Persistent diazinon induced neurotoxicity: The effect on inhibitory avoidance memory performance, amyloid precursor proteins, and TNF-α levels in the prefrontal cortex of rats.

INTRODUCTION: Organophosphate pesticides (Ops) like diazinon (DZN) have well-known neurotoxic effects and low-level chronic exposure has been linked to detrimental neurobehavioral impairments and memory deficits. However, it's not entirely clear how DZN-induced biological changes, particularly in the prefrontal cortex (PFC) contribute to these effects. The purpose of this study is to investigate the impact of DZN exposure on inhibitory avoidance (IA) memory function, amyloid precursor expression (APP), and proinflammatory tumor necrosis factor-α (TNF-α) levels in the rat cortex. MATERIALS AND METHODS: Rats were divided into 4 groups and recived 2 mg/kg DZN for 5-days or 12-weeks and two control groups recived the same volume of vehicle. IA memory was assesed using the shuttle box apparatus. Rats were sacrificed and the prefrontal cortex PFC were removed. Real-time PCR and Western blotting were used to messure TNF-α, and amyloid protein precursors gene expression and protein levels. RESULTS: Our findings indicated that DZN caused body weight loss and a notable decline in performance on the IA memory. Additionally, 5-days exposure increased APP and APLP2 protein levels in the PFC, while 12-weeks exposure decreased these levels. Furthermore, expression of APP and APLP2 gens were decreased in PFC. TNF-α levels increased as a result of 5-days exposure to DZN, but these levels dropped to normal after 12-weeks administration, and this observation was significant. CONCLUSION: Taken together, exposure to low doses of DZN leads to disturbances in IA memory performance and also alternations in amyloid beta precursors that can be related to increased risk of Alzheimer's disease.

Metabolomic analysis of the serum and urine of rats exposed to diazinon, dimethoate, and cypermethrin alone or in combination.

BACKGROUND: Multiple pesticides are often used in combination for plant protection and public health. Therefore, it is important to analyze the physiological changes induced by multiple pesticides exposure. The objective of this study was to investigate the combined toxicity of the widely-used organophosphorus and pyrethroid pesticides diazinon, dimethoate, and cypermethrin. METHODS: Male Wistar rats were administrated by gavage once daily with the three pesticides individual or in combination for consecutive 28 days. The metabolic components of serum and urine samples were detected by using 1H nuclear magnetic resonance (NMR)-based metabolomics method. Histopathological examination of liver and kidneys and serum biochemical determination were also carried out. RESULTS: The results showed that after the 28-day subacute exposure, serum glutamic transaminase and albumin were significantly increased and blood urea nitrogen was significantly decreased in the rats exposed to the mixture of the pesticides compared with the control rats, suggesting that the co-exposure impaired liver and kidney function. Metabolomics analysis indicated that the indicators 14 metabolites were statistically significant altered in the rats after the exposure of the pesticides. The increase in 3-hydroxybutyric acid in urine or decrease of lactate and N-acetyl-L-cysteine in serum could be a potentially sensitive biomarker of the subchronic combined effects of the three insecticides. The reduction level of 2-oxoglutarate and creatinine in urine may be indicative of dysfunction of liver and kidneys. CONCLUSION: In summary, the exposure of rats to pesticides diazinon, dimethoate, and cypermethrin could cause disorder of lipid and amino acid metabolism, induction of oxidative stress, and dysfunction of liver and kidneys, which contributes to the understanding of combined toxic effects of the pesticides revealed by using the metabolomics analysis of the urine and serum profiles.

Using heat shock protein (HSP) inducers as an approach to increase the viability of sterlet (Pisces; Acipenseridae; Acipenser ruthenus) cells against environmental diazinon toxicity.

