Sulfakinins influence lipid composition and insulin-like peptides level in oenocytes of Zophobas atratus beetles.

Insect sulfakinins are pleiotropic neuropeptides with the homology to vertebrate gastrin/cholecystokinin peptide family. They have been identified in many insect species and affect different metabolic processes. They have a strong influence on feeding and digestion as well as on carbohydrate and lipid processing. Our study reveals that sulfakinins influence fatty acids composition in Zophobas atratus oenocytes and regulate insulin-like peptides (ILPs) level in these cells. Oenocytes are cells responsible for maintenance of the body homeostasis and have an important role in the regulation of intermediary metabolism, especially of lipids. To analyze the lipid composition in oenocytes after sulfakinins injections we used gas chromatography combined with mass spectrometry and for ILPs level determination an immunoenzymatic test was used. Because sulfakinin peptides and their receptors are the main components of sulfakinin signaling, we also analyzed the presence of sulfakinin receptor transcript (SKR2) in insect tissues. We have identified for the first time the sulfakinin receptor transcript (SKR2) in insect oenocytes and found its distribution more widespread in the peripheral tissues (gut, fat body and haemolymph) as well as in the nervous and neuro-endocrine systems (brain, ventral nerve cord, corpora cardiaca/corpora allata CC/CA) of Z. atratus larvae. The presence of sulfakinin receptor transcript (SKR2) in oenocytes suggests that observed effects on oenocytes lipid and ILPs content may result from direction action of these peptides on oenocytes.

New activity of yamamarin, an insect pentapeptide, on immune system of mealworm, Tenebrio molitor.

In insects, two types of the immune responses, cellular and humoral, constitute a defensive barrier against various parasites and pathogens. In response to pathogens, insects produce a wide range of immune agents that act on pathogens directly, such as cecropins or lysozyme, or indirectly by the stimulation of hemocyte migration or by increasing phenoloxidase (PO) activity. Recently, many new immunologically active substances from insects, such as peptides and polypeptides, have been identified. Nevertheless, in the most cases, their physiological functions are not fully known. One such substance is yamamarin - a pentapeptide isolated from the silk moth Antheraea yamamai. This yamamarin possesses strong antiproliferative properties and is probably involved in diapause regulation. Here, we examined the immunotropic activity of yamamarin by testing its impact on selected functions of the immune system in heterologous bioassays with the beetle Tenebrio molitor, commonly known as a stored grains pest. Our results indicate that the pentapeptide affects the activity of immune processes in the beetle. We show that yamamarin induces changes in both humoral and cellular responses. The yamamarin increases the activity of PO, as well as causes changes in the hemocyte cytoskeleton and stimulates phagocytic activity. We detected an increased number of apoptotic hemocytes, however after the yamamarin injection, no significant variations in the antibacterial activity in the hemolymph were observed. The obtained data suggest that yamamarin could be an important controller of the immune system in T. molitor.

The activity of the nonsulfated sulfakinin Zopat-SK-1 in the neck-ligated larvae of the beetle Zophobas atratus.

Insect sulfakinins (SKs) are multifunctional neuropeptides structurally and functionally homologous to the mammalian gastrin/cholecystokinin (CCK). It has been proposed that SKs play a role in modulating energy management in insects by interacting with adipokinetic hormone (AKH), the principle hormone controlling insect intermediary metabolism. To exclude head factors (including AKH) that influence the activity of the nonsulfated sulfakinin Zopat-SK-1 in the larvae of the beetle Zophobas atratus, ligature and in vitro bioassays were used. Our study showed that in the neck-ligated larvae, Zopat-SK-1 evoked a much more pronounced glycogenolytic effect in fat body tissue and a significantly higher hypertrahelosemic effect in hemolymph than in larvae without ligation. We found that the concentration of the sugar trehalose increased under hormonal treatment but no changes in glucose levels were observed. Under in vitro conditions, the maximal glycogenolytic effect of Zopat-SK-1 in fat body was observed at 10 pmol of hormone. Ligature and in vitro bioassays indicated that Zopat-SK-1 activity in the Z. atratus larvae is modulated by head signals and/or factors from the gastrointestinal tract. Our data indicate the existence of a brain-gastrointestinal axis that has a role in controlling of energy (carbohydrate) metabolism in the insect body. Moreover, these results, together with immunological evidence of a cholecystokinin-like (sulfakinin) receptor in the Z. atratus fat body, help us to better understand the SK signaling pathways and its physiological role in insect biology.

