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Excretion
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Excretion
Estivation see Aestivation
Evolution
see Phylogeny of Insects
E Excretion Timothy J. Bradley University of California, Irvine
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xcretion is the elimination of excess ions, water, and metabolic wastes from the body. As in all organisms, excretion in insects serves to promote the appropriate regulation (homeostasis) of the intracellular environment as the key to organismal well-being and survival. Regulation of the intracellular environment depends in turn on appropriate regulation of the extracellular fluids. In insects, these extracellular fluids are contained in the hemolymph and in the interstitial fluids surrounding the cells. Excretion therefore refers to the processes by which wastes, as well as excess ions and water, are eliminated from the hemolymph. Wastes produced in the cells and transported into the hemolymph are concentrated and excreted by the excretory organs.
STRUCTURE OF THE EXCRETORY ORGANS The processes of excretion in insects are carried out largely by the organs of the insect gut. These include the midgut, the Malpighian tubules, and the structures in the hindgut, namely the ileum and rectum (Fig. 1).
FIGURE 1 A diagrammatic representation of the insect gut.
Midgut The midgut is a tubular epithelium. Upon ingestion, food and fluids move through the esophagus and pass directly into the midgut. Because the cells in the midgut epithelium are derived from embryonic endoderm, the midgut is not lined with cuticle. Most of the cells in the midgut are involved in the secretion of digestive fluids and the
absorption of nutrients from the midgut lumen. These secretory and absorptive cells have apical microvilli that greatly increase the surface area available for inward and outward transport. In insects that feed periodically (such as adult mosquitoes), the microvilli shorten during nonfeeding periods and lengthen following ingestion. Many insects have additional cell types termed goblet cells that are thought to be involved in the secretion of fluids that modify the acidity and alkalinity (pH) of the luminal fluid. These goblet cells have been intensively investigated in Lepidoptera, where they serve to produce a markedly alkaline pH. The basal surface of the midgut cells possesses a network of longitudinal and circular muscles that upon contraction can produce peristaltic waves. These contractions serve to move the food along the gut and stir the midgut contents during digestion. Many insects possess globular outpocketings in the anterior region of the midgut, termed ceca. The cells types in the ceca are generally differentiated from those in the midgut proper.
Malpighian Tubules The Malpighian tubules are the site of urine formation in all insects except the Collembola, Thysanura, and aphids. The Malpighian tubules are tubular epithelia that are diverticulae (outpocketed extensions) of the gut itself. The tubules open into the gut near the midgut–hindgut junction, and the lumina of these two tubular epithelia are continuous. The contents of the tubules flow into the gut lumen; the ends of the tubules distal to the gut are closed. Fluid is produced in the Malpighian tubules by secretion; and because the tubules are closed at the distal end, hydrostatic pressure builds up and fluid flows through the tubules into the gut. The number of Malpighian tubules is quite variable depending on the insect species. Bloodsucking Hemiptera (e.g., Rhodnius prolixus) and higher Diptera (e.g., Drosophila melanogaster) have as few as four tubules, whereas the desert locust (Schistocerca gregaria) has hundreds. Attached to the Malpighian tubules of many insects are longitudinal muscles. When these muscles contract, the tubules are waved about in the hemolymph, presumably for the purpose of stirring the fluid adjacent to the tubules and promoting fluid and solute transport. These tubules may also serve the more general function of promoting hemolymph circulation throughout the abdomen. The Malpighian tubules of all species examined to date contain more than one cell type. In some cases, a single epithelial region contains two or more cell types (regions with heterologous cell types) reflecting, presumably, separate physiological roles for each cell type. In other species, the tubules are divided into distinct regions, each consisting of a single cell type (regions with homologous cell types). In these insects, each tubule region has a distinct function in transport. Finally, in many insects, the tubules show regional specialization as well as multiple cell types within a region. It is presumed that each cell type in these tubules performs a distinct function. As an example of cell type heterogeneity, consider the Malpighian tubules of the larvae of the brine fly, Ephydra hians (Fig. 2). The tubules in this insect are differentiated on each side of the body, as well as along their length. On one side of the body is a pair of tubules called the lime gland tubules. The distal ends of these are secretory and contain two regions that can be differentiated on the basis of cell color: one white, one yellow. More proximal to the gut are expanded regions of the tubules that serve to store concentric concretions in the tubule lumen. Finally, two of these cells combine in a common ureter that empties into the gut. On the opposite side, the tubules have only the yellow and white regions of the tubules, with no storage
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Midgut Unmodified tubule (posterior) Hindgut
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Rectum
Proximal tubule of lime gland Storage region of time gland Distal tubule of lime gland
FIGURE 2 A schematic diagram of the Malpighian tubules of E. hians. The Malpighian tubules are differentiated on the left and right side. The hindgut is composed, from anterior to posterior, of the ileum, colon, and rectum. [From Herbst, D. B., and Bradley, T. J. (1989). A Malpighian tubule lime gland in an insect inhabiting alkaline salt lakes. J. Exp. Biol. 145, 63–78.]
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core of microfilaments. Frequently, these microvilli contain fingerlike extensions of the mitochondria and even of the endoplasmic reticulum. The basal surface of the cells exhibits deep infolds, often again closely associated with mitochondria by means of structures termed scalariform junctions. The intercellular spaces are occupied apically by septate or continuous junctions. More basally, the intercellular space contains gap junctions or undifferentiated basolateral membranes indistinguishable from the basal membrane infolds. Numerous other cell types occur in the Malpighian tubules. It is presumed that each histologically distinguishable cell type performs a unique function within the tubules. In addition, distinct functions have been found in some cell types in the absence of histological or ultrastructural differentiation. A common cell type in Malpighian tubules is the stellate or secondary cell. This cell type possesses smaller microvilli than the primary cells, and these microvilli contain no mitochondria. As described later in the section on function, the secondary cells may be involved in modification of the primary urine produced by the primary cells.
Ileum section. This example illustrates the variety of cell types that can exist in a single tubule. The details of transport function in these and other highly complex tubules have not been fully elucidated. In most Malpighian tubules, formation of the primary urine occurs in a cell type often referred to as the primary cell (Fig. 3). This cell type has extensive apical microvilli, often containing a central
Posterior to the midgut, most insects possess a segment of gut referred to as the ileum. Because this region is part of the hindgut, it is covered on the apical surface by cuticle. The cells often show deep apical and basal membrane infoldings reflecting the role of these cells in fluid and solute transport. The basal surface of the cells is covered by layers of longitudinal and circular muscles that serve to generate peristaltic movements of the gut. These muscular contractions move the gut contents through the gut and may also serve an important role in reducing unstirred layers adjacent to both the apical and basal membranes of the epithelium. Historically, the Malpighian tubules and rectum have been assumed to carry out most of the fluid transport in the posterior regions of the gut. The ileal epithelium is smaller in diameter, with less highly developed apical and basal membrane infolds, and a lower mitochondrial density than is observed in rectum. Nonetheless, when the transport properties of the ileum are investigated, this region of the gut is always found to be carrying out important transport functions.
Rectum
FIGURE 3 Model of the transport processes occurring in the Malpighian tubules of adult mosquitoes, based on the work of Klaus Beyenbach using the species A. aegypti. Upon stimulation with mosquito natriuretic peptide (MNP), rates of transepithelial fluid secretion increase from an unstimulated rate of 0.4–2.8 nl min1. In parallel, Na concentrations in the secreted fluid rise, and K concentrations fall. Electrophysiological studies reveal that MNP, working via cyclic AMP, induces an increase in basolateral membrane Na conductance, presumably through the actions of Na channels in the basolateral membrane of principal cells. The hyperpolarization of the transepithelial voltage and the decrease in transepithelial resistance are consistent with the activation of Na channels in the basolateral membrane of principal cells. (Figure and legend provided by Klaus Beyenbach.)
