The recent study described the neurobehavioral toxicity of acute buprofezin exposure however it is previously well established that buprofezin (BPFN) frequently used as a moulting inhibitor insecticide all over the world to eradicate pests like leafhoppers, mealy bugs and whitefly (Chang et al., 2015), infiltrating the fruit crops, leafy crops and citrus crops. Its metabolic compounds are potentially hazardous to the neighboring milieu (EFSA, 2007). It has been proved extremely toxic to aquatic environment (EFSA, 2010: Ku et al., 2015) as in embryo of zebra fish reactive oxygen species (ROS) have been detected following the exposure to buprofezin and nickel. Administration of embryos and larvae of African catfish (Clarias gariepinus) to various doses of buprofezin consequences into death of embryos when its amount of dose rises to 5–100 mg/L. In African catfish dose < 5 mg/L also carry out numerous hazardous effects during embryogenesis and development of larva. These effects comprise asymmetrical head, bleeding from pericardium, inward curvature of the lumbar and cervical regions, arcuate in body, ulcerates and accumulation of fluid in yolk sac (Marimuthu et al., 2013).
In our study we have explored that the acute oral dose of buprofezin 87.9 mg/kg/day induced wide range of neurobehavioral toxic effects. Acute intoxication of buprofezin induce a wide range of neurobehavioral toxicity including damage of pyramidal cells of hippocampal CA1 and CA3, region neurons and behavioral impairments for example, loss of motor coordination, locomotor activity, fear loss, hearing, heat shock, sensorimotor, cognitive and spatial navigation impairment following the acute exposure in adult Sprague dowley male rats. We have also found that acute intoxication of Buprofezin is potentially reversed by pre-administration of Atropine. The complete molecular and biochemical mechanism of Buprofezin neurobehavioral toxicity is not elucidated so for however we suggested that it inhibit the synthesis and release of AChE in synapse as in our experiment Buprofezin intoxicated rat was suddenly dead after the administration of neostigmine (30µl/kg/day i.p) a blocker of AChE. It put forward that a small concentration of AChE in synapse was dominantly occupied by a small concentration of neostigmine consequently tremendously elevated the ACh level in synapse leading to tremor and death of rats. This hypothesis was also supported by previous studies as activity of cytochrome and TCA cycle enzyme was rendered by buprofezin that interfere the energy metabolism and inhibit production of ATP. (Ji et al., 2016; Binukumar et al., 2010; Shan et al., 2013). Atropine function as a physiological antagonist and competitively block the acetylcholine action of muscarinic receptors and acts as antidote for excessive parasympathetic activation arised as consequence of inhibition of AChE (Johnson et al., 2000). Acetylcholinesterase (AChE) being serine hydrolase enzyme that cleaves rapidly terminate the cholinergic transmission in synapse by breakdown of Ach into choline and acetate (Soreq and Seidman, 2001). AChE also described as key player in activation of glial cells, brain blood flow, amyloid pathway, phosphorylation of tau protein, also function as adhesion protein for maintenance and development of synapse (Ballard et al., 2005). Arsenic exposure in animal model induces behavioral alteration, abnormality in nervous system shaping and development, inflammation and neuron death, (Yen et al., 2011; Flora et al., 2012). In addition, arsenic could induce toxicity in HAPI microglia (Mao et al., 2016), granular neurons of cerebellum (Liu et al., 2013) and snail neurons (Lu et al., 2009).
