[Wild reservoirs and vectors of cruzi. LX. Attempts to cross Rhodnius prolixus Stal, 1859 and Rhodnius neglectus Lent, 1954 (Hemiptera, Reduviidae)].
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The fate of calcium in the diet of Rhodnius prolixus: storage in concretion bodies in the Malpighian tubules.
We have investigated the fate of the large amounts of calcium ingested by Rhodnius prolixus in its meals of blood. 45Ca2+ injected into the haemolymph or fed to fifth-stage Rhodnius reared on rabbits is accumulated at high concentrations in the cells of the upper Malpighian tubules; very little is excreted from the body This 45Ca2+ accumulation goes on continuously for at least 12 days and the rate of uptake is increased several-fold within 3-4 days of a meal. The extent of calcium accumulation in tubule cells is correlated with the presence of intracellular membrane-bound concretion bodies, which are therefore likely sites of calcium deposition. X-ray diffraction showed that the calcium deposits are non-crystalline. Tubules from rabbit-fed fifth-stage Rhodnius contain 410 mmol l-1 calcium; in those from chicken-fed insects the calcium concentration is over 1 mol l-1; and in those fed in vitro on heparinised low-K+ sheep blood the calcium concentration is only 21 mmol l-1. The concentration of calcium in the haemolymph in all these insects was 8 mmol l-1 and its activity determined by an ion-selective electrode was 2.5 mmol l-1. 45Ca2+ deposited in the tubules is readily exchangeable, but the efflux preferentially passes to the haemolymph side of the tubule epithelium. The ability to sequester calcium in the Malpighian tubules may prevent calcium from interfering with reabsorptive processes in the rectum.
Animals↗
Distribution of FMRFamide-related peptides in the blood-feeding bug, Rhodnius prolixus.
Immunohistochemistry was used to study the distribution of FMRFamide-like material in the central and peripheral nervous systems and visceral tissues of 5th instar Rhodnius prolixus. Over 200 immunoreactive cell bodies and their processes as well as extensive neuropile regions were distributed throughout the nervous system. Immunoreactive processes were seen over the cephalic aorta, corpus cardiacum/corpus allatum complex, and in neurohaemal sites on the abdominal nerves. In visceral tissues, immunoreactive processes were seen innervating the salivary glands, the foregut, and the hindgut. Immunoreactive cells were also found in the anterior midgut (i.e., the crop and the anterior intestine). A radioimmunoassay specific for "RFamide" carboxy-terminal peptides was used to quantify the amount and the distribution of FMRFamide-like material. Reversed-phase high performance liquid chromatography of nervous tissue extracts revealed several peaks of immunoreactive material. The results suggest the existence of a number of FMRFamide-related peptides in Rhodnius which may have roles in both central and peripheral transmission, may be released as neurohormones and may have endocrine functions in the gut.
Amino Acid Sequence↗
Isozyme variability and differentiation between Rhodnius prolixus, R.robustus and R.pictipes, vectors of Chagas disease in Venezuela.
Enzyme polymorphism in triatomine bugs of the genus Rhodnius (Hemiptera: Reduviidae), vectors of Chagas disease, is analysed using both starch and polyacrylamide gel electrophoresis. Out of forty-five enzymes assayed, the electromorphs of seventeen of them: AO, CA, DIA, ES, ES-A, FH, GPD, G6PD, GPI, MDH, ME, 6PGD, PGM, ACON, ACPH, LAP and SOD, involving twenty-two putative structural loci, were scorable. These gene-enzyme systems were therefore selected for routine characterization of R.prolixus Stal adults from different strains. The first thirteen enzymes, involving sixteen structural loci, were also analysed in first instar nymphs of the three species, R.prolixus, R.robustus Larrousse and R.pictipes Stal. Allelic frequencies were calculated for three R.prolixus strains: three to five loci appeared to be polymorphic. The proportion of polymorphic loci (22%) and the average heterozygosity (0.06) indicated low genetic variability, with significant differences between the strains at individual loci. Rhodnius prolixus and R.robustus were found to have identical isozymic patterns. R.pictipes was genetically well differentiated, with twelve diagnostic loci.
Alleles↗
Diuresis from left atrial receptors: effect of plasma on the secretion of the Malpighian tubules of Rhodnius prolixus.