All insects possess an enlarged chamber called the rectum near the posterior end of the gut. The structure of the rectum can vary substantially from species to species. The rectal lumen is covered by a thick cuticle. Posterior to the rectum, insects possess an anal canal through which the feces and urine are eliminated. The strong bands of muscle surrounding the rectum contract during defecation, expelling the feces and urine into the external environment through the anus. The recta of terrestrial insects are large and very active organs. Regions within the rectum are highly differentiated and contain cells with deep membrane folds. If (as in many orthopterans and lepidopterans) these cells are contained in thickened ridges that extend into the rectal lumen, they are referred to as rectal pads. If (as in many adult Diptera) the cells extend into the lumen as fingerlike structures, often on a thin stalk, they are referred to as rectal papillae. The rectal pads and the papillae are the major sites of fluid resorption and urine concentration. The cells in the rectal pads have a complicated array of intercellular junctions associated with the active sites of ion secretion and resorption. The evolution of a rectal structure permitting the formation of a urine hyperosmotic to the
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hemolymph was, along with the evolution of a waxy cuticle, a major adaptive event permitting insects to invade drier terrestrial habitats. Both rectal pads and rectal papillae are covered with thick cuticle to protect the underlying epithelial cells from abrasion by the fecal material. Between the rectal pads, the rectum of terrestrial insects possesses a thin, cuticle-lined epithelium that, upon unfolding and stretching, permits rectal swelling during feces and fluid accumulation. In E. hians, the rectum is merely a thin, distensible epithelium that expands upon filling with feces prior to defecation. In this insect, the task of modifying the urine is carried out by the colon, which lies just anterior to the rectum. The presence of a rectum without transport capabilities is very unusual. In most insects the rectum is the major organ responsible for osmotic regulation of the urine.
Excretion outside the Gut Storage excretion of concentric concretions containing calcium salts of urate and carbonate occurs in the fat body cells of most insects. Some insects contain nephrocytes, cells in the head that also store wastes and toxic elements. Finally, in collembolans and thysanurans, cephalic nephridial glands, sometimes termed labial glands, are responsible for excretory function, including the excretion of nitrogenous waste.
FUNCTION OF THE EXCRETORY ORGANS Midgut TRANSPORT The primary function of the midgut in insects is the digestion and uptake of ingested nutritive materials. There are two processes that occur in the midgut and contribute to excretion: pH regulation and storage excretion. The midguts of many insects secrete fluids that contribute to extreme alkalinity or acidity in the gut. In Lepidoptera, for example, the midgut epithelium consists of goblet and columnar epithelial cells. The goblet cells are responsible for transporting ions and creating a highly alkaline solution in the midgut lumen. The columnar cells contribute to digestion of the food and uptake of ions. The apical membrane of the goblet cell contains hydrogen pumps (H-ATPases) that use adenosine triphosphate (ATP) as their source of energy. These pumps transport hydrogen ions into the cuplike apical crypt of the goblet cell. The resulting electrical and pH gradient is used in an exchange process (antiporter) to exchange potassium ions for the hydrogen ion. Potassium ions associated with hydroxyl ions remain in the lumen following the hydrogen/potassium exchange process. These ions diffuse from the goblet cavity into the midgut lumen, with the result that the potassium hydroxide (KOH) causes the lumen to be very alkaline. The high potassium concentration in the lumen is, in turn, used as an energy source for the uptake of amino acids from the digested foodstuffs. This uptake occurs by means of transporters (symporters) in the apical cell membranes of the columnar cells that use the potassium gradient to cotransport potassium ions and amino acids from the lumen into the cell interior. Various functions have been proposed for the highly alkaline pH in the midgut of Lepidoptera. Clearly, such an extreme pH would serve to kill pathogens and to saponify lipids in the ingested food material. It has also been proposed that this pH serves to reduce the solubility and activity of tannic acids in the food of Lepidoptera, protecting herbivorous larvae from the high concentrations of these toxic compounds found in the leaves of many plants. In other insects, for example, mosquito larvae, low-pH (acid) conditions are observed in the midgut. The precise mechanisms by which this pH is produced and maintained remain to be elucidated,
but there is evidence in mosquitoes that the midgut epithelium is also energized by the H-ATPase. STORAGE EXCRETION IN THE MIDGUT The columnar midgut cells of insects often contain concentric mineralized concretions. The major cation in these concretions has been shown to be calcium. The anion can be quite variable and has been found to be largely urate or carbonate, depending on the cell type in which the concretions are located. In the midgut, the concretions have also been shown to play a role in the storage excretion of toxic cations such as copper, zinc, iron, and selenium. Each of these elements is toxic in high concentrations but plays a critical role in metabolism in trace amounts. The concretions may therefore play both a protective and a homeostatic role in insects by regulating the free concentration of these ions and metals in the body. They reduce toxic concentrations and serve as a reservoir for these elements, which can be used for physiological purposes when required. Storage excretion in the form of concentric concretions also occurs in the Malpighian tubules.
Malpighian tubules TRANSPORT The production of urine in insects occurs by the active transport of ions across the epithelium from hemolymph to tubule lumen. This process generates an osmotic concentration that drives the movement of water across the epithelium as well. Generally, Malpighian tubules have a high permeability coefficient for water (a low osmotic resistance), and as a result water moves rapidly across the epithelium in response to relatively low osmotic concentration gradients. These concentration differences across the epithelium are indeed so low that they have been difficult to measure. Recent experimental results have led to a general consensus, however, that an osmotic gradient of a few milliosmoles is sufficient to account for the observed rates of water movement across Malpighian tubules. Let us consider first the active transport of ions across the epithelium, and then the passive movement of water that follows. In insects, potassium is the predominant cation transported across the epithelium of the Malpighian tubules. Insects evolved as a distinct clade on land, feeding on plants and detritus. It may be that their dependence on potassium as the major cation used for fluid transport evolved at this time as well. Certainly, animals of marine origin, such as the vertebrates and crustaceans, rely on sodium as the principal cation for driving fluid movements. In the cell interior, the Malpighian tubule cells have a negative electrical charge relative to the hemolymph. This electrical potential facilitates the entry of potassium into the cells. Thermodynamically speaking, therefore, the most energetically costly transmembrane movement for potassium in the epithelium occurs as this cation crosses the apical membrane. The process by which this occurs has been very difficult to unravel, but in recent years it has been demonstrated that a very active hydrogen ATPase, related to vacuolar H-ATPase found widely in eukaryotic cells, is located on the apical membrane of Malpighian tubule cells. This transporter moves hydrogen ions from the cell interior into the tubule lumen, thereby setting up a large electrochemical gradient for hydrogen ions. Depending on the circumstances, this electrochemical gradient can be predominantly expressed as a pH gradient or as an electrical gradient. In either circumstance, this electrochemical gradient is thought to serve as an energy reservoir that can be subsequently used for a variety of coupled transport processes.
