Insects are found in habitats ranging from the driest deserts to aquatic habitats of diverse ionic composition. In many species, the environment changes dramatically during metamorphosis. Their survival depends on the ability to keep their tissues and cells moist, and to regulate the composition of their body fluids. Body fluid composition is challenged whenever exchanges of materials with the environment take place, and such exchanges are unavoidable. Routes of gain include ingestion, osmosis, active uptake, and diffusion. During feeding or drinking, an insect may ingest material that is significantly different in composition from the body fluids. Materials are lost through the excretory system and in the feces, as well as by osmotic, evaporative, or diffusive losses across the body surface. Insects respond to perturbations in body water composition by using a combination of hormonal and autonomous mechanisms to vary the amount and composition of fluid excreted by the excretory system.
STRUCTURE AND FUNCTION OF THE EXCRETORY SYSTEM
The excretory system consists of the Malpighian tubules and the hindgut (Fig. 1. . The Malpighian tubules form the primary urine by active secretion of ions into the lumen. Water follows passively
FIGURE 1 The larval mosquito gut and excretory system. The five Malpighian tubules form the primary urine by active secretion of ions, with water following the resulting osmotic gradient. Fluid formed by the Malpighian tubules enters the alimentary canal at the midgut-hindgut junction. This primary urine can be either shunted forward into the midgut and reabsorbed, or passed backward into the hindgut for modification and eventual elimination. Evidence suggests that all three regions, the midgut, Malpighian tubules, and hindgut, contribute to water and ion balance and are under hormonal control.
by osmosis, drawing with it small hemolymph solutes. This primary urine is then modified as it passes through more proximal regions of the Malpighian tubules and through the hindgut. The Malpighian tubules and the hindgut each frequently consists of several functionally distinct regions and/or distinct cell types, and each region contributes by specific mechanisms to the overall formation and processing of the urine.
Water balance can be adjusted by altering the rate of formation and/or composition of primary urine, and by changing the activity of mechanisms that modify the primary urine. Primary urine secreted by Malpighian tubules can be modified in more proximal regions of the tubules, passed forward into the midgut and reabsorbed, or passed back through the hindgut for final modification prior to elimination. Evidence suggests that Malpighian tubule, midgut, and hind-gut transport processes are each regulated by hormones.
REGULATION OF THE EXCRETORY SYSTEM
A number of chemical factors have been isolated that alter fluid and/or ion transport rates of particular regions of the excretory systems of various insect species. The chemical structures of these factors, and their actions at the cellular level, have received considerable attention. In contrast, far fewer studies have attempted to identify their specific roles in the maintenance of hemolymph water and ion balance at the level of the organism. Evidence to date suggests that hormones regulate several related parameters, including hemolymph volume, hemolymph ionic composition, and clearance of metabolic wastes and toxins from the hemolymph. Regulation of water and ion balance in any species involves several different hormonal factors, each acting on only a portion of the excretory system or even on a specific group of cells within an organ. At least some regions appear to be regulated by multiple hormones, with a specific mechanism of action and with combinations of hormones producing responses that are distinct from the actions of the individual hormones acting alone. Adding to the complexity, different concentrations of a single hormone may trigger qualitatively distinct responses in the same organ.
Hormonal Regulation of Malpighian Tubule Transport
Both diuretic and antidiuretic responses of Malpighian tubules to hormones have been reported. Endocrine factors involved in the regulation of Malpighian tubule function include several distinct structurally related groups of peptides, and the biogenic amine serotonin (5-hydroxytryptamine). Diuretic peptides identified to date include corticotropin-releasing factor (CRF)-related peptides, calcitonin-like peptides, kinins, tachykinins, and type 2b cardioacceleratory pep-tides (CAP2b). Kinins are small (6- to 15-amino-acid) peptides with a C-terminal amide. Those identified to date have similar C-terminal sequences of Phe-Xxx-Yyy-Trp-Gly-amide, where Xxx can be Asn, Ser, His, Phe, or Tyr and Yyy can be Pro, Ser, or Ala. The kinins have been isolated from several insect species but have not been detected in other animals. A single species may contain several kinins. The CRF-related diuretic peptides range from 30 to 46 amino acids in length and show structural homology to a family of peptides found in vertebrates that includes CRF (hence the name), sauvagine, and urotensin. Available evidence suggests that a single species produces more than one CRF-related diuretic peptide. Serotonin stimulates Malpighian tubule secretion in a number of insect species and is known to hormo-nally regulate excretory function in the bloodsucking bug Rhodnius prolixus and in larval mosquitoes (Aedes aegypti) (see below).
Short-term hormonal adjustments in Malpighian tubule transport rates in response to immediate challenges may be superimposed upon longer term increases or decreases in basal and maximal transport capacities. These long-term responses may be mediated at least in part by prostaglandins.
Hormonal Regulation of Hindgut Transport
Hormones acting on the hindgut appear to stimulate recovery of fluid and ions from the primary urine, leading to cycling of materials through the excretory system and hemolymph, and clearance of wastes. The majority of our information about regulation of hindgut transport comes from studies on the locust. The locust hindgut consists of two distinct regions, a more anterior ileum and a more posterior rectum. Two peptide factors isolated from the corpus cardiacum are ion transport peptide (ITP), acting on the ileum, and chloride transport stimulating hormone (CTSH), acting on the rectum. The amino acid sequence of these peptides has been determined. Other factors that stimulate water reabsorption by the hindgut include neu-roparsins A and B. In addition, a number of factors modulate contractions by the hindgut musculature.
