Manganese increases Aß and Tau protein levels through proteasome 20S and heat shock proteins 90 and 70 alteration, leading to SN56 cholinergic cell death following single and repeated treatment.

Manganese (Mn) produces cholinergic neuronal loss in basal forebrain (BF) region that was related to cognitive dysfunction induced after single and repeated Mn treatment. All processes that generate cholinergic neuronal loss in BF remain to be understood. Mn exposure may produce the reduction of BF cholinergic neurons by increasing amyloid beta (Aß) and phosphorylated Tau (pTau) protein levels, altering heat shock proteins' (HSPs) expression, disrupting proteasome P20S activity and generating oxidative stress. These mechanisms, described to be altered by Mn in regions different than BF, could lead to the memory and learning process alteration produced after Mn exposure. The research performed shows that single and repeated Mn treatment of SN56 cholinergic neurons from BF induces P20S inhibition, increases Aß and pTau protein levels, produces HSP90 and HSP70 proteins expression alteration, and oxidative stress generation, being the last two effects mediated by NRF2 pathway alteration. The increment of Aß and pTau protein levels was mediated by HSPs and proteasome dysfunction. All these mechanisms mediated the cell decline observed after Mn treatment. Our results are relevant because they may assist to reveal the processes leading to the neurotoxicity and cognitive alterations observed after Mn exposure.

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.

Molecular mechanisms of amitraz mammalian toxicity: a comprehensive review of existing data.

Amitraz is a formamidine pesticide widely used as an insecticide and acaricide. Amitraz poisoning cases in humans and animals are still being described to date, which is a cause of concern for health authorities. Amitraz was reported not to pose unreasonable risks or adverse effects to humans or the environment unlike the other commercialized member of the formamidine family, chlordimeform, which was removed from the market because of carcinogenic effects in animal studies. Amitraz was classified as a nonquantifiable "Suggestive Evidence of Carcinogenicity" and not genotoxic, but recently, it has been reported that it could induce genotoxic effects. Moreover, ever since the previously published evaluations made by the Environmental Protection Agency (EPA) and the Joint Meeting of Pesticide Residues (JMPR) there have been new reported data on amitraz toxicity related to genotoxicity, oxidative stress, cell death, immunotoxicty, endocrine disruption, and developmental toxicity which indicate that the risk of this compound could be underestimated. Furthermore, there is missing information about the dose-response relationship for some mechanisms and toxic effects described for amitraz and its metabolites, the mechanism of action by which several toxic effects are produced, and amitraz pharmacokinetics on different species. According to this, the new information reported should be taken into account, and more studies should be performed to fill in the gaps of missing information for a complete hazard identification and therefore an exhaustive risk assessment of amitraz. This review is aimed at updating the current knowledge on molecular mechanisms of amitraz mammalian toxicity, pointing out the missing information, providing some possible explanation of the mechanism by which some toxic effects observed are produced, and suggesting future direction of its research. To our knowledge, this is the first review on the molecular mechanisms of amitraz toxicity.

Assessment of cardiotoxicity and plasma ropivacaine concentrations after serratus intercostal fascial plane block in an experimental model.

Serratus intercostal fascial plane block (SIFPB) has emerged as an alternative to paravertebral block in breast surgery. It involves the administration of high volumes and doses of local anesthetics (LA) that can potentially reach toxic levels. Ropivacaine is widely used in thoraco-fascial blocks; however, there is no information on the plasma concentrations attained after SIPFB and whether they are associated with cardiotoxicity. Plasma concentrations of ropivacaine and its electrophysiological effects were evaluated in eight pigs after bilateral SIFPB with ropivacaine in doses of 3 mg/kg. Plasma concentrations, electrophysiological and hemodynamic parameters were measured sequentially for the following 180 min until the end of the study. The area under the curve, the maximum plasma concentration (Cmax) and the time to reach Cmax (tmax) were calculated. The median arterial ropivacaine concentration Cmax was, 2.34 [1.40 to 3.74] µg/ml. The time to reach the highest concentration was 15 [10 to 20] min. Twenty-five percent of the animals had arterial concentrations above the lower limit concentration of ropivacaine for LA systemic toxicity (3.4 µg/ml). No alterations were observed in the electrophysiological or electrocardiographic parameters except for a prolongation of the QTc interval, from 489 ± 30 to 544 ± 44 ms (Δ11.38 ± 6%), P = 0.01. Hemodynamic parameters remained in the physiological range throughout the study. SIFPB with ropivacaine in doses of 3 mg/kg has reached potentially toxic levels, however, it has not been associated with adverse electrophysiological or hemodynamic effects.

