Environmental Toxicology
Volume 29, Issue 6 pp. 665-671
Research Article
Full Access

Diphenyl diselenide protects against metabolic disorders induced by acephate acute exposure in rats

Carmine Inês Acker, 

Corresponding Author

Carmine Inês Acker

Departamento de Química, Centro de Ciências Naturais e Exatas, Universidade Federal de Santa Maria, Santa Maria, CEP 97105-900, RS, Brasil

Departamento de Química, Centro de Ciências Naturais e Exatas, Universidade Federal de Santa Maria, Santa Maria, CEP 97105-900, RS, Brasil. E-mail: [email protected]Search for more papers by this author
Cristina Wayne Nogueira, 

Cristina Wayne Nogueira

Departamento de Química, Centro de Ciências Naturais e Exatas, Universidade Federal de Santa Maria, Santa Maria, CEP 97105-900, RS, Brasil

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First published: 10 July 2012
https://doi.org/10.1002/tox.21793
Citations: 6

Abstract

The present study investigated the effect of diphenyl diselenide [(PhSe)2] on metabolic disorders induced by acephate acute exposure in rats. We also investigated a possible mechanism of action of (PhSe)2 against hyperglycemia induced by acephate. (PhSe)2 was administered to rats at a dose of 10 or 30 mg/kg by oral gavage (p.o.) 1 hour prior to acephate administration (140 mg/kg; p.o.). Glucose and corticosterone levels as well as the lipid status were determined in plasma of rats. Cardiovascular risk factors and the atherogenic index were calculated. Glycogen levels as well as tyrosine aminotransferase (TAT) and glucose-6-phosphatase (G6Pase) activities were determined in livers of rats. Cerebral acetylcholinesterase (AChE) activity was assayed. Acephate induced an increase in glucose and corticosterone levels as well as in TAT and G6Pase activities. AChE activity was inhibited by acephate. Triglyceride (TG) levels and the cardiovascular risk factor TG/high-density lipoprotein-cholesterol (HDL) were increased by acephate. (PhSe)2 was effective against the metabolic disorders induced by acephate acute exposure in rats. © 2012 Wiley Periodicals, Inc. Environ Toxicol 29: 665–671, 2014.

Abbreviations

  • AChE
  • acetylcholinesterase
  • AI
  • atherogenic index
  • GC-FID
  • gas chromatography with flame ionization detection
  • HDL
  • high-density lipoprotein-cholesterol
  • HPA
  • hypothalamus-pituitary-adrenal
  • LDL
  • low-density lipoprotein-cholesterol
  • OPs
  • organophosphate insecticides
  • pHBA
  • p-hydroxybenzaldehyde
  • TAT
  • tyrosine aminotransferase
  • TC
  • total cholesterol
  • TCA
  • trichloroacetic acid
  • TG
  • triglycerides

    • INTRODUCTION

    Organophosphate insecticides (OPs) constitute one of the most widely used classes of pesticides being employed for both agricultural and domestic pest control. The use of OPs has increased considerably due to their low toxicity and low persistence in the mammalian system compared to organochlorine pesticides (Costa, 2006). The wide application of OPs was accompanied by potentially hazardous impact on humans, animals, and environment (water, air, soil, and food), causing severe acute and chronic poisoning (Kanbur et al., 2008; Soltaninejad and Abdollahi, 2009). OPs are primarily recognized for their ability to induce toxicity in mammals through inhibition of acetylcholinesterase (AChE) activity and subsequent activation of cholinergic receptors (Aardema et al., 2008).

    Hyperglycemia has been investigated as another facet of OPs toxicity. Studies with animals have shown altered glucose homeostasis following acute and chronic exposure to OPs (Kamath and Rajini, 2007; Lasram et al., 2008). The strength of evidence provided by a systematic review indicates the role of glycogenolysis, gluconeogenesis, and the activation of hypothalamus-pituitary-adrenal (HPA) axis in the mechanisms of OPs-induced hyperglycemia (Rahimi and Abdollahi, 2007). Accordingly, some studies have reported the activation of HPA axis and hepatic gluconeogenesis enzymes, such as tyrosine aminotransferase (TAT) and glucose-6-phosphatase (G6Pase) after OPs exposure in rats (Joshi and Rajini, 2009, 2012).

