We have spent a little bit of time considering aspects of movement. We've gotten an idea of the contractile process in muscle, a process driven by hydrolysis of ATP. We saw that ATP is generated as an end product of energy metabolism and considered gas exchange as a requisite for energy metabolism. Flight is very important to many aspects of insect biology, and I purposefully emphasize flight. Let us also emphasize the broader biological issue in flight. Flight can be taken under the mantle of motility, or movement. In this view, flight is one of many kinds of movement, all fundamental processes in biology.

Let us now turn attention to still another are of physiology confronting nearly every life form: osmotic balance. Osmotic regulation is the business of maintaining optimal salt and water concentrations in an organism. We begin by considering the biological significance of this. About 50 to 90% of the body weight of an insect, or nearly any other animal, is water. Some of this fluid is inside the various cells that make up the body while the remainder is in the spaces outside of the cells. As a unit, these spaces are regarded as a collective known as the extracellular space. The extracellular space is filled with fluid, for obvious reasons called the extracellular fluid. Among the dissolved constituents of the extracellular fluids are the ions and the nutrients needed by the cells for maintenance of life. The extracellular fluid is in constant motion throughout the body, and it is rapidly mixed to near homogeneity by hemolymph circulation and by diffusion. Because of these mixing processes, we can say that all cells in the body live in essentially the same environment. The great French physiologist, Bernard, called the extracellular fluid the melieu int`erieur or what we would call the internal environment.

The point is to recognize two main fluid compartments in an insect body. One is the intracellular compartment and the other is the extracellular compartment. To appropriate a teaching device from Daniel Matzia, the famed cell biologist retired from UC Berkeley, we can regard the extracellular fluids as the out environment, that is, the environment inside the body that cells experience. The intracellular environment, then, could be the in environment. However, we know eucaryotic cells have organells and compartments within them. Within cells, the compartment inside any particular organelle, such as a nucleus or mitochondrion, could be the in-in environment. At the organismal level, the world we live in is the out-out environment. This language allows us to appreciate the world in terms of environments that things experience: the in-in, the in, the out and the out-out environments. Cells experience the out environment, the same thing that Bernard called the melieu int`erieur.

Some percentage of the total body fluids of any organism are intracellular fluids. The fluid within each cell is made up of its own individual mixture of various constituents, such as ions, amino acids, proteins, sugars and so forth. On the other hand, within an organism, the concentrations of these things appear to be reasonably similar from cell to cell. For our discussion, the intracellular fluids of all of the different cells of an organism can be considered to be one large fluid compartment: the in environment or the intracellular compartment. Regardless of the particular language or expression that is easiest to keep in mind, we want to come to the point of view that an insect body has two major fluid compartments: the intracellular and extracellular compartments. The extracellular compartment is easy to visualize as the circulating hemolymph.

While the extracellular environment is substantially different in composition from the intracellular environment, the two compartments are in osmotic balance. That is to say that there is rarely a net flow of water from one compartment to another. There are various mechanisms to handle the maintenance of the correct concentrations of the intracellular constituents, including active transport and facilitated diffusion. These mechanisms are a related, but different matter. The thrust for our purposes is that water always tends to diffuse by a process called osmosis across cellular membranes from the side with fewer total dissolved particles to the side with a greater number of dissolved particles.

Let us consider some possibilities. First, if a cell in placed in a solution with far fewer osmotic particles than the cell, water will diffuse into the cell. This can result in considerable swelling, even to the point of bursting the cell. Alternatively, if a cell is placed into a solution with a far greater number of particles, water tends to diffuse, once again, down its osmotic gradient, in this case, out of the cell. A badly shrunken cell that may or may not survive may result from this flow. We gather the significance of a physical phenomenon, osmosis. Osmosis explains the importance of maintaining a proper balance of water and of salts in the extracellular compartment or the melieu int`erieur. During times when, for whatever reason, the extracellular compartment is not correctly regulated, the physiological situation is exactly like placing cells into a solution that is too dilute or too concentrated. This is a common problem faced by insects and most other organisms living in freshwater, salt water or in terrestrial environments.

