Adaptive response of rat pancreatic ß-cells to insulin resistance induced by monocrotophos: Biochemical evidence.

Our previous findings clearly suggested the role of duration of exposure to monocrotophos (MCP) in the development of insulin resistance. Rats exposed chronically to MCP developed insulin resistance with hyperinsulinemia without overt diabetes. In continuation of this vital observation, we sought to delineate the biochemical mechanisms that mediate heightened pancreatic ß-cell response in the wake of MCP-induced insulin resistance in rats. Adult rats were orally administered (0.9 and 1.8mg/kgb.w/d) MCP for 180days. Terminally, MCP-treated rats exhibited glucose intolerance, hyperinsulinemia, and potentiation of glucose-induced insulin secretion along with elevated levels of circulating IGF1, free fatty acids, corticosterone, and paraoxonase activity. Biochemical analysis of islet extracts revealed increased levels of insulin, malate, pyruvate and ATP with a concomitant increase in activities of cytosolic and mitochondrial enzymes that are known to facilitate insulin secretion and enhanced shuttle activities. Interestingly, islets from MCP-treated rats exhibited increased insulin secretory potential ex vivo compared to those isolated from control rats. Further, MCP-induced islet hypertrophy was associated with increased insulin-positive cells. Our study demonstrates the impact of the biological interaction between MCP and components of metabolic homeostasis on pancreatic beta cell function/s. We speculate that the heightened pancreatic beta cell function evidenced may be mediated by increased IGF1 and paraoxonase activity, which effectively counters insulin resistance induced by chronic exposure to MCP. Our findings emphasize the need for focused research to understand the confounding environmental risk factors which may modulate heightened beta cell functions in the case of organophosphorus insecticide-induced insulin resistance. Such an approach may help us to explain the sharp increase in the prevalence of type II diabetes worldwide.

Effect of monocrotophos, an organophosphorus insecticide, on the striatal dopaminergic system in a mouse model of Parkinson's disease.

Our earlier study had shown that low concentrations of monocrotophos (MCP) elicited dopaminergic features of Parkinson's disease (PD) in the nematode Caenorhabditis elegans In the present study, the effect of low doses of MCP on the striatal dopaminergic neurons was investigated using the mouse model system. MCP was initially screened for its ability to cause any neurobehavioral deficits and alterations in the dopaminergic system in Swiss albino mice, aged 8 weeks and weighing 25-30 g, with repeated doses at 0.3 and 0.6 mg/kg body weight (b.w.)/day for 7 days and 30 days. Mice were treated with four intraperitoneal injections for every 2 h with 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP) at the dosage of 14 mg/kg b.w. MCP was administered to these mice at the above-mentioned doses for 7 days. Mice administered with MCP alone revealed a significant (p < 0.05) reduction in the dopamine (DA) content at both 7 and 30 days and showed a significant (p < 0.05) increase in neurobehavioral deficits. Interestingly, when MCP was administered for 7 days to MPTP-treated mice, further significant decrease in both DA content and increase in neurobehavioral deficits were apparent. The extent of reactive oxygen species and lipid peroxidation were markedly increased, while the ratio of reduced to oxidized glutathione levels were significantly decreased (p < 0.05) in the treated mice as compared to the control. Significant histopathological alterations and a marked reduction in the number of tyrosine hydroxylase positive cells were evident in striatum of mice treated with higher doses of MCP. These changes were comparable to that seen in mice treated with MPTP and post-administered lower doses of MCP. Our findings suggest that MCP per se has the propensity to induce pathological changes in the dopaminergic neurons as well as augment the degeneration in a compromised nigrostriatal system such as that in PD.

Organophosphorus insecticide, monocrotophos, possesses the propensity to induce insulin resistance in rats on chronic exposure.

