midbrain area, only facilitated inhibitory avoidance. Preadministration of WAY-100635 into the DH blocked the behavioral effect of intra-MRN injection of WAY-100635, but not of kainic acid. These results indicate that DH 5-HT1A receptors mediate the anxiogenic effect induced by the selective stimulation of 5-HT neurons in the MRN. (c) 2007 Elsevier B.V. and ECNP. All rights reserved.

Cannabidiol inhibits the hyperphagia induced by cannabinoid-1 or serotonin-1A receptor agonists

Delta 9-THC is a component of Cannabis sativa that increases food intake in animals and humans, an effect prevented by selective CB1 receptor antagonists. Cannabidiol (CBD) is another constituent of this plant that promotes several opposite neuropharmacological effects compared to Delta 9-THC. CBD mechanisms of action are still not clear, but under specific experimental conditions it can antagonize the effects of cannabinoid agonists, block the reuptake of anandamide and act as an agonist of 5-HT1A receptors. Since both the cannabinoid and serotoninergic systems have been implicated in food intake control, the aim of the present work was to investigate the effects caused by CBD on hyperphagia induced by agonists of CB1 or 5-HT1A receptors. Fed or fasted Wistar rats received intraperitoneal (i.p.) injections of CBD (1, 10 and 20 mg/kg) and food intake was measured 30 min later for 1 h. Moreover, additional fed or fasted groups received, after pretreatment with CBD (20 mg/kg) or vehicle, i.p. administration of vehicle, a CBI receptor agonist WIN55,212-2 (2 mg/kg) or a 5-HT1A receptor agonist 8-OH-DPAT (1 mg/kg) and were submitted to the food intake test for 1 h. CBD by itself did not change food intake in fed or fasted rats. However, it prevented the hyperphagic effects induced by WIN55,212-2 or 8-OH-DPAT. These results show that CBD can interfere with food intake changes induced by a CB1 or 5-HT1A receptor agonist, suggesting that its role as a possible food intake regulator should be further investigate. (C) 2011 Elsevier Inc. All rights reserved.

Involvement of dorsal raphe nucleus and dorsal periaqueductal gray 5-HT receptors in the modulation of mouse defensive behaviors

Previous findings point to the involvement of the dorsal raphe nucleus (DRN) and dorsal periaqueductal gray (dPAG) serotonergic receptors in the mediation of defensive responses that are associated with specific subtypes of anxiety disorders. These studies have mostly been conducted with rats tested in the elevated T-maze, an experimental model of anxiety that was developed to allow the measurement, in the same animal, of two behaviors mentioned: inhibitory avoidance and one-way escape. Such behavioral responses have been respectively related to generalized anxiety disorder (GAD) and panic disorder (PD). In order to assess the generality of these findings, in the current study we investigated the effects of the injection of 5-HT-related drugs into the DRN and dPAG of another rodent species, mouse, on the mouse defense test battery (MDTB), a test of a range of defensive behaviors to an unconditioned threat, a predator. Male CD-1 mice were tested in the MDTB after intra-DRN administration of the 5-HT(1A) receptor antagonist WAY-100635 or after intra-dPAG injection of two serotonergic agonists, the 5-HT1A receptor agonist 8-OH-DPAT and the 5-HT(2A/2C) receptor agonist DOI. Intra-DRN injection of WAY-100635 did not change behavioral responses of mice confronted with a rat in the MDTB. In the dPAG, both 8-OH-DPAT and DOI consistently impaired mouse escape behavior assessed in the MDTB. Intra-dPAG infusion of 8-OH-DPAT also decreased measures of mouse risk assessment in the rat exposure test. In conclusion, the current findings are in partial agreement with previous results obtained with rats tested in the elevated T-maze. Although there is a high level of similarity between the behavioral effects obtained in rats (elevated T-maze) and mice (MDTB and RET) with the infusion of 5-HT agonists into the dPAG, the same is not true regarding the effects of blockade of DRN 5-HT(1A) receptors in these rodent species. These data suggest that there may be differences between mice and rats regarding the involvement of the DRN in the mediation of defensive behaviors. (C) 2010 Elsevier B.V. and ECNP. All rights reserved.

