Attachment and Survival of Salmonella enterica and Listeria monocytogenes on Tomatoes (Solanum lycopersicum) as Affected by Relative Humidity, Temperature, and Storage Time.

ABSTRACT: Tomatoes (Solanum lycopersicum) are one of the most commonly consumed fruits worldwide. The fruit can become contaminated with Salmonella and Listeria monocytogenes at various stages of the production and supply chain, and these pathogens may survive under various storage conditions. The effects of relative humidity, temperature, and duration of storage on the attachment and survival of both pathogens on the surface of tomatoes were investigated. Fresh whole Roma tomatoes were inoculated with a cocktail of Salmonella or L. monocytogenes strains and stored at 5, 12, 25, 30, or 35°C for up to 10 days. Every day during storage, relative humidity and temperature were measured and tomatoes were removed to enumerate pathogens cells that were loosely attached (LA; cells were detached from the tomato surface by rinsing) and strongly attached (SA; sonication was required to detach cells from the tomato surface). The attachment strength (SR) was calculated to express the proportion of surviving SA cells on the tomato surface. The initial levels of Salmonella and L. monocytogenes on the tomato surface after inoculation were 6.6 and 6.5 log CFU per tomato for LA cells and 5.1 and 5.6 log CFU per tomato for SA cells, respectively. For both pathogens, the LA levels were higher (P < 0.05) than the SA levels. The LA and SA levels differed significantly as a function of temperature, relative humidity, and duration of storage. The SR for Salmonella was affected by storage time but not temperature, whereas the SR for L. monocytogenes was affected by storage time and temperature and relative humidity (P < 0.05). An understanding of the attachment and survival of Salmonella and L. monocytogenes on tomatoes stored under various temperature conditions may be useful for preventing or reducing the establishment of pathogens and for designing improved decontamination methods.

Effect of Different Wheat-Based Diets and Corn Meal Addition on Development Parameters of Ephestia kuehniella (Zeller, 1879) (Lepidoptera: Pyralidae).

The expansion of Integrated Pest Management (IPM), including biological control, has had several positive consequences for the agricultural environment and participants in the production chain. To enable successful operation and applications of biological control, production of insects used for rearing natural enemies (parasitoids and predators) must be optimized to reduce time and costs and improve production both qualitatively and quantitatively. The present study evaluated the effect of wheat varieties, the main component of artificial diets for Ephestia kuehniella, on the reproductive performance and biological parameters of this flour moth, which is used for mass production of Trichogramma spp. (Hymenoptera: Trichogrammatidae) and other parasitoids and predators. Four varieties of wheat were compared: BRS 327, BRS Marcante, BRS Parrudo, and KBR, with and without the addition of corn E. kuehniella reared on 97% BRS 327 wheat flour + 3% nutritional yeast had the best biological parameters and substitution of corn for about half of the wheat increased the number of eggs per female.

Measurements meet perceptions: rheology-texture-sensory relations when using green, bio-derived emollients in cosmetic emulsions.

OBJECTIVE: Product aesthetics and sensory performance can strongly influence a cosmetic product's acceptance by consumers. However, classic sensory analysis is time-consuming, expensive and does not provide information on the target group's preference. In the previous phase of this project, we had untrained consumers evaluate six cosmetic emulsions based on their aesthetics using a check-all-that-apply (CATA) survey. In this project, our goals were to quantitatively characterize the rheology and textural properties of the six cosmetic emulsions containing green, bio-derived emollients and identify statistical relationships between the consumers' description of products and the instrumental measurements. METHODS: Six emulsions were prepared-three with olive oil and three with heptyl undecylenate as an emollient. Four sensory-like attributes, namely firmness, work of shear, stickiness and adhesiveness, were tested using a texture analyser. Rheological characterization included continuous flow testing and oscillatory measurements. Droplet size and stability were also evaluated. Statistical relationships were quantified between measurements in this study and sensory survey results published previously. RESULTS: The textural and rheological results indicated that the emulsions were different-as designed. The texture and rheology measurements had analogous grouping outcomes to the consumers' discrimination. Emulsions 1 and 2 were the firmest, hardest to spread, stickiest and had the highest viscosity, while Emulsions 5 and 6 were the least firm, easiest to spread, less sticky than Emulsions 1 and 2, and had the lowest viscosity. Emulsions 3 and 4 fell in between the other two groups. Using olive oil instead of heptyl undecylenate as an emollient increased firmness, spreading, stickiness, viscosity and droplet size of the emulsions in every case-when comparing emulsions within each pair. All six emulsions had a shear-thinning behaviour. Viscosity and firmness directly correlated for the emulsions. Emulsions were visually stable at room temperature over the course of 6 months and viscosity remained relatively constant over this period also. CONCLUSION: Certain sensory attributes can be reliably predicted with instrumental measurements. Identifying and quantifying sensory-texture-rheology relationships can contribute to achieving appropriate product characteristics tailored to suit market needs.

