Higher than 20-fold levels of expression (p < 0.01) was sustained in LD 10–87 VERO cells at p250 and

in A4497 (p > 200) VERO cells, which are tumorigenic in both newborn and adult nude mice [10]. Three of the six miRNAs (miR-376a, miR-543 and miR-299-3p) were overexpressed more than 4-10 fold compared with pAGMK control cells and the LD 10–87 VERO cell passages before the expression of the tumorigenic phenotype was detected at p194 ( Table 1 and Fig. 1A). These results suggest that these miRNA-based biomarkers may be capable of predicting the pre-tumor stages of neoplastic development in VERO cells. To verify the accuracy and specificity of these results, we assessed the six miRNAs in HD VERO cells that were passaged independently at higher, confluent densities. The trend in the alteration of miRNA expression was generally similar S3I-201 supplier between the LD 10–87 VERO cell lines and the HD 10–87 VERO cell lines. When compared with the pAGMK controls, five of these six miRNAs were over-expressed by greater than 4-fold in the tumorigenic BLZ945 HD 10–87 VERO cells at p183, and all six were

over-expressed by 6- to >50-fold at p250 ( Table 2). To further evaluate the ability of individual miRNA to reflect the expression of the tumorigenic phenotype in VERO cells, we examined three miRNA data sets (miR-376a, miR-654-3P, and miR-543) from experiments shown in Table 1 and Table 2. The expression pattern of each of these miRNA followed the progression of neoplastic development and peaked at p194 (Fig. Isotretinoin 4A) where the ability of LD 10–87 VERO cells

to form tumors was detected (Fig. 1). In HD 10–87 VERO cells, the same association between elevated expression levels of the same miRNAs and tumorigenicity was observed at p183; however, the expression levels in cells at p250 increased by an additional 4-fold compared with cells at p183 (Fig. 4B). Together, regardless of how the tumor-forming cells were established, whether by passaging at low density or high density, the individual miRNA expression pattern correlated with the detection of the tumorigenic phenotype. Therefore, these six miRNAs appeared to be biomarkers for this property of VERO cells. Managing the threats posed by emerging and re-emerging infectious diseases, such as pandemic influenza, call for the rapid production of large, possibly unprecedented, amounts of vaccines to immunize populations worldwide [31], [32] and [33]. Current production methods may be insufficient to meet these demands in the short period required to manage pandemics successfully [33]. Cell-culture technology based on immortalized cell substrates provides a possible method for increasing the efficiency of vaccine manufacture and improving vaccine efficacy [1], [3], [6], [8], [31], [32], [34], [35], [36] and [37]. Regulatory agencies have recommended that the tumorigenic potential of immortalized cell substrates proposed for human vaccine production be evaluated (21 Code of Federal Regulations 610.18).

These peaks presumably represent multimers of VP1, 2 and/or 3. Two peaks

at 11.3 and 14.0 kDa are present in purified FMDV O1 Manisa ( Fig. 2c) that do not correspond to predicted FMDV structural proteins. FMDV O1 Manisa that was not purified by ultracentrifugation (Fig. 2d) shows many additional peaks that presumably represent substances of non-viral origin that are present in the FMDV antigen preparations. A peak at about 67 kDa could represent bovine serum albumin that originates from the foetal bovine serum used as a medium supplement during growth of BHK-21 cells. A repetitive pattern of peaks differing in molecular mass by 44 Da is present in the range of 6–7 kDa (Fig. 2e). This most likely represents PEG6000 molecules Pomalidomide chemical structure that were used in downstream processing of the antigen since the repeating unit corresponds exactly to the expected differences in molecular

mass of PEG molecules. We next analysed FMDV O1 Manisa that was immunocaptured using three FMDV binding VHHs (M3, M8 and M23) that recognize independent antigenic sites. As a control we used the K609 VHH that does not bind FMDV. These VHHs were covalently coupled to RS100 arrays that were subsequently incubated with FMDV O1 Manisa that was not purified by ultracentrifugation. The spectral peaks previously identified as VP1, VP2 and VP1–VP2 dimers were also observed using the three FMDV binding VHHs (Fig. 3b–d) but not using the control VHH (Fig. 3a), confirming their identification. However, the spectral AZD4547 research buy peak at about 9.0 kDa previously identified below as VP4 was only observed using M8 or M23 but not using M3 (nor the control VHH). Two spectral peaks of low height at 6.6 and 7.5 kDa were also specifically observed using the three FMDV binding VHHs (Fig. 3b–d), suggesting their viral origin. However, these peaks do not match a