Sodium bicarbonate reverts electrophysiologic cardiotoxicity of ropivacaine faster than lipid emulsions in a porcine model.

Ropivacaine has been described as a safer local anaesthetic (LA); however, serious cardiotoxic accidents have been reported. Intravenous-lipid-emulsion (ILE) therapy during LA intoxication seems to act as an antidote. Sodium bicarbonate is the standard treatment for sodium channel blocker drug toxicity. We compared both antidotes on the reversion of electrophysiologic toxicity induced by ropivacaine. Ropivacaine 5 mg kg-1 was administered in 24 pigs, and 3 min later, the animals received ILE: 1.5 ml kg-1  + 0.25 ml kg-1  min-1 (ILE group); sodium bicarbonate: 2 mEq kg-1  + 1 mEq kg-1  h-1 (NaHCO3 group); saline solution (CTL group). Electrophysiological parameters were evaluated for 30 min. The area under the curve (AUC) for the first 5 or 30 min was compared between groups. Ropivacaine induced a lengthening of the PR interval by 17% (P = 0.0001), His-ventricle-interval by 58% (P = 0.001), sinus QRS complex by 56% (P = 0.0001), paced QRS at 150 bpm by 257% (P = 0.0001), and at 120 bpm by 143% (P = 0.0001) in all groups. At 5 min after treatment, sinus QRS in the NaHCO3 group was shorter than that in the CTL group (AUCQRS5 , P = 0.003) or ILE group (AUCQRS5 , P = 0.045). During the first minute, seven of the animals in the NaHCO3 group vs. two in the ILE or 0 in the CTL group recovered more than 30% of the sinus QRS previously lengthened by ropivacaine (P = 0.003). Sodium bicarbonate reversed the electrophysiological toxicity of ropivacaine faster than ILE and control groups.

Cadmium-promoted thyroid hormones disruption mediates ROS, inflammation, Aß and Tau proteins production, gliosis, spongiosis and neurodegeneration in rat basal forebrain.

Cadmium (Cd) produces cognition decline following single and repeated treatment, although the complete mechanisms are still unrevealed. Basal forebrain (BF) cholinergic neurons innervate the cortex and hippocampus, regulating cognition. Cd single and repeated exposure induced BF cholinergic neuronal loss, partly through thyroid hormones (THs) disruption, which may cause the cognition decline observed following Cd exposure. However, the mechanisms through which THs disruption mediate this effect remain unknown. To research the possible mechanisms through which Cd-induced THs deficiency may mediate BF neurodegeneration, Wistar male rats were treated with Cd for 1- (1 mg/kg) or 28-days (0.1 mg/kg) with or without triiodothyronine (T3, 40 µg/kg/day). Cd exposure promoted neurodegeneration, spongiosis, gliosis and several mechanisms related to these alterations (increased H202, malondialdehyde, TNF-α, IL-1ß, IL-6, BACE1, Aß and phosphorylated-Tau levels, and decreased phosphorylated-AKT and phosphorylated-GSK-3ß levels). T3 supplementation partially reversed the effects observed. Our results show that Cd induces several mechanisms that may be responsible for the neurodegeneration, spongiosis and gliosis observed in the rats' BF, which are partially mediated by a reduction in THs levels. These data may help to explain the mechanisms through which Cd induces BF neurodegeneration, possibly leading to the cognitive decline observed, providing new therapeutic tools to prevent and treat these damages.