Diazinon (DZN) is an organophosphate pesticide frequently used in agriculture and released into aquatic environments. In this study, sterlet sturgeon cells were exposed to DZN to investigate possible defense mechanisms via HSP induction (HSPi). Liver, kidney, and gill cells of Acipenser ruthenus were isolated and cultured and then treated with HSPi (Pro-Tex®, amygdalin, and a novel pirano-piranazole-based synthesized compound: SZ) in the presence and absence of DZN. MTT assays were used to evaluate the effects of different HSPis and their combinations with DZN. Western blotting analysis was conducted to evaluate HSP27, HSP70, and HSP90 expression patterns in each group. The highest rates of caspase-3 and caspase-8 activities were found in the DZN group, whereas HSPi treatment resulted in the lowest rates. The combination of HSPi+DZN resulted in increased HSP levels and antioxidant parameters but decreased cortisol, immune parameters, and metabolic enzymes. Many of the studied parameters (caspases, acetylcholinesterase, antioxidant, immune, and metabolic parameters) showed significant correlations with HSP expression, indicating that HSPs may be associated with markers of sterlet cell health. The results of this study demonstrate that using HSP inducers may be a powerful and reliable way to increase A. ruthenus resistance prior to exposure to DZN.

The novel hepatoprotective effects of silibinin-loaded nanostructured lipid carriers against diazinon-induced liver injuries in male mice.

In the current study, silibinin-loaded nanostructured lipid carriers (Sili-NLCs) was synthesized, and the hepatoprotective effectiveness of Sili-NLCs against diazinon (DZN)-induced liver damage in male mice was evaluated. The emulsification-solvent evaporation technique was applied to prepare Sili-NLCs, and characterized by using particle size, zeta potential, entrapment efficacy (EE %), in vitro drug release behavior, and stability studies. In vivo, studies were done on male mice. Hepatotoxicity in male mice were induced by DZN (10 mg/kg/day, i.p.). Four groups treated with silibinin and Sili-NLCs with the same doses (100 and 200 mg/kg, p.o.). On 31th days, serum and liver tissue samples were collected. Alanine (ALT) and aspartate (AST) aminotransferase levels, oxidative stress biomarkers, inflammatory cytokines, and histopathological alterations were assessed. The Sili-NLCs particle size, zeta potential, polydispersity index (PDI), and EE % were obtained at 220.8 ± 0.86 nm, -18.7 ± 0.28 mV, 0.118 ± 0.03, and 71.83 ± 0.15%, respectively. The in vivo studies revealed that DZN significantly increased the serum levels of AST, ALT, hepatic levels of lipid peroxidation (LPO), and tumor necrosis factor-α (TNF-α), while decreased the antioxidant defense system in the mice's liver. However, Sili-NLCs was more effective than silibinin to return the aforementioned ratio toward the normal situation, and these results were well correlated with histopathological findings. Improvement of silibinin protective efficacy and oral bioavailability by using NLCs caused to Sili-NLCs can be superior to free silibinin in ameliorating DZN-induced hepatotoxicity in male mice.

Malathion or diazinon exposure and male reproductive toxicity: a systematic review of studies performed with rodents.

Malathion and diazinon are pesticides commonly used in agriculture to avoid insects that damage crops; however, they may cause impairment to the male genital system of exposed humans. The present work carried out a systematic review of the literature concerning the primary studies that assessed the reproductive effects resulting from male rats and mice exposed to malathion or diazinon. The search for articles was performed on the databases PubMed, LILACS, Scopus, and SciELO, using different combinations of the search terms "malathion," "diazinon," "mice," "rats," "male reproduction," "fertility," and "sperm," followed by the Boolean operators AND or OR. The results obtained indicate that both pesticides act as reproductive toxicants by reducing sperm quality, diminishing hormonal concentrations, inducing increased oxidative stress, and provoking histopathological damage in reproductive organs. Then, the exposure to malathion and diazinon may provoke diminished levels of testosterone by increasing acetylcholine stimulation in the testis through muscarinic receptors, thus, providing a reduction in steroidogenic activity in Leydig cells, whose effect is related to lower levels of testosterone in rodents, and consequently, it is associated with decreased fertility. Considering the toxic effects on the male genital system of rodents and the possible male reproductive toxicity in humans, it is recommended the decreased use of these pesticides and their replacement for others that show no or few toxic effects for non-target animals.

Serum butyrylcholinesterase activity in healthy dogs with and without exposure to diazinon.