    Acephate (O,S-dimethyl acetylphosphoramidothioate) is an OP widely used in horticulture, household gardens, and for crop protection. Acephate and its primary metabolite, methamidophos, are toxic to various species (Rao et al., 2006). The toxic effects of acephate on experimental animal models, such as neurotoxicity in rats (Chen et al., 2003), genotoxicity in mice (Rahman et al., 2002), and oxidative stress in rats (Datta et al., 2010) have been demonstrated. Furthermore, it was reported that acephate causes a temporary hyperglycemia in rats by activation of the HPA axis and gluconeogenesis pathway (Joshi and Rajini, 2009).

    Considering that the exposure to OPs, including acephate, has been related to the development of hyperglycemia, the development of anti-hyperglycemic drugs are of potential interest to reduce hyperglycemia caused by OPs exposure. In this context, the organoselenium compound diphenyl diselenide [(PhSe)2] has numerous pharmacological properties (Nogueira and Rocha, 2010). Of particular importance are the anti-hyperglycemic (Barbosa et al., 2006), anti-diabetic (Barbosa et al., 2008), anti-hyperlipidemic (da Rocha et al., 2011), hepatoprotective (Borges et al., 2008), antiulcer (Savegnago et al., 2006) and antioxidant (Luchese et al., 2007; Prigol et al., 2009) properties demonstrated in different experimental models.

    Considering that acephate exposure causes metabolic disorders and (PhSe)2 has anti-hyperglycemic property, the aim of the present study was to investigate the protective effect of (PhSe)2 on metabolic disorders induced by acephate acute exposure in rats. We also investigated a possible mechanism of action of (PhSe)2 against hyperglycemia induced by acephate.

    MATERIAL AND METHODS

    Chemicals

    Acephate (Orthene 750 BR, Arysta Lifescience do Brasil Indústria Química e Agropecuária LTDA) was obtained from commercial grade. The purity of acephate commercial pesticide (74.9%) was determined by gas chromatography with flame ionization detection (GC-FID) according to Dobrat and Martijn (1998). (PhSe)2 was prepared in our laboratory according to Paulmier (1986) and the chemical purity (99.9%) was determined by GC/MS. Analysis of 1H and 13C NMR spectra showed analytical and spectroscopic data in full agreement with its assigned structure. (PhSe)2 and acephate were dissolved in ethanol and saline, respectively. (PhSe)2 was dissolved in ethanol to avoid a possible interference of another vehicle (oil, for example) in the biochemical determinations related to glucose and lipid metabolism. All other chemicals were obtained from standard commercial suppliers.

    Animals

    Male adult Wistar rats, weighing 200–300 g, were obtained from our own breeding colony (Federal University of Santa Maria, Brazil). Animals were kept in a separate animal room, on a 12 h light/12 dark cycle with lights on at 7:00 a.m., in an air-conditioned room (22°C ± 2°C). Commercial diet (GUABI, RS, Brasil) and tap water were supplied ad libitum. Animals were used according to the guidelines of the Committee on Care and Use of Experimental Animal Resources and with the approval of the Animal Use Committee (23081.017070/2011-19), Federal University of Santa Maria, Brazil.

    Experimental Procedure

    Rats were divided into six groups of six animals each. Following overnight fasting (12 hours), (PhSe)2 was administered to rats at a dose of 10 or 30 mg/kg by oral gavage (p.o.) 1 hour prior to p.o. administration of acephate (140 mg/kg of active ingredient). The dose of 10 mg/kg of (PhSe)2, which does not cause toxicity in rodents, was chosen based on our previous study (Barbosa et al., 2008), which demonstrated that (PhSe)2 has anti-hyperglicemic effect on alloxan-induced diabetes in rats. The dose of 30 mg/kg of (PhSe)2 was tested because a dose of 10 mg/kg was not effective against all parameters evaluated in the current study. The dose of 140 mg/kg of acephate was selected based on a previously published study (Joshi and Rajini, 2009), which demonstrated that acephate caused metabolic disorders in rats.