Effects of intravenous lipid emulsions on the reversal of pacing-induced ventricular arrhythmias and electrophysiological alterations in an animal model of ropivacaine toxicity.

INTRODUCTION: Ropivacaine is considered to have a wider margin of cardiovascular safety. However, several reports of ventricular arrhythmias (VA) due to ropivacaine toxicity have been documented. Intravenous lipid emulsions (ILEs) have recently been used successfully in the treatment of local anesthetic intoxication. The main objective of the present study was to evaluate the efficacy of the ILEs in the prevention of pacing-induced-VA and electrophysiological alterations in an animal model of ropivacaine toxicity. METHODS: Nineteen pigs were anesthetized and instrumentalized. A baseline programmed electrical ventricular stimulation protocol (PEVSP) to induce VA was performed. Ropivacaine (5 mg·kg-1 + 100 µg·kg-1·min-1) followed by normal saline infusion (control group n = 8) or intralipid 20% (1.5 mL·kg-1 + 0.25 mL·kg-1·min-1) for the ILE group (n = 8), were administered three minutes after the ropivacaine bolus. PEVSP was repeated 25 min after the onset of ropivacaine infusion. Pacing-induced VA and electrophysiological abnormalities were assessed in both groups. A sham-control group (n = 3) without ropivacaine infusion was included. RESULTS: Most of the electrophysiological parameters evaluated were affected by ropivacaine: PR interval by 28% (p = 0.001), AV interval by 40% (p = 0.001), sinus QRS by 101% (p = 0.001), paced QRS at a rate of 150 bpm by 258% (p = 0.001), and at 120 bpm by 241% (p = 0.001). Seven animals (87.5%) in the control group and eight animals (100%) in the ILE group developed sustained-VA (p = 0.30). Successful resuscitation occurred in 100% of animals in the ILE group vs. 57% of animals in the control group, p = 0.038. Pacing-induced-VA terminated at the first defibrillation attempt in 75% of the animals in the ILE group vs. 0% in the control group, p = 0.01. CONCLUSION: Ropivacaine strongly altered the parameters of ventricular conduction, thus facilitating the induction of VA. ILEs did not prevent pacing-induced VA. However, facilitated resuscitation and termination of VA were delivered at the first defibrillation attempt compared to the control group.

Cadmium-induced neurotoxic effects on rat basal forebrain cholinergic system through thyroid hormones disruption.

Cadmium (Cd) single and repeated exposure produces cognitive dysfunctions. Basal forebrain cholinergic neurons (BFCN) regulate cognitive functions. BFCN loss or cholinergic neurotransmission dysfunction leads to cognitive disabilities. Thyroid hormones (THs) maintain BFCN viability and functions, and Cd disrupts their levels. However, Cd-induced BFCN damages and THs disruption involvement was not studied. To research this we treated male Wistar rats intraperitoneally with Cd once (1 mg/kg) or repetitively for 28 days (0.1 mg/kg) with/without triiodothyronine (T3, 40 µg/kg/day). Cd increased thyroid-stimulating-hormone (TSH) and decreased T3 and tetraiodothyronine (T4). Cd altered cholinergic transmission and induced a more pronounced neurodegeneration on BFCN, mediated partially by THs reduction. Additionally, Cd antagonized muscarinic 1 receptor (M1R), overexpressed acetylcholinesterase S variant (AChE-S), downregulated AChE-R, M2R, M3R and M4R, and reduced AChE and choline acetyltransferase activities through THs disruption. These results may assist to discover cadmium mechanisms that induce cognitive disabilities, revealing a new possible therapeutic tool.