BACKGROUND: Cholinesterase is a biomarker for poisonings by anticholinesterase agents, but its reference values are scarce, and possible interaction with collars containing parasiticides has not been studied. OBJECTIVES: We aimed to evaluate the serum cholinesterase activity of healthy dogs without a history of contact with anticholinesterase agents and healthy animals exposed to commercial collars containing organophosphate. METHODS: Ninety-nine dogs were used and included healthy animals without recent exposure to anticholinesterase agents and healthy animals previously exposed to diazinon collars. Serum quantification of the enzyme butyrylcholinesterase (BuchE) through spectrophotometry was conducted on all samples. In experiment 1, BuchE activity was quantified at time 0 and 7 days after, a time when the samples were kept at -18°C. In experiment 2, sampling times were 0, 7, 14, 21, 28, and 56 days. RESULTS: Time 0 values were 4622.38 ± 1311.53 U/L. After 7 days, a significant decay was observed, with a mean of 3934.45 ± 1430.45 U/L. Spearman's test was performed, finding a weak correlation between ALT, creatinine, total plasma proteins, age, weight, red blood cells, platelets, leukocytes, and BuchE activities. In experiment 2, the mean at time 0 was 4753 ± 454.8 U/L. With exposure to the collar, there was a decay of up to 93% after 14 days. CONCLUSIONS: Normality values of serum BuchE in healthy dogs without a history of exposure to anticholinesterase agents were 4360.8-4883.96 U/L. Freezing serum caused a decrease in BuchE activity. Exposure to commercial collars containing diazinon also reduced BuchE activity without clinical signs, indicating that previously exposed animals should be evaluated carefully.

Protective effects of HSP Inducer on diazinon-exposed stellate sturgeon (Acipenser stellatus) fry: Insights on HSP70 gene expression, immune response, and enzyme indices.

Aquatic environments face frequent exposure to organophosphate pesticides, such as diazinon, which are frequently utilized in agriculture. The goal of this study was to evaluate the effects of diazinon exposure on fish and to investigate the potential of the HSP inducer (HSPi) in developing a defense mechanism. To achieve this, several factors were analyzed, including the HSP70 gene expression, levels of immunity markers (lysozyme, IgM, and C3), antioxidant status, and the activity of acetylcholine esterase (AChE). Stellate sturgeon (Acipenser stellatus) fry, was exposed to diazinon (25, 50, and 75% of 96h-LC50) for 6 days after pre-treatment with an HSP inducer (HSPi), TEX-OE® (a prickly pear cactus extract), for 4 hours. Two HSPi concentrations, 100 and 200 mg.L-1, were used. Pre-treatment with HSPi significantly enhanced HSP70 gene expression in the gill and liver, as well as immune markers in the blood of Acipenser stellatus. Diazinon-treated groups exhibited higher antioxidant activities of SOD, CAT, and T-AOC. Increased activity also observed in control fish pre-treated with HSPi. However, stellate sturgeon receiving both diazinon and HSPi+diazinon experienced a significant decrease in AChE activity in comparison with control group. Cortisol levels were elevated in the fish that were subjected to diazinon. Those subjected to diazinon after receiving HSPi showed a significant decrease in cortisol levels. In conclusion, the study suggests that HSPi-mediated HSP70 induction may have a protective effect. The presence of an HSP inducer offers a potential strategy to mitigate the consequences of diazinon exposure in stellate sturgeon.

24-Epibrassinolide alleviates diazinon oxidative damage by escalating activities of antioxidant defense systems in maize plants.

Excessive use of pesticides against pests has contaminated agricultural crops and raised global concerns about food safety. This research investigates the alleviation effects of 24-epibrassinolide (EBL) seed priming on diazinon (DZ) pesticide toxicity. The experiment was conducted with eight groups including control, DZ, EBL (10 µM), EBL (0.1 µM), EBL (0.01 µM), EBL (10 µM) + DZ, EBL (0.1 µM) + DZ, and EBL (0.01 µM) + DZ. Plants grown with the lowest concentration of EBL (0.01 µM) exhibited an upward increase in the activity of SOD, CAT, POD, APX, GR, and GST enzymes under DZ toxicity stress. In contrast, higher concentrations of EBL showed some inhibitory effects on the activity of antioxidant enzymes. In addition, low concentrations of EBL elevated the free radical scavenging capacity (DPPH), iron-reducing antioxidant power (FRAP), photosynthesis rate (Pn), stomatal conductance (Gs) and proline, and protein contents. EBL also reduced lipid peroxidation (MDA levels) in the DZ-exposed plants, leading to membrane integrity. The favorable effects of EBL were more evident when plants were exposed to pesticides than normal growth conditions. The results indicated that EBL seed priming intensifies the antioxidant enzymes system activity, and helps maize plants against toxic effects of DZ under proper concentration.

