The SEEN comprehensive clinical survey of adult obesity: Executive summary. / Abordaje clínico integral SEEN de la obesidad en la edad adulta: resumen ejecutivo.

Obesity is one of the great challenges in healthcare nowadays with important implications for health so requiring comprehensive management. This document aims to establish practical and evidence-based recommendations for the diagnosis and management of in Spain, from the perspective of the clinical endocrinologist. A position statement has been made that can be consulted at www.seen.es, and that has been agreed by the Obesity Group of the Spanish Society of Endocrinology and Nutrition (GOSEEN), together with the Nutrition Area (NutriSEEN) and the Working Group of Endocrinology, Nutrition and Physical Exercise (GENEFSEEN).

Therapeutic drug monitoring of anti-tumour necrosis factor-α agents in inflammatory bowel disease: Limits and improvements.

AIMS: Since the publication of the American Gastroenterological Association's recommendations in 2017, there have been no significant changes in the biological monitoring recommendations in inflammatory bowel disease. Possible limitations are the lack of evidence to recommend proactive therapeutic drug monitoring (pTDM) over reactive TDM (rTDM), and the limited information about individualized dosing methods. This article aims to review the TDM strategy updates and the use of individualized dosing methods. METHODS: For the analysis of the TDM strategies and individualized dosing method, a search was carried out in PubMed and Cochrane Central. In the TDM case, since August 2017. RESULTS: A total of 263 publications were found, but only 7 related to proactive TDM. Five of these publications directly compared pTDM vs rTDM and 2 were randomized clinical trials. Six studies found benefits of pTDM and 1 found no differences. Regarding the individualized dosing method, 229 distinct results were found. Population pharmacokinetics was the most widely used method to develop individual dosage models and to analyse the influence of factors on drug concentrations (albumin concentration, weight, presence of anti-drug antibodies etc). CONCLUSION: We have found no major changes in TDM strategies. There is a growing trend towards the use of pTDM because it has shown a longer duration of treatment response, lower rates of discontinuation and relapses. However, the available evidence is limited and of low quality. Despite the common use of population pharmacokinetic methods to analyse pharmacokinetic factors, they are not commonly used for personalized dosing.

Terapias avanzadas / Advanced therapies

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Automated therapy preparation of isoleucine formulations using 3D printing for the treatment of MSUD: First single-centre, prospective, crossover study in patients.

Maple syrup urine disease (MSUD) is a rare metabolic disorder with a worldwide prevalence of 1 in every 185,000 live births. However, certain populations display a significant overexpression of the disorder where incidence is reported to be 1 in every 52,541 new-borns. The first-line therapy for MSUD involves a strict dietary leucine restriction and oral supplementation of isoleucine and valine. The dose administered to patients requires strict tailoring according to age, weight and blood levels. In current clinical practice, however, practitioners still have to prepare extemporaneous formulations due to the lack of suitable oral treatments for MSUD. Herein, we evaluate the first time use of 3D printing in a hospital setting for the preparation of personalised therapies with the aim of improving safety and acceptability to isoleucine supplementation in paediatric patients suffering from MSUD. This investigation was a single-centre, prospective crossover experimental study. Four paediatric patients with MSUD (aged 3-16 years) were treated at the Clinic University Hospital in Santiago de Compostela, Spain which is a MSUD reference hospital in Europe. The primary objective was to evaluate isoleucine blood levels after six months of treatment with two types of formulations; conventional capsules prepared by manual compounding and personalised chewable formulations prepared by automated 3D printing. A secondary investigation was to evaluate patient acceptability of 3D printed formulations prepared with different flavours and colours. Isoleucine blood levels in patients were well controlled using both types of formulations, however, the 3D printed therapy showed mean levels closer to the target value and with less variability (200-400 µM). The 3D printed formulations were well accepted by patients regarding flavour and colour. The study demonstrates for the first time that 3D printing offers a feasible, rapid and automated approach to prepare oral tailored-dose therapies in a hospital setting. 3D printing has shown to be an effective manufacturing technology in producing chewable isoleucine printlets as a treatment of MSUD with good acceptability.

