Pesticides were economically important chemicals in agriculture. Their use has permitted agricultural progress, through the eradication of harmful insect and the fight against vectors of disease. However, several studies question the beneficial effects of organophosphorus compounds, showing that their deregulated use causes various problems of environmental pollution and human health. The present study shows that chronic exposure to a subtoxic dose of dimethoate is likely to affect cognitive and behavioral functions of rats (both males and females). Our results show that exposure to dimethoate affects both short and long-term memory capacities. The short-term memory results are more pronounced. Treatment with nettle extract allowed a significant improvement in cognitive and behavioral performance of the rats after their exposure to dimethoate.
It was in the 1940s that the first synthetic pesticides appeared on the market with very positive results in increasing agricultural yields. Twenty years later, the first accusations of harm to people’s health and the environment were heard [
The debate about the risks and benefits of chemical control has since been extended and a great deal of research has been carried out to better understand the impact of pesticides on the environment.
The mechanism of acute neurotoxic action of Dimethoate is typically cholinergic. It involves inhibition of Acetylcholinesterase (AChE) of neuronal tissue through its active metabolic form; Omethoate which is 10 times more toxic than dimethoate (WHO, 2003).
Omethoate rapidly binds to the hydroxyl group of the active site of AChE, and undergoes a double displacement reaction involving the hydroxyl-serum and dimethylphosphoryl groups of AChE. Thus, phosphorylated AChE is stable and irreversible.
Inhibition of AChE causes an accumulation of acetylcholine released in the synaptic cleft upon nerve stimulation, leading to hyperstimulation of the nicotinic and muscarinic receptors. Consequently, the passage of nerve information is disrupted [
In this study, we will highlight the effect of sub-chronic intoxication by dimethoate on short-term and long-term memory. The poisoning took place on male and female Wistar rats. These intoxicated rats will undergo a trial of treatment by phytochelation. Extract of a medicinal plant, nettle, was obtained by decoction.
Chelation therapy has started as a branch of medicine to remove heavy metals and other toxins from the body by introducing powerful chemical agents that associate with toxins and are then excreted in the urine.
From the point of view of a medical use, several clinical trials carried out in this context confirmed these pharmacological properties in humans.
Urtica dioica L., is a ubiquitous plant of the Urticaceae family, renowned for its unpleasant irritating effects, the nettle is actually rich in vitamins, minerals and numerous medicinal virtues [
Indeed, put into practice in vitro and in vivo in animals, these studies have proved that this plant possesses numerous pharmacological properties in terms of antiproliferative, anti-inflammatory, antioxidant, analgesic, antiulcer, immunostimulating, anti-Infectious, anti-hypertensive and protective against cardiovascular disease. In addition, and in view of its richness in protein, minerals and vitamins, nettle provides a proven nutritional interest. In terms of toxicology, nettle remains innocuous and significant doses, administered orally to humans, showed no side effects.
Nettle extracts have a neutralizing role for reactive oxygen species (ERO). Their anti-free radical activity, with respect to the superoxide anion O2˚, the hydroxyl radical OH˚ and the nitric oxide radical NO˚, was determined by spectrophotometry. Numerous studies have shown that the methanolic and ethanolic extracts of the leaves have a remarkable antioxidant effect with respect to the 1,1-diphenyl-2-picrylhydrazyl radical (DPPH) [
Nettle is very rich in polyphenols and is located in different parts of plants according to the plant species and the polyphenolic group considered. These compounds group together a multitude of molecules and represent one of the most important groups present in the plant kingdom. Although they are very diversified, they all have in common the presence of one or more benzene rings carrying one or more hydroxyl functions. Polyphenols are divided into several categories, including flavonoids [SFA, 2005].
Flavonoids act mainly as primary antioxidants, stabilizing peroxide radicals, but they can also deactivate reactive oxygen species, inhibit lipoxygenase or chelate metals [
Flavonoids are in vitro enzyme inhibitors of: histidine decarboxylase (by queretol and naringenin), elastase, hyaluronidase (by flavones and proanthocyanidols) and catechol-O-methyltransferase.
The animals used in this study are wistar albino rats (male and female), and are obtained from the Faculty of Science animal house. The animals are reared in transparent plastic cages (32 × 15 × 13 cm), covered by a stainless steel grill. The breeding takes place under standard conditions of ventilation, light (cycle of alternation 12H light/12H darkness), temperature (21 ± 2 degrees C) and relative humidity (65% ± 15%). Rats have ad libitum access to fresh water and secalin-based food (SNV, Temara, Morocco).