predicted FMDV protein. Several spectral peaks occur in the range of 4–14 kDa using the control VHH, as well as the three FMDV binding VHHs, indicating that these peaks do not represent FMDV proteins. This includes the peaks at 11.3 and 14.0 kDa identified in the previous section as being of non-viral origin. A closer view of VP1 shows that it actually consists of two peaks differing in mass by 0.2 kDa (Fig. 3e). Similarly, VP4 consists of 8 peaks differing by 14–17 Da (Fig. 3e). Such VP1 and VP4 heterogeneity was consistently found in all spectra (results not shown). We next analysed trypsin-treated FMDV O1 Manisa by immunocapture with M8 (Fig. 3f) or M23 VHHs (Fig. 3g and h). The spectra obtained with both capturing VHHs were essentially identical. Therefore, we only compared the spectral peaks observed with M23 to predicted trypsin cleavage fragments (Table 1). Trypsin treatment abolishes both VP1 peaks at about 23.4 kDa (Fig. 3h) and appears to reduce the height of the VP2 peak at 24.5 kDa. Due to the absence of VP1 a shoulder at 24.0 kDa on the VP2 peak at 24.5 kDa is now visible (Fig. 3h). This could represent VP3.

25:1 and 0.5:1, and mixed

in a mechanical stirrer. The prepared mixture was then degassed under vacuum for 10 min. The resulting dispersion was dropped through a 26G syringe needle into 1%w/v of calcium chloride solution containing 10% v/v glacial acetic acid. The solution containing the suspended beads was stirred with a magnetic stirrer for 10 min, to improve the mechanical strength of the beads and it was allowed to complete the reaction to produce gas inside the beads. The formulated beads were separated by filtration, washed with ethanol and distilled water, and freeze-dried.17 Angle of repose method was employed to assess the flowability. Beads were allowed to fall freely through the funnel fixed at 2 cm above the horizontal Metabolism inhibitor flat surface until the apex of conical pile just touched the tip of the funnel. The angle of repose (θ) was determined by formula. θ = tan−1 (h/r) where, h – cone height of beads, r – radius of circular base formed by the beads on the ground. 18 and 19 The average diameter of twenty dry beads was determined randomly

using a caliper in triplicate. 20 Accurately selleck inhibitor weighed quantities of approximately 300 mg of beads were placed in 25 ml of 0.1 N HCl. The solution was centrifuged using the centrifuge at 4200 rpm for 30 min; the supernatant layer of the liquid was assayed by UV-spectroscopy at 266 nm. The encapsulation efficiency was determined by the following equation.17 and 21 Encapsulationefficiency=%Drugofformulation×TotalweightofthedriedbeadsAmountofdrugloaded−Druglossinthegelationmedia Drug content was performed to check dose uniformity in the formulation. Randomly ten tablets were weighed and powdered. A quantity equivalent to 300 mg of zidovudine was added in to a 100 ml

volumetric flask and dissolved in 0.1 N HCL, shaken for 10 min and made up the volume up to the mark and filtered. After suitable dilutions the drug content was determined by UV spectrophotometer at 266 nm against blank (Using UV–VIS Spectrophotometer, Shimadzu 1700).21 Swelling studies for beads was performed in dissolution media (0.1 N HCl). The swelling index was calculated using the formula: swelling index = (Wg − Wo)/Wo × 100, where Wo was the initial weight of beads and Wg was the weight of beads in the swelling medium. 17 Fifty beads were placed in 500 ml of 0.1 N HCl media. most The floating properties of beads were evaluated in a dissolution vessel [USP Type II dissolution tester]. Paddle rotation speeds of 0 and 100 revolutions per minute were tested. Temperature was maintained at 37 ± 0.5 °C. The percentage of floating samples was measured by visual observation.17 The in-vitro dissolution studies were carried out using USP XXIV Dissolution Apparatus No.2 (type) at 50 rpm. The dissolution medium consisted of 0.1 N HCL for 12 h (900 ml) maintained at 37 ± 0.50. The release studies were conducted in triplet.