1. Stimulation of left atrial receptors by distension of a balloon in the lumen of the left atrium of anaesthetized dogs was shown to result in an increase in urine flow. Plasma samples obtained from these dogs during control periods and during periods of stimulation were applied to the Malpighian tubules of Rhodnius prolixus. 2. It was found that the tubules suspended in test plasma secreted at a significantly lower rate than those suspended in control plasma. 3. These differences were also evident in extracts of plasma prepared using the solvent n-butanol. 4. Cutting or cooling the cervical vagi abolished these differences along with the increase in urine flow. It is argued that this preparation of the Malpighian tubule of Rhodnius prolixus could be used as a means of detecting the diuretic agent responsible for the increase in urine flow.
Animals↗
Distribution of lipid in the lamellate endocuticle of Rhodnius prolixus (Hemiptera).
The lamellate appearance of the cuticle in the abdomen of the Rhodnius larva conforms to the conception of Bouligand in being an optical artifact which results from the spiral arrangement of successive layers of oriented fibrils. But superimposed on this structure is an actual lamination of bound lipid with the same spacing. The relation of the lipid layers to the optical lamination changes with the aspect from which the system is viewed. There must therefore be a cyclical secretion of lipid by the epidermal cells. Since the period of this cycle agrees with the cycle of rotation of the fibrous layers, which is supposedly inherent in the chemistry of the system, it is possible that it is the lipid which controls or initiates this helicoidal 'cholesteric crystallization'. There is evidence of a cyclical change in the secretion of lipid by the microvilli; it is suggested that there may be alternating cycles of eccrine and apocrine secretion, and that the lipid laminae represent the apocrine phases. The pore canals in Rhodnius are roughly cylindrical in cross-section, with lipid-impregnated walls. The contents of the lumen become slightly more electron opaque before the cuticle is stretched by feeding. There is probably some enzymic dissolution of the cuticle which precedes stretching; and this may concern particularly the lipid fraction. After the great distension and expansion of the cuticle which occur at feeding, lipid laminae can no longer be demonstrated in the old cuticle.
Animals↗
5-Hydroxytryptamine: a second diuretic hormone in Rhodnius prolixus.
Bioassays of 5-hydroxytryptamine (5-HT) in fifth-instar Rhodnius prolixus haemolymph using Calliphora salivary glands indicate that: (1) biologically active 5-HT is present, (2) in unfed animals there is not enough 5-HT to stimulate Malpighian tubule fluid secretion, and (3) there is enough 5-HT soon after the initiation of feeding to stimulate rapid tubule secretion. The 5-HT receptor antagonists ketanserin and spiperone reversibly and selectively inhibit 5-HT-induced fluid secretion, indicating the presence of specific 5-HT receptors on Rhodnius Malpighian tubules. The data provide evidence that 5-HT is a naturally occurring hormone acting with a previously described peptide hormone to regulate diuresis in this species.
Animals↗
[Demonstration of an enzyme inhibiting coagulation of blood ingested by the blood-sucking insect, Rhodnius prolixus (Heteroptera, Reduviidae)].
A new fibrinolytic enzyme, "the prolixase", was isolated and purified from the haemolymph of Rhodnius Prolixus. The mode of action of the enzyme was studied and compared to that of extracts from salivary glands and midgut of the insect. It appears that the prolixase is involved in inhibiting the coagulation of the blood ingested by Rhodnius during its blood meal. The mode of action of the enzyme in this phenomenon is discussed.
Animals↗
A FMRFamide-like peptide is associated with the myotropic ovulation hormone in Rhodnius prolixus.
The protocerebral neurosecretory cells previously shown to be the source of the myotropin controlling ovulation in Rhodnius prolixus react in an immunocytochemical assay using an antiserum against FMRFamide. When the same antiserum was injected into fed mated females at the appropriate time the timing of oviposition was delayed, but the total number of eggs developed was unaffected compared to controls injected with pre-immune rabbit serum. The titer of FMRFamide-like peptide (assayed by RIA) in the hemolymph of mated and virgin females was found to fluctuate with the egg laying cycle, and to reflect earlier determinations of the titer of myotropic activity. Western blots of SDS-PAGE revealed a FMRFamide-immunoreactive peptide of approximately 8.5 kDa in both hemolymph and extracts of the ovulation hormone cells.
Animals↗
Role of phospholipids in the lipophorin particles of Rhodnius prolixus.
