Cockroach Blaberus Discoidalis Research Articles

Contrasting modulations of gene expression by a juvenile hormone analog

Decapitation altered the pattern of in vitro protein synthesis by the fat body of adult female Blaberus discoidalis cockroaches. Several polypeptides showed marked diminution, whereas two polypeptides showed markedly increased synthesis. The polypeptide profile was restored to normal in decapitated cockroaches by treatment with the juvenile hormone analog (JHA) methoprene. Cloned cDNAs that were isolated represented two fat body transcripts: one transcript was strongly stimulated and the other strongly suppressed by JHA. JHA acted on fat bodies in vitro to modulate the levels of the two transcripts. These studies indicate a system where juvenile hormone may provoke vividly contrasting gene expressions in the same tissue at the same time.

Characterisation of neuropeptides of the AKH/RPCH-family from corpora cardiaca of Coleoptera.

Extracts of corpora cardiaca from two members of the family Tenebrionidae. Zophobas rugipes and Tenebrio molitor, from one member of the Chrysomelidae, Leptinotarsa decemlineata, and from three members of the Scarabaeidae, Pachnoda marginata, P. sinuata and Melolontha hippocastani, were assayed for adipokinetic and hypertrehalosaemic activity in acceptor locusts (Locusta migratoria) and cockroaches (Periplaneta americana), respectively. All corpus cardiacum material tested, except that from the cockchafer, M. hippocastani, gave positive bioassay results. Biological activities of corpus cardiacum extracts from all species investigated can be resolved on reversed-phase high performance liquid chromatography (RP-HPLC). Gland extracts from the two tenebrionid species each show a single peak of biological activity associated with a single peak of UV absorbance having an identical retention time in both species. The two biologically active fractions from the corpora cardiaca of the potato beetle, L. decemlineata, coelute with exogenous (synthetic) hypertrehalosaemic hormones I and II of the American cockroach. The two species of the genus Pachnoda contain two active compounds in their glands; compound I of each species is more abundant and elutes just ahead of the (synthetic) hypertrehalosaemic hormone of the cockroach Blaberus discoidalis. The gland material of M. hippocastani exhibits an absorbance peak with the same retention time as the major peak from the Pachnoda-species; however, this peak material does not elicit biological activity in the assays used here. After fractionation by RP-HPLC the main biologically active compounds were subjected to amino acid analyses. All factors are peptidic and contain 8 amino acid residues.(ABSTRACT TRUNCATED AT 250 WORDS)

Neuroendocrine and juvenile hormone effects on fat body protein synthesis during the reproductive cycle in female Blaberus discoidalis cockroaches

Ovarian protein content and fat body protein synthesis were measured during the first gonotrophic cycle in virgin female Blaberus discoidalis cockroaches. Protein systhesis was measured for in vitro fat bodies from animals treated with combinations of a juvenile hor mone analog (JHA = methoprene) and corpora cardiaca (CC) extracts. Ovarian protein content began to increase on Day 5 of adult life and reached its maximum at Days 20–22. Synthesis of proteins secreted by the in vitro fat body increased by 12-fold between emergence and its maximum on Day 18, then declined to nearly its original level by Day 33. Synthesis of nonsecreted, fat body proteins increased by 4-fold between emergence and a maximum on Day 21, then declined. CC extracts and JHA were administered to decapitated females to determine their regulative effects on fat body protein synthesis. The synthesis by the fat body of nonsecreted proteins was increased by both JHA and CC extracts. In contrast, synthesis of secreted proteins increased only in the presence of JHA. CC extracts, alone, had no effects on the synthesis of secreted proteins, but administration of CC extracts in combination with JHA increased the synthesis of the secreted proteins by 55% above that observed with JHA alone. SDS-PAGE of proteins secreted into the medium by the in vitro fat body demonstrated that JHA stimulated the synthesis of specific polypeptides, whereas CC extracts did not affect the synthesis of specific-secreted polypeptides. These results suggest that JH regulates specific protein synthesis by the fat body of B. discoidalis; neurohormones elevate the general capacity of the fat body for protein synthesis and amplify the specific effects of JH.