Present results are obviously similar to the results explained above which shows that bilirubin level increases due to malarial infection. Present study also shows that hypoglycemia is more common in sever malaria patients. 68% of patients were found with hypoglycemia. We detected hypoglycemia in nearly 11% of the patients with sever falciparum malaria. Shah et al 11 reported hypoglycemia in 2 out of 20 cases (10%) of severe falciparum malaria from Karachi (Pakistan). The occurrence of malaria in adults is due to mal-absorption of glucose from intestine. Thai adults with sever malaria had

greatly reduced absorption capacity for sugar transport both actively and passively. 12 Most of our patients have hypoglycemia before quinine administration. selleckchem This suggests that other causes may also be responsible for hypoglycemia. 13 All authors have none to declare. We are very thankful to Professor Dr. Salman Akbar Malik Chairman

Department of Biochemistry, QAU Islamabad, Pakistan for his valuable suggestions. “
“Multi-particulate (MP) modified release drug delivery systems have several performance advantages over single unit dosage forms. MP formulations generally have a more reliable in vivo dissolution performance, resulting in more uniform bioavailability and clinical effect. 1 Pelletization is an agglomeration process that converts fine powders or granules of bulk drugs and excipients into small, free flowing, spherical or semi spherical Thymidine kinase units, referred to as pellets. 2 Pellets offer a high degree of flexibility and can be divided into desired dose strengths without formulation or process changes. 3 Pellets are in check details a size range between 0.5 and 1.5 mm and are produced primarily for the purpose of oral controlled release dosage forms having gastro resistant or sustained release properties or the capability of site-specific drug delivery. 4 The pelletized products can improve the safety and efficacy of the active agent, showing a number of advantages over the single unit dosage

system. 5 Extended release formulations are designed to allow at least twofold reduction in dosing frequency or significant increase in patient compliance or therapeutic performance when compared to a conventional immediate release dosage form. 6 Sustained release pharmaceutical pellet is one of the most popular approaches among the various types of extended release dosage forms as it offers several manufacturing and biopharmaceutical advantages. 7 Pellets are also less affected by gastric emptying. 8 After administration, the coated pellets spread uniformly throughout the gastrointestinal tract resulting in a consistent drug release with reduced risk of local irritation and dose dumping of the drug can be avoided. NSAIDs are a group of drugs of diverse chemical composition and different therapeutic potentials.9 Most NSAIDs are weak acids, with a pKa values in the range of 3.0–5.0 and contain hydrophilic groups and lipophilic ones.

009) and CD8+ (P = 0.02) cells after booster vaccination than after prime vaccination. The concentration of IFN-γ, a cytokine which is one of the main indicators of the formation of Th1 and a cytotoxic cellular immune response, was also determined. As shown in Fig. 2, significant (P < 0.0001) accumulation of IFN-γ after stimulation with Brucella L7/L12 and Omp16 proteins was observed in the samples from the animals vaccinated with the viral constructs vaccine formulation only, as well as its combination with Montanide Gel01, or the B. abortus S19 vaccine

as compared to the control samples (without stimulation). Significant accumulation of IFN-γ was not observed in the samples from the group of animals vaccinated with Flu-L7/L12-Omp16-chitosan. click here It should be noted that the highest levels of IFN-γ accumulation after stimulation with Brucella antigens was observed in the samples from animals TGFbeta inhibitor vaccinated with Flu-L7/L12-Omp16-MontanideGel01; the IFN-γ levels for this group were significantly higher (P = 0.01 or P = 0.0003) than the other experimental groups (28 days after the prime vaccination) and even slightly

superior (P = 0.12 or P = 0.22) to that of the positive control group vaccinated with B. abortus S19. Booster immunization did not significantly (P = 0.09 to P = 0.99) increase the concentration of IFN-γ in the samples from the animals in the experimental groups. As shown in Fig. 3, the highest level of protection was achieved with Flu-L7/L12-Omp16-MontanideGel01; the effectiveness of vaccination and index of infection for this group were 100% and 0, respectively. Good why results were also obtained with Flu-L7/L12-Omp16, which had a similar effectiveness of vaccination (60%), index of infection and number of cultured Brucella (P = 0.99 or P > 0.99) to the group vaccinated with the B. abortus S19 vaccine. The lowest effectiveness of

vaccination (40%) was observed for Flu-L7/L12-Omp16-chitosan. Despite this, the number of Brucella cultured from the lymph nodes and index of infection in this group was significantly lower (P = 0.02 or P = 0.007) than that of the negative control group (PBS), and not significantly different to the other experimental groups (from P = 0.29 to P = 0.98) or the positive control group (P = 0.62 or P = 0.92) groups. After challenge with B. abortus 544, the body temperature of the animals in the experimental groups remained within the normal range (37.5–39.5 °C) during the entire period of observation (30 days), while the body temperature of the animals in the negative and positive control groups increased to 40.0 °C on days 1–3 and day 2 post-challenge, respectively. The present work is a continuation of a series of studies aimed at developing an effective vaccine against B. abortus. As previously stated, a number of candidate vaccines against B. abortus have been prepared to date, most of which are DNA vaccines and live recombinant vaccines.