Ferulago angulata extract alleviates testicular toxicity in male mice exposed to diazinon and lead.

In this study, we investigated the protective effects of Ferulago angulata extract (FAE) against the reproductive toxicants Diazinon (DZN) and Lead (Pb) in mice. These pollutants are known to induce oxidative stress (OS), while FAE acts as a natural antioxidant. Adult male NMRI mice were exposed to DZN, Pb, and DZN+Pb, with or without FAE treatment for six weeks. We evaluated OS markers, testicular histology, and expression of mRNA related to enzymatic antioxidants. Exposure to DZN and Pb led to increased levels of thiobarbituric acid reactive substance (TBARS) and nitric oxide (NO) in the testes, along with a decrease in the total antioxidant capacity (TAC). Furthermore, the mRNA expression of antioxidant enzymes such as superoxide dismutase 1 (SOD1) and glutathione peroxidase 4 (GPX4) was altered. However, when FAE was administered concurrently, it restored the biochemical parameters to normal levels, reduced the toxic effects of DZN and Pb, and provided protection against testicular histopathological injury. These findings suggest that FAE has the potential to serve as a protective agent against oxidative damage caused by contaminants in reproductive organs, specifically in the testes.

Ecotoxicity of Diazinon and Atrazine Mixtures after Ozonation Catalyzed by Na+ and Fe2+ Exchanged Montmorillonites on Lemna minor.

The toxicity of two pesticides, diazinon (DAZ) and atrazine (ATR), before and after montmorillonite-catalyzed ozonation was comparatively investigated on the duckweed Lemna minor. The results allowed demonstrating the role of clay-containing media in the evolution in time of pesticide negative impact on L. minor plants. Pesticides conversion exceeded 94% after 30 min of ozonation in the presence of both Na+ and Fe2+ exchanged montmorillonites. Toxicity testing using L. minor permitted us to evaluate the change in pesticide ecotoxicity. The plant growth inhibition involved excessive oxidative stress depending on the pesticide concentration, molecular structure, and degradation degree. Pesticide adsorption and/or conversion by ozonation on clay surfaces significantly reduced the toxicity towards L. minor plants, more particularly in the presence of Fe(II)-exchanged montmorillonite. The results showed a strong correlation between the pesticide toxicity towards L. minor and the level of reactive oxygen species, which was found to depend on the catalytic activity of the clay minerals, pesticide exposure time to ozone, and formation of harmful derivatives. These findings open promising prospects for developing a method to monitor pesticide ecotoxicity according to clay-containing host-media and exposure time to ambient factors.

The relationship between the cholinergic mechanism of toxicity and oxidative stress in rats during subacute diazinon poisoning.

Diazinon is an organophosphate pesticide (OP) that has significant potential for accidental and intentional poisoning of wildlife, domestic animals and humans. The aim of the study is to investigate the correlation between cholinesterase activity and oxidative stress parameters in liver and diaphragm by continuous monitoring as a function of time during prolonged use of diazinon. Wistar rats were treated orally with diazinon (55 mg/kg/day): 7, 14, 21 and 28 days. At the end of each period, blood, liver and diaphragm were collected to examine cholinesterase activity and enzymatic/non-enzymatic oxidative stress parameters: superoxide dismutase 1 (SOD1), catalase (CAT), thiobarbituric acid substances (TBARS), protein carbonyl groups. In all four time periods, there was a significant change in acetylcholinesterase (AChE) in erythrocytes and butyrylcholinesterase (BuChE) in blood plasma, CAT in liver and diaphragm and SOD1 in diaphragm. Parameters significantly altered during the cholinergic crisis included: cholinesterases and TBARS in liver and diaphragm and partially SOD1 in liver. Protein carbonyl groups in liver and diaphragm were significantly altered outside the cholinergic crisis. In the liver, there was a very strong negative correlation between BuChE and TBARS in all four time periods and BuChE and CAT on day 7. In the diaphragm, a very strong negative correlation was found between AChE and TBARS at days 7 and 14, and a very strong positive correlation between AChE and SOD1 at days 14, 21 and 28. A better understanding of the relationship between cholinergic overstimulation and oxidative stress may help to better assess health status in prolonged OPs intoxication.