BACKGROUND: Our earlier studies had shown that monocrotophos (MCP), an organophosphorus insecticide (OPI), has the propensity to augment the secondary complications associated with type-1 diabetes. The present study investigates whether rats exposed for prolonged periods to monocrotophos would develop insulin resistance mediated by alteration in glucose homeostasis. METHODS: Male rats were administered sublethal doses of monocrotophos daily for 180 days. Interim blood samples were collected to measure alteration in blood glucose and lipid profile. Rats were also subjected to glucose and insulin tolerance test and fasting blood glucose and insulin levels were measured to calculate insulin resistance by HOMA-IR method. After 180 days, the rats were also evaluated for pancreatic histology and activities of hepatic gluconeogenetic enzymes. RESULTS: Monocrotophos elicited a gradual and sustained increase in blood glucose and insulin resistance in rats with concomitant glucose intolerance and reduced insulin sensitivity. MCP exposure was also associated with increase in weights of key white adipose pads, activities of gluconeogenesis enzymes and increase in pancreatic islet diameter, all of which led to hyperglycemia, hyperinsulinemia and dyslipidaemia. CONCLUSION: Long-term exposure of rats to MCP resulted in glucose intolerance with hyperinsulinemia, a hallmark of insulin resistance. Our data suggest that chronic exposure to low doses of monocrotophos, might lead to development of insulin resistance by altering lipid profile and glucose homeostasis.

Biochemical and physiological responses in Caenorhabditis elegans exposed to sublethal concentrations of the organophosphorus insecticide, monocrotophos.

The objective of this study was to investigate the impact of sublethal concentrations of MCP on definitive markers of toxicity and oxidative balance in the nematode, Caenorhabditis elegans. Exposure of worms to 0.85, 1.7 and 3.4mM (1/40, 1/20 and 1/10 LC50; LC50=34 mM) for 4h at 20°C induced significant perturbations in physiological parameters such as decreased brood size (47-73%), increased paralysis (47-85%) and inhibition of the activities of acetylcholinesterase (75-86%) and carboxylesterase (76-81%). These changes were accompanied by distinct oxidative impairments as evidenced by increased reactive oxygen species, decline in glutathione content and decrease in superoxide dismutase activity. Our results clearly demonstrate that low concentrations of MCP may alter the physiological and biochemical status in the nematode, thereby affecting the organism's fitness. Our findings on C. elegans provide an easy diagnosis for OPI contamination and may become helpful in evaluating the ecotoxicological effects of OPI in the aquatic environment near agricultural fields.

Estimation of plasma triglycerides with correction for free glycerol by orlistat inhibition of lipoprotein lipase activity.

This report presents a modification of the enzymatic method (lipoprotein lipase/glycerol kinase/glycerol-3-phosphate oxidase/peroxidase) for determination of plasma triglyceride levels in order to achieve correction for the free glycerol content. The strategy is based on elimination of lipoprotein lipase activity from the single "multienzyme reagent" by use of orlistat, thereby allowing formation of quinoneimine chromophore from free glycerol. Orlistat was found to abolish the lipoprotein lipase activity (triolein was used as substrate) without any impact on the glycerol-driven rate of quinoneimine generation. Conditions were standardized for estimation of both triglyceride and glycerol content of mice plasma using a microplate reader.

Elicitation of dopaminergic features of Parkinson's disease in C. elegans by monocrotophos, an organophosphorous insecticide.