Dans les « Standards Options Recommandations » de 2003 [2], 20 à 50 % des 9007 patients analysés

(sur 36 études) étaient douloureux au moment du diagnostic de cancer et la prévalence de la douleur augmentait au cours de l’évolution de la maladie avec 55 à 95 % de patients douloureux. Dans l’étude de Breivik et al., regroupant 5084 patients cancéreux adultes contactés entre 2006 et 2007 dans onze pays européens (dont 642 France) et en Israël, la prévalence globale de la douleur était de 84 % et de 75 % en France [3]. Parmi ces patients, 56 % avaient une douleur modérée à sévère et pour 573 patients this website tirés au sort, 41 % recevaient un traitement opioïde fort, 69 % mentionnaient un retentissement de la douleur sur la qualité de vie et 50 % avaient Crenolanib chemical structure le sentiment que la qualité de vie n’était pas une priorité pour les professionnels de santé. La prévalence de la douleur était particulièrement élevée (plus de 85 %) pour les patients qui avaient un cancer du pancréas, des os, du cerveau, de la tête et du cou et les patients porteurs de lymphome. Une enquête nationale, réalisée en

2010, sous l’égide de l’INCa (Institut national du cancer) en collaboration avec l’Institut BVA, a été menée auprès de 1507 patients atteints de cancer traités en ambulatoire. L’objectif principal était de préciser l’état des lieux concernant les modalités de prise en charge de la douleur du cancer en France [4]. Ce document s’inscrit

dans la mise en œuvre du Plan cancer 2009–2013, à savoir « renforcer la qualité des prises en charge pour tous les malades atteints de cancer », et plus précisément la mesure 19.1 du plan cancer : « généraliser l’accès aux mesures transversales lancées par le Plan cancer précédent, améliorant la qualité de toute prise en charge en cancérologie ». Cette enquête visait à décrire la douleur des patients en phase de traitement Cell press curatif, en situation de cancer avancé et également à distance des traitements (en phase de surveillance ou de rémission), à individualiser la douleur neuropathique, les crises douloureuses et leurs prises en charge. Sur les 1507 patients interrogés, 28 % étaient en phase de traitement curatif, 53 % en situation de cancer avancé, 18 % en phase de surveillance ou de rémission avec, pour la majorité d’entre eux, un recul de plus d’un an par rapport à la fin de la chimiothérapie. La prévalence déclarée de la douleur dans cette enquête est identique à celle des données de la littérature, la douleur étant présente chez 53 % des patients interrogés. Une douleur chronique (présente depuis plus de trois mois) est rapportée par 30 % des patients douloureux en situation de cancer avancé, mais aussi par 25 % des patients douloureux à distance de tout traitement ou bien en rémission.

Group G-D was inoculated with 107 C6/36 derived RVFV. Group G-E was inoculated with 105 PFU of Vero E6 RVFV stock, and re-inoculated IV with the same inoculum at 1 dpi. Group G-F was inoculated with 105 PFU of C6/36 derived RVFV, and re-inoculated

IV with the same inoculum at 1 dpi. Group G-G was inoculated with 107 C6/36 derived RVFV and re-inoculated SC with the same inoculum at 1 dpi. All goats were kept for four weeks following the inoculation to monitor an antibody development. Serum samples collected at 0, 4, 5, 6, 7, 14, 21 and 28–30 dpi were analyzed for presence of neutralizing antibodies. Differences in susceptibility to RVFV infections were observed between sheep and goats, and also between breeds of sheep. In the first study,

conducted LY2109761 cost in Suffolk-cross sheep, all animals developed viremia at 3 dpi, both by virus isolation and RNA detection when inoculated with 105 PFU of virus produced in Vero cells. However, when the Rideau Arcott cross lambs were inoculated via the same route and the same inoculum, only three out of four animals had detectable RVFV RNA in their blood and only two developed viremia (Fig. 1). Subsequently different inoculation approaches were tested to obtain a more reliable viremia model. Genomic sequences of the inocula were verified prior to the start of the animal inoculations. Concurrently with the infection experiments, characterization on protein level of RVFV Verteporfin chemical structure generated in Vero E6 cells or the C6/36 was taking place. There was no difference in genome of RVFV generated in Vero E6 cells