The Amphibian Short-Term Assay: Evaluation of a New Ecotoxicological Method for Amphibians Using Two Organophosphate Pesticides Commonly Found in Nature-Assessment of Behavioral Traits.

Neurotoxic pesticides are used worldwide to protect crops from insects; they are recognized to impact nontarget organisms that live in areas surrounded by treated crops. Many biochemical and cell-based solutions have been developed for testing insecticide neurotoxicity. Nevertheless, such solutions provide a partial assessment of the impact of neurotoxicity, neglecting important phenotypic components such as behavior. Behavior is the apical endpoint altered by neurotoxicity, and scientists are increasingly recommending including behavioral endpoints in available tests or developing new methods for assessing contaminant-induced behavioral changes. In the present study, we extended an existing protocol (the amphibian short-term assay) with a behavioral test. To this purpose, we developed a homemade device along with an open-source computing solution for tracking trajectories of Xenopus laevis tadpoles exposed to two organophosphates insecticides (OPIs), diazinon (DZN) and chlorpyrifos (CPF). The data resulting from the tracking were then analyzed, and the impact of exposure to DZN and CPF was tested on speed- and direction-related components. Our results demonstrate weak impacts of DZN on the behavioral components, while CPF demonstrated strong effects, notably on speed-related components. Our results also suggest a time-dependent alteration of behavior by CPF, with the highest impacts at day 6 and an absence of impact at day 8. Although only two OPIs were tested, we argue that our solution coupled with biochemical biomarkers is promising for testing the neurotoxicity of this pesticide group on amphibians. Environ Toxicol Chem 2023;42:1595-1606. © 2023 The Authors. Environmental Toxicology and Chemistry published by Wiley Periodicals LLC on behalf of SETAC.

Genomic organization and transcription of superoxide dismutase genes (sod1, sod2, and sod3b) and response to diazinon toxicity in platyfish (Xiphophorus maculatus) by using SOD enzyme activity.

The aim of this study is to determine the effects of 50% of 96 h LC50 (5.25 ppm) diazinon on the expression of superoxide dismutase (SOD) enzyme genes (sod1, sod2, and sod3b) and SOD enzyme activity at the end of 24, 48, 72, and 96 h in platyfish liver and gill tissues. To this end, we determined the tissue-specific distribution of sod1, sod2, and sod3b genes and performed in silico analyses in platyfish (Xiphophorus maculatus). It was determined that malondialdehyde (MDA) level and SOD enzyme activity were increased in the liver [(43.90 EU mg protein-1 (control), 62.45 EU mg protein-1 (24 h), 73.17 EU mg protein-1 (48 h), 82.18 EU mg protein-1 (72 h), 92.93 EU mg protein-1 (96 h)] and gill [(16.44 EU mg protein-1 (control), 33.47 EU mg protein-1 (24 h), 50.38 EU mg protein-1 (48 h), 64.62 EU mg protein-1 (72 h), 74.04 EU mg protein-1 (96 h)] tissues of platyfish exposed to diazinon, while the expression of the sod genes was down-regulated. The tissue-specific distribution of the sod genes varied, with the tissues and the sod genes expression were being predominant in the liver (628.32 in sod1, 637.59 in sod2, 888.5 in sod3b). Thus, the liver was considered a suitable tissue for further gene expression studies. Based on the phylogenetic analyses, platyfish sod genes can be reported to be orthologs of sod/SOD genes from other vertebrates. Identity/similarity analyses supported this determination. Conserved gene synteny proved that there are conserved sod genes in platyfish, zebrafish, and humans.