Further our study demonstrated the impairment of motor coordination and fore limb and hind limb grip strength as Occupational exposure to acrylamide leads to cumulative but reversible neurotoxicity described by axanopathy of peripheral nerves, ataxia, muscle weakness, tingling of hands and feet and cognitive deficiency. Because it inhibits kinesin transport, decrease the neurotransmitters and inhibition of transmission (Exon, 2006). Crofton and colleagues stated decreased grip strength in a 30-day acrylamide administration study at 15 and 20 mg/kg/days because it causes peripheral axonopathy (Crofton et al., 1996) after exposure to high oral doses of buprofezin clinical signs appear as reduced locomotor activity, tremble, runny nose, abnormal movement and urinary discontinuous urination. (EFSA, 2007). In present study impairment in the grip strength and motor coordination was assessed by using rotarod, horizontal and parallel bars. Similar impairments in motor activity, abnormal gait, and cognitive deficiency were detected following exposure to bifenthrin due to oxidative stress (Farah et al., 2017) A study on earthworm described that reduction in growth rate by combined exposure of buprofezin, Lufenuron, and Triflumuron pesticide-exposed worms was observed by dose-dependent over the 28-day treated duration, which was accompanied by a decline in activity of AChE and GST. The lowest activity of AChE was noted at the highest dose of buprofezin (300 mg/kg soil) following two weeks of exposure as compared to control. The activity of AChE was intensely repressed by lufenuron subsequently by buprofezin, and then triflumuron in descendant. (Badawy et al., 2013). Similarly, our study showed that acute exposure of buprofezin inhibit the synthesis and release of AChE in synapse due to limited supply of ATP described by other study that buprofezin efficiently repressed the cytochrome c oxidase activity by binding to SCO1 active pockets and COX17, which increased the concentration of reactive oxygen species. Additionally, administration with an ROS inhibitor (N-acetyl-L-cysteine) (NAC) counteract the decreased level of ATP and cytochrome c oxidase activity, which also showed that ROS contributed in buprofezin-induced conversion of energy metabolism. After sub lethal treatment of buprofezin, the levels of the end product metabolism (ATP), end product of glycolysis (lactate) and (pyruvate) a component in initial stage of the TCA cycle were evaluated. The higher concentration of these factors after buprofezin exposure reveals the BPFN induced inhibition of TCA cycle. Pesticides can decrease the ATP concentration in HepG2 cells revealed by in vivo and in vitro studies. (Binukumar et al., 2010; Shan et al., 2013).
Pre- treated atropine counteracts the poisoning by blocking muscarinic receptors and it counteracted over parasympathetic activity. At single oral dose (24 h) of chlorpyrifos reduced the activity of plasma butyrylcholinesterase (BChE) and rats (AChE) activity in hippocampus, striatum and prefrontal cortex. The acute chlorpyrifos toxicity can be counteracted by the atropine antidote (10 mg/kg i.p.) and/or pralidoxime (40 mg/kg; i.p.) treated one hour following toxicity (Alciene Almeida Siqueira et al., 2019). In pre-clinical and clinical experiments muscarinic antagonists exhibit antidepressant effects (Drevets et al., 2013; Mancinelliet al., 1988; Witkin et al., 2014). Acute buprofezin decreases the synthesis and release of AChE in synapse. Furthermore (ip) neostigmine injection in buprofezin intoxicated rats cause tremor, and sudden death of rat also proposed that minor amount of AChE is predominately blocked by neostigmine extremely elevated the ACh level in synapse subsequent acute exposure and is counteracted by pre-treated atropine.
Buprofezin exposure impair the passive avoidance as MSK1 knockout affects numerous various forms of hippocampus-dependent memory, as evaluated by fear conditioning, Morris water maze and passive avoidance (Wilson et al.,2007). Pre-treated atropine showed counteract effect not reported in any previous study.
The behavioral analysis revealed that SA (sodium arsenide) treatment cause loss in learning and memory in passive avoidance as well as motor activity and balance. Additionally, acute or chronic administration to SA as revealed by other experiments induce abnormalities of CNS containing slowing of cognitive development, decreased psychomotor speed, loss of learning due to decreased number and apoptosis of pyramidal cells and was mitigated by ellagic acid. (Franzblau et al., 1989; Mathew et al., 2010; Tsai et al., 2003). Our finding also agreed with similar behavior abnormalities and loss of learning and memory due to pyramidal neuron damage in hippocampus was attenuated by pre –treated atropine. Buprofezin induced decreased in the step- through latency was reversed by pre- treated atropine as the SA exposure has showed significant reduction in the step-through latency comparison to the control due to oxidative stress (Mehdi et al., 2018). The study reported that MSK1 knock-out animals can process the sensory information of foot shock and memorize it in association with contextual and the auditory stimulus, but loss this memory in 24h. (Wilson et al 2007). Our study reported the impairment in long tern contextual fear memory and rapid loss of sensory stimulus of foot shock in acute buprofezin exposure and was slightly counteracted by pre-treated atropine.