Excretion
For example, if the energy contained in the hydrogen ion gradient is used in an antiporter (a transporter that couples ion movement in one direction to ion movement in the opposite direction) that exchanges hydrogen for potassium at the apical membrane, the result of hydrogen transport with subsequent hydrogen exchange for potassium is a net active transport of potassium. Potassium is moved against its electrochemical gradient by the antiporter, using the energy contained in the transmembrane hydrogen ion gradient. In some Malpighian tubules, similar mechanisms may exist for the coupled transport of other cations (e.g., sodium, calcium, or magnesium). In bloodsucking insects such as adult mosquitoes and the reduviid R. prolixus, the plasma portion of the blood meal also provides the insect with a substantial intake of sodium. Any amount of sodium that exceeds the physiological needs of the insect must be excreted. In these insects, a substantial active transport of sodium occurs accords the Malpighian tubules. This process is thought to be driven across the apical membrane by a hydrogen–sodium exchange mechanism. Cation transport across the epithelium also requires a process for entry of the ions into the cytoplasm from the hemolymph. In most insects, the basal membrane contains an enzyme (Na–K ATPase) that uses the energy in ATP to transport sodium actively out of the cell and potassium actively in. As a result, the cytoplasm is greatly enriched in potassium. The passive outward diffusion of this ion through barium-sensitive channels produces an electrical potential across the basal membrane, the inside of which is negatively charged. This potential can be used for a variety of transport functions; one that seems to be almost universally present in Malpighian tubules is the bumetanide-sensitive Na/K/2Cl, cotransporter. This transporter uses the energy contained in the sodium gradient to move one sodium, one potassium, and two chloride ions simultaneously from the hemolymph into the cytoplasm. This process serves to provide chloride to the cell interior, as well as sodium in the cell types to which sodium is moved transepithelially. The movement of anions across the epithelium involves one or more of three distinct transport pathways, depending on the characteristics of the tubules. In the first, chloride is thought to move across the same cells as those in which the cations are transported. As mentioned earlier, the interior of the cells is electrically negative relative to the cell exterior. The movement of chloride into the cell is therefore a thermodynamically active process. As already discussed, it may be driven by the sodium electrochemical gradient in a cotransport process. In other insect species in which the Malpighian tubule cells in a single region of the tubule are differentiated into two or more cell types, chloride ions have been shown to enter the lumen via a cell type distinct from that involved in active cation transport. For example, in D. melanogaster, the fluid-transporting segment of the tubules contains both primary and stellate cells. The former cells are the sites of active potassium transport, the latter the site of chloride flux from the hemolymph to the tubule lumen. Although this movement of chloride into the tubule lumen is thermodynamically downhill, the precise mechanism of chloride transport is presently unknown. A third process has been described in the Malpighian tubules of adult mosquitoes. Although these insects also possess primary and stellate cells in the Malpighian tubules, it has been proposed that chloride moves into the lumen of the tubules via the intercellular junctions. In fact, this process has been shown to be under hormonal control. The movements of chloride into the Malpighian tubules may therefore be quite variable depending on cell types. The movements of anions are much less well characterized at this time, with regard to the molecules
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that drive the process, than are the movements of cations. A model of ion transport at both the apical and basal membrane of the Malpighian tubules of adult mosquitoes is shown in Fig. 3. Regardless of the mechanisms by which cations and anions enter the Malpighian tubules, it is clear that the types of ions transported can vary greatly with the species of insect and will depend on an individual’s physiological needs and demands. Thus since bloodsucking insects ingest a large amount of sodium compared with other insects, the Malpighian tubules of bloodsuckers contain specific mechanisms designed to reduce the large sodium load. Species of mosquitoes whose larvae can survive in saltwater must ingest the medium and eliminate the ions as means of obtaining water. Those species that have been investigated can excrete magnesium and sulfate via the Malpighian tubules. The larvae of brine flies (ephydrids) generate crystals in the lumina of the tubules that are rich in calcium and carbonate. Both these ions must be transported across the epithelium, although the combination forms insoluble crystals that reduce the activity of these ions in the lumen. The Malpighian tubules of insects are also the site of excretion of the waste products of energy and nitrogen metabolism. Acid byproducts of energy metabolism have been shown to be actively transported into the lumen from the hemolymph. The precise molecular mechanisms of the process remain unclear, but the process is of paramount importance for the insects in maintaining acid/base balance and energy homeostasis. The by-products of nitrogen metabolism are also excreted by the Malpighian tubules. In aquatic insects, ammonia may be excreted, but in most insects and certainly in terrestrial forms, urea and uric acid predominate. Both these compounds are actively removed from the hemolymph by the Malpighian tubules. The transport of uric acid has been investigated in R. prolixus, in which the blood meal provides a very protein-rich meal requiring intense capacity for the elimination of nitrogenous waste. In Rhodnius, the primary urine is produced in the most upstream portions of the Malpighian tubules, the upper tubule. This urine is modified in the downstream section (the lower tubule) through the resorption of potassium and chloride. This process serves to return potassium to the hemolymph, and to remove waste from the hemolymph, through the retrieval of an isosmotic fluid. Thus hemolymph volume is retained and the sodium in the urine is concentrated. In addition, uric acid is transported in the lower tubule from hemolymph to urine. Potassium urate is fairly insoluble, particularly at neutral to acid pH. As a result, crystals of uric acid form in the urine. This process further removes osmotically active compounds from the urine, allowing the additional movement of water from the urine to the hemolymph by osmosis. CONCENTRIC CONCRETIONS IN THE MALPIGHIAN TUBULES Concentric concretions occur in the midgut, where they are thought to contribute to excretion by storage in an insoluble form of salts containing calcium, magnesium, manganese, copper, cadmium, and zinc. Identical concretions are found intracellularly in the Malpighian tubules. These concretions are thought to perform an identical function, namely storage of ions in an insoluble form either for subsequent use or as a means of removing the ions from the body. In the Malpighian tubules, however, these concretions also appear in the tubule lumen, a location from which they can move into the gut and be eliminated with the excrement. It has been suggested by many authors that the intracellular concretions in the cells of the Malpighian tubules can be transported by exocytosis into the lumen of the tubules. Although there are occasionally physiological conditions in which the concretions disappear from
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the cells and appear in the lumen, it has not been unambiguously demonstrated that the crystals move from one location to the other intact. Instead, it is likely that the crystals are dissolved within the cells of the Malpighian tubules, that the soluble ions are transported into the lumen, and that the crystals are formed anew in the tubule lumen. Crystals are formed in some tubule segments (e.g., in the lower tubule of R. prolixus) where no crystals exist in the cells. The crystals in the midgut, fat body, and Malpighian tubules are concentric and perfectly round. This is in marked contrast to the natural structure of the crystals formed by the same salts in solution. Uric acid crystals, for example, have sharp corners and sometimes take a needlelike form. It is thought that the concentric concretions avoid acicularity through the activity of organic compounds that are known to be a substantial component of the concretions. The compounds are thought to nucleate and direct crystal formation, leading to the formation of round concretions. This spherical shape is less damaging to the cells of the tubules and can be excreted from the tubules and gut with little tissue damage. Ultrastructurally identical concretions are observed in the urine of birds, which is rich in uric acid. It has been proposed that organic compounds are excreted into the tubule lumen, where they nucleate and guide the formation of the concretions. The ions contained in the concretions can vary greatly, ranging from potassium urate in some tissues to calcium carbonate in others. Even though such crystals should be quite distinct in shape, the concretions produced by the insects all have the same distinct concentric, spherical shape. This set of properties argues that the organic compounds have a profound effect on crystal form and formation.
Ileum The principal function of the ileum is to act as a tubular epithelium that serves to transport to the rectum the undigested remains of the food from the midgut and fluid from the Malpighian tubules. This transport occurs by peristaltic movements of the circular and longitudinal muscles surrounding the ileum. The ileum also engages in important transepithelial transport functions. This has been investigated in considerable detail in the locust Schistocerca gregaria. In this species, potassium and chloride ions are transported from the lumen of the ileum into the hemolymph. This transport is isoosmotic. It therefore does not contribute directly to osmotic regulation but serves instead to reduce the volume of the urine and to retain valuable ions and water in the hemolymph. This transport is under hormonal control, presumably to allow the insect to modulate the return of water to the hemolymph depending on whether osmotic condition of the animal dictates a diuretic or an antidiuretic response. In some insects, an additional segment of the hindgut exists, which is termed the colon. Although this segment is hard to distinguish with the unaided eye, it is functionally and histologically distinct from the ileum. In larvae of E. hians, for example, an ileal segment occurs near the midgut, while a colonic segment of the hindgut lies between the ileum and the rectum. It has been shown that active ion transport occurs in the colon. The colon has a relatively low osmotic permeability, thus allowing the secretion in this segment of a fluid that is strongly hyperosmotic to the hemolymph. Production of hyperosmotic excreta is crucial for the osmotic regulation in this species because the insects live in the waters of a saline lake, the osmotic concentration of which is six times more concentrated than the hemolymph. The ions transported in the colon include sodium, chloride, and sulfate. Sulfate ions are large in comparison
to other transported ions; therefore the transport of sulfate through an epithelium capable of maintaining a substantial osmotic gradient is unusual. The larvae of the blow fly, Sarcophaga bullata, have also been shown to engage in active transport in the colon. In this species, the colon is a major site for the excretion of nitrogenous waste in the form of ammonium ion. Because these larvae feed in rotting flesh, the active transport of ammonium is a critical adaptive feature in the physiology of the species.