The pro-apoptotic action of the peptide hormone Neb-colloostatin on insect haemocytes.

The gonadoinhibitory peptide hormone Neb-colloostatin was first isolated from ovaries of the flesh fly Neobellieria bullata. This 19-mer peptide is thought to be a cleaved product of a collagen-like precursor molecule that is formed during remodelling of the extracellular matrix. In this study, we report that upon injection of picomolar and nanomolar doses, this peptide exerts a pro-apoptotic action on haemocytes of Tenebrio molitor adults, as visualized by changes in morphology and viability. The F-actin cytoskeleton was found to aggregate into distinctive patches. This may be responsible for the observed inhibition of adhesion of haemocytes and for the stimulation of filopodia formation. However, Neb-colloostatin injection did not induce the formation of autophagic vacuoles. Our results suggest that physiological concentrations of Neb-colloostatin play an important role in controlling the quantity and activity of haemocytes in insect haemolymph. They also suggest that during periods in which Neb-colloostatin is released, this peptide may cause a weakening of the insects' immune system. This is the first report that exposure to a peptide hormone causes apoptosis in insect haemocytes.

Synthesis, cardiostimulatory, and cardioinhibitory effects of selected insect peptides on Tenebrio molitor.

The subject of these studies was a search for proctolin antagonists among peptides originating from insect species because the proctolin antagonists constantly pose a problem. During these studies we performed the synthesis of the following peptides: a native decapeptide from Manduca sexta Mas-MT-I and its 11 analogs with shortened sequences at the N-end as well as a growth suppressor, a pentapeptide isolated from Antheraea yamamai, Any-GS and its 10 analogs, modified at position 1 and with a shortened peptide chain. Biological effects were evaluated by the cardiotropic test on the semi-isolated heart of the insect species Tenebrio molitor. Mas-MT-I and six analogs stimulate the heartbeat frequency, especially [6-10]-Mas-MT-I, whereas the [4-10]-Mas-MT-I analog shows a strong inhibition of the heartbeat frequency, if insect. The Any-GS and the analogs [Gln(1)]- and [Gly(1)]-Any-GS also show a strong cardioinhibitory effect.

Gonadoinhibitory effects of Neb-colloostatin and Neb-TMOF on ovarian development in the mealworm, Tenebrio molitor L.

The gonadostatic action of the peptides Neb-colloostatin (SIVPLGLPVPIGPIVVGPR) and Neb-TMOF (NPTNLH) from Neobellieria bullata was studied in female mealworm Tenebrio molitor. Both peptides potently inhibit ovarian development and terminal oocyte maturation of mated females during their first reproductive cycle. Injection of 4 mug of Neb-colloostatin or Neb-TMOFNeb-TMOF reduced, at day 4 of the cycle, the size of the terminal oocytes to about half or one third of the normal size in saline-injected controls. In addition, follicular patency was arrested. The injections of Neb-colloostatin and Neb-TMOF also caused a delay to the first ovulation and oviposition as well as a reduction of the number of eggs by about 50% in the first 3 days of the oviposition period. At 4 days after adult emergence, none of the peptides had caused significant changes in protein concentration or composition of the haemolymph. However, both peptides reduced total protein content in ovaries and induced qualitative changes in ovarian protein patterns. Electrophoretic analyses indicated that Neb-colloostatin and Neb-TMOF caused a loss of two proteins (150, 180 kDa) and a drastic reduction of 4 others (39, 43, 47, 130 kDa), which are the most abundant ones in ovaries of control females. On the other hand, they increased the concentration of 2 other polypeptides (65, 70 kDa), which normally occur in insignificant quantities in ovaries. Our results indicate that both peptides have a very similar mode of action despite the differences in their amino acid sequence. They seem to interfere with vitellogenin production by the fat body as well as with vitellogen uptake by the oocytes through modification of patency.