Positive correlations have been suggested between usage of pesticides and the incidence of Parkinson's disease (PD) through epidemiological as well as few experimental evidences. Organophosphorus insecticides (OPI), which are extensively used in agricultural and household insect control, have been the subject of increasing concern in the past decades due to their neurotoxic potential. However, very few studies have demonstrated the potentials of OPI to induce features of PD in model organisms. In the present study, Caenorhabditis elegans was selected as the model organism to evaluate the potential of monocrotophos (MCP), an OPI, to elicit dopaminergic features of Parkinson's disease in terms of dopamine content, basic movement and integrity of dopaminergic neurons along with its effect on acetylcholinesterase (AChE) activity and life span. All the responses elicited by MCP were compared with that elicited by 1-methyl-4-phenyl- 1, 2, 3, 6-tetrahydropyridine (MPTP) in both N2 and BZ555 worms. N2 worms were exposed to varying concentrations of MCP (50, 100 and 200 µM) or MPTP (200, 300 and 400 µM) for 48 hours and locomotory rate, as measured by the number of body bends made in 20 seconds, was enumerated. Worms subjected to the same dose paradigms were also analyzed for the dopamine content by HPLC. The results indicated a significant reduction in the dopamine levels in the worms that were treated with MCP/MPTP and this correlated with the changes in locomotion compared to untreated worms. Worms treated with MCP also exhibited significant reduction in AChE activity. Both MPTP and MCP caused a marked reduction in life span in the worms. Transgenic worms (BZ555, which has GFP tagged to its 8 dopaminergic neurons) exposed to MCP and MPTP at the above concentrations showed a dose-dependent reduction in the number of green pixels in CEP and ADE neurons which also correlated with the neurodegeneration as visualized by decreased fluorescence in photomicrographs. Taken together, our data demonstrate that low levels of MCP elicits dopaminergic features of PD in C. elegans.

Insights into the mechanisms mediating hyperglycemic and stressogenic outcomes in rats treated with monocrotophos, an organophosphorus insecticide.

The present investigation provides mechanistic insights into the hyperglycemic and stressogenic effects of monocrotophos, an organophosphorus insecticide. Pre-treatment of rats with mifepristone (glucocorticoid receptor antagonist) prevented induction of liver tyrosine aminotransferase activity (TAT), but was ineffective in attenuating hyperglycemia induced by monocrotophos. Pre-treatment with propranolol (ß-adrenergic receptor antagonist) and phentolamine (α-adrenergic receptor antagonist) were effective in abrogating monocrotophos-induced hyperglycemia. Interestingly, while propranolol offered partial protection against hyperglycemia, phentolamine completely abolished the same. However, monocrotophos-induced hyperlactacidemia was completely abolished by propranolol. Both the adrenoreceptor antagonists, however, failed to attenuate the stressogenic potential of monocrotophos. Hyperglycemia and hyperlactacidemia induced by monocrotophos were abolished by pre-treatment with atropine. Exogenous epinephrine was associated with hyperglycemia and hyperlactacidemia. The impact of adrenergic antagonists on epinephrine-induced hyperglycemia and hyperlactacidemia were remarkably similar to that of monocrotophos-induced hyperglycemia and hyperlactacidemia. Further, hydrazine sulfate (a gluconeogenesis inhibitor) abolished hyperglycemia in monocrotophos-treated rats. From our data, it can be hypothesized that excessive stimulation of adrenoreceptors, probably elicited by increased plasma epinephrine, mediates hyperglycemic outcomes induced by monocrotophos. Pattern of changes in plasma lactate suggests that ß-adrenergic activation mediates monocrotophos-induced hyperlactacidemia, while α-adrenergic receptor mediates lactate utilization, leading to hyperglycemia. Induction of liver TAT activity is attributable to glucocorticoid receptor activation as a result of hypercorticosteronemia.

Evaluation of sublethal effects of dichlorvos upon Caenorhabditis elegans based on a set of end points of toxicity.

The primary objective of this study was to examine a possible correlation among the three endpoints of toxicity, namely, stress gene expression (hsp16), feeding, and acetylcholinesterase (AChE) activity in transgenic C. elegans (hsp16-lacZ) exposed to sublethal concentrations of dichlorvos, an organophosphorus insecticide. Worms exposed to dichlorvos (at 5, 40, and 80 microM) exhibited a concentration-dependent inhibition in feeding with total cessation in feeding occurring beyond 4 h of exposure. Concomitantly, marked and dose-dependent inhibition (69%-90%) of AChE was also evident after 4 h of exposure. Induction of heat shock protein (Hsp) was evident after 4 h of exposure (as seen from quantitative analysis), although maximum expression of Hsp was evident only after 24 h of exposure (as evident from qualitative analysis). Interestingly, the Hsp induction was restricted only to the pharyngeal region. Significant correlation was discernible between the three evaluated end points suggesting their possible interrelated role in the physiological dysfunctions evoked by sublethal concentrations of dichlorvos.