Anodonta cygnea, a freshwater swan mussel, exposed to diazinon: toxicity thresholds in behaviour and physiology.

Swan mussels (Anodonta cygnea) have been suggested as suitable bioindicators for the presence of pollutants in the environment. Application of the physiological and behavioral markers in these sessile species can be beneficial for environmental monitoring. The present study aimed to investigate the relationship between the behavioral disorders of movement and siphoning associated with the inhibition of tissue Acetylcholinesterase (AChE). For experiments, overally 120 bivalves of Anodonta cygnea (mean total length 80.33 ± 6.7 mm) were transported from the agricultural drains and canals in Sari county (Mazandaran Province, Iran) to our laboratory. First, the LC50-96 h of diazinon was estimated according to the Organization for Economic Co-operation and Development (OECD 1992) guideline with static water conditions. The sub-lethal toxicity pesticide experiments were conducted on the basis of the lowest observed effect concentration (LOEC) and the maximum acceptable toxicant concentration (MATC). The LC50-96 h, LOEC, and MATC values of diazinon were 85.2, 42.1, and 8.5 mg L- 1, respectively. Based on the observations of mussels' movement, the burrowing and displacement decreased with the concentration of toxicant in water. Moreover, the presence of diazinon in water and its exposure to experimental animals significantly reduces their siphoning rate. The RDA showed that the AChE activity had a higher correlation with the siphoning behavior than the movement behavior. The comparison of enzyme activity at different exposure and recovery times showed that there was a significant difference among the groups affected by the consumed pesticide (p = 0.001, between contrasts). The most remarkable morphometric characteristic was the siphon opening that was inversely correlated with the enzymatic activity. Studies in bioethics might benefit from paying attention to these traits that are directly related to the level of toxicity and behavioral adaptations required for animal survival.

The efficacy of 2-formyl benzoic acid in reactivating diazinon inhibited murine cholinesterase.

Oximes, as classical acetylcholinesterase (AChE) reactivators, have some pharmacokinetics/pharmacodynamics disadvantages. During the synthesis of non-oxime compounds, we encountered the compound 2-formylbenzoic acid (2-FBA) with promising in vitro and in vivo cholinesterase (ChE) reactivating properties in the acute exposure to diazinon (DZN). For in vitro experiments, the healthy mice serum and brain homogenate were freshly prepared and exposed to DZN (160 µg/mL). After 10 minutes, 2-FBA was added to the poisoned samples, and ChE activity was measured afterward. For the in vivo assay, the mice were poisoned with DZN subcutaneous (SC) injection (50 mg/kg), and after 1 hour, either 2-FBA or Pralidoxime (2-PAM) was injected intravenously (IV). After 3 h, ChE activity was measured in the serum and brain homogenate samples. The LD50 (IV) for 2-FBA in mice was measured as well. 2-FBA effectively reactivated the inhibited ChE in serum and brain homogenate samples in vitro. In the in vivo experiments, while 2-FBA could significantly reactivate the brain ChE even better than 2-PAM, they failed to reactivate the serum ChE by single IV injection. LD50 of 2-FBA was calculated to be 963 mg/kg. There were no general toxicity signs in any treatment groups. The in silico results support the potential ability of 2-FBA efficacy via possibly Witting reaction mechanism. Our findings indicate that 2-FBA seems to be a suitable non-oxime candidate for AChE reactivation with minimal side effects. Further toxicokinetic studies on this compound are strongly recommended to be performed before conducting the clinical trial in humans.

Effects of maternal diazinon exposure on frontal cerebral cortical development in mouse embryo.