Properties of an in vitro bioassay for hypertrehalosemic hormone of Blaberus discoidalis cockroaches

The ability of the fat body to respond to the hypertrehalosemic action of corpora cardiaca-allata (CC + CA) extracts was measured in vitro with tissue from adult, male Blaberus discoidalis cockroaches. Only trehalose increased in the hemolymph when gland extracts were injected in vivo; likewise, only trehalose levels responded when fat bodies were exposed to gland extracts in vitro. Although fat bodies from 0-day-old animals gave the least response to gland extracts, all animals 5 or more days old showed elevated responses. A linear dose-response occurred between 0.02 and 0.08 CC + CA pair. Fat bodies did not show an in vitro, hypertrehalosemic response to insulin, glucagon, proctolin, dopamine, epinephrine, norepinephrine, serotonin, octopamine, or tyramine. A strong hypertrehalosemic response was found to both synthetic adipokinetic hormone and red-pigment concentrating hormone; however, reversed-phase, thin-layer chromatography of CC + CA extracts and adipokinetic hormone indicated that the hypertrehalosemic effect and the adipokinetic hormone were separate. This in vitro system comprises a relatively rapid, simple, and accurate bioassay for the hypertrehalosemic neurohormone of insects.

Endocrine regulations of ovarian maturation in the cockroach Blaberus discoidalis

1. 1. The first gonotrophic cycle of oocyte maturation was defined for virgin Blaberus discoidalis cockroaches. 2. 2.A high correlation was found between growth of the basal oocytes and increased ovarian protein content. 3. 3.Allatectomy blocked ovarian protein formation, whereas JH-III administration compensated for the loss of the corpora allata. 4. 4.Experiments involving neuro-endocrine surgery and hormone replacement therapy indicated that JH, alone, could stimulate ovarian protein formation; however, the addition of neuroendocrine extracts enhanced the JH effects. 5. 5.The data suggest that both JH and neurohormones stimulate aspects of ovarian protein formation in this insect.

Isolation of neurosecretory granules containing cytochromogenic hormone activity from the corpora cardiaca of Blaberus discoidalis cockroaches

Neurosecretory granules (NSG) were isolated by differential centrifugation from a homogenate of the corpora cardiaca-corpora allata complex of newly emerged, adult, female Blaberus discoidalis cockroaches. By centrifuging at 11,000 g, the procedure yielded a final pellet that was enriched in small, membrane-bound vesicles (103 nm mean diameter). This pellet contained 61% of the cytochromogenic activity present in the orginal homogenate at an 11-fold increase in concentration. A pellet formed at 4000 g contained a mixture of mitochondria and larger NSG (146 nm mean diameter) and most of the hindgut-stimulating hormone activity. This procedure provided a quick, simple method for purifying neurosecretory granules and separating them into two categories of size and endocrine content.

Reverse heteropyknosis between euchromatin and heterochromatin of the X chromosome in meiotic prophase of the cockroach Blaberus discoidalis

The X chromosome and all autosomes of the cockroach Blaberus discoidalis (2n = 37, XO) contain large segments of constitutive heterochromatin (C-bands). From spermatogonial metaphases, the C-bands are found to be located in the middle of most chromosomes, including the X. The C-bands show bright fluorescence when stained with Hoechst 33258. In pachytene, autosomal heterochromatin can be easily identified in acetic orcein squash preparations as condensed segments. In the same preparations the X chromosome exhibits a "closed" appearance with a lightly stained middle loop and two heavily stained terminal segments which lie side-by-side as if they are paired. In C-banded preparations, an opposite reaction is found, i.e., the loop is heavily stained while the tips are lightly stained. In Hoechst 33258 preparations, the loop is brightly fluorescent while the tips are less brightly stained. Thus, in pachytene of conventional orcein preparations the heteropyknotic behavior between euchromatin and heterochromatin of the X chromosome is the reverse of the usual staining pattern; instead of condensed heterochromatin and decondensed euchromatin, euchromatin is condensed whereas heterochromatin is decondensed. The "paired" euchromatic tips may suggest autologous homology between the original X and the original Y which might have been translocated onto the X in the course of karyological evolution.