Cardiorespiratory effects of epidurally administered ketamine or lidocaine in dogs undergoing ovariohysterectomy surgery: a comparative study.

BACKGROUND: Analgesic and hemodynamic effects of ketamine in subanesthetic doses during surgical anesthesia and postoperative, are due to the action on the N-methyl-D-aspartate receptors (NMDAR). AIMS: To evaluate the intraoperative cardiorespiratory effects provided by ketamine compared to lidocaine, both administered epidurally, in bitches submitted to ovariohysterectomy. METHODS: Thirty-six dogs of different breeds were used in a randomized, prospective, and blinded clinical trial. Two groups were formed: GKET (ketamine 3 mg/kg, n=18) and GLIDO (lidocaine 4 mg/kg, n=18). Animals were premedicated with acepromazine 0.05 mg/kg intravenous. Anesthesia was induced with propofol 5 mg/kg intravenous. Anesthetic maintenance was performed with isoflurane in 100% oxygen. Every 5 min during surgery, heart rate (HR), respiratory rate (RR), esophageal temperature (°C), oxygen saturation (SPO2), end tidal carbon dioxide (ETCO2) and mean arterial pressure (MAP) were monitored. RESULTS: Cardiorespiratory variables during anesthesia were within normal ranges. Heart rate was significantly higher at 5 (108 ± 12 vs 95 ± 11) and 10 (110 ± 11 vs 97 ± 11) min in GKET compared to GLIDO after the start of surgery (P=0.03 and P=0.01, respectively). Mean arterial pressure was higher in GKET, (100 ± 23, 105 ± 35, and 103 ± 35 mmHg) in comparison with GLIDO (66 ± 7, 74 ± 10, and 67 ± 9 mmHg) at 20, 25 and 30 min (P=0.01, P=0.004, and P=0.002, respectively). Mild hypothermia at 25 (36.5 1.3C) and 30 (36.5 1.4C) min in the GKET was recorded. CONCLUSION: Epidural administration of ketamine provides better hemodynamic stability, compared to the use of epidural lidocaine.

The Enemy is Outside: Releasing the Parasitoid Tamarixia radiata (Hymenoptera: Eulophidae) in External Sources of HLB Inocula to Control the Asian Citrus Psyllid Diaphorina citri (Hemiptera: Liviidae).