    The protocol of rat treatments is given below:
    • Group I: ethanol (1 mL/kg; p.o.) plus saline 0.9% (1 mL/kg; p.o.);
    • Group II: (PhSe)2 (10 mg/kg; p.o.) plus saline 0.9% (1 mL/kg; p.o.);
    • Group III: (PhSe)2 (30 mg/kg; p.o.) plus saline 0.9% (1 mL/kg; p.o.);
    • Group IV: ethanol (1 mL/kg; p.o.) plus acephate (140 mg/kg; p.o.);
    • Group V: (PhSe)2 (10 mg/kg; p.o.) plus acephate (140 mg/kg; p.o.);
    • Group VI: (PhSe)2 (30 mg/kg; p.o.) plus acephate (140 mg/kg; p.o.).

    (PhSe)2 was administered at 8:00 a.m. and acephate at 9:00 a.m. Two hours after the acephate administration, all rats were anesthetized for blood collection (2 mL) by heart puncture. Plasma was separated by centrifugation at 2400 × g for 10 min and stored at −20°C for biochemical analyzes (hemolyzed plasma was discharged). After this procedure, the rats were killed, and the livers of animals were removed, dissected, and kept on ice until the time of assay. The livers were kept on ice no more than three hours before assay.

    The liver samples were homogenized in 50 mM Tris-HCl (pH 7.4; 1:10 w/v) for TAT assay and in 250 mM sucrose containing 1 mM EDTA (pH 7.0; 1:10 w/v) for G6Pase assay and centrifuged at 2400 × g for 10 min. The low-speed supernatants (S1) were separated and used for TAT and G6Pase assays. The brains of rats were removed, kept on ice, homogenized in 0.25 M sucrose buffer (1/20, w/v), and centrifuged at 2400 × g for 10 min. The low-speed supernatants (S1′) were used for AChE assay.

    Biochemical Determinations

    Plasma Glucose Levels Determination

    Plasma glucose levels were determined by an enzymatic method based on the oxidase/peroxidase system using a commercial kit (LABTEST, Diagnostica S.A., Minas Gerais, Brasil). Plasma glucose levels were expressed as mg/dL.

    Plasma Corticosterone Levels Determination

    Corticosterone levels were estimated by the fluorescence method previously described by Zenker and Bernstein (1958). Corticosterone in plasma aliquot (200 μL) was extracted with 2 mL of chloroform. The volume into tubes was completed to 3 mL with distilled water. The tubes were shaken for 15 seconds, centrifuged at 2400 × g for 5 minutes, and the aqueous layer was discharged. Then, 1 mL of NaOH 0.1 M was added to tubes which were shaken again for 15 seconds and centrifuged at 2400 × g for 5 minutes. The aqueous layer of each tube was discharged as above. Aliquots of 1 mL of chloroform layer were transferred to other tubes containing 3 mL of fluorescence reagent (2.4 parts concentrated H2SO4 + 1.0 part 50% ethanol). The tubes were shaken for 15 seconds, centrifuged at 2400 × g for 5 minutes, and the chloroform layer was discharged. The tubes with sulfuric acid layer were allowed to stand at room temperature for 2 hours. After that, the fluorescence intensity emission was recorded at 540 nm (with 257 nm excitation). Corticosterone levels were expressed as μg corticosterone/dL plasma.

    Plasma Lipid Levels Determination

    Total cholesterol (TC), high-density lipoprotein-cholesterol (HDL), and triglycerides (TG) levels were determined by enzymatic colorimetric methods using commercial kits (LABTEST, Diagnostica S.A., Minas Gerais, Brazil). Low-density lipoprotein-cholesterol (LDL) values were obtained by the difference between TC and HDL levels. Plasma lipid levels were expressed as mg/dL.

    To explore the lipid metabolism, we calculated the cardiovascular risk factors TC/HDL and TG/HDL and the atherogenic index (AI) [(TC-HDL)/HDL] (Reaven, 2003).

    Hepatic Glycogen Levels Determination

    The hepatic glycogen content was assayed by the method described by Krisman (1962). Briefly, a known amount of liver was digested in 2 mL of 30% KOH solution. Followed by 10 minutes in boiling water bath, 2 mL of ethanol was added to the tubes to precipitate glycogen. After precipitation, glycogen was resuspended in 0.2 mL 5 M HCl and 0.8 mL distilled water. The glycogen content was measured with iodine reagent at 460 nm and expressed as g of glycogen/100 g of liver.