BACKGROUND: Diazinon including organophosphate (OP) that is widely used in agriculture and animal husbandry industry and the risk of human infection with the toxin and their toxicity. METHODS: Pregnant balb/c mice (30-35 g) were randomly divided into five groups of five: the control group (no intervention), two sham groups (emulsifier 0.52, and 5.2 liters/volume). From the seventh to the eighteenth day of pregnancy, two experimental groups received diazinon inhaled 1.3 (EXP1) and 13 liters/volume (EXP2) for 40 min every other day, respectively. On the 18th day of pregnancy, the animals were killed and their embryos were removed to appraisal the growth of fetus and development of the frontal cortex. A computer-assisted morphometric quantitative images analysis were performed on the frontal cerebral cortex (FCC) including cortical plate (CP), intermediate zone (IZ) and matrix (proliferative) zone (MZ) of the mouse embryos. FINDINGS: The average of crown-rump length and weight of the embryos in the experimental groups were increased without any significant difference. The mean fetal FCC thickness in the EXP2 group was significantly reduced compared to the control group, CP thickness was remarkably increased in fetuses exposed to diazinon. Comparing the mean thickness of MZ and IZ in EXP groups with the sham and control groups indicated a significant decrease. The positive K-67 cells in the FCC of the EXP2 group were significantly reduced. DISCUSSION: Exposing diazinon during pregnancy can reduce brain development and would be neurotoxic to the developing brain and can lead to behavioral changes in the offspring.

Organophosphate insecticides disturb neuronal network development and function via non-AChE mediated mechanisms.

Exposure to organophosphate (OP) insecticides has been related to several adverse health effects, including neurotoxicity. The primary insecticidal mode of action of OP insecticides relies on (irreversible) binding to acetylcholine esterase (AChE), with -oxon metabolites having a much higher potency for AChE inhibition than the parent compounds. However, OP insecticides can also have non-AChE-mediated effects, including changes in gene expression, neuroendocrine effects, disruption of neurite outgrowth and disturbance of the intracellular calcium (Ca2+) homeostasis. Since Ca2+ is involved in neurotransmission and neuronal development, our research aimed to assess the effects of two widely used OP insecticides, chlorpyrifos (CPF) and diazinon (DZ) and their respective -oxon metabolites, on intracellular Ca2+ homeostasis in human SH-SY5Y cells and rat primary cortical cultures. Furthermore, we assessed the acute and chronic effects of exposure to these compounds on neuronal network maturation and function in rat primary cortical cultures using microelectrode array (MEA) recordings. While inhibition of AChE appears to be the primary mode of action of oxon-metabolites, our data indicate that both parent OP insecticides (CPF and DZ) inhibit depolarization-evoked Ca2+ influx and neuronal activity at concentrations far below their sensitivity for AChE inhibition, indicating that inhibition of voltage-gated calcium channels is a common mode of action of OP insecticides. Notably, parent compounds were more potent than their oxon metabolites, with exposure to diazinon-oxon (DZO) having no effect on both neuronal activity and Ca2+ influx. Human SH-SY5Y cells were more sensitive to OP-induced inhibition of depolarization-evoked Ca2+ influx than rat cortical cells. Acute exposure to OP insecticides had more potent effects on neuronal activity than on Ca2+ influx, suggesting that neuronal activity parameters are especially sensitive to OP exposure. Interestingly, the effects of DZ and chlorpyrifos-oxon (CPO) on neuronal activity lessened after 48 h of exposure, while the potency of CPF did not differ over time. This suggests that neurotoxicity after exposure to different OPs has different effects over time and occurs at levels that are close to human exposure levels. In line with these results, chronic exposure to CPF during 10 days impaired neuronal network development, illustrating the need to investigate possible links between early-life OP exposure and neurodevelopmental disorders in children and highlighting the importance of non-AChE mediated mechanisms of neurotoxicity after OP exposure.

Epigallocatechin gallate improves meiosis maturation against Diazinon exposure in porcine oocytes.

Diazinon (DZN) is a refractory organophosphorus pesticide (OP) in the surrounding environment due to its overuse in agriculture. The antioxidant activity of Epigallocatechin gallate (EGCG) from green tea is at least 100 times greater than that of vitamin C. This study aimed to study the effects of DZN on the meiotic maturation of porcine oocytes, as well as the protective roles of EGCG. Firstly, the effects of DZN and EGCG on meiotic nuclear maturation of porcine oocytes were detected, and then embryonic development was investigated by chemical parthenogenetic activation. Next, the spindle assembly, mitochondrial membrane potential (MMP), reactive oxygen species (ROS), DNA damage, and finally the early apoptosis of oocytes were examined by immunofluorescence staining. The results revealed that DZN exposure significantly reduced the quality of porcine oocytes, such as failure of nuclear and cytoplasmic maturation, evidenced by abnormal spindle assembly, disordered chromosome alignment, low MMP, observably increased ROS, severe DNA damage, and early apoptosis. Appropriate EGCG could significantly reduce all these defects caused by DZN. In conclusion, EGCG can help prevent the harm that DZN exposure can do. These findings offer convincing support for enhancing the oocyte quality from EGCG through daily ordinary beverages.