Huanglongbing (HLB), the most destructive citrus disease worldwide, was first recorded in Brazil in 2004, and since then, more than 50 million trees identified with this disease have been eliminated. The disease is managed mainly by controlling the psyllid vector Diaphorina citri Kuwayama, 1908 (Hemiptera: Liviidae). Although the presence of the insect in commercial citrus groves is low, HLB infection rates increase in areas bordering the groves. The disease is transmitted by psyllids from host citrus plants in areas outside the managed groves, such as abandoned or organic groves and residential trees, and from orange jasmine plants in urban settings. In order to provide information to support HLB control, this study evaluated the biotic and abiotic variables that affect the dynamics of D. citri populations after releases of the parasitoid wasp Tamarixia radiata (Waterston, 1922) (Hymenoptera: Eulophidae) in external sources of HLB inocula. The study was divided into two parts. After releasing the parasitoids in non-commercial areas, we determined the following: (a) the variables that significantly affected the number of nymphs collected on shoots in the same non-commercial area; (b) the variables that significantly affected the number of adult psyllids collected in a neighboring commercial citrus area. Our results indicated that the number of nymphs in external areas was affected only by the host plant and rainfall. However, periodic parasitoid releases significantly reduced the number of adult psyllids collected in the commercial area. The results indicate that the release of parasitoids in external sources of inocula has the potential to maximize actions for D. citri control, contributing to the reduction of psyllid populations in commercial areas. Consequently, this strategy may help to manage the disease infection without an increase in insecticide use.

Interpretive search of optimal isocratic and gradient separations in micellar liquid chromatography in extended organic solvent domains.

Micellar liquid chromatography (MLC) is a reversed-phase mode with mobile phases containing an organic solvent and a micellised surfactant. Most procedures developed in MLC are implemented in the isocratic mode, since the general elution problem in chromatography is less troublesome. However, gradient elution may be still useful in MLC to analyse mixtures of compounds within a wide range of polarities, in shorter times. MLC using gradients is attractive to determine by direct injection moderate to low polar compounds in physiological samples. In these analyses, the use of initial micellar conditions (isocratic or gradient) with a fixed amount of surfactant above the critical micellar concentration, keeping the organic solvent content low, will provide better protection of the column against the precipitation of the proteins in the physiological fluid. Once the proteins are swept away, the elution strength can be increased using a positive gradient of organic solvent to reduce the analysis time. This may give rise to the transition from the micellar to the submicellar mode, since micelles are destroyed at sufficiently high concentration of organic solvent. In this work, several retention models covering extended solvent domains in MLC are developed and tested, and applied to investigate the performance in isocratic, linear and multi-linear gradient separations. The study was applied to the screening of ß-adrenoceptor antagonists in urine samples, using mobile phases prepared with sodium dodecyl sulphate and 1-propanol. Predicted chromatograms were highly accurate in all situations, although suffered of baseline problems and minor shifts for peaks eluting close to a steep gradient segment. Two columns (C18 and C8) were investigated, with the C8 column being preferable owing to the smaller amount of adsorbed surfactant.

Prevalence of eosinophilic esophagitis: A multicenter study on a pediatric population evaluated at thirty-six Latin American gastroenterology centers. / Prevalencia de esofagitis eosinofílica: estudio multicéntrico en población pediátrica evaluada en 36 centros de gastroenterología de América Latina.

INTRODUCTION AND OBJECTIVE: Eosinophilic esophagitis is a chronic, immune-mediated disease described in case series and publications worldwide. Over the past twenty years, the authors of different studies have attempted to evaluate its incidence and prevalence. The objetive of the present study was to estimate the prevalence of eosinophilic esophagitis in a group of children seen at 36 pediatric gastroenterology centers in ten Latin American countries. MATERIALS AND METHODS: A multicenter, observational, and cross-sectional study was conducted that estimated the period prevalence of eosinophilic esophagitis in children seen at outpatient consultation and that underwent diagnostic upper gastrointestinal endoscopy for any indication at 36 centers in 10 Latin American countries, within a 3-month time frame. RESULTS: Between April and June 2016, 108 cases of eosinophilic esophagitis were evaluated. Likewise, an average of 29,253 outpatient consultations and 4,152 diagnostic upper gastrointestinal endoscopies were carried out at the 36 participating centers. The period prevalence of eosinophilic esophagitis in the population studied (n=29,253) was 3.69 cases×1,000 (95% CI: 3.04 to 4.44), and among the children that underwent routine upper gastrointestinal endoscopy (n=4,152), it was 26x1,000 (95% CI: 22.6 to 29.4). CONCLUSIONS: The general period prevalence of eosinophilic esophagitis in a group of children evaluated at 36 Latin American pediatric gastroenterology centers was 3.69×1,000, and in the children that underwent endoscopy, it was 26×1,000. There was important prevalence variability between the participating countries and centers. The present analysis is the first study conducted on the prevalence of pediatric eosinophilic esophagitis in Latin America.