    Hepatic TAT Activity Assay

    TAT was assayed by the method described by Diamondstone (1966). The S1 was diluted in 50 mM Tris-HCl (pH 7.4; 1:2 v/v). The reaction mixture contained 6 mM L-tyrosine, 9.4 mM α-ketoglutarate, 4 mM diethyldithiocarbamate, and 40 μM pyridoxal-5-phosphate in a final volume of 3.2 mL. The reaction was initiated by the addition of 0.025 mL of S1. The samples were incubated at 37°C for 10 min and the incubation was stopped adding 200 μL of 10 M NaOH. After a stabilization period of 30 min at room temperature, the absorbance of samples was measured at 331 nm. TAT activity was expressed as nmol p-hydroxybenzaldehyde (pHBA)/min/mg protein.

    Hepatic G6Pase Activity Assay

    G6Pase activity was assayed based on the method reported by Ricketts (1963). The S1 was diluted in 250 mM sucrose containing 1 mM EDTA (pH 7.0; 1:5 v/v) and incubated (0.1 mL) with 50 mM glucose-6-phosphate for 30 minutes at 37°C. The incubation was stopped by adding 1.0 mL of 10% trichloroacetic acid (TCA). The samples were centrifuged at 2400 × g for 5 min. Inorganic phosphate (Pi) levels in supernatants were measured at 650 nm as described by Fiske and Subbarow (1925). G6Pase activity was expressed as nmol Pi/min/mg protein.

    Cerebral AChE Activity Assay

    AChE activity assay was carried out according to the method reported by Ellman et al. (1961), using acetylthiocholine as substrate. The activity of AChE was spectrophotometrically measured at 412 nm and expressed as μmol/h/mg protein.

    Protein Determination

    Protein concentration was measured by the method of Bradford (1976), using bovine serum albumin as standard.

    Statistical Analysis

    Statistical analysis was performed using a two-way analysis of variance (ANOVA), followed by the Duncan's Multiple Range Test. Main effects are presented only when the higher second-order interaction was nonsignificant. Data were expressed as means ± S.E.M. of six animals. Values of p < 0.05 were considered statistically significant.

    RESULTS

    Plasma Glucose Levels

    As shown in Table 1, plasma glucose levels in control rats were 91.46 ± 4.25 mg/dL. Acephate-exposed rats presented glucose levels of 229.30 ± 12.88 mg/dL while in acephate-exposed rats pre-treated with (PhSe)2 (10 or 30 mg/kg) glucose levels were 132.20 ± 11.78 and 121.80 ± 9.59 mg/dL, respectively. The two-way ANOVA of glucose levels demonstrated a significant (PhSe)2 × acephate interaction (F2,20= 44.51, p < 0.0001). Post-hoc comparisons revealed that acephate increased glucose levels of rats if compared to those of the control group. (PhSe)2 pretreatment at doses of 10 and 30 mg/kg attenuated the increase of glucose levels caused by acephate.