Rectum In most insects, the rectum is the most active ion-transporting organ on a per-gram basis. All fluids and solids deriving from the midgut and Malpighian tubules pass through the ileum and enter the rectum before being excreted. The rectum is therefore the last location in the gut in which the ionic and osmotic concentration of the excreta can be modified to meet the regulatory needs of the insect. In terrestrial animals, the requirements for osmotic homeostasis vacillate between the production of a dilute excreta (diuresis) and the production of a concentrated excreta (antidiuresis). Control of the rectum is therefore a critical element in the maintenance of osmotic homeostasis in the hemolymph. None of the other elements of the excretory system discussed thus far are capable of producing a fluid differing in osmotic concentration from that of the hemolymph. The role of the rectum in terrestrial insects has been most intensively studied in S. gregaria. In this insect, the cells in the rectal pads serve to transport a hypo-osmotic fluid from the lumen into the hemolymph. This serves to produce excreta with a very high osmotic concentration and, in the process, conserve water in the hemolymph. The process by which the locust transports a hypo-osmotic fluid is complicated and, unlike a functionally analogous process in the kidney of mammals, it requires cells of only a single type, the cuticle-covered rectal pad cells. In the parts of the rectum that are differentiated into rectal pads, the cells underlying the cuticle have deep apical infolds associated with numerous mitochondria. The rectal epithelium in the regions of the rectal pads is thick, meaning that the rectal pad cells comprise a tall, columnar epithelium. The intercellular junctions in these cells are highly convoluted and contain open spaces or intercellular swellings in the clefts between the cells. The process of fluid resorption from the lumen begins with the active transport of ions across the apical membrane (i.e., from the lumen to the intracellular compartment). Once in the cytoplasm, the ions are transported across the intercellular membrane into the enlarged spaces in the intercellular clefts. The compounds transported are principally potassium and chloride, although other compounds including acetate and proline are actively transported out of the lumen as well. These transported compounds produce a fluid with high osmotic concentration. It is thought that water is drawn from the lumen into the intercellular clefts, probably through the apical septate junctions. As a result, fluid accumulates in the intercellular clefts and in the open spaces in the intercellular regions. From here, the fluid flows extracellularly between the cells in a basal direction toward the hemolymph. It is thought that as this fluid flows, transporters within the lateral cell membranes remove ions. If these membranes have a low osmotic permeability, ions can move across with little water following. As a result, ions are removed faster than water can follow, resulting in a fluid that is hypo-osmotic not only to the lumen but also to the hemolymph. Under conditions in which the insect is well hydrated (e.g., after eating lush vegetation), the rectum removes ions from the rectal
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lumen but little water follows, presumably because either the site or the rate of transport in the more lateral and basal membranes has been modified. This produces a dilute urine, the excretion of which serves the osmotic needs of the insect. Aquatic insects are similarly dependent on the rectum for the final modification of the urine prior to excretion. In freshwater insects, the fluid derived from the midgut and Malpighian tubules is iso-osmotic to the hemolymph. Excretion of this fluid would lead to rapid loss of ions and the death of the animal. The rectum serves to transport ions from this primary urine back into the hemolymph. Transport of potassium and chloride has been documented for a number of freshwater insects. These transport mechanisms are relatively easy to demonstrate because the fluid entering the rectum from the Malpighian tubules is enriched in these two ions, and the excreted urine leaving the rectum much depleted. Rectal function has also been investigated in aquatic insects residing in hyperosmotic media, for example, in saline-tolerant dipteran larvae inhabiting coastal and desert saline waters. In species of Aedes inhabiting these waters, the rectum is differentiated into two segments. The anterior rectal segment is identical in function to the rectum of freshwater species and serves to remove ions from the urine under conditions in which the larvae find themselves in hypoosmotic media (i.e., fresh water). When the larvae hatch in saltwater, or when the medium becomes concentrated because of evaporation, the posterior rectal segment becomes active. This segment has a single cell type, which is characterized by deep apical and basal infolds associated with numerous mitochondria. The cells actively transport ions from the hemolymph into the rectal lumen. Because the epithelium has a low osmotic permeability, ions are transported faster than water can follow. As a result, a concentrated urine is produced by secretion in this segment, which has been called the salt gland. The ions transported in the posterior rectal segment vary with the environment in which the larvae occur. In seawater, sodium, magnesium, and chloride predominate. In bicarbonate-rich waters, a concentrated fluid is secreted, and the urine is rich in sodium and bicarbonate. The precise molecular mechanisms of ion transport in the rectum, as well as their neuronal or hormonal control, are poorly known for aquatic insects.
See Also the Following Articles Digestion ■ Fat Body ■ Hemolymph ■ Water and Ion Balance
Further Reading Beyenbach, K. W. (1995). Mechanisms and regulation of epithelial transport across Malpighian tubules. J. Insect Physiol. 41, 197–207. Bradley, T. J. (1985). The excretory system: Structure and physiology. In “Comprehensive Insect Physiology, Biochemistry and Pharmacology,” (G. A. Kerkut, and L. I. Gilbert, eds.), pp. 421–465. Pergamon Press, Oxford, U.K. Bradley, T. J. (1998). Malpighian tubules. In “Microscopic Anatomy of the Invertebrates. “Insecta” (M. Locke, and F. W. Harrison, eds.), Vol. XI, pp. 809–829. Liss, New York. Chapman, R. F. (1998). “The Insects: Structure and Function.” Cambridge University Press, Cambridge, U.K. Herbst, D. B., and Bradley, T. J. (1989). A Malpighian tubule lime gland in an insect inhabiting alkaline salt lakes. J. Exp. Biol. 145, 63–78. Phillips, J. E., and Audsley, N. (1995). Neuropeptide control of ion and fluid transport across locust hindgut. Am. Zool. 35, 503–514. Wigglesworth, V. B. (1965). “The Principles of Insect Physiology.”, 6th ed. Methuen, London.
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Exopterygota Ring T. Cardé University of California, Riverside
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xopterygota is a division of the class Insecta in the phylum Arthropoda. The orders of insects in this division have wings that develop externally during the maturation of the larva (which is variously referred to as a larva, nymph, or naiad). There are two superorders in this division: the Orthopteroidea (which includes the orders Blattodea, Dermaptera, Embiidina, Grylloblattodea, Isoptera, Mantodea, Mantophasmatodea, Orthoptera, Phasmatodea, and Plecoptera) and the Hemipteroidea (Hemiptera, Phthiraptera, Psocoptera, Thysanoptera, and Zoraptera). Except for the developed wings and genitalia, there is a strong morphological resemblance between larvae and adults (although habitats and biology may differ greatly).
Exoskeleton Svend O. Andersen The Royal Danish Academy of Sciences and Letters
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he exoskeleton is noncellular material that is located on top of the epidermal cell layer and constitutes the outermost part of the integument. The local properties and appearance of the exoskeleton are highly variable, and nearly all visible features of an insect result from the exoskeleton. The exoskeleton serves as a barrier between the interior of the insect and the environment, preventing desiccation and penetration of microorganisms. Muscles governing the insect’s movements are attached to the exoskeleton. Although the exoskeleton is a continuous structure, its mechanical properties differ from region to region. Sometimes the transition between regions is gradual, but often it is quite abrupt; pliant and elastic regions can border on hard and heavily sclerotized regions. Most exoskeletal regions of soft-bodied larvae, such as larvae of moths and flies, are soft and pliant, and only restricted regions of their exoskeletons, such as legs, head capsule, and mandibles, are hard and stiff. Most of the body surface of adult, winged insects is covered by a stiff exocuticle, which can be somewhat flexible and bendable but also serves as a hard protective armor. The exoskeleton covering the dorsal abdomen of many beetle species is thin and easily flexed, whereas the ventral abdominal exoskeleton of the same animals is hard and resistant. The mechanical properties of all exoskeletal regions are precisely adapted to be optimal for the lifestyle of the insect.
FORMATION OF THE EXOSKELETON The exoskeleton is produced and modified by the epidermal cell layer, and each cell in the epidermis must have the necessary information for producing and depositing the right amount of the right cuticular components at the right time; later some of them will modify the secreted products to give a mature material. The timing of the various events is often hormonally controlled, but the quantitative information on how much to produce must be inherent in individual epidermal cells.