SN56 neuronal cell death after 24 h and 14 days chlorpyrifos exposure through glutamate transmission dysfunction, increase of GSK-3ß enzyme, ß-amyloid and tau protein levels.

Chlorpyrifos (CPF) is an organophosphate insecticide described to induce cognitive disorders, both after acute and repeated administration. However, the mechanisms through which it induces these effects are unknown. CPF has been reported to produce basal forebrain cholinergic neuronal cell death, involved on learning and memory regulation, which could be the cause of such cognitive disorders. Neuronal cell death was partially mediated by oxidative stress generation, P75NTR and α7-nAChRs gene expression alteration triggered through acetylcholinesterase (AChE) variants disruption, suggesting other mechanisms are involved. In this regard, CPF induces Aß and tau proteins production and activation of GSK3ß enzyme and alters glutamatergic transmission, which have been related with basal forebrain cholinergic neuronal cell death and development of cognitive disorders. According to these data, we hypothesized that CPF induces basal forebrain cholinergic neuronal cell death through induction of Aß and tau proteins production, activation of GSK-3ß enzyme and disruption of glutamatergic transmission. We evaluated this hypothesis in septal SN56 basal forebrain cholinergic neurons, after 24 h and 14 days CPF exposure. This study shows that CPF increases glutamate levels, upregulates GSK-3ß gene expression, and increases the production of Aß and phosphorylated tau proteins and all these effects reduced cell viability. CPF increases glutaminase activity and upregulates the VGLUT1 gene expression, which could mediate the disruption of glutamatergic transmission. Our present results provide new understanding of the mechanisms contributing to the harmful effects of CPF, and its possible relevance in the pathogenesis of neurodegenerative diseases.

Cadmium induced ROS alters M1 and M3 receptors, leading to SN56 cholinergic neuronal loss, through AChE variants disruption.

Cadmium, an environmental neurotoxic compound, produces cognitive disorders, although the mechanism remains unknown. Previously, we described that cadmium induces a more pronounced cell death on cholinergic neurons from basal forebrain (BF). This effect, partially mediated by M1 receptor blockade, triggering it through AChE splices variants alteration, may explain cadmium effects on learning and memory processes. Cadmium has been also reported to induce oxidative stress generation leading to M2 and M4 muscarinic receptors alteration, in hippocampus and frontal cortex, which are necessary to maintain cell viability and cognitive regulation, so their alteration in BF could also mediate this effect. Moreover, it has been reported that antioxidant treatment could reverse cognitive disorders, muscarinic receptor and AChE variants alterations induced by cadmium. Thus, we hypothesized that cadmium induced cell death of BF cholinergic neurons is mediated by oxidative stress generation and this mechanism could produce this effect, in part, through AChE variants altered by muscarinic receptors disruption. To prove this, we evaluated in BF SN56 cholinergic neurons, whether cadmium induces oxidative stress and alters muscarinic receptors, and their involvement in the induction of cell death through alteration of AChE variants. Our results show that cadmium induces oxidative stress, which mediates partially the alteration of AChE variants and M2 to M4 muscarinic receptors expression and blockage of M1 receptor. In addition, cadmium induced oxidative stress generation by M1 and M3 receptors alteration through AChE variants disruption, leading to cell death. These results provide new understanding of the mechanisms contributing to cadmium harmful effects on cholinergic neurons.

Primary hippocampal neuronal cell death induction after acute and repeated paraquat exposures mediated by AChE variants alteration and cholinergic and glutamatergic transmission disruption.