    Table 1. Effect of (PhSe)2 on biochemical parameters of rats exposed to acephate (AC)
    Parameters Experimental Groups
    Control (PhSe)2 (10) (PhSe)2 (30) AC (PhSe)2 (10) + AC (PhSe)2 (30) + AC
    Glucose 91.46 ± 4.25 93.11 ± 7.26 104.30 ± 7.01 229.30 ± 12.88 132.20 ± 11.78 121.80 ± 9.59
    G6Pase 38.50 ± 1.88 42.13 ± 1.40 37.47 ± 1.64 64.22 ± 3.47 54.55 ± 2.54 46.53 ± 4.68
    TAT 18.60 ± 1.16 21.35 ± 1.05 17.42 ± 1.37 31.84 ± 1.48 32.36 ± 2.76 26.02 ± 1.65
    Corticosterone 29.92 ± 2.47 31.63 ± 3.52 34.47 ± 0.67 56.33 ± 3.41 52.10 ± 1.05 52.97 ± 1.82
    AChE 6.25 ± 0.30 5.81 ± 0.30 7.07 ± 0.85 2.50 ± 0.27 3.31 ± 0.63 2.46 ± 0.26
    TC 90.50 ± 8.16 90.17 ± 10.33 75.00 ± 4.81 86.83 ± 8.22 88.50 ± 5.24 72.83 ± 8.37
    HDL 42.00 ± 3.87 37.00 ± 2.54 34.00 ± 2.37 37.17 ± 3.72 37.67 ± 1.84 37.50 ± 2.22
    LDL 37.90 ± 5.65 43.27 ± 7.86 33.47 ± 5.17 38.83 ± 5.38 41.43 ± 3.90 29.60 ± 6.63
    TG 44.67 ± 3.40 49.83 ± 4.59 37.67 ± 2.90 62.50 ± 4.17 47.00 ± 2.87 48.67 ± 1.20
    TG/HDL 1.13 ± 0.16 1.35 ± 0.10 1.12 ± 0.08 1.76 ± 0.19 1.27 ± 0.12 1.31 ± 0.07
    • a Plasma glucose, TC, HDL, LDL and TG levels are expressed as mg/dL.
    • b Hepatic G6Pase activity is expressed as nmol Pi/min/mg protein.
    • c Hepatic TAT activity is expressed as nmol pHBA/min/mg protein.
    • d Plasma corticosterone levels are expressed as μg corticosterone/dL plasma.
    • e Cerebral AChE activity is expressed as μmol/h/mg protein.
    • f Denotes p < 0.05 as compared to the control group.
    • g Denotes p < 0.05 as compared to the AC group (two-way ANOVA/Duncan's multiple range test).
    • Data are reported as means ± S.E.M. of six animals per group.

    (PhSe)2 at doses of 10 and 30 mg/kg did not alter glucose levels in plasma of rats (93.11 ± 7.26 and 104.30 ± 7.01 mg/dL, respectively) (Table 1).

    Hepatic Glycogen Levels

    The two-way ANOVA showed that neither (PhSe)2 nor acephate administration alter glycogen levels in livers of rats (data not shown).

    Hepatic G6Pase Activity

    Hepatic G6Pase activity in control rats was 38.50 ± 1.88 nmol Pi/min/mg protein. Acephate-exposed rats presented G6Pase activity of 64.22 ± 3.47 nmol Pi/min/mg protein, whereas in acephate-exposed rats pretreated with (PhSe)2 (10 or 30 mg/kg) G6Pase activity were 54.55 ± 2.54 and 46.53 ± 4.68 nmol Pi/min/mg protein, respectively. The two-way ANOVA of G6Pase activity yielded a significant (PhSe)2 x acephate interaction (F2,20 = 6.91, p < 0.05). Post-hoc comparisons showed that acephate increased G6Pase activity if compared to that of rats from the control group. (PhSe)2 pretreatment at doses of 10 and 30 mg/kg partially and completely protected against the increase of G6Pase activity caused by acephate, respectively (Table 1).

    (PhSe)2 at doses of 10 and 30 mg/kg did not alter G6Pase activity in livers of rats (42.13 ± 1.40 and 37.47 ± 1.64 nmol Pi/min/mg protein, respectively) (Table 1).

    Hepatic TAT Activity

    Hepatic TAT activity in control rats was 18.60 ± 1.16 nmol pHBA/min/mg protein. Acephate-exposed rats presented TAT activity of 31.84 ± 1.48 nmol pHBA/min/mg protein while in acephate-exposed rats pretreated with (PhSe)2 (10 or 30 mg/kg) TAT activity were 32.36 ± 2.76 and 26.02 ± 1.65 nmol pHBA/min/mg protein, respectively. The two-way ANOVA of TAT activity revealed a significant main effect of acephate (F1,20 = 58.40, p < 0.0001) and (PhSe)2 (F2,20= 5.99, p < 0.05). Post-hoc comparisons demonstrated that acephate increased TAT activity when compared to the control group. (PhSe)2 pretreatment at a dose of 30 mg/kg partially protected against the increase of TAT activity resulting from acephate exposure (Table 1).