Single and repeated bisphenol A treatment induces ROS, Aß and hyperphosphorylated-tau accumulation, and insulin pathways disruption, through HDAC2 and PTP1B overexpression, leading to SN56 cholinergic apoptotic cell death.

Bisphenol-A (BPA), a polymer component extensively used, produces memory and learning alterations after acute and long-term exposure. However, the mechanisms are not well known. Cortex and hippocampus neuronal networks control cognitive functions, which are innervated by basal forebrain cholinergic neurons (BFCN), and their neurodegeneration induces cognitive dysfunctions. Wild type or protein tyrosine phosphatase 1B (PTP1B), histone deacetylase 2 (HDAC2), tau or ß amyloid precursor protein (ßAPP) silenced SN56 cells treated with BPA (0.001 µM-100 µM) with or without N-acetylcysteine (NAC; 1 mM), following 1 and 14 days, were used, as a model of BFCN to determine the insulin pathway dysfunction, oxidative stress (OS) generation and amyloid-ß (Aß) and tau proteins accumulation involvement in the BCFN cell death induction, as a possible mechanism that could produce the cognitive disorders reported. BPA-induced BFCN cell death, after 24 h and 14 days of treatment, through insulin pathway dysfunction, OS generation, mediated by NRF2 pathway downregulation, and Aß and tau proteins accumulation, which were in turn induced by HDAC2 and PTP1B overexpression. This is relevant information to explain the BFCN neurodegeneration mechanisms that could trigger the neurodegeneration in the rest of the regions innerved by them, leading to cognitive disorders.

Bisphenol A single and repeated treatment increases HDAC2, leading to cholinergic neurotransmission dysfunction and SN56 cholinergic apoptotic cell death through AChE variants overexpression and NGF/TrkA/P75NTR signaling disruption.

Bisphenol-A (BPA), a widely used plasticizer, induces cognitive dysfunctions following single and repeated exposure. Several studies, developed in hippocampus and cortex, tried to find the mechanisms that trigger and mediate these dysfunctions, but those are still not well known. Basal forebrain cholinergic neurons (BFCN) innervate hippocampus and cortex, regulating cognitive function, and their loss or the induction of cholinergic neurotransmission dysfunction leads to cognitive disabilities. However, no studies were performed in BFCN. We treated wild type or histone deacetylase (HDAC2), P75NTR or acetylcholinesterase (AChE) silenced SN56 cholinergic cells from BF with BPA (0.001 µM-100 µM) with or without recombinant nerve growth factor (NGF) and with or without acetylcholine (ACh) for one- and fourteen days in order to elucidate the mechanisms underlying these effects. BPA induced cholinergic neurotransmission disruption through reduction of ChAT activity, and produced apoptotic cell death, mediated partially through AChE-S overexpression and NGF/TrkA/P75NTR signaling dysfunction, independently of cholinergic neurotransmission disruption, following one- and fourteen days of treatment. BPA mediates these alterations, in part, through HDAC2 overexpression. These data are relevant since they may help to elucidate the neurotoxic mechanisms that trigger the cognitive disabilities induced by BPA exposure, providing a new therapeutic approach.

Chlorpyrifos induces cell proliferation in MCF-7 and MDA-MB-231 cells, through cholinergic and Wnt/ß-catenin signaling disruption, AChE-R upregulation and oxidative stress generation after single and repeated treatment.