Development and endocrine effects on the synthesis of mitochondrial proteins in the fat body of the cockroach, Blaberus discoidalis

The pattern of mitochondrial protein synthesis was observed in the fat body during postemergence maturation of adult male Blaberus discoidalis cockroaches. The incorporation rate of [ 14C]-leucine into mitochondrial proteins was high between days 0 and 4 of adult life then decreased by about 40% to a lower, stable rate between days 6 and 30. Cycloheximide-insensitive, intramitochondrial protein synthesis paralleled this pattern. The half life of [ 14C-guanidino]-arginine was 36.2 days in mitochondrial proteins which suggests that the mitochondria are stable organelles in the fat body. Cervical ligation lowered the rate of mitochondrial protein synthesis by 60%, but attempts to return the synthetic rate to normal by injections of corpora cardiaca extracts were inconclusive.

Cytochromogenic factor: a newly discovered neuroendocrine agent stimulating mitochondrial cytochrome synthesis in the insect fat body.

Abstract Evidence is presented for a neurohormone that stimulated by fivefold the synthesis of cytochromes aa 3 + b in the fat body of young, adult, male Blaberus discoidalis cockroaches. This newly discovered agent is designated as the “cytochromogenic factor” (CGF). CGF was concentrated in the corpora cardiaca (CC). A CGF bioassay was developed with decapitated animals that had a threshold response at 0.01 CC pair and showed a linear dose response up to a maximum of 0.08 CC pair. A 24- to 48-hr latent period was needed for the maximum in vivo response to CGF. CGF activity was heat labile and was inactivated by chymotrypsin; partially inactivated by carboxypeptidases A and Y and aminopeptidase M; but not by trypsin. Other hormones such as juvenile hormone (III), 20-hydroxyecdysone, proctolin, and triiodothyronine failed to elicit the cytochromogenic response.

Age and endocrine effects on fat body metabolite composition in adult male Blaberus discoidalis cockroaches

AbstractEndocrine‐and age‐related changes in metabolite contents were used as a measure of fat body development and function in adult male Blaberus discoidalis cockroaches. Fat body dry weight, protein, glycogen, and lipid levels coordinately decreased at days two to three of adult life, increased at day four, then declined to lower stable levels by days six to seven. The metabolite contents remained stable from days seven through ten and were still at this level on day 30. Long‐term endocrine deficiencies resulting from corpora cardiaca‐allata extirpation did not change the amounts of metabolites present at 30 days of age. The fat body DNA content remained constant regardless of age or endocrine balance. Unlike other metabolites, the RNA content increased to twice its original level during the first six days of adult life and was unaffected by either age or endocrine balance thereafter. The findings suggest that the adult fat body changes from a storage function at the time of the nymphal‐adult molt to a biosynthetic function by day six. The transition is independent of endocrine control by the corpora cardiaca‐allata complex.

Development and endocrine regulation of mitochondrial cytochrome biosynthesis in the insect fat body: δ-[ 14C]aminolevulinic acid incorporation