Again, terrestrial insects are faced with the problem of regulating their levels of body fluids and ionic concentrations such that the extracellular and intracellular fluid compartments are maintained in osmotic balance. Failure in this respect would lead to this situation: the body's cells are effectively placed in a fluid that would cause them to swell or shrink to a point of dysfunction or death. Physiological solutions to the problem of water balance have tremendous evolutionary and ecological significance. First, it can be argued that the spectacular evolutionary radiation of insects occurred after a host of adaptations to terrestrial life. We saw some of these adaptations during our consideration of movement. Physiological regulation of water balance is another adaptation that supports terrestrial life. The biogeographical distribution of insects reflects the broad range of terrestrial habitats in which insects live and thrive. Many of these habitats are extremely arid and very hot. Physiological adaptations to these sorts of environments have permitted exploitation of such habitats that would otherwise be unavailable to insects. The focus on terrestrial life is not meant to overlook aquatic insects, which we will discuss in future. Furthermore, aquatic insects arose from their terrestrial counterparts. Hence, adaptations to aquatic life can be thought of as a component of successful terrestrialization. Two main factors may work to upset the essential osmotic equilibrium within an insect's body:

1) Loss of water by transpiration across the cuticle, or a very large gain of water by drinking or eating.

2) The intake of food whose concentration of inorganic salts is quite different from the ionic composition of the hemolymph.

Today we are going to begin with the first point, the regulation of water balance, which may be insulted by substantial loss or gain of water. Insects are generally small creatures, which tends to put them in an unfavorable setting with respect to the ratio of body surface area to body volume. The idea is to recognize a relatively large surface across which water can be lost by transpiration. We will see that water loss across the body surface can not be entirely stopped; however, if it were not restricted to a level that can be balanced by water uptake, insects would not survive in the more arid environments. The first line of defense was the development of a cuticle relatively impermeable to water.

The outer layer of an insect is the integument, which is made up of the epidermis and the cuticle. The epidermis is the outer cell layer. The epidermis lies on a basement membrante, and secretes the cuticle, which so far as I know is illustrated in virtually every book written on insect biology.

There are three major sections of cuticle, endocuticle, exocuticle and epicuticle. In standard drawings, we note the pore canal, cells of the epidermis and the basement membrane. The endocuticle provides the extensibility of the integument and combines toughness with flexibility. The rigidity of the hard parts of an insect, the appendages and so on, comes from the properties of the exocuticle. The relative impermeability of the cuticle to water is due to the presence of the epicuticle.

The epicuticle is a complex structure that differs among various insects. There is a cuticulin layer made of two components: a thin, dense outer layer and a thicker, still denser inner layer. Cuticulin is generally made of lipoprotein. A waxy layer lies atop the cuticulin. This is made of long-chain hydrocarbons, and esters of fatty acids and alcohols. There is an inner wax layer composed of strongly oriented long-chain molecules that generally form a monolayer only one molecule thick. The strong orientation provides the waterproof properties of the epicuticle. The outer wax layer is composed of wax molecules that are randomly orientated; on top of this is the cement layer. People think that this is a shellac-like substance that may serve to protect the wax.

Terrestrial insects lose water by evaporation from the general body surface - a process called transpiration. The rate of water loss from the surface on any given insect is a function of both wind speed and also of the humidity of air. Transpiration is relatively independent of body temperature, up to some point. Water is lost from the body surface at a constant, but rather low, rate through a broad range of temperatures. At some temperature the cuticle reaches a temperature point called the transition point. This point varies from insect to insect, depending upon the chemical composition of the hydrocarbons and waxes in the epicuticle. Insects living in hot, arid habitats tend to have wax layers with higher transition temperatures than do insects living in cooler or more humid habitats. Some insects have two transitions points.