The lipophorin of Rhodnius prolixus metabolically labelled with 32P exclusively in the phospholipid moiety was purified on a potassium bromide gradient and treated with phospholipase A2 in the presence of an excess of fatty acid-free albumin. The treatment completely removed the phospholipids from the particles and generated [32P]-lysophosphatidylcholine, [32P]-lysophosphatidylethanolamine, and free fatty acids that remained bound to albumin. The phospholipid-depleted lipophorin particles remained soluble, indicating that phospholipids are not essential in maintaining the stability of the particles in aqueous solution. Complete removal of phospholipids did not affect the association of apolipophorin III with lipophorin particles. Lipophorin density increased slightly from 1.120 to 1.134 g/ml after treatment. The phospholipid-depleted particles also retained their ability to be recognized and loaded in vitro with phospholipids delivered by the fat body, thus supporting the concept of lipophorin's role as a reusable lipid shuttle for phospholipids.
Animals↗
Identification of yolk platelet-associated hydrolases in the oocytes of Rhodnius prolixus.
The yolk platelets from Rhodnius prolixus, a blood-sucking bug, are composed mostly of vitellin and here are shown to contain at least two hydrolytic enzymes, a phosphatase and a cathepsin D-like proteinase. Both the proteinase and the phosphatase have an acid pH optimum. No hydrolytic activity was observed under alkaline or neutral conditions. Among several proteinase inhibitors tested, only pepstatin could abolish vitellin breakdown in vitro. The proteinase appears to be bound to the yolk platelet membranes. The phosphatase activity, using p-nitrophenyl phosphate as substrate, was enhanced after disruption of the platelet membrane by Triton X-100. This activity could be inhibited by tartrate but not by p-cloromercuribenzoate.
Acid Phosphatase↗
Tyrosine hydroxylase-like immunoreactivity in the brain of fifth instar Rhodnius prolixus Stål (Hemiptera: Reduviidae).
The distribution of tyrosine hydroxylase-like immunoreactivity was mapped in whole-mount preparations of the brain of fifth instar Rhodnius prolixus Stål. Immunoreactivity was limited to neuronal cell bodies and processes, which were distributed over both ventral and dorsal surfaces of the CNS. The brain, excluding the optic lobes, contained about 160 tyrosine hydroxylase-like immunoreactive cells. Each optic lobe contained two groups of small round cell bodies, which were too numerous to count. The wide distribution of immunoreactivity suggests that tyrosine hydroxylase is present in neurons with diverse central functions. Tyrosine hydroxylase is the rate-limiting enzyme in catecholamine synthesis in vertebrates. A comparison of a map of the distribution of catecholamine-induced fluorescence obtained using the glyoxylic-acid technique (Flanagan; J. Insect Physiol. 30(9):697-704, 1984) with that generated for tyrosine hydroxylase reveals considerable overlap between the two systems, suggesting that tyrosine hydroxylase is used in the catecholamine pathway in this insect. The mapping of these reactive neurons is an important step for identification of unique tyrosine hydroxylase-containing neurons, and is our initial step in the analysis of identified catecholamine-containing neurons in R. prolixus.
Animals↗
Morphology of the dorsal vessel in the abdomen of the blood-feeding insect Rhodnius prolixus.
The dorsal vessel (DV) in the abdomen of the blood-feeding insect Rhodnius prolixus was divided functionally into two regions, the heart, into which haemolymph entered the DV through four pairs of ostia located in abdominal segment VII, and the aorta, along which the haemolymph was propelled from abdominal segment VI to the thorax. Osmium-fixed whole mounts revealed the DV to consist of spirally arranged striated muscle fibers and to possess two rows of ventrally attached longitudinal fibers extending the length of the abdomen. Seven pairs of alary muscles were found attached to the DV in the posterior abdominal segments. Contractions of the alary muscles attached to the ventral surface of abdominal segments VII and VIII served to expand the heart. Electron microscopy revealed the DV to consist of a thin layer of contractile elements surrounded by an inner (intima) and outer (adventitia) connective tissue layer. Embedded in the intima along each lateral side of the DV were two large groups of endocardial cells extending the length of the DV. A small group of pericardial cells was embedded in the adventitia along the mid-ventral side of the DV, and clusters of pericardial cells were found attached to the alary muscles. Nerve terminals were found only on the heart: they contained agranular synaptic vesicles approximately 30 nm in diameter and densely stained granules approximately 100-120 nm in diameter. These structural components are discussed in relation to the role of the DV in circulation.
Abdomen↗
Development and interactions of Trypanosoma rangeli in and with the reduviid bug Rhodnius prolixus.