Cardioacceleratory action of tachykinin-related neuropeptides and proctolin in two coleopteran insect species.

Several cardioactive peptides have been identified in insects and most of them are likely to act on the heart as neurohormones. Here we have investigated the cardioactive properties of members of a family of insect tachykinin-related peptides (TRPs) in heterologous bioassays with two coleopteran insects, Tenebrio molitor and Zophobas atratus. Their effects were compared with the action of the pentapeptide proctolin. We tested the cardiotropic activity of LemTRP-4 isolated from the midgut of the cockroach Leucophaea maderae, CavTK-I and CavTK-II isolated from the blowfly Calliphora vomitoria. The semi-isolated hearts of the two coleopteran species were strongly stimulated by proctolin. We observed a dose dependent increase in heartbeat frequency (a positive chronotropic effect) and a decrease in amplitude of contractions (a negative inotropic effect). In both beetles the TRPs are less potent cardiostimulators and exert lower maximal frequency responses than proctolin. LemTRP-4 applied at 10(-9)-10(-6) M was cardiostimulatory in both species inducing an increase of heart beat frequency. The amplitude of contractions was stimulated only in Z. atratus. CavTK-I and CavTK-II also exerted cardiostimulatory effects in Z. atratus at 10(-9)-10(-6) M. Both peptides stimulated the frequency, but only CavTK-II increased the amplitude of the heart beat. In T. molitor, however, the CavTKs induced no significant effect on the heart. Immunocytochemistry with antisera to the locust TRPs LomTK-I and LomTK-II was employed to identify the source of TRPs acting on the heart. No innervation of the heart by TRP immunoreactive axons could detected, instead it is possible that TRPs reach the heart by route of the circulation. The likely sources of circulating TRPs in these insects are TRP-immunoreactive neurosecretory cells of the median neurosecretory cell group in the brain with terminations in the corpora cardiaca and endocrine cells in the midgut. In conclusion, LemTRP-4, CavTK-I and CavTK-II are less potent cardiostimulators than proctolin and also exert stimulatory rather than inhibitory action on amplitude of contractions. The differences in the responses to proctolin and TRPs suggest that the peptides regulate heart activity by different mechanisms.

[Polish multicenter study on type I diabetes incidence in the age group 15-29 between 1998 and 1999]. / Wieloosrodkowe badania nad zapadalnoscia na cukrzyce typu 1 w grupie wiekowej 15-29 lat w Polsce w latach 1998-1999.

The increase in diabetes type 1 incidence observed in various centers in Poland and the need for a centralized study covering a large population have resulted in the construction of a standardized registry of type 1 diabetes in 1998 within the Polish Multicenter Study in Diabetes Epidemiology. The aim of the study was to evaluate the incidence of type 1 diabetes in the age group 15-29 in 5 distinct regions of Poland (Krakow, Warsaw, Bialystok, Rzeszow and Olsztyn centers) with over 15% of the Polish population at risk in 1998 and 1999. The data for the standardized registry were obtained prospectively from hospital departments and diabetes outpatient units. The incidence rates calculated in 1998 showed the highest value of 11.2/100,000 for Krakow and its region, and the lowest (4.4/100,000) for Bialystok and its region. In 1999 the highest value of 12.3/100,000 was noted in Olsztyn and its region and the lowest (3.4/100,000) in Warsaw. There were significant differences in the incidence rates between the study centers were found. Incidence rates in the whole study area were significantly higher among males as compared with females in 1998 and 1999 (8.9/100,000 vs. 4.9/100,000; p = 0.0001), marked in the age groups 15-19 and 20-24 (p = 0.001 and p = 0.002 respectively). A significant increase in diabetes type 1 incidence (from 4.6/100,000 to 6.9/100,000) was found as compared with results of the "Three Cities Study" (1986-1988).