Paraquat (PQ) is a widely used non-selective contact herbicide shown to produce memory and learning deficits after acute and repeated exposure similar to those induced in Alzheimer's disease (AD). However, the complete mechanisms through which it induces these effects are unknown. On the other hand, cholinergic and glutamatergic systems, mainly in the hippocampus, are involved on learning, memory and cell viability regulation. An alteration of hippocampal cholinergic or glutamatergic transmissions or neuronal cell loss may induce these effects. In this regard, it has been suggested that PQ may induce cell death and affect cholinergic and glutamatergic transmission, which alteration could produce neuronal loss. According to these data, we hypothesized that PQ could induce hippocampal neuronal loss through cholinergic and glutamatergic transmissions alteration. To prove this hypothesis, we evaluated in hippocampal primary cell culture, the PQ toxic effects after 24h and 14 consecutive days exposure on neuronal viability and the cholinergic and glutamatergic mechanisms related to it. This study shows that PQ impaired acetylcholine levels and induced AChE inhibition and increased CHT expression only after 14days exposure, which suggests that acetylcholine levels alteration could be mediated by these actions. PQ also disrupted glutamate levels through induction of glutaminase activity. In addition, PQ induced, after 24h and 14days exposure, cell death on hippocampal neurons that was partially mediated by AChE variants alteration and cholinergic and gultamatergic transmissions disruption. Our present results provide new view of the mechanisms contributing to PQ neurotoxicity and may explain cognitive dysfunctions observed after PQ exposure.

61.º Congreso de la SEFH. Nutrición parenteral libre de aminoácidos ramificados para el manejo urgente de una descompensación grave de la enfermedad del jarabe de arce / No disponible

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61.º Congreso de la SEFH. Complicaciones asociadas con medicamentos en un postoperatorio tórpido de trasplante hepático / No disponible

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Amitraz changes NE, DA and 5-HT biosynthesis and metabolism mediated by alterations in estradiol content in CNS of male rats.

Amitraz is a formamidine insecticide/acaricide that alters different neurotransmitters levels, among other neurotoxic effects. Oral amitraz exposure (20, 50 and 80 mg/kg bw, 5 days) has been reported to increase serotonin (5-HT), norepinephrine (NE) and dopamine (DA) content and to decrease their metabolites and turnover rates in the male rat brain, particularly in the striatum, prefrontal cortex, and hippocampus. However, the mechanisms by which these alterations are produced are not completely understood. One possibility is that amitraz monoamine oxidase (MAO) inhibition could mediate these effects. Alternatively, it alters serum concentrations of sex steroids that regulate the enzymes responsible for these neurotransmitters synthesis and metabolism. Thus, alterations in sex steroids in the brain could also mediate the observed effects. To test these hypothesis regarding possible mechanisms, we treated male rats with 20, 50 and 80 mg/kg bw for 5 days and then isolated tissue from striatum, prefrontal cortex, and hippocampus. We then measured tissue levels of expression and/or activity of MAO, catechol-O-metyltransferase (COMT), dopamine-ß-hydroxylase (DBH), tyrosine hydroxylase (TH) and tryptophan hydroxylase (TRH) as well as estradiol levels in these regions. Our results show that amitraz did not inhibit MAO activity at these doses, but altered MAO, COMT, DBH, TH and TRH gene expression, as well as TH and TRH activity and estradiol levels. The alteration of these enzymes was partially mediated by dysregulation of estradiol levels. Our present results provide new understanding of the mechanisms contributing to the harmful effects of amitraz.

Pharmacotherapy follow-up of patients under treatment with biologic agents for chronic inflammatory systemic conditions: an agreement among hospital pharmacists for the standardized collection of a minimum set of data / Seguimiento farmacoterapéutico en pacientes en tratamiento con agentes biológicos por enfermedades sistémicas inflamatorias crónicas: acuerdo entre farmacéuticos hospitalarios para la recopilación estandarizada de un conjunto mínimo de datos