Facilitation of 5-HT(2A/2C)-mediated neurotransmission in the ventromedial hypothalamic nucleus decreases anxiety in the elevated T-maze

Previous evidence has shown that facilitation of GABA/benzodiazepine-mediated neurotransmission in the ventromedial hypothalamus (VMH) inhibits both escape and inhibitory avoidance responses generated in the elevated T-maze test of anxiety (ETM). These defensive behaviors have been associated with panic and generalized anxiety, respectively. Aside from GABA/benzodiazepine receptors, the VMH also contains a significant number of serotonin (5-HT) receptors, including 1A, 2A and 2C subtypes. The purpose of the present study was to investigate the effect of the activation of 5-HT(1A) and 5-HT(2A/2C) receptors in the VMH on defensive behavioral responses in rats submitted to the ETM. For that, male Wistar rats were treated intra-VMH with the 5-HT(1A) agonist 8-OH-DPAT, with the 5-HT(2A/2C) agonist DOI, with the 5-HT(2C) selective agonist MK-212, or with the 5-HT(2A/2C) antagonist ketanserin and 10 min after were submitted to the ETM. Results showed that both DOI and MK-212 significantly decreased avoidance measurements, an anxiolytic-like effect, without altering escape. 8-OH-DPAT and ketanserin were without effect, although the last drug attenuated the effects of DOI. None of the drugs altered locomotor activity in an open field. These results suggest that 5-HT(2A/2C) receptors of the VMH are involved in the regulation of inhibitory avoidance and might be of relevance to the physiopathology of generalized anxiety. (C) 2010 Elsevier B.V. All rights reserved.

Dorsal raphe nucleus regulation of a panic-like defensive behavior evoked by chemical stimulation of the rat dorsal periaqueductal gray matter

Electrical or chemical stimulation of the dorsal periaqueductal gray matter (DPAG) evokes escape, a defensive behavior that has been related to panic attacks. Injection of 5-HT(1A) or 5-HT(2A) receptor agonists into this midbrain area inhibits this response. It has been proposed that the impairment of 5-HT mechanisms controlling escape at the level of the DPAG may underlie the susceptibility to panic attacks that characterizes the panic disorder. In this study we evaluated the effects of the pharmacological manipulation of the dorsal raphe nucleus (DRN), which are the main source of 5-HT input to the DPAG, on the escape response evoked in rats by the intra-DPAG injection of the nitric oxide donor SIN-1. The results showed that DRN administration of the 5-HT(1A) receptor agonist 8-OH-DPAT which inhibits the activity of 5-HT neurons favored the expression of escape induced by SIN-1. Intra-DRN injection of the excitatory amino acid kainic acid or the 5-HT(1A) receptor antagonist WAY-100635 did not change escape expression. However, both compounds fully blocked the escape reaction generated by intra-DPAG injection of the excitatory amino acid D,L-homocysteic acid (DLH). Overall, the results indicate that 5-HT neurons in the DRN exert a bidirectional control upon escape behavior generated by the DPAG. Taking into account the effect of WAY-100635 on DLH-induced escape, they also strengthen the view that DRN 5-HT(1A) autoreceptors are under tonic inhibitory influence by 5-HT. (C) 2010 Elsevier B.V. All rights reserved.

Cannabidiol injected into the bed nucleus of the stria terminalis modulates baroreflex activity through 5-HT(1A) receptors

Cannabidiol (CBD) is a non-psychotomimetic constituent of the Cannabis sativa plant that inhibits behavioral and cardiovascular responses to aversive situations. facilitating 5-HT(1A)-mediated neurotransmission. Previous results from our group suggest that the bed nucleus of the stria terminalis (BNST) may be involved in CBD`s anti-aversive effects. To investigate whether the cardiovascular effects of the CBD could involve a direct drug effect on the BNST, we evaluated the effects of CBD microinjection into this structure on baroreflex activity. We also verified whether these effects were mediated by the activation of 5-HT(1A) receptors. Bilateral microinjection of CBD (60 nmol/100 nL) into the BNST increased the bradycardiac response to arterial pressure increases. However, no changes were observed in tachycardiac responses evoked by arterial pressure decreases. Pretreatment of the BNST with the selective 5-HT(1A) receptor antagonist WAY100635 (0.37 nmol/100 nL) prevented CBD effects on the baroreflex activity. Moreover, microinjection of the 5-HT(1A) receptor agonist 8-OH-DPAT (4 nmol/100 nL) caused effects that were similar to those observed after the microinjection of CBD, which were also blocked by pretreatment with WAY100635. In conclusion, the present studies show that the microinjection of CBD into the BNST has a facilitatory influence on the baroreflex response to blood pressure increases, acting through the activation of 5-HT(1A) receptors. (C) 2010 Elsevier Ltd. All rights reserved.