Myotropic effects of new proctolin analogues modified in the position 5 of peptide chain in insects.

To explain the role of the Thr5 residue of proctolin (Arg-Tyr-Leu-Pro-Thr) in the myotropic activity of this insect neuropeptide, we synthesized two groups of its analogues: 1) Arg-Tyr-Leu-Pro-X-OH with X = Val (1), D-Val (2), Ile (3), D-Ile (4), Ala (5), D-Ala (6), Asn (7), Gln (8), Ser (9), Pro (10), Phe (11), Asp (12), Glu (13), Arg (14), D-Arg (15), Lys (16) and Gly (17) and 2) Arg-Tyr-Leu-Pro-R', where R' = isobutylamine (18), S-1-methyl-1-phenylmethylamine (19), R-1-methyl-1-phenylmethylamine (20), R-2-amino-1-propanol (21), S-2-amino-1-propanol (22), R-1-amino-2-propanol (23), S-2-amino-1-propanol (24), 3-amino-1-propanol (25). Decapeptide proctolylproctolin (H-Arg-Tyr-Leu-Pro-Thr-Arg-Tyr-Leu-Pro-Thr-OH) (26) was synthesized. Syntheses of these peptides were carried out by solid-phase method. All peptides were bioassayed in vitro on the semi-isolated hearts of Tenebrio molitor using a cardioexcitatory test and on the foregut of locust (Schistocerca gregaria). Peptides 1, 3, 5, 9, 13, 14, 16, 22, and 23 retained about 30-50% of the cardioexcitatory activity in T. molitor. Analogues 1 and 3 preserved about 50% and analogue 8 about 80% of the myotropic activity, whereas compound 4 and 9 showed a very weak contractile activity in S. gregaria.

Synthesis and biological evaluation of selected insect neuropeptide analogs modified by D- or L-phenylglycine derivatives.

Novel analogs, modified by L- or D- phenylglycine and p-substituted derivatives, of the neuromodulator proctolin (Arg-Tyr-Leu-Pro-Thr) and of the Trypsin Modulating Oostatic Factor from the gray flesh fly Neobellieria bullata (Neb-TMOF-Asn-Pro-Thr-Asn-Leu-His) were synthesized and checked for activity. Proctolin analogs were modified at position 2: Arg-Phg-Leu-Pro-Thr (I), Arg-D-Phg-Leu-Pro-Thr (II), Arg-Phg(p-OH)-Leu-Pro-Thr (III), Arg-D-Phg(p-OH)-Leu-Pro-Thr (IV), Arg-Phg(p-NO2)-Leu-Pro-Thr (V) Arg-D-Phg(p-NO2)-Leu-Pro-Thr (VI), Arg-Phg(p-NH2)-Leu-Pro-Thr (VII), Arg-D-Phg(p-NH2)-Leu-Pro-Thr (VIII), Arg-Phg(p-N,N-di-Me)-Leu-Pro-Thr (IX), Arg-D-Phg(pp-N,N-di-Me)-Leu-Pro-Thr (X) while analogs of Neb-TMOF underwent modifications at position 6: Asn-Pro-Thr-Asn-Leu-Phg(p-NO2) (XI), Asn-Pro-Thr-Asn-Leu-D-Phg(p-NO2) (XII), Asn-Pro-Thr-Asn-Leu-Phg(p-NH2) (XIII), Asn-Pro-Thr-Asn-Leu-D-Phg(p-NH2) (XIV), Asn-Pro-Thr-Asn-Leu-Phg(p-N,N-di-Me) (XV), Asn-Pro-Thr-Asn-Leu-D-Phg(p-N,N-di-Me) (XVI). Earlier studies on proctolin demonstrated that the presence of the -CH2- group between C-alpha and the phenyl ring at position 2 of the peptide chain is important for the myotropic activity. Based on these results, we replaced Tyr at position 2 by different phenylglycine derivatives, lacking the methylene group at the side chain. Myotropic activity of the proctolin analogs was assayed in vitro on the semi-isolated heart of the mealworm Tenebrio molitor and on the foregut of the locust Schistocerca gregaria. All analogs (I-X) were practically inactive. For Neb-TMOF, it was previously demonstrated that the exchange of His-6 by p-substituted Phe-derivatives, especially by Phe(p-NH2), an amino acid containing a basic function, results into analogs which inhibit trypsin biosynthesis in the gray fleshfly. For this reason these new Neb-TMOF analogs with L- or D-phenylglycine p-substituted derivatives at position 6, were developed and tested (in vivo) in the trypsin biosynthesis assay of the gray fleshfly N. bullata. Only analogs XV and XVI slightly inhibited trypsin biosynthesis in the midgut. Because more than 50% of the injected animals died and none of the surviving animals ate much of the liver meal, the lower trypsin level in the gut might be a indirect effect. Other peptides (XI-XIV) had no effect on the level of trypsin biosynthesis in the midgut.