Chlorpyrifos (CPF) biocide, is associated with breast cancer. The processes underlying this association have not been elucidated to date. CPF increases MCF-7 and MDA-MB-231 cell proliferation after acute and long-term treatment, partially through KIAA1363 overexpression and aryl-hydrocarbon receptor activation but also through estrogen receptor-alpha activation after 24 h exposure in MCF-7 cells, suggesting other mechanisms may be involved. CPF induces reactive oxygen species (ROS) generation, acetylcholine accumulation, and overexpression of acetylcholinesterase-R/S (AChE-R/S) variants, while it also alters the Wnt/ß-catenin pathway, both in vitro and in vivo, in processes different from cancer. These latter mechanisms are also linked to cell proliferation and could mediate this effect induced by CPF. Our results show that CPF (0.01-100 µM), following one-day and fourteen-days treatment, respectively, induced ROS generation and lipid peroxidation, and acetylcholine accumulation due to AChE inhibition, Wnt/ß-catenin up- or downregulation depending on the CPF treatment concentration, and AChE-R and AChE-S overexpression, with the latter being mediated through GSK-3ß activity alteration. Finally, CPF promoted cell division through ACh and ROS accumulation, AChE-R overexpression, and Wnt/ß-catenin signaling disruption. Our results provide novel information on the effect of CPF on human breast cancer cell lines that may help to explain its involvement in breast cancer.

Dysregulation of prostaglandine E2 and BDNF signaling mediated by estrogenic dysfunction induces primary hippocampal neuronal cell death after single and repeated paraquat treatment.

Paraquat (PQ) produces hippocampal neuronal cell death and cognitive dysfunctions after unique and continued exposure, but the mechanisms are not understood. Primary hippocampal wildtype or ßAPP-Tau silenced cells were co-treated with PQ with or without E2, N-acetylcysteine (NAC), NS-398 (cyclooxygenase-2 inhibitor), MF63 (PGES-1 inhibitor) and/or recombinant brain-derived neurotrophic factor (BDNF) during one- and fourteen-days to studied PQ effect on prostaglandin E2 (PGE2) and BDNF signaling and their involvement in hyperphosphorylated Tau (pTau) and amyloid-beta (Aß) protein formation, and oxidative stress generation, that lead to neuronal cell loss through estrogenic disruption, as a possible mechanism of cognitive dysfunctions produced by PQ. Our results indicate that PQ overexpressed cyclooxygenase-2 that leads to an increase of PGE2 and alters the expression of EP1-3 receptor subtypes. PQ induced also a decrease of proBDNF and mature BDNF levels and altered P75NTR and tropomyosin receptor kinase B (TrkB) expression. PQ induced PGE2 and BDNF signaling dysfunction, mediated through estrogenic disruption, leading to Aß and pTau proteins synthesis, oxidative stress generation and finally to cell death. Our research provides relevant information to explain PQ hippocampal neurotoxic effects, indicating a probable explanation of the cognitive dysfunction observed and suggests new therapeutic strategies to protect against PQ toxic effects.

Primary hippocampal estrogenic dysfunction induces synaptic proteins alteration and neuronal cell death after single and repeated paraquat exposure.

The extensively utilized herbicide Paraquat (PQ) was reported to generate cognitive disorders and hippocampal neuronal cell death after unique and extended exposure. Although, most of the mechanisms that mediate these actions remain unknown. We researched whether PQ induces synaptic protein disruption, Tau and amyloid beta protein formation, oxidative stress generation, and hippocampal neuronal cell loss through anti-estrogen action in primary hippocampal neurons, after day and two weeks PQ treatment, as a probable mechanism of such learning and memory impairment. Our results reveal that PQ did not alter estrogen receptors (ERα and ERß) gene expression, yet it decreased ER activation, which led to synaptic proteins disruption and amyloid beta proteins generation and Tau proteins hyperphosphorylation. Estrogenic signaling disruption induced by PQ also downregulated the NRF2 pathway leading to oxidative stress generation. Finally, PQ exposure induced cell death mediated by ER dysfunction partially through oxidative stress and amyloid beta proteins generation and Tau proteins hyperphosphorylation. The results presented provide a therapeutic strategy to protect against PQ toxic effects, possibly giving an explanation for the learning and memory impairment generated following PQ exposure.

Chlorpyrifos-induced cell proliferation in human breast cancer cell lines differentially mediated by estrogen and aryl hydrocarbon receptors and KIAA1363 enzyme after 24 h and 14 days exposure.