The rate of incorporation of [ 14C]aminolevulinic acid (ALA) into cytochrome hemes was used to measure mitochondrial cytochrome synthesis in the fat body of adult male Blaberus discoidalis cockroaches. The hemes of cytochromes aa 3+ b and c+ c 1, were chemically separated to observe differential rates in their synthesis and regulation. [ 14C]ALA was linearly incorporated into cytochrome hemes for at least 8 h. No significant pool of endogenous ALA was detected relative to the amount of administered [ 14C]ALA. Peak cytochrome synthesis occurred 4 to 6 days after adult emergence. Endocrine disruption by corpora cardiaca-corpora allata extirpation or cervical ligation eliminated the 4-day developmentally related increase in the rate of cytochrome aa 3+ b synthesis but had no effect on the production of cytochromes c+ c 1. Injections of corpora cardiaca extracts into cervically ligated animals stimulated the rate of production of cytochromes aa 3+ b by 2.5 times but did not affect cytochromes c+ c 1. By comparison, juvenile hormone injections did not affect the rate of synthesis of either cytochrome fraction. These findings indicate that a neurohormone regulates the rate of synthesis of cytochromes a+ b in insect fat body mitochondria.

Development and endocrine regulation of mitochondrial cytochrome biosynthesis in the insect fat body: δ-Aminolevulinic acid synthase

Aminolevulinic acid (ALA) synthase activity was measured in fat body mitochondria from adult male Blaberus discoidalis cockroaches. The enzyme reached its maximum activity at 4 to 6 days of adult age and then dropped to a minimal level which was maintained throughout the remainder of the study period. ALA synthase activity was doubled by allylisopropylacetamide and showed a half-life of about 6 h at 25 °C. Enzyme activity was depressed by long-term allatectomy. However, juvenile hormone administration in vivo did not significantly stimulate the enzyme relative to appropriate controls, and endocrine regulation of fat body ALA synthase remains inconclusive. Hemin inhibited ALA synthase activity, suggesting that fat body heme synthesis could be regulated by end-product inhibition.

Development and endocrine regulation of cytochrome levels in fat body mitochondria of the cockroach, Blaberus discoidalis

Cytochromes aa 3 , b, c, and c 1 each showed distinct developmental patterns in fat body mitochondria of adult, male Blaberus discoidalis cockroaches. Cytochrome b content reached a maximum by day 2 of adult life; whereas, the other cytochromes reached their maxima by day 6. Once developed, the quantities of all the measured cytochromes remained constant through 30 days of age. Endocrine deficiencies changed the cytochrome levels relative to sham operated controls. Alletectomy increased cytochromes aa 3 and b, somewhat decreased cytochrome c 1 and left cytochrome c unchanged. Cardiacectomy-allatectomy significantly reduced the levels of cytochromes c and c 1 but had little effect on cytochromes a and b.

Proprioceptive indentation of the campaniform sensilla of cockroach legs

1. Individual proprioceptive campaniform sensilla of tibial Group 6 in the cockroachBlaberus discoidalis were stimulated by combinations of proprioceptive and punctate stimuli. 2. Punctate stimulation during a proprioceptive discharge increased the response rather than decreasing it, indicating that proprioceptive stimuli indent, rather than bulge, the cap of the sensillum. 3. The magnitude of the indentation associated with a strong proprioceptive discharge was estimated from compliance and sensitivity measurements to be in the range 10 to 50 nanometers. 4. The excitatory strain of the tibial cuticle appears to be compression perpendicular to the long axis of the cap of the sensillum. We suggest that the elaborate structure of the cap cuticle serves to amplify the tibial cuticular strain to squeeze the terminal of the dendritic sensory process.

Feeding effects on the neuroendocrine regulated development of fat body mitochondria in the cockroach Blaberus discoidalis

Fat body mitochondria in male Blaberus discoidalis showed a stepwise three-fold development in succinoxidase activity during the first 10 days of adult life. Long-term corpora cardiaca deficiency or prevention of feeding by oral blockage prevented the third step in succinoxidase development. Starvation alone did not prevent the complete development. Administration of corpora cardiaca extracts to orally blocked animals stimulated the final development of succinoxidase activity. It is speculated that the mechanical acts of feeding stimulate neurosecretory activity which increases the metabolic capacity of the fat body through effects on mitochondriogenesis.

