The effect of adipokinetic hormone on midgut characteristics in Pyrrhocoris apterus L. (Heteroptera) Dalibor Kodrík a,b,⇑ , Konstantin Vinokurov a , Aleš Tomc ˇala c , Radomír Socha a a Institute of Entomology, Biology Centre, Academy of Sciences, Branišovská 31, 370 05 C ˇ eské Bude ˇjovice, Czech Republic b Faculty of Science, University of South Bohemia, Branišovská 31, 370 05 C ˇ eské Bude ˇjovice, Czech Republic c Institute of Organic Chemistry and Biochemistry, Academy of Sciences, Flemingovo Sq. 2, 166 10, Praha 6, Czech Republic article info Article history: Received 5 September 2011 Received in revised form 9 November 2011 Accepted 10 November 2011 Available online 18 November 2011 Keywords: Adipokinetic hormone AKH Digestion Enzyme activity Midgut ESI-MS abstract Digestive processes and the effect of adipokinetic hormone (Pyrap-AKH) on the amount of nutrients (lip- ids, proteins, and carbohydrates), and on the activity of digestive enzymes (lipases, peptidases, and car- bohydrases) were studied in the midgut of the firebug, Pyrrhocoris apterus. The analyses were performed on samples of anterior (AM), middle (MM) and posterior (PM) midgut parts. The results revealed that the digestion of lipids, carbohydrates and proteins take place in the acidic milieu. The Pyrap-AKH treatment increased significantly the level of lipids and proteins in the midgut, and also the level of triacylglycerols (TGs) predominantly in the AM, and the level of diacylglycerols (DGs) in the MM. The increase was not uniform for all present TG and DG species – those containing the linoleic fatty acid were predominant. No hormonal effect on lipase activity was recorded, while peptidase and glucosidase activity was increased in the MM and PM. All these facts indicate that the Pyrap-AKH probably stimulates digestion by more intensive food ingestion or turnover, and perhaps by the stimulation of metabolite absorption; the activation of digestive enzymes seems to be secondary or controlled by other mechanisms. Ó 2011 Elsevier Ltd. All rights reserved. 1. Introduction Feeding of insects represents a complex of physical, chemical and nutritional processes. The regulation of feeding in insects in- volves central excitation that is positively augmented by food as well as by non-food stimuli, both within and outside the animal, and is modulated by deterrent stimuli and by feedback from peripheral systems, such as stretch receptors on the gut wall, hor- mones and blood composition (Simpson and Bernays, 1983). The insect alimentary canal is basically made up of specialised com- partments of various structures depending on their systematic po- sition, life strategy, developmental stage and feeding habits. Generally it consists of the foregut with adjacent salivary glands, midgut and hindgut. The midgut is predominantly involved in the digestive enzymes secretion and nutrient absorption. In insects, possible mechanisms controlling the release of diges- tive enzymes are supposed to be hormonal – paracrine or prandial (food-stimulated enzyme synthesis and secretion) (Lehane et al., 1995); direct neural regulation of this process is considered to be unlikely (Lehane et al., 1996; Sehnal and Z ˇ itn ˇ an, 1996; Chapman, 1998). The midgut endocrine cells express genes for allatostatin, leucomyosuppressin, neuropeptide F and others kinds of neuro- peptides (Audsley and Weaver, 2009), which support the idea that some of them play a pivotal role as non-neural, gut regulatory fac- tors. It is generally accepted that the gut motility is also modulated by neurohormones (FMRF amides, allatostatins, allatotropins, tachykinis, proctolin, diuretic peptides etc.) produced in (or trans- ported by) the stomatogastric nervous system (Audsley and Weaver, 2009). The direct neurohormonal effect on the insect digestive enzymes was described previously for cockroaches. It was shown in Diplop- tera puctata that allatostatin and leucomyosuppressin stimulated both invertase and a-amylase activity in the lumen of isolated mid- gut (Fusé et al., 1999). Similar effect was described for crustacean cardioactive peptide (CCAP) in the American cockroach Periplaneta americana. Correlation was found between the increase of the a- amylase and peptidase activity after the initiation of feeding, and the CCAP level in the midgut endocrine cells. Correspondingly, the 0022-1910/$ - see front matter Ó 2011 Elsevier Ltd. All rights reserved. doi:10.1016/j.jinsphys.2011.11.010 Abbreviations: AEBSF, 4-(2-aminoethyl)benzenesulfonyl fluoride hydrochloride; AKH, adipokinetic hormone; AM, anterior midgut; DGs, diacylglycerols; DNS, 3,5- dinitrosalicylic acid reagent; DTT, dithiotreitol; E-64, L-trans-epoxysuccinyl-L- leucylamido(4-guanidino) butane; EDTA, ethylene diamine tetraacetate; 4-MU butyrate, 4-methylumbelliferyl butyrate; 4-MU, 4-methylumbelliferone; MM, middle midgut; PM, posterior midgut; Pyrap-AKH, Pyrrhocoris apterus adipokinetic hormone; TGs, triacylglycerols; UB, universal buffer. ⇑ Corresponding author at: Institute of Entomology, Biology Centre, Academy of Sciences, Branišovská 31, 370 05 C ˇ eské Bude ˇjovice, Czech Republic. Tel.: +420 387 775 271; fax: +420 385 310 354. E-mail address: kodrik@entu.cas.cz (D. Kodrík). Journal of Insect Physiology 58 (2012) 194–204 Contents lists available at SciVerse ScienceDirect Journal of Insect Physiology journal homepage: www.elsevier.com/locate/jinsphys