Organophosphate biocide chlorpyrifos (CPF) is involved with breast cancer. However, the mechanisms remain unknown. CPF increases cell division in MCF-7 cells, by estrogen receptor alpha (ERα) activation, although it is a weak ERα agonist, suggesting other mechanisms should be involved. Aromatic hydrocarbon receptor (AhR) activation increases cell division in human breast cancer cells, and CPF strongly activates it. Finally, the KIAA1363 enzyme, which is regulated by CPF, is overexpressed in cancer cells. Accordingly, we hypothesized that CPF or its metabolite chlorpyrifos-oxon (CPFO) could induce cell viability promotion in MCF-7 and MDA-MB-231 cell lines, through mechanisms related to ERα, AhR, and KIAA1363, after 24 h and 14 days treatment. Results show that, after acute and long-term treatment, CPF and CPFO alter differently KIAA1363, AhR, ER and cytochrome P450 isoenzyme 1A1 (CYP1A1) expression. In addition, they induced cell proliferation through ERα activation after 24 h exposure in MCF-7 cells and through KIAA1363 overexpression and AhR activation in MCF-7 and MDA-MB-231 cells after acute and long-term treatment. The results obtained in this work provide new information relative to the mechanisms involved in the CPF toxic effects that could lead to breast cancer disease.

Manganese induced ROS and AChE variants alteration leads to SN56 basal forebrain cholinergic neuronal loss after acute and long-term treatment.

Manganese (Mn) induces cognitive disorders and basal forebrain (BF) cholinergic neuronal loss, involved on learning and memory regulation, which could be the cause of such cognitive disorders. However, the mechanisms through which it induces these effects are unknown. We hypothesized that Mn could induce BF cholinergic neuronal loss through oxidative stress generation, cholinergic transmission and AChE variants alteration that could explain Mn cognitive disorders. This study shows that Mn impaired cholinergic transmission in SN56 cholinergic neurons from BF through alteration of AChE and ChAT activity and CHT expression. Moreover, Mn induces, after acute and long-term exposure, AChE variants alteration and oxidative stress generation that leaded to lipid peroxidation and protein oxidation. Finally, Mn induces cell death on SN56 cholinergic neurons and this effect is independent of cholinergic transmission alteration, but was mediated partially by oxidative stress generation and AChE variants alteration. Our results provide new understanding of the mechanisms contributing to the harmful effects of Mn on cholinergic neurons and their possible involvement in cognitive disorders induced by Mn.

Cadmium alters heat shock protein pathways in SN56 cholinergic neurons, leading to Aß and phosphorylated Tau protein generation and cell death.

Cadmium, a neurotoxic environmental compound, produces cognitive disorders, although the mechanism remains unknown. Cadmium induces a more pronounced cell death on cholinergic neurons from basal forebrain (BF), mediated, in part, by increase in Aß and total and phosphorylated Tau protein levels, which may explain cadmium effects on learning and memory processes. Cadmium downregulates the expression of heat shock proteins (HSPs) HSP 90, HSP70 and HSP27, and of HSF1, the master regulator of the HSP pathway. HSPs proteins reduce the production of Aß and phosphorylated Tau proteins and avoid cell death pathways induction. Thus, we hypothesized that cadmium induced the production of Aß and Tau proteins by HSP pathway disruption through HSF1 expression alteration, leading to BF cholinergic neurons cell death. Our results show that cadmium downregulates HSF1, leading to HSP90, HSP70 and HSP27 gene expression downregulation in BF SN56 cholinergic neurons. In addition, cadmium induced Aß and total and phosphorylated Tau proteins generation, mediated partially by HSP90, HSP70 and HSP27 disruption, leading to cell death. These results provide new understanding of the mechanisms contributing to cadmium harmful effects on cholinergic neurons.

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.

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.

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.

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.