Fiber analysis of an insect peripheral nerve: Light vs. electron microscopy

This paper describes the morphology of one of the two major femoral nerve trunks (nerve V of Dresden and Nijenhuis (1958)) of the cockroach Blaberus discoidalis as revealed by light microscopy, scanning electron microscopy and transmission electron microscopy. Fiber analysis of light and electron micrographs of thick and thin sections taken from adjacent areas of the same nerve yield significantly different data about the number and size distribution of the component axons of nerve V. The light microscopic image indicates one example of nerve V contains 2030 individual axons with a mean diameter of 1.4 μ; the electron image indicates the nerve contains 5794 axons with a mean diameter of 0.56 μ. The disparity between the two types of data is due to the large number of very small axons (0.2 μ or less in diameter) that course through the femoral nerve.

Characterization of insect fat body mitochondria isolated by a rapid procedure

1.1. A rapid method is described for isolating mitochondria from the fat body tissue of the cockroach Blaberus discoidalis.2.2. Oxidative activity was comparable to that of conventional procedures with a recovery of 50 per cent of the initial total respiratory activity. With succinate, ADP:O = 1.6 and respiratory control ratios were between 3 and 4.3.3. Electron microscopy showed isolated fat body mitochondria to be about 1.0 μm in dia. with a condensed ultrastructure. Only slight amounts of membraneous contamination were found in pellets, and the symbiotic micro-organisms found in fat body mycetocytes were rare.

Biogenesis of mitochondria: Neuroendocrine effects on the development of respiratory functions in fat body mitochondria of the cockroach, Blaberus discoidalis

Fat body mitochondria from male Blaberus discoidalis cockroaches underwent a hormone-independent respiratory development during the first 5 days of adult life followed by a neurohormone-dependent development during the next 5 days. Increases of two- to threefold were shown for oxygen uptake and for succinate-cytochrome c reductase and cytochrome c oxidase activities during this period. Also, an increased capacity for respiratory control by ADP was evident. No further respiratory changes occurred between 10 and 30 days of adult age. Respiratory enzyme activities and respiratory control failed to develop beyond the 5 day level after disruption of neurosecretory functions by extirpation of the corpora cardiaca. Conversely, daily administration of corpora cardiaca extracts stimulated precocious 10 day levels of respiration by 5 days of adult age. Uncoupling with 2,4-DNP indicated electron transport as the rate-limiting process of respiratory activity. Total incorporation of 14Camino acids into fat body mitochondria increased twofold during the first 5 days of adult life. This level of amino acid incorporation was maintained thereafter and corpora cardiaca removal had no effect. The fat body of Blaberus appears useful for studies on the hormonal regulation of mitochondrial development in higher eucaryotic animals.

Effects of DDT on biochemical functions of fat body and muscle mitochondria isolated from the cockroach Blaberus discoidalis

Respiratory activity, ADP:O ratios and respiratory control by ADP were examined for fat body and muscle mitochondria isolated from the cockroach Blaberus discoidalis Serville following administration of a lethal dose of DDT. Both types of mitochondria were unaffected by the poisoning. Using 14C-DDT it was found that 76% of the injected label accumulated in the fat body within 1 hr with 2% associated with the mitochondrial fraction. In vitro administration of DDT to fat body mitochondria decreased both the respiratory control by ADP and the ADP:O ratio. These findings indicate mitochondria are not affected by the levels of DDT reaching the tissues after poisoning. Therefore, the respiratory stimulus of DDT-poisoned insects is not the result of uncoupling oxidation and phosphorylation.

Insect hormones: Evidence for a neuroendocrine factor affecting respiratory metabolism

Gland extirpation studies on adult, male Blaberus discoidalis cockroaches provide new evidence for the hormonal regulation of respiratory metabolism in insects. These studies indicate the active agent is a brain neurosecretion stored and released by the c. cardiaca which affects the respiratory capacity of fat body mitochondria. The depressed Q 02 of fat body mitochondria from cardiacectomized-allatectomized cockroaches is returned to normal by daily injections of c. cardiaca extracts although the extracts have no direct action on oxygen uptake when added to fat body mitochondria in vitro .