compared to virus generated in C6/36 cells, including the stock viruses used in experimental PDK4 inoculations, and the sequences corresponded with sequences published for RVFV ZH501 in Gen Bank. Both viruses had functional NSm and NSs coding genes, as immunoblots of infected cell lysates indicated that all proteins from the M and S segments were expressed. The viruses however differed in protein composition of virions, with the mosquito-cell generated RVFV having an additional large glycoprotein (78 kDa) incorporated into virions [23]. Subcutanous inoculation was used in all primary inoculation. Two doses (105 or 107 PFU/animal) and two different inocula (prepared either in Vero E6 or in C6/36 cells) were tested. The titer of inoculum was confirmed by back-titration at the time of inoculation, and stayed within 0.5 log10 difference from the targeted dose. In specific groups, attempts were made to increase the viremia by re-inoculation, either by the subcutaneous or by the intravenous route at 1 dpi. A summary of the experimental groups is presented in Table 1. Using the same mode of inoculation as for the Suffolk breed (group S-A), the 105 PFU dose of Vero E6 produced RVFV in Rideau Arcott cross lambs (group S-B) lead to development of viremia only in three out of four animals at 2 dpi.

Page 5327, Table 2 • Row “Geometric mean titer + S.D. 581 + 3380, 474 + 1830, 4076 + 7058”, at the month 2, month 6 and month 7 columns. “
“Neisseria meningitidis is a gram-negative diplococcus that causes severe invasive disease including septicemia and meningitis [1]. Most invasive disease is the result of infection with one of five groups (A, B, C, Y, W-135) as characterized by their capsular polysaccharide [2]. Epidemic group A disease occurs in sub-Saharan Africa, the Middle East and in some areas of Asia [3], [4] and [5]. Endemic group B and C disease predominates in Europe and North America; an increase in group Y disease has been reported over HKI-272 manufacturer the last 20 years in the United States [6]. Outbreaks of W-135 disease have been reported

Histone Methyltransferase inhibitor in the Middle East and Africa [4] and [7]. Meningococcal disease is seen in all age groups including children 2–10 years of age; in the US, groups A, C, Y and W-135 account for approximately 60% of meningococcal disease [8]. Using similar conjugation technology that led to the development of effective vaccines against Haemophilus influenzae type b and pneumococcal diseases in infants and young children [9] and [10], group C meningococcal conjugate vaccines (MenC) were

developed that led to dramatic decreases in invasive disease caused by N. meningitidis group C in European countries and Australia where universal immunization programs were implemented [11], [12], [13] and [14]. By chemically conjugating capsular polysaccharide to a protein carrier, the polysaccharide antigen is converted from a T-cell independent antigen to a T-cell dependent antigen with the resultant induction in immune memory in all ages after immunization and improved immunogenicity in infants [15], [16] and [17]. A quadrivalent meningococcal conjugate vaccine was developed in an attempt to improve upon the quadrivalent meningococcal polysaccharide vaccine that has been available for decades. Menactra® (MCV4; Sanofi Pasteur, Swiftwater, PA) was licensed for use in the United States January

17, 2005, for individuals 11–55 years of age and October 19, 2007, for children 2–10 years of age, and is recommended for universal use as a preadolescent dose [18] and for children 2–10 years of age with increased risk of meninogococcal infection [19] and [20]. Menveo® (MenACWY-CRM; Novartis Vaccines and Diagnostics, Cambridge, all MA), a quadrivalent meningococcal conjugate vaccine, was recently licensed in the United States February 19, 2010, for individuals 11–55 years of age and in Canada on May 21, 2010 for individuals 11 years and older; further studies were undertaken to support its use in infants [21], [22] and [23] and younger children [24]. The purpose of this study was to compare the safety and immunogenicity of MenACWY-CRM to the licensed MCV4 vaccine in children 2–10 years of age. The investigational quadrivalent meningococcal conjugate vaccine (MenACWY-CRM; Menveo®, Novartis Vaccines and Diagnostics, Cambridge, MA) contained (per 0.