Hormonal Regulation of Midgut Transport
Exchanges of materials between the midgut lumen and the hemolymph have important consequences for hemolymph composition, yet few studies have addressed the role of endocrine regulation of midgut transport in hemolymph homeostasis. In Rhodnius, serotonin stimulates both uptake of fluid across the midgut into the hemolymph and removal of that fluid from the hemolymph by the Malpighian tubules. Evidence suggests that a similar process may occur in larval mosquitoes. Acid/base exchanges between the hemo-lymph and the midgut lumen of larval Lepidoptera, creating conditions favorable to nutrient digestion and absorption, also appear to be regulated. Hormones inhibit ion transport across the lepidopteran midgut, and serotonin stimulates both acid/base transport and rhythmic muscular contractions of the larval mosquito midgut. Evidence for regulation of more than one midgut transport mechanism by multiple chemical signals suggests that the regulation of midgut transport may play an important role in hemolymph water and ion homeostasis.
BIOLOGICAL ROLES OF HORMONAL REGULATION OF EXCRETORY FUNCTION
Orthoptera: Grasshoppers
Locusts ingest considerable volumes of fluid with their food, and produce copious, moist feces when feeding. When deprived of food and water, Malpighian tubules secrete fluid at a low rate, and the feces produced are very dry due to the actions of antidiuretic factors acting on the hindgut. This recovery of ions and water by the hindgut is mediated by a hormone or hormones (ITP and CTSH, mentioned earlier). Upon feeding, a CRF-related diuretic hormone appears in the hemolymph, stimulating fluid secretion by the Malpighian tubules.
Orthoptera: Crickets
Cricket Malpighian tubules consist of distinct distal, mid, and proximal regions. These regions may be regulated independently, and secretion rates of a region may be either stimulated or inhibited. Achetakinin (a member of the kinin family of peptides) appears in hemolymph at levels sufficient to stimulate Malpighian tubule secretion in response to starvation, at a time when the insect is conserving water, rather than during feeding or water loading. These data suggest that achetakinin may act to stimulate fluid cycling through the excretory system, leading to clearance of materials from the hemol-ymph, rather than stimulating diuresis.
Heteroptera: Rhodnius
Most of our information concerning regulation of water balance in Heteroptera comes from studies on the bloodsucking bug Rhodnius. The physiology of this animal is unusual in that Rhodnius seldom feeds, feeds only on blood, and ingests a volume of blood up to 12 times its own body mass when feeding. At this time, serotonin released into the hemolymph causes the cuticle to become more plastic (allowing accommodation of the meal), stimulates anterior midgut fluid and ion absorption, and acts synergistically with a diuretic peptide to cause a 1000-fold increase in Malpighian tubule secretion rates. This leads to rapid removal from the midgut lumen and hemolymph of the fluid and ion load ingested with the meal.
Diptera: Mosquitoes
Larval mosquitoes are aquatic, and regulate water, ion, and acid/ base status very effectively in a wide range of aquatic habitats. Adults are terrestrial, and adult females of most species consume large volumes of blood in order to obtain protein to support egg production. Larvae and adults thus face very different challenges in water and ion homeostasis. Drinking rates, and hemolymph serotonin levels, of larval A. aegypti increase in response to increased ambient salinity. The serotonin stimulates secretion of fluid and ions by the Malpighian tubules, and alters transport rates across the midgut, leading to clearance of ingested material from the hemolymph. Regulation of adult mosquito water and ionic balance is best understood in the context of blood feeding. Females of many species consume blood meals that may triple their body mass within a few minutes, and severely compromise flight. Even while feeding, they begin to eliminate the bulk of the fluid and NaCl ingested with the meal. This response is mediated by mosquito natriuretic peptide, identified as either a CRF-like or a calcitonin-like diuretic peptide. Recent work in the housefly Musca domestica has shown that abdominal stretch receptors trigger release of diuretic hormone from the thoracic-abdominal ganglion mass, a response mediated by the brain. This suggests a primary role in volume regulation.
Lepidoptera: Butterflies and Moths
Lepidopteran larvae are able to adjust to diets of different water content by regulating the amount of water lost in the feces. This regulation appears to endocrine. Both CRF-related diuretic pep-tides and kinins stimulate Malpighian tubule secretion i n vitro, but appear to have very different biological roles. CRF-related diuretic peptide (Mas-DH) has an antidiuretic effect: it stimulates fluid (and presumably ion) recovery from the cryptonephric complex. In contrast, kinins increase the fluid content of the feces. Allatotropins and FLRFamides have been found to inhibit ion transport across the midgut epithelium, but the physiological significance of this response in terms of water and ion balance is not known. Upon emerging from the chrysalis, butterflies discharge a considerable volume of fluid containing metabolic wastes accumulated during the pupal stage. This diuresis is regulated by unidentified hormones.
Desert Beetles
Beetles of the family Tenebrionidae are generally very tolerant of dry conditions, to the extent that larvae of at least some species can remove water vapor from unsaturated air across the rectum. Despite this, factors that stimulate Malpighian tubule fluid secretion to high rates have been found in some of the most drought-tolerant species of this family. When injected into the animals, these factors stimulate fluid secretion by Malpighian tubules; but rather than being passed back through the hindgut and eliminated, the secreted fluid is passed forward into the midgut. The water and valuable hemolymph components are recovered, leaving toxins and metabolic wastes in the gut where they can be eliminated. These actions have led to the suggestion that some factors regulating excretory function may be more properly called “clearance hormones” than “diuretic hormones.” In fact, regulation of ionoregulation and clearance of wastes are likely to be more important than diuresis in many insect species due to their high surface area to volume ratios, and the desiccating conditions in which they live.