Further proctolin analogues modified in the position 2 of the peptide chain and their myotropic effects in insects Tenebrio molitor and Schistocerca gregaria.

We have extended our studies on the structure-activity relationship in neuropeptide proctolin (Arg-Tyr-Leu-Pro-Thr) by evaluating the effects of a series of proctolin analogues modified in position 2 of the peptide chain, including: [Phe(p-Cl)2]- (1), [D-Phe(p-Cl)2]- (2), [N-Me-Tyr2]- (3), [D-Phe(p-NH2)2]- (4), [D-Phe(p-N,N-di-Me)2]- (5), [N-Me-Tyr(OMe)]- (6), [D-3-Pal2]- (7), [L-Nal2]- (8), [D-Nal2]- (9), [Lys(Nic)2]- (10), [D-Lys(Nic)2]- (11), [D-Phe-(p-NO2)2]- (12). These peptides were evaluated for myotropic activity on the heart of Tenebrio molitor and contractile activity of the foregut of Schistocerca gregaria. Analogues 1-5, 7-9, and 12 retained a weak cardiotropic activity in Tenebrio molitor while peptides 1, 8 and 12 preserved 15-25% of the locust-gut contracting activity of proctolin. Peptides 2, 4 and 7 showed weak inhibitory activity in Schistocerca gregaria foregut, whereas only peptides 4 and 7 reduced the maximum response to applied proctolin by 64% and 49% respectively, at the 10(-6) M concentration.

Proctolin, an insect neuropeptide.

Synthetic, biological and conformational studies on the insect neuropeptide proctolin (Arg-Tyr-Leu-Pro-Thr) and some of its analogues are reviewed.

New proctolin analogues and their myotropic effects on heart of yellow mealworm Tenebrio molitor L. and foregut of locust-Schistocerca gregaria L.

We have extended our work on structure/activity relationship of neuropeptide proctolin (H-Arg-Tyr-Leu-Pro-Thr-OH) by evaluating the effects of the following proctolin analogues: H-X1-Tyr-Leu-Pro-Thr-OH, where X1 = D-Arg (1), N-Me-Arg (2), Can (3), D-Tyr2, D-Leu3, D-Thr5]-proctolin (12). In analogues 1-9, the N-terminal Arg-residue was replaced by basic amino acid derivatives with peptides containing amino acid residues with an isosteric system on the back side chain relative to Arg (compounds 3, 5 and 6) or homo-Arg (compound 7). Analogues 1-12 were evaluated for myotropic action on in vitro heart preparation of Tenebrio molitor, whereas peptides 2, 5 and 7-12 were tested for contractile action on isolated foregut of Schistocerca gregaria. Peptides 2 and 3 retained full cardiotropic activity in Tenebrio molitor while peptides 5 and 7 preserved 40% and 15%, respectively, locust-gut contracting activity of proctolin. Peptides 11 and 12 showed antagonistic activity in Schistocerca gregaria foregut.