Background and objective: The objective of this study was to reach a consensus on the minimum set of data that would allow to optimize the pharmacotherapy follow-up of patients on biologic agents for chronic systemic inflammatory conditions, through structured and standardized collection with an electronic tool in the hospital pharmacy. Materials and method: A scientific committee was formed (n = 5 hospital pharmacists). The Delphi Technique was used, 2 rounds of consultation by e-mail for hospital pharmacists. A structured questionnaire was used, based on a bibliographic review and recommendations by the scientific committee; 37 statements were assessed with the Likert 5-point scale (1= ‘Strongly Disagree’; 5= ‘Strongly Agree’). Consensus was reached when 75% or more of panel members assigned a score of 1-2 (rejection consensus) or 4-5 (agreement consensus) to the matter reviewed. Descriptive statistical analyses were conducted. Results: The study included 21 hospital pharmacists (70 were invited, there was 70% response). Consensus was reached for 100% of statements. The minimum set of data was agreed upon, as well as the recommendations that the pharmacist had to collect and make during visits: to document the health status, health-related quality of life, changes in treatment compliance and in patient autonomy, as well as the conditions to make feasible the systematic collection of the minimum data set. Conclusions: There is consensus among hospital pharmacists about a minimum data set to be collected, through an electronic tool, which will order, standardize and structure the pharmacotherapy follow-up of patients with chronic inflammatory conditions on treatment with biologic agents in the spanish public health system (AU)

Fundamento y objetivo: El objetivo de este estudio fue consensuar un conjunto mínimo de datos cuya recopilación sistemá- tica y estandarizada, mediante una herramienta electrónica en la farmacia hospitalaria, permitiera optimizar el seguimiento farmacoterapéutico de los pacientes tratados con agentes biológicos por enfermedades sistémicas inflamatorias crónicas. Material y método: Se constituyó un comité científico (n = 5 farmacéuticos hospitalarios). Se empleó la técnica Delphi, 2 rondas de consulta, por correo electrónico entre farmacéuticos hospitalarios. Se utilizó un cuestionario estructurado basado en una revisión bibliográfica y en recomendaciones del comité científico, valorándose 37 afirmaciones en una escala Likert de 5 puntos (1 = ‘En total desacuerdo’; 5 = ‘Totalmente de acuerdo’). Se alcanzó consenso cuando el 75% o más de los panelistas puntuaron 1-2 (consenso-rechazo) o 4-5 (consenso-acuerdo) la cuestión planteada. Se realizaron análisis estadísticos descriptivos. Resultados: Participaron 21 farmacéuticos hospitalarios (70 invitados, 70% respuesta). Se logró consenso en el 100% de las afirmaciones. Se acordó el conjunto mínimo de datos y de recomendaciones que el farmacéutico debe recoger y hacer en las visitas; documentar el estado de salud, la calidad de vida relacionada con la salud, los cambios en la adherencia al tratamiento y en la autonomía de los pacientes, así como las condiciones para hacer factible la recopilación sistemática del conjunto mínimo de datos. Conclusiones: Existe consenso entre los farmacéuticos hospitalarios en un conjunto mínimo de datos cuya recopilación, mediante una herramienta electrónica, ordenará, estandarizará y sistematizará el seguimiento farmacoterapéutico de los pacientes con enfermedades inflamatorias crónicas en tratamiento con agentes biológicos en el entorno sanitario público español (AU)

Pharmacotherapy follow-up of patients under treatment with biologic agents for chronic inflammatory systemic conditions: an agreement among hospital pharmacists for the standardized collection of a minimum set of data.