    Hepatic TAT was not altered by (PhSe)2 pretreatment at doses of 10 mg/kg and 30 mg/kg (21.35 ± 1.05 and 17.42 ± 1.37 nmol pHBA/min/mg protein, respectively) (Table 1).

    Plasma Corticosterone Levels

    As shown in Table 1, plasma corticosterone levels in control rats were 29.92 ± 2.47 μg corticosterone/dL plasma. Acephate-exposed rats presented corticosterone levels of 56.33 ± 3.41 μg corticosterone/dL plasma while in acephate-exposed rats pretreated with (PhSe)2 (10 or 30 mg/kg) corticosterone levels were 52.10 ± 1.05 and 52.97 ± 1.82 μg corticosterone/dL plasma, respectively. The two-way ANOVA of corticosterone levels data showed a significant main effect of acephate (F1,20= 93.62, p < 0.0001). Post-hoc comparisons revealed that acephate increased corticosterone levels when compared to those of rats from the control group. (PhSe)2 pretreatment at both doses did not protect against the increase of corticosterone levels caused by acephate.

    Corticosterone levels remained unaltered in plasma of rats which received (PhSe)2 at doses of 10 and 30 mg/kg (31.63 ± 3.52 and 34.47 ± 0.67 μg corticosterone/dl plasma, respectively) (Table 1).

    Cerebral AChE Activity

    Cerebral AChE activity in control rats was 6.25 ± 0.30 μmol/h/mg protein. Acephate-exposed rats presented AChE activity of 2.50 ± 0.27 μmol/h/mg protein, while in acephate-exposed rats pretreated with (PhSe)2 (10 or 30 mg/kg) AChE activity were 3.31 ± 0.63 and 2.46 ± 0.26 μmol/h/mg protein, respectively. The two-way ANOVA of AChE activity yielded a significant main effect of acephate (F1,20= 72.81, p < 0.0001). Post-hoc comparisons demonstrated that acephate inhibited AChE activity in brains of rats if compared to those of the control group. (PhSe)2 pre-treatment at both doses did not protect against the inhibition of AChE activity resulting from acephate exposure (Table 1).

    (PhSe)2 at doses of 10 and 30 mg/kg did not alter AChE activity in brains of rats (5.81 ± 0.30 and 7.07 ± 0.85 μmol/h/mg protein, respectively) (Table 1).

    Plasma Lipid Status

    TC, HDL, and LDL levels were not altered in plasma of rats administered with (PhSe)2 and/or acephate (Table 1).

    Plasma TG levels in control rats were 44.67 ± 3.40 mg/dL. Acephate-exposed rats presented TG levels of 62.50 ± 4.17 mg/dL, whereas in acephate-exposed rats pretreated with (PhSe)2 (10 or 30 mg/kg) TG levels were 47.00 ± 2.87 and 48.67 ± 1.20 mg/dL, respectively. The two-way ANOVA of TG levels demonstrated a significant (PhSe)2 × acephate interaction (F2,20= 7.32, p < 0.05). Post-hoc comparisons revealed that acephate increased TG levels of rats if compared to those of the control group. (PhSe)2 pre-treatment at doses of 10 and 30 mg/kg completely protected against the increase of TG levels caused by acephate (Table 1).

    (PhSe)2 at doses of 10 and 30 mg/kg did not alter TG levels in plasma of rats (49.83 ± 4.59 and 37.67 ± 2.90 mg/dL, respectively) (Table 1).

    The cardiovascular risk factor TG/HDL in control rats was 1.13 ± 0.16. Acephate-exposed rats presented TG/HDL of 1.76 ± 0.19, while in acephate-exposed rats pre-treated with both doses of (PhSe)2 TG/HDL were 1.27 ± 0.12 and 1.31 ± 0.07. The two-way ANOVA of the cardiovascular risk factor TG/HDL data yielded a significant (PhSe)2 × acephate interaction (F2,20= 5.96, p < 0.05). Post-hoc comparisons showed that acephate increased TG/HDL ratio when compared to the control group. (PhSe)2 pretreatment at doses of 10 and 30 mg/kg was effective against the increase of TG/HDL ratio caused by acephate (Table 1).

    (PhSe)2 at doses of 10 and 30 mg/kg did not alter TG/HDL ratio of rats (1.35 ± 0.10 and 1.12 ± 0.08, respectively) (Table 1).