Two strains of Trypanosoma rangeli and three strains of Rhodnius prolixus were used in various combinations to transmit the trypanosomes via the reduviid bug. A persisting infection in the midgut lumen posterior to the stomach resulted in all 2,500 bugs being third and fourth instars. Infectious, metacyclic forms developed exclusively in the salivary glands; forms excreted with bug feces were noninfectious to mice. The midgut epithelium was the main barrier to transmission of the parasite. In only 2%-5% of the infected bugs was it penetrated, and no correlation with any parameter tested could be found. The trypanosomes passed through the gut epithelium by an intracellular route; they were contained in a parasitophorous vacuole. The salivary glands of R. prolixus were found to be infected whenever the hemolymph was infected, after a successful penetration of the gut epithelium, or after injection of T. rangeli. On the other hand, hemolymph infections in Triatoma infestans were eliminated within a few days, and the salivary glands of this bug were never invaded. Pathogenic effects of T. rangeli on R. prolixus could be seen in midgut epithelial cells by a loss of cytoplasm and in muscle and salivary gland cells by very high parasite densities. However, as the penetration rate of the gut is low, T. rangeli is not likely to prove to be efficient in natural control of R. prolixus.
Animals↗
Proteolytic enzymes in the Rhodnius prolixus midgut.
Sequential chromatographic fractionation of Rhodnius prolixus midgut homogenate yielded only one endopeptidase, but revealed the presence of carboxypeptidase-A and B-like enzyme, di- and tripeptidases, as well as aminopeptidase activities.
Aminopeptidases↗
Cell proliferation and rearrangement in the development of the malpighian tubules of the hemipteran, Rhodnius prolixus.
The pattern of cell activities resulting in the generation of a simple tubular epithelium, the Malpighian tubules of Rhodnius prolixus, is examined during embryogenesis. The anlage of the tubules is shown to be of ectodermal origin. An evolving pattern of cycling cells in the primordia is revealed using the immunocytochemical localization of a substituted nucleotide, BUdR. A cell unique to each tubule is shown not to enter the cell cycle but to be required for the proliferation of the remaining cells in the tubules. Furthermore, the activity of this cell imposes on each tubule a clear proximo-distal axis.
Animals↗
Cyclic AMP in the Malpighian tubule fluid and in the urine of Rhodnius prolixus.
We demonstrate the presence of cyclic adenosine monophosphate (cAMP) in the fluid secreted by isolated Malpighian tubules of Rhodnius prolixus. In addition, we show that fifth-instar R. prolixus excrete cAMP in the urine after a meal of human blood. Nonstimulated isolated Malpighian tubules secrete small amounts of cAMP that increase about 10-fold after the addition of 5-hydroxytryptamine (5-HT). 5-HT is known to mimic R. prolixus diuretic hormone. The present findings demonstrate that 5-HT also acts via cAMP. The presence of cAMP in the rectal sac of the insect could be of importance in the differentiation of Trypanosoma cruzi and in the cycle of Chagas disease.
Animals↗
Circadian regulation of synthesis of ecdysteroids by prothoracic glands of the insect Rhodnius prolixus: evidence of a dual oscillator system.
Rhythmic synthesis of moulting hormones (ecdysteroids) by prothoracic glands (PGs) of the insect Rhodnius prolixus during the last larval instar was studied in vitro following explantations every 4-5 hr for up to 96 hr. Ecdysteroid synthesis was measured by radioimmunoassay as the quantity of ecdysteroid produced during 4 hr in vitro. A massive daily rhythm is seen, with synthesis at night being three- to fivefold higher than during the day. This rhythm of ecdysteroid synthesis by PGs free-runs in continuous darkness with a temperature-compensated period length close to 24 hr and is therefore controlled by a circadian system. This is the first report of circadian regulation of synthesis of an invertebrate hormone. The synthesis rhythm also free-runs in continuous light, but with an inverted phase and shorter period length. It is argued that the circadian system controlling synthesis comprises two oscillators which free-run in antiphase, occupy different anatomical locations, and are coupled by a humoral factor, possibly prothoracicotropic hormone. The ecdysteroid synthesis rhythm in PGs appears to drive the previously reported circadian rhythm in the haemolymph ecdysteroid titre. It is concluded that the circadian system controlling synthesis of ecdysteroids constitutes a pacemaker which drives various rhythms in the target cells of ecdysteroids via the rhythm in the haemolymph titre. Ecdysteroids are viewed as "hormonal Zeitgebers," imposing temporal order on development.
Animals↗