BACKGROUND AND OBJECTIVE: The objective of this study was to reach a consensus on the minimum set of data that would allow to optimize the pharmacotherapy follow-up of patients on biologic agents for chronic systemic inflammatory conditions, through structured and standardized collection with an electronic tool in the hospital pharmacy. MATERIALS AND METHOD: A scientific committee was formed (n = 5 hospital pharmacists). The Delphi Technique was used, 2 rounds of consultation by e-mail for hospital pharmacists. A structured questionnaire was used, based on a bibliographic review and recommendations by the scientific committee; 37 statements were assessed with the Likert 5-point scale (1= "Strongly Disagree"; 5= "Strongly Agree"). Consensus was reached when 75% or more of panel members assigned a score of 1-2 (rejection consensus) or 4-5 (agreement consensus) to the matter reviewed. Descriptive statistical analyses were conducted. RESULTS: The study included 21 hospital pharmacists (70 were invited, there was 70% response). Consensus was reached for 100% of statements. The minimum set of data was agreed upon, as well as the recommendations that the pharmacist had to collect and make during visits: to document the health status, health-related quality of life, changes in treatment compliance and in patient autonomy, as well as the conditions to make feasible the systematic collection of the minimum data set. CONCLUSIONS: There is consensus among hospital pharmacists about a minimum data set to be collected, through an electronic tool, which will order, standardize and structure the pharmacotherapy follow-up of patients with chronic inflammatory conditions on treatment with biologic agents in the spanish public health system.

Fundamento y objetivo: El objetivo de este estudio fue consensuar un conjunto mínimo de datos cuya recopilación sistemática y estandarizada, mediante una herramienta electrónica en la farmacia hospitalaria, permitiera optimizar el seguimiento farmacoterapéutico de los pacientes tratados con agentes biológicos por enfermedades sistémicas inflamatorias crónicas. Material y método: Se constituyó un comité científico (n = 5 farmacéuticos hospitalarios). Se empleó la técnica Delphi, 2 rondas de consulta, por correo electrónico entre farmacéuticos hospitalarios. Se utilizó un cuestionario estructurado basado en una revisión bibliográfica y en recomendaciones del comité científico, valorándose 37 afirmaciones en una escala Likert de 5 puntos (1 = "En total desacuerdo"; 5 = "Totalmente de acuerdo"). Se alcanzó consenso cuando el 75% o más de los panelistas puntuaron 1-2 (consenso-rechazo) o 4-5 (consenso- acuerdo) la cuestión planteada. Se realizaron análisis estadísticos descriptivos. Resultados: Participaron 21 farmacéuticos hospitalarios (70 invitados, 70% respuesta). Se logró consenso en el 100% de las afirmaciones. Se acordó el conjunto mínimo de datos y de recomendaciones que el farmacéutico debe recoger y hacer en las visitas; documentar el estado de salud, la calidad de vida relacionada con la salud, los cambios en la adherencia al tratamiento y en la autonomía de los pacientes, así como las condiciones para hacer factible la recopilación sistemática del conjunto mínimo de datos. Conclusiones: Existe consenso entre los farmacéuticos hospitalarios en un conjunto mínimo de datos cuya recopilación, mediante una herramienta electrónica, ordenará, estandarizará y sistematizará el seguimiento farmacoterapéutico de los pacientes con enfermedades inflamatorias crónicas en tratamiento con agentes biológicos en el entorno sanitario público español.

Toxicogenomic profile of apoptotic and necrotic SN56 basal forebrain cholinergic neuronal loss after acute and long-term chlorpyrifos exposure.

Chlorpyrifos (CPF) is an organophosphate insecticide reported to induce, both after acute and repeated exposure, learning and memory dysfunctions, although the mechanism is not completely known. CPF produces basal forebrain cholinergic neuronal loss, involved on learning and memory regulation, which could be the cause of such cognitive disorders. This effect was reported to be induced through apoptotic process, partially mediated by AChE overexpression, although neuronal necrosis was also described after CPF exposure. Accordingly, we hypothesized that CPF induces apoptotic and necrotic basal forebrain cholinergic cell death. We evaluated, in septal SN56 basal forebrain cholinergic neurons, the CPF effect after 24h and 14days exposure on apoptosis and necrosis induction and the apoptotic and necrotic gene expression pathways. This study shows that CPF induces, after acute and long-term exposure, apoptosis and necrosis, partially mediated through AChE overexpression. Evaluation of cell death pathways supports the necrosis and apoptosis data and revealed that some genes are altered at lower concentrations than those at which the effects observed are produce and below the No Observed Adverse Effect Level (NOAEL). The present finding suggests that the use of gene expression profile could be a more sensitive and accurate way to determine the CPF's NOAEL.