Histological changes in the rat brain and spinal cord following prolonged intracerebroventricular infusion of cerebrospinal fluid from amyotrophic lateral sclerosis patients are similar to those caused by the disease. / La infusión intracerebroventricular prolongada de líquido cefalorraquídeo procedente de pacientes con esclerosis lateral amiotrófica provoca cambios histológicos en el cerebro y la médula espinal de la rata similares a los hallados en la enfermedad.

INTRODUCTION: Cerebrospinal fluid (CSF) from amyotrophic lateral sclerosis (ALS) patients induces cytotoxic effects in in vitro cultured motor neurons. MATERIAL AND METHODS: We selected CSF with previously reported cytotoxic effects from 32 ALS patients. Twenty-eight adult male rats were intracerebroventricularly implanted with osmotic mini-pumps and divided into 3 groups: 9 rats injected with CSF from non-ALS patients, 15 rats injected with cytotoxic ALS-CSF, and 4 rats injected with a physiological saline solution. CSF was intracerebroventricularly and continuously infused for periods of 20 or 43days after implantation. We conducted clinical assessments and electromyographic examinations, and histological analyses were conducted in rats euthanised 20, 45, and 82days after surgery. RESULTS: Immunohistochemical studies revealed tissue damage with similar characteristics to those found in the sporadic forms of ALS, such as overexpression of cystatinC, transferrin, and TDP-43 protein in the cytoplasm. The earliest changes observed seemed to play a protective role due to the overexpression of peripherin, AKTpan, AKTphospho, and metallothioneins; this expression had diminished by the time we analysed rats euthanised on day 82, when an increase in apoptosis was observed. The first cellular changes identified were activated microglia followed by astrogliosis and overexpression of GFAP and S100B proteins. CONCLUSION: Our data suggest that ALS could spread through CSF and that intracerebroventricular administration of cytotoxic ALS-CSF provokes changes similar to those found in sporadic forms of the disease.

Delimiting Strategic Zones for the Development of Fall Armyworm (Lepidoptera: Noctuidae) on Corn in the State of Florida.

The fall armyworm, Spodoptera frugiperda (J.E. Smith) (Lepidoptera: Noctuidae), cannot survive prolonged periods of freezing temperatures, thereby limiting where it can overwinter in North America. Climate change is anticipated to reduce the frequency of freeze days in Florida over the decades, with the potential consequence of a significant expansion of the overwintering range, whose northern limit in North America was assessed between 27 and 28°N in the last century. To assess this possibility, the development of the fall armyworm on corn leaves, one of the main host plants in the United States, was determined at five constant temperatures ranging from 14 to 30°C. Based on the development time, the thermal constant and the lower threshold temperature were used to estimate the number of generations of fall armyworm at 42 locations in the state of Florida, from 2006 to 2016. Maps were constructed to provide a visual description of the interpolated data, using GIS (Geographic Information System). The highest number of generations was observed in the counties farther south, an area that showed the highest temperatures during the years and plays a strategic role in maintaining fall armyworm populations in corn fields. Additionally, we conclude that in the absence of freeze periods, the northern limit for fall armyworm overwintering should be between 28 and 29°N.

Geostatistics and Geographic Information System to Analyze the Spatial Distribution of the Diversity of Anastrepha Species (Diptera: Tephritidae): the Effect of Forest Fragments in an Urban Area.