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Estivation see Aestivation
Evolution
see Phylogeny of Insects
E Excretion Timothy J. Bradley University of California, Irvine
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xcretion is the elimination of excess ions, water, and metabolic wastes from the body. As in all organisms, excretion in insects serves to promote the appropriate regulation (homeostasis) of the intracellular environment as the key to organismal well-being and survival. Regulation of the intracellular environment depends in turn on appropriate regulation of the extracellular fluids. In insects, these extracellular fluids are contained in the hemolymph and in the interstitial fluids surrounding the cells. Excretion therefore refers to the processes by which wastes, as well as excess ions and water, are eliminated from the hemolymph. Wastes produced in the cells and transported into the hemolymph are concentrated and excreted by the excretory organs.
STRUCTURE OF THE EXCRETORY ORGANS The processes of excretion in insects are carried out largely by the organs of the insect gut. These include the midgut, the Malpighian tubules, and the structures in the hindgut, namely the ileum and rectum (Fig. 1).
FIGURE 1 A diagrammatic representation of the insect gut.
Midgut The midgut is a tubular epithelium. Upon ingestion, food and fluids move through the esophagus and pass directly into the midgut. Because the cells in the midgut epithelium are derived from embryonic endoderm, the midgut is not lined with cuticle. Most of the cells in the midgut are involved in the secretion of digestive fluids and the
absorption of nutrients from the midgut lumen. These secretory and absorptive cells have apical microvilli that greatly increase the surface area available for inward and outward transport. In insects that feed periodically (such as adult mosquitoes), the microvilli shorten during nonfeeding periods and lengthen following ingestion. Many insects have additional cell types termed goblet cells that are thought to be involved in the secretion of fluids that modify the acidity and alkalinity (pH) of the luminal fluid. These goblet cells have been intensively investigated in Lepidoptera, where they serve to produce a markedly alkaline pH. The basal surface of the midgut cells possesses a network of longitudinal and circular muscles that upon contraction can produce peristaltic waves. These contractions serve to move the food along the gut and stir the midgut contents during digestion. Many insects possess globular outpocketings in the anterior region of the midgut, termed ceca. The cells types in the ceca are generally differentiated from those in the midgut proper.
Malpighian Tubules The Malpighian tubules are the site of urine formation in all insects except the Collembola, Thysanura, and aphids. The Malpighian tubules are tubular epithelia that are diverticulae (outpocketed extensions) of the gut itself. The tubules open into the gut near the midgut–hindgut junction, and the lumina of these two tubular epithelia are continuous. The contents of the tubules flow into the gut lumen; the ends of the tubules distal to the gut are closed. Fluid is produced in the Malpighian tubules by secretion; and because the tubules are closed at the distal end, hydrostatic pressure builds up and fluid flows through the tubules into the gut. The number of Malpighian tubules is quite variable depending on the insect species. Bloodsucking Hemiptera (e.g., Rhodnius prolixus) and higher Diptera (e.g., Drosophila melanogaster) have as few as four tubules, whereas the desert locust (Schistocerca gregaria) has hundreds. Attached to the Malpighian tubules of many insects are longitudinal muscles. When these muscles contract, the tubules are waved about in the hemolymph, presumably for the purpose of stirring the fluid adjacent to the tubules and promoting fluid and solute transport. These tubules may also serve the more general function of promoting hemolymph circulation throughout the abdomen. The Malpighian tubules of all species examined to date contain more than one cell type. In some cases, a single epithelial region contains two or more cell types (regions with heterologous cell types) reflecting, presumably, separate physiological roles for each cell type. In other species, the tubules are divided into distinct regions, each consisting of a single cell type (regions with homologous cell types). In these insects, each tubule region has a distinct function in transport. Finally, in many insects, the tubules show regional specialization as well as multiple cell types within a region. It is presumed that each cell type in these tubules performs a distinct function. As an example of cell type heterogeneity, consider the Malpighian tubules of the larvae of the brine fly, Ephydra hians (Fig. 2). The tubules in this insect are differentiated on each side of the body, as well as along their length. On one side of the body is a pair of tubules called the lime gland tubules. The distal ends of these are secretory and contain two regions that can be differentiated on the basis of cell color: one white, one yellow. More proximal to the gut are expanded regions of the tubules that serve to store concentric concretions in the tubule lumen. Finally, two of these cells combine in a common ureter that empties into the gut. On the opposite side, the tubules have only the yellow and white regions of the tubules, with no storage
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Midgut Unmodified tubule (posterior) Hindgut
Ureter
Rectum
Proximal tubule of lime gland Storage region of time gland Distal tubule of lime gland
FIGURE 2 A schematic diagram of the Malpighian tubules of E. hians. The Malpighian tubules are differentiated on the left and right side. The hindgut is composed, from anterior to posterior, of the ileum, colon, and rectum. [From Herbst, D. B., and Bradley, T. J. (1989). A Malpighian tubule lime gland in an insect inhabiting alkaline salt lakes. J. Exp. Biol. 145, 63–78.]
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core of microfilaments. Frequently, these microvilli contain fingerlike extensions of the mitochondria and even of the endoplasmic reticulum. The basal surface of the cells exhibits deep infolds, often again closely associated with mitochondria by means of structures termed scalariform junctions. The intercellular spaces are occupied apically by septate or continuous junctions. More basally, the intercellular space contains gap junctions or undifferentiated basolateral membranes indistinguishable from the basal membrane infolds. Numerous other cell types occur in the Malpighian tubules. It is presumed that each histologically distinguishable cell type performs a unique function within the tubules. In addition, distinct functions have been found in some cell types in the absence of histological or ultrastructural differentiation. A common cell type in Malpighian tubules is the stellate or secondary cell. This cell type possesses smaller microvilli than the primary cells, and these microvilli contain no mitochondria. As described later in the section on function, the secondary cells may be involved in modification of the primary urine produced by the primary cells.
Ileum section. This example illustrates the variety of cell types that can exist in a single tubule. The details of transport function in these and other highly complex tubules have not been fully elucidated. In most Malpighian tubules, formation of the primary urine occurs in a cell type often referred to as the primary cell (Fig. 3). This cell type has extensive apical microvilli, often containing a central
Posterior to the midgut, most insects possess a segment of gut referred to as the ileum. Because this region is part of the hindgut, it is covered on the apical surface by cuticle. The cells often show deep apical and basal membrane infoldings reflecting the role of these cells in fluid and solute transport. The basal surface of the cells is covered by layers of longitudinal and circular muscles that serve to generate peristaltic movements of the gut. These muscular contractions move the gut contents through the gut and may also serve an important role in reducing unstirred layers adjacent to both the apical and basal membranes of the epithelium. Historically, the Malpighian tubules and rectum have been assumed to carry out most of the fluid transport in the posterior regions of the gut. The ileal epithelium is smaller in diameter, with less highly developed apical and basal membrane infolds, and a lower mitochondrial density than is observed in rectum. Nonetheless, when the transport properties of the ileum are investigated, this region of the gut is always found to be carrying out important transport functions.
Rectum
FIGURE 3 Model of the transport processes occurring in the Malpighian tubules of adult mosquitoes, based on the work of Klaus Beyenbach using the species A. aegypti. Upon stimulation with mosquito natriuretic peptide (MNP), rates of transepithelial fluid secretion increase from an unstimulated rate of 0.4–2.8 nl min1. In parallel, Na concentrations in the secreted fluid rise, and K concentrations fall. Electrophysiological studies reveal that MNP, working via cyclic AMP, induces an increase in basolateral membrane Na conductance, presumably through the actions of Na channels in the basolateral membrane of principal cells. The hyperpolarization of the transepithelial voltage and the decrease in transepithelial resistance are consistent with the activation of Na channels in the basolateral membrane of principal cells. (Figure and legend provided by Klaus Beyenbach.)