    The cardiovascular risk factor TC/HDL and the atherogenic index were not altered in rats administered with (PhSe)2 and/or acephate (data not shown).

    DISCUSSION

    In the current study, we reported the protective effect of (PhSe)2 on metabolic disorders induced by acephate acute exposure in rats. Acephate acute exposure induced hyperglycemia in rats, which was demonstrated by increased glucose levels in plasma. Furthermore, acephate caused an increase of corticosterone levels and hepatic TAT and G6Pase activities, associated with an inhibition of cerebral AChE activity. In addition, acephate induced an increase in plasma TG levels and of the cardiovascular risk factor TG/HDL in rats. (PhSe)2 attenuated these alterations induced by acephate, except for the increase of corticosterone levels and AChE activity inhibition.

    With agreement with Joshi and Rajini (2009) in our study, we demonstrated the hyperglycemic effect of acephate after 2 hours of a single acute administration in rats. (PhSe)2 attenuated the increase of glucose levels caused by acephate, indicating its anti-hyperglycemic property. Accordingly, similar effect of (PhSe)2 was reported when this compound was administered to diabetic rats either induced by alloxan or streptozotocin (Barbosa et al., 2008; Kade et al., 2009).

    Our results showed that acephate stimulates the HPA axis as demonstrated by the increase of corticosterone levels in plasma of rats. Similarly, other authors have demonstrated this effect after acephate and monocrotophos administration in rats (Spassova et al., 2000; Joshi and Rajini, 2009, 2012). Moreover, the activation of HPA axis seems to be related to the inhibition of cerebral AChE activity since the accumulation of ACh stimulates the hypothalamus to release the corticotropin-releasing hormone (Bugajski et al., 2001). In the present study, (PhSe)2 was not effective against the AChE activity inhibition and consequently did not protect against the increase of corticosterone levels. These results suggest that the modulation of the HPA axis is not involved in the anti-hyperglycemic effect of (PhSe)2.

    Besides to the activation of HPA axis, our results showed increased TAT and G6Pase activities in livers of rats exposed to acephate. These results probably are a consequence of the increase of corticosterone levels. (PhSe)2 at a dose of 30 mg/kg partially protected against the increase of TAT activity and completely protected against the increase of G6Pase. These results indicate that the modulation of the gluconeogenic enzyme activities is one of the mechanisms involved in the anti-hyperglycemic effect of (PhSe)2.

    However, one cannot rule out the involvement of other mechanisms in the anti-hyperglycemic effect of (PhSe)2. In fact, in the current study we found that (PhSe)2 at doses of 10 and 30 mg/kg similarly attenuated the rise in glucose levels and differently protected against the increase of TAT and G6Pase activities resulting from acephate exposure. (PhSe)2 at a dose of 10 mg/kg did not protect against the increase of TAT activity and partially protected against the increase of G6Pase activity. Conversely, at a dose of 30 mg/kg (PhSe)2 partially and completely protected against the increase of TAT and G6Pase activities, respectively. Taken these results collectively one can suggest that other mechanisms, besides the modulation of the activity of these gluconeogenic enzymes, may be involved in the anti-hyperglycemic effect of (PhSe)2. However, these mechanisms remain to be elucidated.

    Furthermore, in the present study acephate caused a significant increase in plasma TG levels and of the cardiovascular risk factor TG/HDL. (PhSe)2 completely protected against the increase of plasma TG levels and of the TG/HDL ratio demonstrating its anti-hyperlipidemic and cardioprotective activity. Accordingly, the anti-hyperlipidemic property of (PhSe)2 was demonstrated in other experimental protocols (da Rocha et al., 2009, 2011). However, the exact mechanism by which (PhSe)2 acts as an anti-hyperlipidemic drug remains to be elucidated.

    In conclusion, this study demonstrated that pretreatment with (PhSe)2 was effective against metabolic disorders induced by acephate acute exposure in rats. Further studies are needed to elucidate the exact mechanism by which (PhSe)2 exerted its anti-hyperglycemic and anti-hyperlipidemic properties, since this organoselemium compound could be a promising alternative to minimize metabolic disorders associated with OPs exposure.

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