Water loss from the body surface is held to a low rate by the oriented wax layer on the outside of the cuticulin. When the temperature of the cuticle rises to such a point that the van der Waals forces holding the wax molecules together are broken, the molecules move slightly apart and water is allowed to escape. Below the transition temperatures the wax molecules are close together, and at a slight angle so that water loss is restricted. At temperatures above the the transition point, the molecules part, which usually leads to a quick and probably pleasing and comfortable death due to dehydration.

Then we see that osmotic balance in the melieu int`erieur is in part maintained by reduction of water loss from the body surface. Another important avenue of water loss in terrestrial insects is from the respiratory surfaces. In the orthopteran Gastrimargus, for example, 70% of the normal water loss is from the trachael system. We have already seen that water loss from the respiratory surfaces is restricted by the invagination, or internalization, of the trachael system such that it is inside the body. Water loss is also reduced by control of the spiracular valves. Internalization of the respiratory surfaces in a major adaptation to terrestrial life in vertebrates as well as in invertebrates.

We can model an experiment designed to demonstrate the influence of spiracles on water loss. Consider the rate of water loss from Tenebrio larvae as a function of time. Tenebrio is well adapted to arid conditions, and its water loss rates are typically quite low. If we impose the condition of open spiracles, by holding the animals in an atmosphere of 5% carbon dioxide, water loss would be far higher than what could be easily balanced by an insect living in an arid habitat. Physiological regulation of the spiracles plays an important part in reducing potential water loss.

Some insects live through long periods of quiescence during which they do not feed or drink water. Control of the spiracles in these species is aimed at keeping them closed as much as possible to minimize water loss while facilitating the gas exchange necessary to support metabolic needs. One strategy is called cyclic release of carbon dioxide. This has been well studied in pupae of Hyalopohora cecropia, a large insect suitable for study in this regard. Carbon dioxide is allowed to build up in the trachael system for a long as seven hours, then is released in bursts during which the spiracles are wide open. Between the bursts of carbon dioxide release the spiracles are not closed as tight as possible, but rather flutter so as to allow a continuous uptake of oxygen.

While the cuticle and control of the spiracles serve to restrict the loss of water, a certain amount of water is inevitably lost. Then we can identify three avenues of water loss:

1) Transpiration across the cuticular surfaces

2) Transpiration across respiratory surfaces

3) Loss of water in the excreta

Water that is lost must be balanced by an appropriate level of water uptake. Most insects take in sufficient water with the food they eat. In his book on insect structures and function, Chapman thought some insects may select food with a higher water content under certain circumstances. Still other insect species may consume more food than they actually need in order to realize the potential water in the food. Moreover, many insects drink water. The dipteran Phormia, for example, has specific water receptors in the tarsal and labellar sensilla. I've often observed wasps drinking water in desert pools in Mexico.

Some insects have evolved slightly more bizarre adaptations to gain water. Larvae of the dipteran, Epistrophe, can extrude an anal pipilla into a water droplet and absorb water. The cockroach Periplaneta is among many insects that can absorb water from water droplets on the cuticle. The cuticle is asymmetrical with respect to water movement: water passes inwards more readily than outwards. Also, a few insects can absorb water from water vapor in the air when they are desiccated. Tenebrio, for example, can absorb water from air with relative humidities greater than 90%. Prepupa of the flea Xinopsylla can absorb water from air at relative humidity above 50%.

A final source of water is called metabolic water. As we saw during our look at metabolism, the final electron acceptor at the end of the electron transport chain is oxygen. Electrons and their hydrogens combine with oxygen at this point, yielding water. The amount of water that can be gained by metabolism depends on the metabolic substrate. A hundred grams of the fatty acid palmitate (16:0) yields 112 grams of water while 100 grams of glycogen yields 56 grams of water. Grain insects, such as Tribolium, normally obtain a very large proportion if not all of their water from metabolism of their food. We can guess that metabolism of lipid may be adaptive with respect to water balance during long periods of hibernation in several insect species.