Evaluation of the in vitro ocular toxicity of the fortified antibiotic eye drops prepared at the Hospital Pharmacy Departments.

The use of parenteral antibiotic eye drop formulations with non-marketed compositions or concentrations, commonly called fortified antibiotic eye drops, is a common practice in Ophthalmology in the hospital setting. The aim of this study was to evaluate the in vitro ocular toxicity of the main fortified antibiotic eye drops prepared in the Hospital Pharmacy Departments. We have conducted an in vitro experimental study in order to test the toxicity of gentamicin, amikacin, cefazolin, ceftazidime, vancomycin, colistimethate sodium and imipenem-cilastatin eye drops; their cytotoxicity and acute tissue irritation have been evaluated. Cell-based assays were performed on human stromal keratocytes, using a cell-based impedance biosensor system [xCELLigence Real-Time System Cell Analyzer (RTCA)], and the Hen's Egg Test for the ocular irritation tests. All the eye drops, except for vancomycin and imipenem, have shown a cytotoxic effect dependent on concentration and time; higher concentrations and longer exposure times will cause a steeper decline in the population of stromal keratocytes. Vancomycin showed a major initial cytotoxic effect, which was reverted over time; and imipenem appeared as a non-toxic compound for stromal cells. The eye drops with the highest irritating effect on the ocular surface were gentamicin and vancomycin. Those antibiotic eye drops prepared at the Hospital Pharmacy Departments included in this study were considered as compounds potentially cytotoxic for the ocular surface; this toxicity was dependent on the concentration used.

El uso de reformulaciones de antibioticos parenterales en forma de colirios de composicion o concentraciones no comercializadas, comunmente denominados colirios antibioticos reforzados, es una practica habitual en oftalmologia a nivel hospitalario. El objetivo del presente trabajo ha consistido en evaluar la toxicidad ocular in vitro de los principales colirios antibioticos reforzados elaborados en los Servicios de Farmacia Hospitalaria. Hemos realizado un estudio experimental in vitro para evaluar la toxicidad de los colirios de gentamicina, amikacina, cefazolina, ceftazidima, vancomicina, colistimetato de sodio e imipenem- cilastatina en el que se ha evaluado su citotoxicidad y la irritacion tisular aguda. Los ensayos celulares se realizan sobre queratocitos estromales humanos, mediante la utilizacion de un sistema biosensor de impedancia celular [(xCELLigence Real- Time System Cell Analyzer (RTCA)] y los ensayos de irritacion ocular mediante el ensayo Hen/s Egg Test. Todos los colirios, excepto vancomicina e imipenem, han mostrado un efecto citotoxico de concentracion y tiempo dependiente, siendo las concentraciones mas altas y los tiempos mas prolongados los que provocan un descenso mas pronunciado en la poblacion de queratocitos estromales. La vancomicina muestra un importante efecto citotoxico inicial que revierte con el transcurso del tiempo y el imipenem se muestra como un compuesto no toxico para las celulas estromales. Los compuestos con mayor efecto irritante para la superficie ocular son la gentamicina y la vancomicina. Los colirios antiinfecciosos elaborados en los Servicios de Farmacia Hospitalaria estudiados se muestran como compuestos potencialmente citotoxicos para la superficie ocular, siendo esta toxicidad dependiente de la concentracion utilizada.

Muscarinic M1 receptor partially modulates higher sensitivity to cadmium-induced cell death in primary basal forebrain cholinergic neurons: A cholinesterase variants dependent mechanism.