Fruit flies are among the most damaging insect pests of commercial fruit in Brazil. It is important to understand the landscape elements that may favor these flies. In the present study, spatial data from surveys of species of Anastrepha Schiner (Diptera: Tephritidae) in an urban area with forest fragments were analyzed, using geostatistics and Geographic Information System (GIS) to map the diversity of insects and evaluate how the forest fragments drive the spatial patterns. The results indicated a high diversity of species associated with large fragments, and a trend toward lower diversity in the more urbanized area, as the fragment sizes decreased. We concluded that the diversity of Anastrepha species is directly and positively related to large and continuous forest fragments in urbanized areas, and that combining geostatistics and GIS is a promising method for use in insect-pest management and sampling involving fruit flies.

A novel functionalization method for carbon nanotubes to repel ox-LDL in treatments after stent placement.

Carbon nanotubes (CNT) can be chemically modified by doping or functionalization to change the chemical and surface properties. These characteristic makes to CNT candidates for multiple applications including medical field in cardiovascular area. A novel method to CNT functionalization by formation of two compounds: α-bromoacid and the organic compound 2-(methacryloyloxy) ethyl phosphorylcholine (MPC), will be discussed in this article. According to results, CNT are suggested like candidates to repel oxidized low-density lipoproteins (ox-LDL) to prevent restenosis. The electronegative character on surface of functionalized CNT (F-CNT) is shown by wettability analysis observing a repellent behaviour in contact with ox-LDL after functionalization route. Here we analyse the toxicity of CNT and F-CNT on HepG2 cell line and find no damage to the cell membrane of HepG2 cells in concentration at doses below 1mg/ml.

A Theoretical Approach to Analyze the Parametric Influence on Spatial Patterns of Spodoptera frugiperda (J.E. Smith) (Lepidoptera: Noctuidae) Populations.

Studies of the influence of biological parameters on the spatial distribution of lepidopteran insects can provide useful information for managing agricultural pests, since the larvae of many species cause serious impacts on crops. Computational models to simulate the spatial dynamics of insect populations are increasingly used, because of their efficiency in representing insect movement. In this study, we used a cellular automata model to explore different patterns of population distribution of Spodoptera frugiperda (J.E. Smith) (Lepidoptera: Noctuidae), when the values of two biological parameters that are able to influence the spatial pattern (larval viability and adult longevity) are varied. We mapped the spatial patterns observed as the parameters varied. Additionally, by using population data for S. frugiperda obtained in different hosts under laboratory conditions, we were able to describe the expected spatial patterns occurring in corn, cotton, millet, and soybean crops based on the parameters varied. The results are discussed from the perspective of insect ecology and pest management. We concluded that computational approaches can be important tools to study the relationship between the biological parameters and spatial distributions of lepidopteran insect pests.

Cerebrospinal fluid cytotoxicity does not affect survival in amyotrophic lateral sclerosis.

OBJECTIVES: Cerebrospinal fluid (CSF) from some patients with amyotrophic lateral sclerosis (ALS) has been demonstrated to significantly reduce the neuronal viability of primary cell cultures of motor neurons. We aimed to study the potential clinical consequences associated with the cytotoxicity of CSF in a cohort of patients with ALS. METHODS: We collected CSF from thirty-one patients with ALS. We analysed cytotoxicity by incubating it into the primary cultures of motor cortex neurons. Neural viability was quantified after 24 hours using the colorimetric MTT reduction assay. All patients were followed up from the moment of diagnosis to death, and a complete evaluation during disease progression and survival was performed, including gastrostomy and respiratory assistance. RESULTS: Twenty-one patients (67.7%) presented a cytotoxic CSF. There were no significant differences between patients with and without cytotoxicity regarding mean time from symptom onset to the diagnosis, from the diagnosis to death, from the diagnosis to respiratory assistance with BIPAP, from diagnosis to gastrostomy and from the onset of symptoms to death. In Cox regression analysis, bulbar onset, but not cytotoxicity, gender or age at onset, was associated with a lower risk of survival. CONCLUSIONS: Cerebrospinal fluid cytotoxicity was not associated with differential survival rates. This suggests that the presence of cytotoxicity in CSF, measured through neuronal viability in primary cultures of motor cortex neurons, could reflect different mechanisms of the disease, but it does not predict disease outcome.