Protective effects of Biebersteinia multifida on sub-chronic toxicity of DZN in male Wistar rats: biochemical, hematological, and oxidative stress indices.

The protective effect of Biebersteinia Multifida on diazinon-induced toxicity in male Wistar rats was investigated over 8 weeks. Impacts of diazinon (10 mg/kg daily), Biebersteinia Multifida (500 mg/kg daily), and coadministration of them on oxidative stress parameters besides hematological and biochemical indices were assessed in various groups. The gas chromatography-mass spectrometry analysis was performed to identify the antioxidant components of plant extract by comparing the mass spectra and retention indices with those given in the literature. Pseudocholinesterase level demonstrated a significant attenuation in the Biebersteinia Multifida+diazinon-treated group in comparison to the diazinon group at the end of the 8th week. Statistical significant differences in hematological and biochemical indices were detectable when the diazinon group was compared to Biebersteinia Multifida+diazinon-treated rats. While diazinon destroyed hepatic and renal functions, Biebersteinia Multifida protected the liver and kidney from diazinon toxic effects by normalizing related function indices at the end of the 8th week. By diminishing malondialdehyde and enhancing the ferric-reducing power, Biebersteinia Multifida minimized the hazardous effect of diazinon-induced oxidative stress. Following these results, the beneficial effects of Biebersteinia Multifida in reducing the toxicity of diazinon should be taken into consideration.

The significance of thymoquinone administration on liver toxicity of diazinon and cholinesterase activity; a recommendation for prophylaxis among individuals at risk.

BACKGROUND: Diazinon (DZN), a widely used chemical herbicide for controlling agricultural pests, is an important organophosphorus pesticide and an environmental pollutant which induces toxic effects on living organisms during long-term exposure. Thymoquinone (TQ) is a phytochemical bioactive compound with antioxidant and anti-inflammatory properties. We aimed to evaluate the protective effects of TQ against DZN-induced hepatotoxicity through alleviating oxidative stress and enhancing cholinesterase (ChE) enzyme activity. METHODS: Rats were randomly divided into six groups (n = 8); a negative control group receiving corn oil; a group only receiving DZN (20 mg/kg/day); a group treated with TQ (10 mg/kg/day), and three treatment groups as TQ + DZN, receiving different doses of TQ (2.5, 5, and 10 mg/kg/day). All experimental animals were orally treated for 28 consecutive days. The levels of superoxide dismutase (SOD), glutathione (GSH), malondialdehyde (MDA), alanine transaminase (ALT), aspartate aminotransferase (AST), alkaline phosphatase (ALP), and lactic acid dehydrogenase (LDH) were determined. In addition, ChE activity and histopathological changes were evaluated. RESULTS: The results showed that DZN decreased GSH level (p < 0.01) and SOD activity (p < 0.01) in parallel to an increase in MDA level (p < 0.01) and increased the activity of AST, ALT, ALP, and LDH (p < 0.01) in comparison to the negative control group. Our findings demonstrated that TQ administration could diminish hepatotoxicity and reduce oxidative damage in DZN-treated rats, which could be linked to its antioxidant and free radical scavenging properties. It was also observed that TQ 10 mg/kg remarkably increased the activity of acetylcholinesterase, butyrylcholinesterase, and SOD enzymes, elevated GSH, decreased MDA, and reduced pathological alternations of the liver induced by DZN. CONCLUSION: Thymoquinone 10 mg/kg increased the activity of plasma and blood cholinesterases and reduced DZN-induced alternations of the liver. Improvement of butyryl- and acetylcholinesterase activity suggests that maybe TQ supplement could be beneficial as pre-exposure prophylaxis among farm workers spraying pesticides.