Cadmium is a toxic compound reported to produce cognitive dysfunctions, though the mechanisms involved are unknown. In a previous work we described how cadmium blocks cholinergic transmission and induces greater cell death in primary cholinergic neurons from the basal forebrain. It also induces cell death in SN56 cholinergic neurons from the basal forebrain through M1R blockage, alterations in the expression of AChE variants and GSK-3ß, and an increase in Aß and total and phosphorylated Tau protein levels. It was observed that the silencing or blockage of M1R altered ChAT activity, GSK-3ß, AChE splice variants gene expression, and Aß and Tau protein formation. Furthermore, AChE-S variants were associated with the same actions modulated by M1R. Accordingly, we hypothesized that cholinergic transmission blockage and higher sensitivity to cadmium-induced cell death of primary basal forebrain cholinergic neurons is mediated by M1R blockage, which triggers this effect through alteration of the expression of AChE variants. To prove this hypothesis, we evaluated, in primary culture from the basal forebrain region, whether M1R silencing induces greater cell death in cholinergic neurons than cadmium does, and whether in SN56 cells M1R mediates the mechanisms described so as to play a part in the cadmium induction of cholinergic transmission blockage and cell death in this cell line through alteration of the expression of AChE variants. Our results prove that M1R silencing by cadmium partially mediates the greater cell death observed on basal forebrain cholinergic neurons. Moreover, all previously described mechanisms for blocking cholinergic transmission and inducing cell death on SN56 cells after cadmium exposure are partially mediated by M1R through the alteration of AChE expression. Thus, our results may explain cognitive dysfunctions observed in cadmium toxicity.

SN56 basal forebrain cholinergic neuronal loss after acute and long-term chlorpyrifos exposure through oxidative stress generation; P75(NTR) and α7-nAChRs alterations mediated partially by AChE variants disruption.

Chlorpyrifos (CPF) is an organophosphates insecticide reported to induce, both after acute and repeated exposure, cognitive disorders and basal forebrain cholinergic neuronal loss, involved on learning and memory regulation, which could be the cause of such cognitive disorders. This neuronal loss was mediated partially by AChE variants alteration, suggesting other mechanisms are involved. In this regard, CPF induces oxidative stress that is implicated in the induction of cognitive deficits, changes in AChE variants expression and neuronal loss. Otherwise, it has been shown that P75(NTR) and the α7-nAChRs expression is altered in basal forebrain of rats after CPF long-term exposure; this alteration has been related with oxidative stress induction, cholinergic cell loss, and disruption of learning and memory processes. According to these data, we hypothesized that CPF induces basal forebrain cholinergic neuronal loss through induction of oxidative stress produced by P75(NTR) and α7-nAChRs altered expression, which could mediate this action in part through AChE variants disruption. We evaluated this hypothesis in septal SN56 basal forebrain cholinergic neurons, after 24h and 14days CPF exposure in vitro. This study shows that CPF upregulated P75(NTR) and downregulated α7-nAChRs expression, which increased H2O2 and malondialdehyde content and reduced cell viability partially through AChE variants induction. Alpha7-nAChRs repression induced oxidative stress and cell death partially through this mechanism, but P75(NTR) overexpression did not produce these effects, although it increased oxidative stress and cell death after CPF treatment, showing that its overexpression increases cell vulnerability. Our present results provide new understanding of the mechanisms contributing to the harmful effects of CPF.

Cadmium-induced cell death of basal forebrain cholinergic neurons mediated by muscarinic M1 receptor blockade, increase in GSK-3ß enzyme, ß-amyloid and tau protein levels.