Widespread blunting of hypothalamic and amygdala-septal activity and behavior in rats with long-term hyperglycemia.

Anxiety and depression in diabetic patients contributes to a poor prognosis, but possible causal relationships have been controversial. Anxiety, fear, and anhedonia are mediated by interactions between different deep structures of the temporal lobe (e.g., amygdala complex and hippocampus) and other forebrain-related structures (e.g., lateral septal nucleus). Connections between these structures and the hypothalamic orexinergic system are necessary for the maintenance of energy and wakefulness. However, few studies have explored the impact of long-term hyperglycemia in these structures on anxiety. We induced long-term hyperglycemia (glucose levels of ∼500mg/dl) in Wistar rats by injecting them with alloxan and simultaneously protecting them from hyperglycemia by injecting them daily with a low dose of insulin (i.e., just enough insulin to avoid death), thus maintaining hyperglycemia and ketonuria for as long as 6 weeks. Compared with controls, long-term hyperglycemic rats exhibited a significant reduction of Fos expression in the lateral septal nucleus and basolateral amygdala, but no differences were found in cerebellar regions. Orexin-A cells appeared to be inactive in the lateral hypothalamus. No differences were found in sucrose consumption or behavior in the elevated plus maze compared with the control group, but a decrease in general locomotion was observed. These data indicate a generalized blunting of the metabolic brain response, accompanied by a decrease in locomotion but no changes in hedonic- or anxiety-like behavior.

Symbolic dynamics marker of heart rate variability combined with clinical variables enhance obstructive sleep apnea screening.

Many sleep centres try to perform a reduced portable test in order to decrease the number of overnight polysomnographies that are expensive, time-consuming, and disturbing. With some limitations, heart rate variability (HRV) has been useful in this task. The aim of this investigation was to evaluate if inclusion of symbolic dynamics variables to a logistic regression model integrating clinical and physical variables, can improve the detection of subjects for further polysomnographies. To our knowledge, this is the first contribution that innovates in that strategy. A group of 133 patients has been referred to the sleep center for suspected sleep apnea. Clinical assessment of the patients consisted of a sleep related questionnaire and a physical examination. The clinical variables related to apnea and selected in the statistical model were age (p < 10(-3)), neck circumference (p < 10(-3)), score on a questionnaire scale intended to quantify daytime sleepiness (p < 10(-3)), and intensity of snoring (p < 10(-3)). The validation of this model demonstrated an increase in classification performance when a variable based on non-linear dynamics of HRV (p < 0.01) was used additionally to the other variables. For diagnostic rule based only on clinical and physical variables, the corresponding area under the receiver operating characteristic (ROC) curve was 0.907 (95% confidence interval (CI) = 0.848, 0.967), (sensitivity 87.10% and specificity 80%). For the model including the average of a symbolic dynamic variable, the area under the ROC curve was increased to 0.941 (95% = 0.897, 0.985), (sensitivity 88.71% and specificity 82.86%). In conclusion, symbolic dynamics, coupled with significant clinical and physical variables can help to prioritize polysomnographies in patients with a high probability of apnea. In addition, the processing of the HRV is a well established low cost and robust technique.

Small synthetic hyaluronan disaccharides afford neuroprotection in brain ischemia-related models.