New proctolin analogues modified by D-amino acids in the peptide chain and their high cardioexcitatory effect on Tenebrio molitor.

The object of our studies was the synthesis and conformational and biological evaluation of the series of 14 analogues of the insect neuropeptide, proctolin. The analogues were obtained by replacement of the native L-amino acids by their D-isomers in one, two, and all positions. Biological effects of the peptides were examined by cardioexcitatory test on the heart of yellow mealworm, Tenebrio molitor, in vitro. In biotest performed on insects, D-Arg-D-Tyr-D-Leu-D-Pro-D-Thr, [D-Arg(N-G-nitro)1,D-Leu3]-, [D-Arg1,D-Leu3]-, [D-Tyr2,D-Thr5]- and [D-Arg1,D-Pro4]-proctolin exert high agonistic activity of proctolin on the heart of insects at 10(-11) - 10(-10) M concentrations. The proctolin analogue containing only D-amino acid residues in the peptide chain unexpectedly shows a much higher cardioexcitatory effect than the native peptide. Moreover, preliminary CD and NMR conformational studies show that proctolin analogues investigated here seem to prefer rather ordered structures, although their conformations differ in some cases.

[Incidence of the most important metabolic disorders in the population of a little town in Poland]. / Czestosc wystepowania najwazniejszych zaburzen metabolicznych wsród ludnosci malego miasta w Polsce.

The study aimed at establishing a risk for metabolic diseases in the randomly selected population in the age over 18 years, inhabitants of the little town in Siedlce Voivodeship. Using comparative tables, in was found that percent of women and men in the investigated group is similar to that in the population of this town. Women prevailed slightly in the investigated group which characterized also relatively low age (42 years) and low diastolic blood pressure. Obesity was found in 40% of the investigated subjects. Over 2.6% had diabetes mellitus and 11.4%--improper glucose tolerance (according to WHO criteria). Every fifth subject has blood lipids disorders. Such disorders were more frequent in the subgroup of men. Additionally, relationships between these findings were analysed. Body weight index markedly correlated with blood triglycerides level and less pronounced with blood total cholesterol and uric acid. Negative correlation between BMI and HDL cholesterol levels was noted. The lack of correlation between glycaemia, cholesterolemia and triglyceridemia results from the fact that there is such a correlation in men but not in women. The obtained results indicate the necessity of preventive measures.

New proctolin analogs modified in position 3 of peptide chain--synthesis and their biological evaluation.

Six proctolin analogs modified in position 3 of peptide chain such as Arg-Tyr-X-Pro-Thr where X = Gly (1), Val (2), Pro (3), Thr (4), Acp (1-aminocyclopentane-1-carboxylic acid residue) (5), and Ach (1-aminocyclohexane-1-carboxylic acid residue) (6) were synthesized by liquid-phase method. Biological effects of the pentapeptides (1-6) were examined in cardiostimulatory test in vitro in respect to two insect species: American cockroach (Periplaneta americana L.) and yellow mealworm (Tenebrio molitor L.). Results thus obtained pointed out that the presence of L-leucine in the position 3 of proctolin skeleton plays important role in its cardiotropic activity in insects.

Insect peptide hormones, an overview of the present literature.