All insects possess an enlarged chamber called the rectum near the posterior end of the gut. The structure of the rectum can vary substantially from species to species. The rectal lumen is covered by a thick cuticle. Posterior to the rectum, insects possess an anal canal through which the feces and urine are eliminated. The strong bands of muscle surrounding the rectum contract during defecation, expelling the feces and urine into the external environment through the anus. The recta of terrestrial insects are large and very active organs. Regions within the rectum are highly differentiated and contain cells with deep membrane folds. If (as in many orthopterans and lepidopterans) these cells are contained in thickened ridges that extend into the rectal lumen, they are referred to as rectal pads. If (as in many adult Diptera) the cells extend into the lumen as fingerlike structures, often on a thin stalk, they are referred to as rectal papillae. The rectal pads and the papillae are the major sites of fluid resorption and urine concentration. The cells in the rectal pads have a complicated array of intercellular junctions associated with the active sites of ion secretion and resorption. The evolution of a rectal structure permitting the formation of a urine hyperosmotic to the
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hemolymph was, along with the evolution of a waxy cuticle, a major adaptive event permitting insects to invade drier terrestrial habitats. Both rectal pads and rectal papillae are covered with thick cuticle to protect the underlying epithelial cells from abrasion by the fecal material. Between the rectal pads, the rectum of terrestrial insects possesses a thin, cuticle-lined epithelium that, upon unfolding and stretching, permits rectal swelling during feces and fluid accumulation. In E. hians, the rectum is merely a thin, distensible epithelium that expands upon filling with feces prior to defecation. In this insect, the task of modifying the urine is carried out by the colon, which lies just anterior to the rectum. The presence of a rectum without transport capabilities is very unusual. In most insects the rectum is the major organ responsible for osmotic regulation of the urine.
Excretion outside the Gut Storage excretion of concentric concretions containing calcium salts of urate and carbonate occurs in the fat body cells of most insects. Some insects contain nephrocytes, cells in the head that also store wastes and toxic elements. Finally, in collembolans and thysanurans, cephalic nephridial glands, sometimes termed labial glands, are responsible for excretory function, including the excretion of nitrogenous waste.
FUNCTION OF THE EXCRETORY ORGANS Midgut TRANSPORT The primary function of the midgut in insects is the digestion and uptake of ingested nutritive materials. There are two processes that occur in the midgut and contribute to excretion: pH regulation and storage excretion. The midguts of many insects secrete fluids that contribute to extreme alkalinity or acidity in the gut. In Lepidoptera, for example, the midgut epithelium consists of goblet and columnar epithelial cells. The goblet cells are responsible for transporting ions and creating a highly alkaline solution in the midgut lumen. The columnar cells contribute to digestion of the food and uptake of ions. The apical membrane of the goblet cell contains hydrogen pumps (H-ATPases) that use adenosine triphosphate (ATP) as their source of energy. These pumps transport hydrogen ions into the cuplike apical crypt of the goblet cell. The resulting electrical and pH gradient is used in an exchange process (antiporter) to exchange potassium ions for the hydrogen ion. Potassium ions associated with hydroxyl ions remain in the lumen following the hydrogen/potassium exchange process. These ions diffuse from the goblet cavity into the midgut lumen, with the result that the potassium hydroxide (KOH) causes the lumen to be very alkaline. The high potassium concentration in the lumen is, in turn, used as an energy source for the uptake of amino acids from the digested foodstuffs. This uptake occurs by means of transporters (symporters) in the apical cell membranes of the columnar cells that use the potassium gradient to cotransport potassium ions and amino acids from the lumen into the cell interior. Various functions have been proposed for the highly alkaline pH in the midgut of Lepidoptera. Clearly, such an extreme pH would serve to kill pathogens and to saponify lipids in the ingested food material. It has also been proposed that this pH serves to reduce the solubility and activity of tannic acids in the food of Lepidoptera, protecting herbivorous larvae from the high concentrations of these toxic compounds found in the leaves of many plants. In other insects, for example, mosquito larvae, low-pH (acid) conditions are observed in the midgut. The precise mechanisms by which this pH is produced and maintained remain to be elucidated,
but there is evidence in mosquitoes that the midgut epithelium is also energized by the H-ATPase. STORAGE EXCRETION IN THE MIDGUT The columnar midgut cells of insects often contain concentric mineralized concretions. The major cation in these concretions has been shown to be calcium. The anion can be quite variable and has been found to be largely urate or carbonate, depending on the cell type in which the concretions are located. In the midgut, the concretions have also been shown to play a role in the storage excretion of toxic cations such as copper, zinc, iron, and selenium. Each of these elements is toxic in high concentrations but plays a critical role in metabolism in trace amounts. The concretions may therefore play both a protective and a homeostatic role in insects by regulating the free concentration of these ions and metals in the body. They reduce toxic concentrations and serve as a reservoir for these elements, which can be used for physiological purposes when required. Storage excretion in the form of concentric concretions also occurs in the Malpighian tubules.
Malpighian tubules TRANSPORT The production of urine in insects occurs by the active transport of ions across the epithelium from hemolymph to tubule lumen. This process generates an osmotic concentration that drives the movement of water across the epithelium as well. Generally, Malpighian tubules have a high permeability coefficient for water (a low osmotic resistance), and as a result water moves rapidly across the epithelium in response to relatively low osmotic concentration gradients. These concentration differences across the epithelium are indeed so low that they have been difficult to measure. Recent experimental results have led to a general consensus, however, that an osmotic gradient of a few milliosmoles is sufficient to account for the observed rates of water movement across Malpighian tubules. Let us consider first the active transport of ions across the epithelium, and then the passive movement of water that follows. In insects, potassium is the predominant cation transported across the epithelium of the Malpighian tubules. Insects evolved as a distinct clade on land, feeding on plants and detritus. It may be that their dependence on potassium as the major cation used for fluid transport evolved at this time as well. Certainly, animals of marine origin, such as the vertebrates and crustaceans, rely on sodium as the principal cation for driving fluid movements. In the cell interior, the Malpighian tubule cells have a negative electrical charge relative to the hemolymph. This electrical potential facilitates the entry of potassium into the cells. Thermodynamically speaking, therefore, the most energetically costly transmembrane movement for potassium in the epithelium occurs as this cation crosses the apical membrane. The process by which this occurs has been very difficult to unravel, but in recent years it has been demonstrated that a very active hydrogen ATPase, related to vacuolar H-ATPase found widely in eukaryotic cells, is located on the apical membrane of Malpighian tubule cells. This transporter moves hydrogen ions from the cell interior into the tubule lumen, thereby setting up a large electrochemical gradient for hydrogen ions. Depending on the circumstances, this electrochemical gradient can be predominantly expressed as a pH gradient or as an electrical gradient. In either circumstance, this electrochemical gradient is thought to serve as an energy reservoir that can be subsequently used for a variety of coupled transport processes.