High molecular weight (HMW) glycosaminoglycanes of the extracellular matrix have been implicated in tissue repair. The aim of this study was to evaluate if small synthetic hyaluronan disaccharides with different degrees of sulfation (methyl 2-acetamido-2-deoxy-3-O-(ß-d-glucopyranosyluronic acid)-O-sulfo-α-d-glucopyranoside, sodium salt (di0S), methyl 2-acetamido-2-deoxy-3-O-(ß-d-glucopyranosyluronic acid)-6-di-O-sulfo-α-d-glucopyranoside, disodium salt (di6S) and methyl 2-acetamido-2-deoxy-3-O-(ß-d-glucopyranosyluronic acid)-4,6-di-O-sulfo-α-d-glucopyranoside, trisodium salt (di4,6S)) could improve cell survival in in vitro and in vivo brain ischemia-related models. Rat hippocampal slices subjected to oxygen and glucose deprivation and a photothrombotic stroke model in mice were used. The three hyaluran disaccharides, incubated during the oxygen and glucose deprivation (15min) and re-oxygenation periods (120min), reduced cell death of hippocampal slices measured as 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide reduction, being the most potent di4,6S; in contrast, high molecular hyaluronan was ineffective. The protective actions of di4,6S against oxygen and glucose deprivation were related to activation of the PI3K/Akt survival pathway, reduction of p65 translocation to the nucleus, inhibition of inducible nitric oxide oxidase induction and reactive oxygen species production, and to an increase in glutathione levels. Administered 1h post-stroke, di4,6S reduced cerebral infarct size and improved motor activity in the beam walk test. In conclusion, di4,6S affords neuroprotection in in vitro and in vivo models of ischemic neuronal damage. Our results suggest that its neuroprotective effect could be exerted through its capability to reduce oxidative stress during ischemia. Its small molecular size makes it a more potential druggable drug to target the brain as compared with its HMW parent compound hyaluronan.

Cellular changes in motor neuron cell culture produced by cytotoxic cerebrospinal fluid from patients with amyotrophic lateral sclerosis.

INTRODUCTION: The neurotoxic effects of cerebrospinal fluid (CSF) from patients with amyotrophic lateral sclerosis (ALS) have been reported by various authors who have attributed this neurotoxicity to the glutamate in CSF-ALS. MATERIAL AND METHODS: Cultures of rat embryonic cortical neurons were exposed to CSF from ALS patients during an incubation period of 24 hours. Optical microscopy was used to compare cellular changes to those elicited by exposure to 100µm glutamate, and confocal microscopy was used to evaluate immunohistochemistry for caspase-3, TNFα, and peripherin. RESULTS: In the culture exposed to CSF-ALS, we observed cells with nuclear fragmentation and scarce or null structural modifications to the cytoplasmic organelles or to plasma membrane maintenance. This did not occur in the culture exposed to glutamate. The culture exposed to CSF-ALS also demonstrated increases in caspase-3, TNFα, and in peripherin co-locating with caspase-3, but not with TNFα, suggesting that TNFα may play an early role in the process of apoptosis. CONCLUSIONS: CFS-ALS cytotoxicity is not related to glutamate. It initially affects the nucleus without altering the cytoplasmic membrane. It causes cytoplasmic apoptosis that involves an increase in caspase-3 co-located with peripherin, which is also overexpressed.

Galantamine elicits neuroprotection by inhibiting iNOS, NADPH oxidase and ROS in hippocampal slices stressed with anoxia/reoxygenation.

Galantamine is a drug currently used to treat Alzheimer's disease (AD); in this group of patients it has been observed that concomitant ischemic brain injury can accelerate their cognitive deficit. We have previously shown that galantamine can afford neuroprotection on in vitro and in vivo models related to brain ischemia. In this context, this study was planned to investigate the intracellular signaling pathways implicated in the protective effect of galantamine on an in vitro brain ischemia-reperfusion model, namely rat hippocampal slices subjected to oxygen and glucose deprivation (OGD) followed by reoxygenation. Galantamine protected hippocampal slices subjected to OGD in a concentration-dependent manner; at 15 µM, cell death was reduced to almost control levels. The neuroprotective effects of galantamine were reverted by mecamylamine and AG490, but not by atropine, indicating that nicotinic receptors and Jak2 participated in this action. Galantamine also prevented p65 translocation into the nucleus induced by OGD; this effect was also linked to nicotinic receptors and Jak2. Furthermore, galantamine reduced iNOS induction and production of NO caused by OGD via Jak2. ROS production by NADPH oxidase (NOX) activation was also inhibited by galantamine. In conclusion, galantamine afforded neuroprotection under OGD-reoxygenation conditions by activating a signaling pathway that involves nicotinic receptors, Jak2 and the consequent inhibition of NOX and NFκB/iNOS. This article is part of a Special Issue entitled 'Post-Traumatic Stress Disorder'.