This study also documents the early incidence of rotavirus disease in India. The percentage of children with dehydrating gastroenteritis who were less than six months of age was as high as 12%. The youngest case recorded was one month old at the time of

hospitalization. www.selleckchem.com/products/SNS-032.html An earlier study from central India showed that rotavirus disease was more common during cooler months, with seasonal peaks matching the lowest temperatures [7]. In this study, a distinct winter peak was seen in the months of December to February during the total 16 months of surveillance across 12 sites in India, especially in northern India which has a distinct winter season from November to February. Interestingly, the sites

in southern India did not demonstrate this trend as the area experiences the least annual variation in temperature of the four regions. The worldwide emergence of the G12 strain in 2005 and its increasing incidence during the past two years parallels the emergence and subsequent spread of G9 strains that occurred approximately a decade ago. In the mid-1990s, G9P[6] http://www.selleckchem.com/products/BI-2536.html and G9P[8] strains were reported in India, Japan, the United Kingdom, and the United States. Subsequently, G9P[8] spread globally, and it currently accounts for 4.1% of all rotavirus infections [8]. In our study, a higher percentage of G12 (17.74%) was observed especially in the Eastern part of India as compared to the rest of India. Various studies have found G12 strains in association with multiple VP4 click here types, namely P[4], P[6], P[8], and P[9], suggesting re-assortment among commonly circulating strains [9] and [10]. The increased reporting of infection with G12 strains may be associated with re-assortment, resulting

in generation of a strain that is better adapted to replication in humans, similar to the events that preceded the spread of G9 strains in the past decade. The emergence of G12 strains highlights the need for a surveillance system to respond rapidly to changes in circulating virus and to ensure that vaccines remain effective against emerging strains. Reported G12 cases from our study provided further evidence of the notion that G12 strains should no longer be considered as unusual or rare strains but that they exhibit a capacity to spread among children just like human rotavirus strains of other commonly seen G types. In addition to the challenges posed by the emergence of new strains in the population under surveillance, we found high levels of circulation of unusual recombinant strains, such as G1P[4], G1P[6], G2P[6], G2P[8], G9P[4], and G9P[6] in different parts of the country. This indicates that there may be both regional and temporal variations in rotavirus strain predominance, which will be important to consider when assessing the impact of vaccination on rotavirus strains.

Cognitive dysfunctions are directly correlated with Aβ oligomers in Tg2576 mice, which start at around 6 months old and are stable until 14 months old [29]. Thus, we first evaluated cognitive Quisinostat functions in both non-tg (n = 18) and Tg2576 mice (n = 24) at the age of 12 months. After the behavioral test, mice were divided into two groups to be treated with rSeV-LacZ or rSeV-Aβ. There is no difference between the two groups in behavioral scores at the age of 12 months. To evaluate the effect of vaccine treatment, each group (rSeV-LacZ-treated non-tg mice, n = 9; rSeV-Aβ-treated non-tg mice, n = 9; rSeV-LacZ-treated

Tg2576 mice, n = 10; rSeV-Aβ-treated Tg2576 mice, n = 14) was subjected to behavioral tests at the age of 15 months. All tests were done according to the methods described previously [30]. 24 h after 10 min-training session following 3 day-habituation, each mouse was placed back into the same box in which one of the familiar objects used during training was replaced with a novel one. The animals

were then allowed to 3-MA in vitro explore freely for 10 min and the time spent exploring each object was recorded. The exploratory preference (%), a ratio of the amount of time spent exploring any one of the two objects (training session) or the novel object (retention session) over the total time spent exploring both objects was used to measure cognitive function. Each mouse was placed at the center of the apparatus and allowed

to move freely through the maze during an 8-min session, and the series of arm entries was recorded visually. Alternation was defined as successive entry into the three arms on overlapping triplet sets. The % alternation was calculated as the ratio of actual alternations to the possible alternations (defined as the number of arm entries minus two) multiplied by 100. The Morris water maze test was conducted in a circular pool (1.2 m in diameter) with a hidden platform (7 cm in diameter) filled with water at a temperature of 22 ± 1 °C. The mice were given two trials 3-mercaptopyruvate sulfurtransferase (one block) for 10 consecutive days during which the platform was left in the same position. The time and distance taken to reach to the escape platform (escape latency and distance moved) was determined in each trial by using the Etho Vision system (Brainscience Co. Ltd., Osaka, Japan). Three hours after the last training trial, the platform was removed, and mice were allowed for 60 s to search the removed platform. For measuring basal levels of freezing response (preconditioning phase), mice were individually placed in a neutral cage for 1 min, and then in the conditioning cage for 2 min. For conditioning, mice were placed in the cage, and an 80 dB tone was delivered for 15 s. During the last 5 s of the tone stimulus, a foot shock of 0.