A comprehensive overview of the recent state of the art of insect peptide hormones with chemical structures is presented. An increased interest in insect neuropeptides and dynamic development of that research area has been influenced by a rapid improvement of instrumentation necessary for isolation and structural characterization. Several research teams have studied the relationships between biological properties of insect and vertebrate peptide hormones. Thus hormones from the AKH family can be considered glucagon counterparts, whereas the myotropic hormones such as proctolin and Lem-PK (LPK) are a substance P equivalent. Insect melanization hormones Bom-MRCH in their structural characteristics and properties resemble those of mammal MSH, and leucosulfakinins Lem-SK-I and -II show some similarities with gastrin II and cholecystokinin. Bombyxin-II (Bom-PTTH-II) reveals a structural homology with human insulin and similar biological properties to adenocorticotropic mammal hormone. Allatostatin (Dip-JHS-I) may be compared to somatostatin as it can be inferred from the observations that this peptide modulates JH secretion in cockroach, Blattella germanica. Determination of the primary structure of eclosion hormones Mas-EH and Bom-EH-II as well as the amino acid sequence of allatotropin and allatostatin is a significant contribution to the understanding of the molecular mechanisms of metamorphosis and insect development.

Further studies on proctolin analogues modified in position 2 of the peptide chain and their influence on heart-beat frequency of insects.

Seven proctolin analogues (I-VII) modified in position 2 of the peptide chain by Phe (p-guanidino) (I), Phe (p-OEt) (II), Tyr (3'-NH2) (III), Tyr (3'-NO2) (IV), Afb (p-OH) (V) (Afb = 3-amino-4-phenyl-L-butyric acid), Afb (p-NH2) (VI), Afb (p-NO2) (VII), and the tetrapeptide Tyr (3'-NH2)-Leu-Pro-Thr (VIII) were synthesized by the classic liquid-phase method. The biological effects of the peptides were investigated in cardioexcitatory tests on two insect species, the cockroach Periplaneta americana L., and the yellow mealworm, Tenebrio molitor L. Within physiological concentrations (10(-9)-10(-7) M) peptides II, III, and IV stimulated the heart action of P. americana like proctolin itself. Under identical conditions, in the case of T. molitor, only peptide III showed cardiostimulatory properties, whereas other compounds (including II and IV) were inactive at concentrations up to 10(-7) M. Results reported here reflect, with reference to the analogues I-VII, selective recognition of receptors on myocardium of both insect species. The tetrapeptide VIII revealed a weak deacceleratory effect on P. americana and T. molitor heart action.

The primary structure of the hypertrehalosemic neuropeptide from tenebrionid beetles: a novel member of the AKH/RPCH family.

A hypertrehalosemic neuropeptide from the corpora cardiac of the two tenebrionid beetle species, Tenebrio molitor and Zophobas rugipes, was purified by high performance liquid chromatography, and its sequence determined by pulsed-liquid phase sequencing employing Edman degradation after deblocking enzymatically the N-terminal pyroglutamate residue. Additionally, the C-terminus of the peptide was blocked as shown by the lack of breakdown using carboxypeptidase. In both species an identical octapeptide, designated Tem-HrTH, with the following amino acid sequence, was found: pGlu-Leu-Asn-Phe-Ser-Pro-Asn-Trp-NH2. This primary sequence has an 88% homology with the hypertrehalosemic hormone I (Pea-CAH-I) from the American cockroach as well as with the red pigment-concentrating hormone (RPCH) of prawns. Injection of the synthetic peptide into larvae or young adults of T. molitor or adult Z. rugipes increases the hemolymph carbohydrate levels in a dose-dependent manner. Thin layer chromatography identified the elevated sugar component of the hemolymph as the disaccharide trehalose. Carbohydrate release from larval fat body in vitro was also shown upon administration of a low concentration of synthetic Tem-HrTH.

Role of guanidine group at the N-terminal proctolin chain in cardioexcitatory effects in insects.

Six proctolin analogues (I-VI) modified in position 1 of the peptide chain by the following amino acids: homo-Arg, Gac, Gav, Gap, Phe (p-guanidino) and Orn, were synthesized by conventional liquid phase method. The myotropic activity of the obtained peptides was investigated in cardioexcitatory test on two insect species, cockroach, Periplaneta americana L., and yellow mealworm, Tenebrio molitor.