Excretion
For example, if the energy contained in the hydrogen ion gradient is used in an antiporter (a transporter that couples ion movement in one direction to ion movement in the opposite direction) that exchanges hydrogen for potassium at the apical membrane, the result of hydrogen transport with subsequent hydrogen exchange for potassium is a net active transport of potassium. Potassium is moved against its electrochemical gradient by the antiporter, using the energy contained in the transmembrane hydrogen ion gradient. In some Malpighian tubules, similar mechanisms may exist for the coupled transport of other cations (e.g., sodium, calcium, or magnesium). In bloodsucking insects such as adult mosquitoes and the reduviid R. prolixus, the plasma portion of the blood meal also provides the insect with a substantial intake of sodium. Any amount of sodium that exceeds the physiological needs of the insect must be excreted. In these insects, a substantial active transport of sodium occurs accords the Malpighian tubules. This process is thought to be driven across the apical membrane by a hydrogen–sodium exchange mechanism. Cation transport across the epithelium also requires a process for entry of the ions into the cytoplasm from the hemolymph. In most insects, the basal membrane contains an enzyme (Na–K ATPase) that uses the energy in ATP to transport sodium actively out of the cell and potassium actively in. As a result, the cytoplasm is greatly enriched in potassium. The passive outward diffusion of this ion through barium-sensitive channels produces an electrical potential across the basal membrane, the inside of which is negatively charged. This potential can be used for a variety of transport functions; one that seems to be almost universally present in Malpighian tubules is the bumetanide-sensitive Na/K/2Cl, cotransporter. This transporter uses the energy contained in the sodium gradient to move one sodium, one potassium, and two chloride ions simultaneously from the hemolymph into the cytoplasm. This process serves to provide chloride to the cell interior, as well as sodium in the cell types to which sodium is moved transepithelially. The movement of anions across the epithelium involves one or more of three distinct transport pathways, depending on the characteristics of the tubules. In the first, chloride is thought to move across the same cells as those in which the cations are transported. As mentioned earlier, the interior of the cells is electrically negative relative to the cell exterior. The movement of chloride into the cell is therefore a thermodynamically active process. As already discussed, it may be driven by the sodium electrochemical gradient in a cotransport process. In other insect species in which the Malpighian tubule cells in a single region of the tubule are differentiated into two or more cell types, chloride ions have been shown to enter the lumen via a cell type distinct from that involved in active cation transport. For example, in D. melanogaster, the fluid-transporting segment of the tubules contains both primary and stellate cells. The former cells are the sites of active potassium transport, the latter the site of chloride flux from the hemolymph to the tubule lumen. Although this movement of chloride into the tubule lumen is thermodynamically downhill, the precise mechanism of chloride transport is presently unknown. A third process has been described in the Malpighian tubules of adult mosquitoes. Although these insects also possess primary and stellate cells in the Malpighian tubules, it has been proposed that chloride moves into the lumen of the tubules via the intercellular junctions. In fact, this process has been shown to be under hormonal control. The movements of chloride into the Malpighian tubules may therefore be quite variable depending on cell types. The movements of anions are much less well characterized at this time, with regard to the molecules
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that drive the process, than are the movements of cations. A model of ion transport at both the apical and basal membrane of the Malpighian tubules of adult mosquitoes is shown in Fig. 3. Regardless of the mechanisms by which cations and anions enter the Malpighian tubules, it is clear that the types of ions transported can vary greatly with the species of insect and will depend on an individual’s physiological needs and demands. Thus since bloodsucking insects ingest a large amount of sodium compared with other insects, the Malpighian tubules of bloodsuckers contain specific mechanisms designed to reduce the large sodium load. Species of mosquitoes whose larvae can survive in saltwater must ingest the medium and eliminate the ions as means of obtaining water. Those species that have been investigated can excrete magnesium and sulfate via the Malpighian tubules. The larvae of brine flies (ephydrids) generate crystals in the lumina of the tubules that are rich in calcium and carbonate. Both these ions must be transported across the epithelium, although the combination forms insoluble crystals that reduce the activity of these ions in the lumen. The Malpighian tubules of insects are also the site of excretion of the waste products of energy and nitrogen metabolism. Acid byproducts of energy metabolism have been shown to be actively transported into the lumen from the hemolymph. The precise molecular mechanisms of the process remain unclear, but the process is of paramount importance for the insects in maintaining acid/base balance and energy homeostasis. The by-products of nitrogen metabolism are also excreted by the Malpighian tubules. In aquatic insects, ammonia may be excreted, but in most insects and certainly in terrestrial forms, urea and uric acid predominate. Both these compounds are actively removed from the hemolymph by the Malpighian tubules. The transport of uric acid has been investigated in R. prolixus, in which the blood meal provides a very protein-rich meal requiring intense capacity for the elimination of nitrogenous waste. In Rhodnius, the primary urine is produced in the most upstream portions of the Malpighian tubules, the upper tubule. This urine is modified in the downstream section (the lower tubule) through the resorption of potassium and chloride. This process serves to return potassium to the hemolymph, and to remove waste from the hemolymph, through the retrieval of an isosmotic fluid. Thus hemolymph volume is retained and the sodium in the urine is concentrated. In addition, uric acid is transported in the lower tubule from hemolymph to urine. Potassium urate is fairly insoluble, particularly at neutral to acid pH. As a result, crystals of uric acid form in the urine. This process further removes osmotically active compounds from the urine, allowing the additional movement of water from the urine to the hemolymph by osmosis. CONCENTRIC CONCRETIONS IN THE MALPIGHIAN TUBULES Concentric concretions occur in the midgut, where they are thought to contribute to excretion by storage in an insoluble form of salts containing calcium, magnesium, manganese, copper, cadmium, and zinc. Identical concretions are found intracellularly in the Malpighian tubules. These concretions are thought to perform an identical function, namely storage of ions in an insoluble form either for subsequent use or as a means of removing the ions from the body. In the Malpighian tubules, however, these concretions also appear in the tubule lumen, a location from which they can move into the gut and be eliminated with the excrement. It has been suggested by many authors that the intracellular concretions in the cells of the Malpighian tubules can be transported by exocytosis into the lumen of the tubules. Although there are occasionally physiological conditions in which the concretions disappear from
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the cells and appear in the lumen, it has not been unambiguously demonstrated that the crystals move from one location to the other intact. Instead, it is likely that the crystals are dissolved within the cells of the Malpighian tubules, that the soluble ions are transported into the lumen, and that the crystals are formed anew in the tubule lumen. Crystals are formed in some tubule segments (e.g., in the lower tubule of R. prolixus) where no crystals exist in the cells. The crystals in the midgut, fat body, and Malpighian tubules are concentric and perfectly round. This is in marked contrast to the natural structure of the crystals formed by the same salts in solution. Uric acid crystals, for example, have sharp corners and sometimes take a needlelike form. It is thought that the concentric concretions avoid acicularity through the activity of organic compounds that are known to be a substantial component of the concretions. The compounds are thought to nucleate and direct crystal formation, leading to the formation of round concretions. This spherical shape is less damaging to the cells of the tubules and can be excreted from the tubules and gut with little tissue damage. Ultrastructurally identical concretions are observed in the urine of birds, which is rich in uric acid. It has been proposed that organic compounds are excreted into the tubule lumen, where they nucleate and guide the formation of the concretions. The ions contained in the concretions can vary greatly, ranging from potassium urate in some tissues to calcium carbonate in others. Even though such crystals should be quite distinct in shape, the concretions produced by the insects all have the same distinct concentric, spherical shape. This set of properties argues that the organic compounds have a profound effect on crystal form and formation.
Ileum The principal function of the ileum is to act as a tubular epithelium that serves to transport to the rectum the undigested remains of the food from the midgut and fluid from the Malpighian tubules. This transport occurs by peristaltic movements of the circular and longitudinal muscles surrounding the ileum. The ileum also engages in important transepithelial transport functions. This has been investigated in considerable detail in the locust Schistocerca gregaria. In this species, potassium and chloride ions are transported from the lumen of the ileum into the hemolymph. This transport is isoosmotic. It therefore does not contribute directly to osmotic regulation but serves instead to reduce the volume of the urine and to retain valuable ions and water in the hemolymph. This transport is under hormonal control, presumably to allow the insect to modulate the return of water to the hemolymph depending on whether osmotic condition of the animal dictates a diuretic or an antidiuretic response. In some insects, an additional segment of the hindgut exists, which is termed the colon. Although this segment is hard to distinguish with the unaided eye, it is functionally and histologically distinct from the ileum. In larvae of E. hians, for example, an ileal segment occurs near the midgut, while a colonic segment of the hindgut lies between the ileum and the rectum. It has been shown that active ion transport occurs in the colon. The colon has a relatively low osmotic permeability, thus allowing the secretion in this segment of a fluid that is strongly hyperosmotic to the hemolymph. Production of hyperosmotic excreta is crucial for the osmotic regulation in this species because the insects live in the waters of a saline lake, the osmotic concentration of which is six times more concentrated than the hemolymph. The ions transported in the colon include sodium, chloride, and sulfate. Sulfate ions are large in comparison
to other transported ions; therefore the transport of sulfate through an epithelium capable of maintaining a substantial osmotic gradient is unusual. The larvae of the blow fly, Sarcophaga bullata, have also been shown to engage in active transport in the colon. In this species, the colon is a major site for the excretion of nitrogenous waste in the form of ammonium ion. Because these larvae feed in rotting flesh, the active transport of ammonium is a critical adaptive feature in the physiology of the species.