In the adult stage, 120 Wistar rats aged 3 months of average weight (160 ± 40 g) are divided into two groups according to sex, 60 males and 60 females. Since the lethal dose of oral dimethoate was 300 mg/kg bw in rats [
After the period of intoxication and behavioral evaluation, rats (intoxicated by gavage or drinking water) received an extract by decoction of nettle. Indeed, 40 g of fresh leaves of Nettle have been roughly cut. After soaking in water for 24 hours, the whole is brought to boiling for 20 to 30 minutes under lid. After cooling the decoction under lid, filtering is carried out. This filtrate can be kept for only a few hours. A dose of 1 ml/100mg was administered orally to each of the intoxicated animals and this for duration of one month. The results of the treatment are finally compared with those of the control and intoxicated rats.
Signs of systemic toxicity such as convulsion, weakness and trembling were monitored. Weight is an important parameter in any experimental study because it is an indicator of physical health. The weight of each animal is monitored weekly with an electronic scale throughout the study.
• Description of the device
The object recognition memory test involves spontaneous exploration behavior in rodents, so it does not require food or water restriction or negative reinforcement of the animals. It is based on the spontaneous tendency of rats to preferentially explore a new object in relation to a familiar object. This device was used for the first time by [
This test takes place in a square open enclosure of 50 × 50 cm2 (L × l), surrounded by four walls of 50 cm2 height. It is made of wood painted with black color, which gives the image of an open cube. The device is illuminated by a light source positioned at a height of one meter.
• Experimental procedure
At the beginning of the test, each animal is allowed to freely explore the device for 5 minutes in the absence of any object. Twenty-four hours later, the animal must explore two identical objects (A1 and A2) placed at the two adjacent corners of the device at 10 cm from the walls, for 5 minutes; this is the “acquisition” session. Exploration is only called when the rat has a nose directed less than one cm to the object, while nibbling and marking are not considered exploration. After a delay of 2 hours (for evaluation of short-term memory effect) or 24 hours (for long-term memory evaluation), the rat is placed again in the device, but this time, one of the two identical objects is replaced by another entirely different B). The animal is allowed to explore the two different objects for 5 minutes. The two sets of objects used were in duplicate. The floor and objects are rinsed with 10% ethanol to remove any trace left by the animal in the device.
• Parameters taken into account
When the memory of the animal is not altered, it will explore the new object more than the old one, since it has memorized it. On the other hand, when the memory is defective, it will give more exploration time to the old object or it will explore both in an identical way. To evaluate the state of the memory, we calculated an index of recognition according to the following equation: Recognition index = time spent exploring the object B / Sigma time spent exploring object A and B [
We performed the statistical analyzes using the SPSS 20.0 software. Parametric ANOVA repeat tests and 1-factor ANOVA were used to compare the data series. When there was a difference between more than two batches, a further analysis was carried out with Tukey’s post hoc test. The normality of distribution was first atested by the Kolmogorov-Smirnov test and homogeneity by the Levens test. The values of p < 0.05 are considered significant.
Although the research has reached its aims, there are some unavoidable limitations.
The main of them are expressed as follows:
・ Lack of prior research studies on the topic.
・ The environmental factor (temperature, light...) influences the behavior of the rats and must be taken into consideration.
・ The sample size is an important feature of any study. The higher the number, the more reliable the results.
・ If the mode of administration is by gavage, the rats should be handled with care to avoid throat irritation.
Signs of systemic toxicity such as weight loss, resting quake, decreased food intake were observed in groups of rats poisoned compared to controls.
In males rats: sub-chronic intoxication with dimethoate for four weeks induces a progressive decrease in body weight in treated rats groups, unlike control rats
In female rats: A decrease in body weight was also observed in the group of
rats intoxicated during the study
In males: Body weight remained steady throughout the study period in intoxicated rats, while it increased steadily in control rats
In females: An increase in body weight was observed in female control rats compared with those poisoned by dimethoate gavage. The statistical analysis also showed a highly significant difference between the two study groups (p < 0.001)
In male rats:
In female rats: Dimethoate poisoning alters the short-term recognition memory in female rats. The expressed IR is highly significant between the two groups studied p < 0.001, either 43% to poisoned
In male rats: Dimethoate poisoning in drinking water induces an alteration of (STM) in rats. Indeed, the index of recognition is highly reduced in intoxicated rats. P < 0.001, compared to controls in
In female rats: There was also a highly significant reduction in the index of recognition in Dimethoate addicts p < 0.001, compared with controls in
In male rats: There was a significant decrease in long-term RI in rats treated with
Dimethoate gavage compared to controls, even though this index was above the 50% recognition threshold. The difference revealed is very statistically significant p < 0.01,
In female rats: the long-term RI of 54% in intoxicated females did not differ significantly with controls p > 0.05
In male rats: As shown in
In female rats: It is also observed that the index of recognition is above the threshold of recognition of the objects in the rats intoxicated with Dimethoate, even if this index differs very significantly from that of the controls p < 0.01,
In male rats: Recognition STM is highly altered in rats treated with Dimethoate, the recognition index is 40% below the recognition threshold with a highly significant difference from that of the control rats p < 0.001. However, treatment with the medicinal plant Nettle Dioica has led to an improvement in this index, which has increased from 40% to 53%, with a very significant difference p < 0.01,
In females: Observations similar to that of males were made. As shown in
In male rats: As shown in
In females: The LTM index is very significantly decreased in Dimethoate addicts p < 0.01. However, it was possible to show a slight increase in this index after treatment with the plant Nettle dioica, this difference is not significant p < 0.05 (
In male rats: In the case of gavage intoxication, it was demonstrated that the recognition STM was very much reduced (37%), compared to controls p < 0.001.