Presynaptic muscarinic receptor subtypes involved in the enhancement of spontaneous GABAergic postsynaptic currents in hippocampal neurons.

We investigated the effects of muscarinic acetylcholine receptor (mAChR) activation on GABAergic synaptic transmission in rat hippocampal neurons. Current-clamp recordings revealed that methacholine produced membrane depolarization and action potential firing. Methacholine augmented the bicuculline-sensitive and GABA(A) -mediated frequency of spontaneous inhibitory postsynaptic currents (sIPSCs); the action of methacholine had a slow onset and longer duration. The increase in methacholine-evoked sIPSCs was completely inhibited by atropine and was insensitive to glutamatergic receptor blockers. Interestingly, methacholine action was not inhibited by intracellular perfusion with GDP-ß-S, suggesting that muscarinic effects on membrane excitability and sIPSC frequency are mainly presynaptic. McN-A-343 and pirenzepine, selective agonist and antagonist of the m1 mAChR subtype, respectively, neither enhanced sIPSCs nor inhibited the methacholine effect. However, the m3-m5 mAChR antagonist 4-DAMP, and the m2-m4 mAChR antagonist himbacine inhibited the methacholine effect. U73122, an IP(3) production inhibitor, and 2APB, an IP(3) receptor blocker, drastically decreased the methacholine effect. Recording of miniature events revealed that besides the effect exerted by methacholine on membrane firing properties and sIPSC frequency, muscarinic receptors also enhanced the frequency of mIPSCs with no effect on their amplitude, possibly modulating the molecular machinery subserving vesicle docking and fusion and suggesting a tight colocalization at the active zone of the presynaptic terminals. These data strongly suggest that by activating presynaptic m2, m3, m4 and m5 mAChRs, methacholine can increase membrane excitability and enhance efficiency in the GABA release machinery, perhaps through a mechanism involving the release of calcium from the endoplasmic reticulum.

Modulation by endogenously released ATP and opioids of chromaffin cell calcium channels in mouse adrenal slices.

Modulation of high-threshold voltage-dependent calcium channels by neurotransmitters has been the subject of numerous studies in cultures of neurons and chromaffin cells. However, no studies on such modulation exist in chromaffin cells in their natural environment, the intact adrenal medullary tissue. Here we performed such a study in voltage-clamped chromaffin cells of freshly prepared mouse adrenal slices under the whole cell configuration of the patch-clamp technique. The subcomponents of the whole cell inward Ca(2+) current (I(Ca)) accounted for 49% for L-, 28% for N-, and 36% for P/Q-type channels. T-type Ca(2+) channels or residual R-type Ca(2+) currents were not seen. However, under the perforated-patch configuration, 20% of I(Ca) accounted for a toxin-resistant R-type Ca(2+) current. Exogenously applied ATP and methionine-enkephalin (Met-enk) inhibited I(Ca) by 33%. Stop-flow and Ca(2+) replacement by Ba(2+), which favored the release of endogenous ATP and opioids, also inhibited I(Ca), with no changes in activation or inactivation kinetics. This inhibition was partially voltage independent and insensitive to prepulse facilitation. Furthermore, in about half of the cells, suramin and naloxone augmented I(Ca) in the absence of exogenous application of ATP/Met-enk. No additional modulation of I(Ca) was obtained after bath application of exogenous ATP and opioids to these already inhibited cells. Augmentation of I(Ca) was also seen upon intracellular dialysis of guanosine 5'-[ß-thio]diphosphate (GDPßS), indicating the existence in the intact slice of a tonic inhibition of I(Ca) in resting conditions. These results suggest that in the intact adrenal tissue a tonic inhibition of I(Ca) exists, mediated by purinergic and opiate receptors.