Rectum In most insects, the rectum is the most active ion-transporting organ on a per-gram basis. All fluids and solids deriving from the midgut and Malpighian tubules pass through the ileum and enter the rectum before being excreted. The rectum is therefore the last location in the gut in which the ionic and osmotic concentration of the excreta can be modified to meet the regulatory needs of the insect. In terrestrial animals, the requirements for osmotic homeostasis vacillate between the production of a dilute excreta (diuresis) and the production of a concentrated excreta (antidiuresis). Control of the rectum is therefore a critical element in the maintenance of osmotic homeostasis in the hemolymph. None of the other elements of the excretory system discussed thus far are capable of producing a fluid differing in osmotic concentration from that of the hemolymph. The role of the rectum in terrestrial insects has been most intensively studied in S. gregaria. In this insect, the cells in the rectal pads serve to transport a hypo-osmotic fluid from the lumen into the hemolymph. This serves to produce excreta with a very high osmotic concentration and, in the process, conserve water in the hemolymph. The process by which the locust transports a hypo-osmotic fluid is complicated and, unlike a functionally analogous process in the kidney of mammals, it requires cells of only a single type, the cuticle-covered rectal pad cells. In the parts of the rectum that are differentiated into rectal pads, the cells underlying the cuticle have deep apical infolds associated with numerous mitochondria. The rectal epithelium in the regions of the rectal pads is thick, meaning that the rectal pad cells comprise a tall, columnar epithelium. The intercellular junctions in these cells are highly convoluted and contain open spaces or intercellular swellings in the clefts between the cells. The process of fluid resorption from the lumen begins with the active transport of ions across the apical membrane (i.e., from the lumen to the intracellular compartment). Once in the cytoplasm, the ions are transported across the intercellular membrane into the enlarged spaces in the intercellular clefts. The compounds transported are principally potassium and chloride, although other compounds including acetate and proline are actively transported out of the lumen as well. These transported compounds produce a fluid with high osmotic concentration. It is thought that water is drawn from the lumen into the intercellular clefts, probably through the apical septate junctions. As a result, fluid accumulates in the intercellular clefts and in the open spaces in the intercellular regions. From here, the fluid flows extracellularly between the cells in a basal direction toward the hemolymph. It is thought that as this fluid flows, transporters within the lateral cell membranes remove ions. If these membranes have a low osmotic permeability, ions can move across with little water following. As a result, ions are removed faster than water can follow, resulting in a fluid that is hypo-osmotic not only to the lumen but also to the hemolymph. Under conditions in which the insect is well hydrated (e.g., after eating lush vegetation), the rectum removes ions from the rectal
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lumen but little water follows, presumably because either the site or the rate of transport in the more lateral and basal membranes has been modified. This produces a dilute urine, the excretion of which serves the osmotic needs of the insect. Aquatic insects are similarly dependent on the rectum for the final modification of the urine prior to excretion. In freshwater insects, the fluid derived from the midgut and Malpighian tubules is iso-osmotic to the hemolymph. Excretion of this fluid would lead to rapid loss of ions and the death of the animal. The rectum serves to transport ions from this primary urine back into the hemolymph. Transport of potassium and chloride has been documented for a number of freshwater insects. These transport mechanisms are relatively easy to demonstrate because the fluid entering the rectum from the Malpighian tubules is enriched in these two ions, and the excreted urine leaving the rectum much depleted. Rectal function has also been investigated in aquatic insects residing in hyperosmotic media, for example, in saline-tolerant dipteran larvae inhabiting coastal and desert saline waters. In species of Aedes inhabiting these waters, the rectum is differentiated into two segments. The anterior rectal segment is identical in function to the rectum of freshwater species and serves to remove ions from the urine under conditions in which the larvae find themselves in hypoosmotic media (i.e., fresh water). When the larvae hatch in saltwater, or when the medium becomes concentrated because of evaporation, the posterior rectal segment becomes active. This segment has a single cell type, which is characterized by deep apical and basal infolds associated with numerous mitochondria. The cells actively transport ions from the hemolymph into the rectal lumen. Because the epithelium has a low osmotic permeability, ions are transported faster than water can follow. As a result, a concentrated urine is produced by secretion in this segment, which has been called the salt gland. The ions transported in the posterior rectal segment vary with the environment in which the larvae occur. In seawater, sodium, magnesium, and chloride predominate. In bicarbonate-rich waters, a concentrated fluid is secreted, and the urine is rich in sodium and bicarbonate. The precise molecular mechanisms of ion transport in the rectum, as well as their neuronal or hormonal control, are poorly known for aquatic insects.
See Also the Following Articles Digestion ■ Fat Body ■ Hemolymph ■ Water and Ion Balance
Further Reading Beyenbach, K. W. (1995). Mechanisms and regulation of epithelial transport across Malpighian tubules. J. Insect Physiol. 41, 197–207. Bradley, T. J. (1985). The excretory system: Structure and physiology. In “Comprehensive Insect Physiology, Biochemistry and Pharmacology,” (G. A. Kerkut, and L. I. Gilbert, eds.), pp. 421–465. Pergamon Press, Oxford, U.K. Bradley, T. J. (1998). Malpighian tubules. In “Microscopic Anatomy of the Invertebrates. “Insecta” (M. Locke, and F. W. Harrison, eds.), Vol. XI, pp. 809–829. Liss, New York. Chapman, R. F. (1998). “The Insects: Structure and Function.” Cambridge University Press, Cambridge, U.K. Herbst, D. B., and Bradley, T. J. (1989). A Malpighian tubule lime gland in an insect inhabiting alkaline salt lakes. J. Exp. Biol. 145, 63–78. Phillips, J. E., and Audsley, N. (1995). Neuropeptide control of ion and fluid transport across locust hindgut. Am. Zool. 35, 503–514. Wigglesworth, V. B. (1965). “The Principles of Insect Physiology.”, 6th ed. Methuen, London.
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Exopterygota Ring T. Cardé University of California, Riverside
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xopterygota is a division of the class Insecta in the phylum Arthropoda. The orders of insects in this division have wings that develop externally during the maturation of the larva (which is variously referred to as a larva, nymph, or naiad). There are two superorders in this division: the Orthopteroidea (which includes the orders Blattodea, Dermaptera, Embiidina, Grylloblattodea, Isoptera, Mantodea, Mantophasmatodea, Orthoptera, Phasmatodea, and Plecoptera) and the Hemipteroidea (Hemiptera, Phthiraptera, Psocoptera, Thysanoptera, and Zoraptera). Except for the developed wings and genitalia, there is a strong morphological resemblance between larvae and adults (although habitats and biology may differ greatly).
Exoskeleton Svend O. Andersen The Royal Danish Academy of Sciences and Letters
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he exoskeleton is noncellular material that is located on top of the epidermal cell layer and constitutes the outermost part of the integument. The local properties and appearance of the exoskeleton are highly variable, and nearly all visible features of an insect result from the exoskeleton. The exoskeleton serves as a barrier between the interior of the insect and the environment, preventing desiccation and penetration of microorganisms. Muscles governing the insect’s movements are attached to the exoskeleton. Although the exoskeleton is a continuous structure, its mechanical properties differ from region to region. Sometimes the transition between regions is gradual, but often it is quite abrupt; pliant and elastic regions can border on hard and heavily sclerotized regions. Most exoskeletal regions of soft-bodied larvae, such as larvae of moths and flies, are soft and pliant, and only restricted regions of their exoskeletons, such as legs, head capsule, and mandibles, are hard and stiff. Most of the body surface of adult, winged insects is covered by a stiff exocuticle, which can be somewhat flexible and bendable but also serves as a hard protective armor. The exoskeleton covering the dorsal abdomen of many beetle species is thin and easily flexed, whereas the ventral abdominal exoskeleton of the same animals is hard and resistant. The mechanical properties of all exoskeletal regions are precisely adapted to be optimal for the lifestyle of the insect.
FORMATION OF THE EXOSKELETON The exoskeleton is produced and modified by the epidermal cell layer, and each cell in the epidermis must have the necessary information for producing and depositing the right amount of the right cuticular components at the right time; later some of them will modify the secreted products to give a mature material. The timing of the various events is often hormonally controlled, but the quantitative information on how much to produce must be inherent in individual epidermal cells.
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