Hormonal enhancement of insecticide efficacy in Tribolium castaneum: Oxidative stress and metabolic aspects Ivana Plavšin a,b,c , Tereza Stašková a,b , Michal Šerý a,d , Vlastimil Smýkal a,b , Branimir K. Hackenberger c , Dalibor Kodrík a,b, ⁎ a Institute of Entomology, Biology Centre, Academy of Sciences, Branišovská 31, 370 05 České Budějovice, Czech Republic b Faculty of Science, University of South Bohemia, Branišovská 31, 370 05 České Budějovice, Czech Republic c Department of Biology, Josip Juraj Strossmayer University of Osijek, Ulica cara Hadrijana 8/A, 31000 Osijek, Croatia d Faculty of Education, University of South Bohemia, Jeronýmova 10, 371 15 České Budějovice, Czech Republic abstract article info Article history: Received 13 January 2015 Received in revised form 26 January 2015 Accepted 27 January 2015 Available online 4 February 2015 Keywords: Adipokinetic hormone Insecticide RNA interference Metabolism Oxidative stress Mortality Insect anti-stress responses, including those induced by insecticides, are controlled by adipokinetic hormones (AKHs). We examined the physiological consequences of Pyrap-AKH application on Tribolium castaneum adults (AKH-normal and AKH-deficient prepared by the RNAi technique) treated by two insecticides, pirimiphos- methyl and deltamethrin. Co-application of pirimiphos-methyl and/or deltamethrin with AKH significantly in- creased beetle mortality compared with application of the insecticides alone. This co-treatment was accompa- nied by substantial stimulation of general metabolism, as monitored by carbon dioxide production. Further, the insecticide treatment alone affected some basic markers of oxidative stress: it lowered total antioxidative ca- pacity as well as the activity of superoxide dismutase in the beetle body; in addition, it enhanced the activity of catalase and glutathione-S-transferase. However, these discrepancies in oxidative stress markers were eliminat- ed/reduced by co-application with Pyrap-AKH. We suggest that the elevation of metabolism, which is probably accompanied with faster turnover of toxins, might be responsible for the higher mortality that results after AKH and insecticide co-application. Changes in oxidative stress markers are probably not included in the mech- anisms responsible for increased mortality. © 2015 Elsevier Inc. All rights reserved. 1. Introduction Alternative control of insect pests plays an important role in modern pest management and helps to reduce the amount of classical pesticides deployed. Several studies have shown that the insect endocrine system, especially various neuropeptides, has the potential to be specifically targeted for pest control (Rayne and Oshea, 1997; Hoffmann and Lorenz, 1998; Roeder, 1999; Gäde and Goldsworthy, 2003; Verlinden et al., 2014). In addition to the neuropeptides themselves, the mecha- nisms responsible for their synthesis, transport, secretion, binding, and various physiological and behavioural actions offer numerous targets and multiple modes of action for a novel, neuropeptide-based insect con- trol strategy. Insect stress/metabolic neurohormones are especially suit- able for this purpose because stress imposes a heavy metabolic cost, and interfering with insect's internal stress-reducing mechanisms may suc- cessfully impede critical physiological functions (e.g. reproduction), resulting in a more manageable insect population. Among metabolic neurohormones, a potentially fruitful target is the adipokinetic hormone (AKH) peptide family, one of the largest neuropeptide families in insects (Gäde et al., 1997). Its members predominantly control metabolism via the mobilization of stored products (similar to mammalian glucagon: Goldsworthy, 1994; Alquicer et al., 2009; Bednářová et al., 2013a). Thus, adipokinetic hormones are potentially suitable for pest control (Gäde and Goldsworthy, 2003; Verlinden et al., 2014); by increasing the general metabolism of insects, they accelerate the exchange rate of metabolites, including toxins, which may then result in the intensifica- tion of the toxins' effects (Kodrík et al., 2010; Velki et al., 2011). AKHs are small peptides, from 8 to 10 amino acids long, containing at least two aromatic amino acids; they are synthesised by neurosecre- tory cells of the corpora cardiaca (CC). The effects of AKHs also include stimulation of the heart, locomotor and immune systems, and even reg- ulation of some reproductive processes (Gäde et al., 1997; Van der Horst, 2003; Kodrík, 2008). Besides their role in metabolism, recent studies on AKH function have further increased their attractiveness as a target for pest control by linking the hormone to insect defence re- sponses against oxidative stress (Kodrík et al., 2007; Večeřa et al., 2007; Krishnan and Kodrík, 2012; Bednářová et al., 2013b,c). Oxidative stress can be defined as the overproduction of reactive oxygen species (ROS) or as an imbalance between the organism's production of ROS and their elimination. Oxidative stress may result in the degradation Comparative Biochemistry and Physiology, Part C 170 (2015) 19–27 ⁎ Corresponding author at: Institute of Entomology, Biology Centre, Academy of Sciences, Branišovská 31, 370 05 České Budějovice, Czech Republic. E-mail address: kodrik@entu.cas.cz (D. Kodrík). http://dx.doi.org/10.1016/j.cbpc.2015.01.005 1532-0456/© 2015 Elsevier Inc. All rights reserved. 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