Role of dorsal raphe nucleus 5-HT(1A) and 5-HT(2) receptors in tonic immobility modulation in guinea pigs

Tonic immobility (TI) is an innate defensive behavior characterized by a state of physical inactivity and diminished responsiveness to environmental stimuli. Behavioral adaptations to changes in the external and internal milieu involve complex neuronal network activity and a large number of chemical neurotransmitters. The TI response is thought to be influenced by serotonin (5-HT) activity in the central nervous system (CNS) of vertebrates, but the neuronal groups involved in the mechanisms underlying this behavior are poorly understood. Owing to its extensive afferents and efferents, the dorsal raphe nucleus (DRN) has been implicated in a great variety of physiological and behavioral functions. in the current study, we investigated the influence of serotonergic 5-HT(1A) and 5-HT(2) receptor activity within the DRN on the modulation of TI behavior in the guinea pig. Microinjection of a 5-HT(1A) receptor agonist (8-OH-DPAT, 0.01 and 0.1 mu g) decreased TI behavior, an effect blocked by pretreatment with WAY-100635 (0.033 mu g), a 5-HT(1A) antagonist. In contrast, activation of 5-HT(2) receptors within the DRN (alpha-methyl-5-HT, 0.5 mu g) increased the TI duration, and this effect could be reversed by pretreatment with an ineffective dose (0.01 mu g) of ketanserine. Since the 5-HT(1A) and 5-HT(2) agonists decreased and increased, respectively, the duration of TI, different serotonin receptor subtypes may play distinct roles in the modulation of TI in the guinea pig. (C) 2009 Elsevier B.V. All rights reserved.

High prevalence of polycystic ovary syndrome in women born small for gestational age

There is evidence that intrauterine growth restriction, resulting in newborn girls that are small for gestational age (SGA), may be related to the onset of polycystic ovary syndrome (PCOS). Thus, we studied whether women born SGA have a higher prevalence of PCOS than women born appropriate for gestational age (AGA). This was a prospective birth cohort study of 384 women born at term between June 1, 1978, and May 31, 1979, in Ribeirao Preto, Brazil. After exclusion, 165 women effectively participated in this study, of whom 43 were SGA and 122 were AGA. The prevalence of PCOS was analysed. At a mean age of 29 years, the women agreed to follow the study protocol, which included: anamnesis, physical examination, serum tests [follicle stimulating hormone, luteinizing hormone, total and free testosterone, dehydroepiandrostenedione sulphate, 17-OH-progesterone, fasting insulin, sex steroid-binding globulin (SHBG) and fasting glucose] and pelvic ultrasound. Data regarding gestational age, birthweight, age at menarche and maternal data were obtained from the files of the cohort. The adjusted relative risk (RR) values of the SGA, insulin resistance, body mass index, maternal smoking and parity variables were analysed using Poisson regression with robust adjustment of variance for the prediction of PCOS. The prevalence of PCOS was higher in the SGA group than in the AGA group [adjusted RR = 2.44, 95% CI (1.39-4.28)]. Hyperandrogenism was more prevalent in the SGA women than in the AGA women (P = 0.011). Circulating SHBG was lower in the SGA women than in the AGA women (P = 0.041), but fasting insulinemia was similar in both groups. The prevalence of PCOS in SGA women was twice as high as in AGA women in our study population.

Comparison of different irrigants on calcium hydroxide medication removal: microscopic cleanliness evaluation

Calcium hydroxide dressing residuals can compromise endodontic sealing. This study evaluated the cleaning efficacy of different endodontic irrigants in removing calcium hydroxide by SEM image analysis. Fifty-four single-rooted mandibular premolars were instrumented to a master apical file #60 and dressed with calcium hydroxide. After 36 hours, the teeth were reopened and Ca(OH)(2) medication was removed by 5 different experimental groups: 0.5% NaOCl (G1), EDTA-C (G2), citric acid (G3), EDTA-T (G4), and re-instrumentation with MAF using NaOCl and lubrificant, followed by EDTA-T (G5). The roots were split in the buccal-lingual direction and prepared for SEM analysis in cervical, middle, and apical thirds (9, 6, and 3 mm from the apex). Five blinded examiners evaluated the wall cleanliness using a scale from 1 to 5. Statistical analysis was performed using Kruskal-Wallis at 5% level of significance. Group G5 had the best results in all thirds, with significant statistical differences compared to all other groups in the middle and coronal third, and to G1 in the apical third. On the other hand, G1, only flushed with NaOCl, had the worst results, with statistical differences in all thirds compared to the other groups. The best cleanliness was achieved by G4 and G5 groups. The recapitulation of MAF in combination with irrigants improved the removal of calcium hydroxide medication better than an irrigant flush alone. (Oral Surg Oral Med Oral Pathol Oral Radiol Endod 2009; 107: 580-584)