Various concentration of orally administered imidacloprid to female rats caused a substantial change in different features of locomotor activity and decreased in ambulation at 90 days of treatment as it inhibits AChE activity (Shipra et al., 2010). Substantial reduction in locomotion in the rats exposed with the acute dose of imidacloprid has suggested that imidacloprid or its metabolic residues has accumulated in brain. Administration of imidacloprid directly in to intraperitonium has shown to accumulated in mouse brain (Chao et al., 1997).
Although the mechanism of action of buprofezin is distinct to some extant results described the similar decline in spontaneous locomotion activity in acute buprofezin administered rats compared to control and pre-treated atropine counteract the toxicity. Similar to high dose of imadacloprid decreases the spontaneous locomotor activity. Another study also supported our results that during the peak of the BGS (brain growth spurt) (PND10) administration of single dose of endosulfan or cypermethrin, cause long lasting spontaneous behavior abnormality in adults due to alteration of protein involved in brain development, variation in locomotion, rearing and total activity without affecting body weight as compared to control mice (Lee et al., 2015).
It has also been stated that the number of hippocampal neurons declined in rats after treatment with sulfite (Akdogan et al., 2011). Additionally, it was revealed that after sulfite exposure the excitability of the spinal reflexes was increased (Küçükatay et al., 2005; Küçükatay et al., 2008). The toxic consequences of sulfite on mesencephalic cell lines have been described, as well (Reist et al., 1998).
Our study in line with these findings as pre-treated atropine has neuroprotective effect against buprofezin toxicity. Pre-treated atropine prevents hippocampal neuron degeneration, spatial memory impairment, working and reference memory loss by preventing over excitability induced neuron exhaustion and preventing the apoptosis of hippocampal neurons. It also prevents the oxidative stress of hippocampal neurons and protect against buprofezin induced cognitive impairment as curcumin inhibit lead-induced loss of memory in rats (Dairam et al., 2007). Curcumin can alleviate the cognitive deficit in diabetic rats (Kuhad et al 2007). Therefore, curcumin may preclude the oxidative stress in CA neurons and, as result, may enhance the synaptic plasticity (Noorafshan et al., 2013; Kuhad et al 2007). It is also stated that curcumin protect the neurodegeneration in Parkinson and Alzheimer disease. (Yadav et al., 2009). Additionally, it is also suggested that curcumin prevent the apoptosis of neuron. (Lin et al., 2011).
A wide range of effects produced by Trichlorethane (TCE), and ether are similar to ethanol and depressant like properties of volatile solvent described in earlier literature (Evan et al.,1996) these involved decreases in activity of CNS (alterations in posture. reduced arousal and rearing), decreases in emotionality of CNS (increased ease of removal), impair of muscle tone (disturbances in gait reduction in forelimb grip strength, increased landing foot splay and loss of psychomotor coordination on the inverted screen test), and decreased sensorimotor activity (reduced response to sensory stimuli). Though TCE and ether at concentrations of 13.300 and 30.000 ppm substantially increased landing foot splay. Flurothyl exposure did not affect increase in landing foot splay at any concentration. TCE and ether also decrease forelimb grip strength but not flurothyl. Additionally, flurothyl produccd handling-induced tremors after after animals was removed from cage, effect which was not caused by TCE, ether or ethanol. Flurothyl cause postictal depression (e.g. an increased inversion latency on the inverted screen test) Characterized by convulsion in testing animal and exhibit not any ethanol like properties (Scott e et al., 1996). The results of our study also agreed with the with these findings as the acute exposure of buprofezin in rat results in decreased brain activity, posture abnormality, decreased activity level, increased lacrimation, salivation, piloerection, abnormal muscle tone, loss of limb grip, ambulation and gait abnormalities, decreased rearing and arousal, loss of time space navigation ability, loss of sensorimotor responses, Redness of nostrils, muscle hypoplasia, decrease in body weight and body temperature. However pre-treated atropine attenuated these variations and significantly reversed these effects.
The farmer studies have various shortcoming as they devoid acute buprofezin neurotoxicity in rat model. Additionally, none of previous study has reported the mechanism underlying its neurotoxicity in rats. Further no therapeutic strategy against buprofezin toxicity was carried out. In contrary our finding has proved that acute exposure of buprofezin induce wide range of neurobehavioral toxicity in adult Sprague dowely male rats and potential antidote against its neurotoxicity