Treatment with the Nettle Dioica plant resulted in an improvement in this index, which rose from 37% to 60% in
In females: As shown in
In male rats: Recovery LTM that was slightly impaired after intoxication was slightly improved after treatment with nettle plant p < 0.001
In females: As shown in
nificantly altered (54%) in intoxicated rats, compared to controls p < 0.05. However, treatment with the Nettle plant allowed a small increase in this index without significant effect p < 0.05.
The effect of the diméthoate on the physical weight gain for both modes of administration behaves appreciably in a different way. Indeed, in the case of the poisoning with diméthoate in drinking water, we observe a highly significant decrease in weight depending on the duration of the treatment for both rats’ sexes, as shown in
Another study of the developmental toxicity in Malathion also demonstrates a low physical weight gain for young rats to the (GD 6-PND 45) stage [
The recognition memory capabilities of rats intoxicated with Dimethoate were highlighted by the NOR (Novel Object) recognition memory test. This methodology allowed to link chronic exposure to OPs insecticides and memory impairment which is a serious health problem for the general population. In this work, short and long-term types of memory have been studied, both of which are very much affected in Alzheimer’s disease [
The test is based on the spontaneous tendency of rodents to explore a new object in relation to another already seen. This cognitive test, evaluating the ability to recognize a previous stimulus, is a fundamental study of human amnesia by animal model [
The results of our study reveal that the sub-chronic exposure to Dimethoate induces a significant alteration of the recognition memory of the objects.
For STM short-term memory: In the case of rats poisoned by drinking-water, the recognition index was significantly decreased, with both sexes with a slightly pronounced effect in females:
To evoke hypotheses concerning the mechanism of action of alteration of memory, it is useful to know the cerebral structures considered as support for this cognitive function. Indeed, the NOR test model is influenced by hippocampal and cortical lesions [
For the long-term LTM recognition memory, our results also support a difference in the recognition index in the evaluation of the long-term recognition memory between the treated and control groups for the two forms of exposure, Even if this index turns out to be above the recognition threshold, as compared with the short-term recognition memory Figures 9-12. This implies that short- term recognition memory is more vulnerable to exposure to dimethoate than to long-term memory. Similar results have been demonstrated in a previous study which showed that malathion-exposed rats have a short-term memory disrupted compared to long-term recognition memory [
Although we have studied the effects of dimethoate intoxication in rats, we have tested the therapeutic effects of a medicinal plant, Nettle, which has antioxidant properties. In order to better support its properties, it is desirable to evoke the pro-oxidizing and proinflammatory effects induced by OPs in general and Dimethoate in particular. Most authors agree that chronic or acute exposure induces oxidative and proinflammatory stress effects in animals [
In addition, dimethoate intoxication has been shown to cause cellular alterations and oxidative stress, leading to lipid peroxidation and the generation of free radicals [
These results suggest that nettle treatment has a positive effect by eliminating free radicals associated with exposure to Dimethoate. In addition, it would be appropriate to investigate the effects of dimethoate on the serotonergic system and on the catecholamines involved in the regulation of memory performance [
Agriculture makes extensive use of pesticides and phytosanitary products. By destroying pests and weeds, these substances improve crops and guarantee the availability, quality, reliability and prices of agricultural products for the benefit of farmers and consumers.
Used improperly, pesticides constitute nevertheless a danger, because most can damage the health and the environment.
In this study, we have demonstrated that poisoning by dimethoate alters the short-term memory functions of wistar rats.
We have also tested the effect of nettle as a chelating treatment trial. An improvement in short-term memory was obtained.
Samih, M., N’Go, P.K., Belaaouja, S., Touhami, A.O. and Ahami, A.O.T. (2017) Effect of Dimethoate on Object Recognition Memory in Wistar Rats and Essay of Treatment with Nettle. Journal of Behavioral and Brain Science, 7, 425-445. https://doi.org/10.4236/jbbs.2017.79031