Cadmium is a neurotoxic compound which induces cognitive alterations similar to those produced by Alzheimer's disease (AD). However, the mechanism through which cadmium induces this effect remains unknown. In this regard, we described in a previous work that cadmium blocks cholinergic transmission and induces a more pronounced cell death on cholinergic neurons from basal forebrain which is partially mediated by AChE overexpression. Degeneration of basal forebrain cholinergic neurons, as happens in AD, results in memory deficits attributable to the loss of cholinergic modulation of hippocampal synaptic circuits. Moreover, cadmium has been described to activate GSK-3ß, induce Aß protein production and tau filament formation, which have been related to a selective loss of basal forebrain cholinergic neurons and development of AD. The present study is aimed at researching the mechanisms of cell death induced by cadmium on basal forebrain cholinergic neurons. For this purpose, we evaluated, in SN56 cholinergic mourine septal cell line from basal forebrain region, the cadmium toxic effects on neuronal viability through muscarinic M1 receptor, AChE splice variants, GSK-3ß enzyme, Aß and tau proteins. This study proves that cadmium induces cell death on cholinergic neurons through blockade of M1 receptor, overexpression of AChE-S and GSK-3ß, down-regulation of AChE-R and increase in Aß and total and phosphorylated tau protein levels. Our present results provide new understanding of the mechanisms contributing to the harmful effects of cadmium on cholinergic neurons and suggest that cadmium could mediate these mechanisms by M1R blockade through AChE splices altered expression.

Acute and long-term exposure to chlorpyrifos induces cell death of basal forebrain cholinergic neurons through AChE variants alteration.

Chlorpyrifos (CPF) is one of the most widely used organophosphates insecticides that has been reported to induce cognitive disorders both after acute and repeated administration similar to those induced in Alzheimer's disease (AD). However, the mechanisms through which it induces these effects are unknown. On the other hand, the cholinergic system, mainly basal forebrain cholinergic neurons, is involved in learning and memory regulation, and an alteration of cholinergic transmission or/and cholinergic cell loss could induce these effects. In this regard, it has been reported that CPF can affect cholinergic transmission, and alter AChE variants, which have been shown to be related with basal forebrain cholinergic neuronal loss. According to these data, we hypothesized that CPF could induce basal forebrain cholinergic neuronal loss through cholinergic transmission and AChE variants alteration. To prove this hypothesis, we evaluated in septal SN56 basal forebrain cholinergic neurons, the CPF toxic effects after 24h and 14 days exposure on neuronal viability and the cholinergic mechanisms related to it. This study shows that CPF impaired cholinergic transmission, induced AChE inhibition and, only after long-term exposure, increased CHT expression, which suggests that acetylcholine levels alteration could be mediated by these actions. Moreover, CPF induces, after acute and long-term exposure, cell death in cholinergic neurons in the basal forebrain and this effect is independent of AChE inhibition and acetylcholine alteration, but was mediated partially by AChE variants alteration. Our present results provide a new understanding of the mechanisms contributing to the harmful effects of CPF on neuronal function and viability, and the possible relevance of CPF in the pathogenesis of neurodegenerative diseases.

Impaired glutamatergic and GABAergic transmission by amitraz in primary hippocampal cells.

Amitraz is a formamidine pesticide that has been reported to be a neurotoxic compound that induces convulsions among other effects. Excitatory and inhibitory neurotransmission is mediated mainly by glutamate and GABA, respectively, so their alteration could be responsible for induction of seizures. In this regard, amitraz α2 adrenergic agonist action, which has been suggested as likely responsible for this effect, could alter these neurotransmitter systems and lead to seizure induction. Moreover, other amitraz mechanisms such as histamine H1 receptor inhibition could be involved. To confirm if amitraz disrupts glutamatergic/GABAergic transmission by these mechanisms, we evaluated, in primary hippocampal neurons, the effect of amitraz (0.01 µM to 100 µM) with or without the α2 adrenergic antagonist idazoxan (1 µM) and/or the H1 receptor agonist n-methylhistaprodifen (30 µM) co-treatment on 4-aminobutyrate aminotransferase, glutamate decarboxylase 65 (GAD 65), succinate-semialdehyde dehydrogenase and glutaminase gene expression and on glutamate and GABA levels after 24h treatment. We observed that amitraz disrupts glutaminase and GAD 65 gene expression, altering glutamatergic and GABAergic transmission. These effects were mediated partially by H1 and α2 receptors suggesting that other mechanisms could be involved. These data could help explain the mechanism by which amitraz induces seizures and provide a therapeutic strategy to protect against this effect in case of poisoning.