RESUMEN

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.

Asunto(s)

RESUMEN

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.

Asunto(s)

Asunto(s)

RESUMEN

The action of activated 27,000 Mr toxin from Bacillus thuringiensis var. israelensis (Bti toxin) on Malpighian tubules of Rhodnius prolixus has been investigated. Its binding to the tubules is slowed by low temperature but is not prevented even at 0 degree C. The binding is less effective at pH 10 than at pH7. Pretreatment of the tubules with 0.1 mmol l-1 ouabain or bumetanide or 1 mumol l-1 5-hydroxytryptamine did not affect the toxicity of the toxin. The toxin causes very large changes in the trans-epithelial potential difference; it changes from 40 mV, lumen negative, often to more than 100 mV, lumen positive. This reflects an initial collapse of the potential of the basal cell membrane, followed by a large positive-going potential change at the luminal cell membrane. Just prior to the effects of the toxin on rapid fluid secretion, the basal cell membrane becomes permeable to sucrose molecules. Raffinose at 170 mmol l-1 in the bathing solution does not protect the tubules from Bti toxin action but dextran, Mr5000, at 60 mmol l-1 significantly delayed failure of fluid secretion and, even more, the onset of staining of the tubule cells with Trypan Blue. Exposing tubules to saline that is calcium-free and/or magnesium-free, or has a composition adjusted to be similar to that of the intracellular milieu, does not affect the time course of failure of fluid secretion induced by the toxin. There is no evidence that effective aggregates of Bti toxin molecules are formed in concentrated solutions.(ABSTRACT TRUNCATED AT 250 WORDS)

Asunto(s)

RESUMEN

Ouabain, at all concentrations higher than 2 x 10(-7) mol l-1, stimulates the rate at which the Malpighian tubules of the insect, Rhodnius, transport sodium ions and fluid into the lumen. An effect on paracellular movement of sodium ions is unlikely because ouabain makes the electrical potential of the lumen more positive, which would slow diffusion of sodium into the lumen. Radioactive ouabain binds to the haemolymph-facing sides of the tubule cells but not to the luminal face. This binding is reduced in the presence of elevated levels of potassium or of non-radioactive ouabain. Bound ouabain is only slowly released on washing in ouabain-free saline. The evidence suggests that there is a Na+/K+-ATPase on the outer (serosal) membranes of the tubules. Such a pump would transport sodium in a direction opposed to the flow of ions and water involved in fluid transport; poisoning it with ouabain would remove this brake, and fluid flow and sodium transport would increase, as observed.

Asunto(s)

RESUMEN

The effects of the 27 X 10(3) Mr insecticidal delta-endotoxin from Bacillus thuringiensis var. israelensis have been studied using, as a model system, isolated insect Malpighian tubules. At all concentrations of the toxin higher than 1 microgram ml-1 (4 X 10(-8) moll-1) applied to the outer surface of the tubules, fluid secretion failed within about 30 min. Except at very high concentrations, where failure always takes at least 30 s, there was an inverse relationship between the concentration of toxin and the time of failure of toxin-treated tubules. During exposure to toxin, the tubules were initially unaffected for a relatively long period and then rapid failure occurred. If the tubules were removed into toxin-free saline just before failure would have occurred, fluid secretion remained normal for at least 2 h, but on return to the origin toxin-containing saline failure was almost immediate. The toxin was found not to bind to the basement membrane. Ultrastructural changes became evident as tubule failure occurred. These initially involved modifications to the basal side of the cells, but later also to the luminal microvilli. Intercellular junctions became disassociated and cytoplasmic vacuolization occurred. The population of intramembranous particles in the basal membranes became reduced with time. Our findings suggest the following hypothesis for the initial stages in the interaction of the toxin with the tubules. Toxin molecules attach to the accessible cell membranes progressively and irreversibly. They do not readily associate by diffusing laterally in the membrane, so that toxic effects develop only when sufficiently large numbers of them attach close together. The molecules may then associate in some way as a complex, perhaps forming a pore in the membrane. Relatively few such pores lead rapidly to cell failure and death.

Asunto(s)

RESUMEN

We have found in an insect tissue, the Malpighian tubules of Rhodnius, instances of single epithelial cells which, as the result of a possible error in development, lie within the epithelium some distance from the main population of similar cells. This spatial separation makes it possible to measure the transport abilities of these cells. Their transport abilities were found to be the same as the cells in the main population. This finding shows that the maintenance of function in individual cells of epithelial tissues may not depend on direct contact with other similar cells.

Asunto(s)