The cuticle is an extracellular layer that covers the complete external surface of insects, as well as the surfaces of their foreguts and hindguts, and acts both as a skeleton for muscle attachment and as a protective barrier between the animal and its environment. The cuticle is an integral part of a complex dynamic tissue, the integument, which also includes the cuticle-producing epidermal cells, and various glands and sense organs.
GENERAL PROPERTIES OF CUTICLES
The cuticular layer varies in thickness from a few micrometers to a few millimeters, depending upon the insect species, developmental stage, and body region, but cuticles are typically between 100 and 300 urn thick. They are highly diverse in their mechanical properties, and they can be divided into two groups: stiff and hard cuticles, and soft and pliant cuticles. Intermediate degrees of stiffness also exist, and some types of cuticle have special properties, such as rubberlike elasticity or extreme extensibility. Cuticles differ in color and in surface sculpturing, but electron microscopy shows that all types are built according to a common plan. The details in structure and properties of the various cuticular regions are such that for each species they are optimal for the functioning of the living insect in its natural surroundings.
EPICUTICLE
The outermost layer of a cuticle is called epicuticle; it forms a continuous layer covering the complete cuticular surface. Seldom more than 2 urn thick, it is responsible for the waterproofing properties of the cuticle. Electron microscopy shows that the epicuticle can be subdivided into several layers, which have been differently named by different authors. The thin, outermost layer, 10-30 nm thick, is according to the nomenclature proposed by M. Locke called the cuticle envelope; it is the first part of the cuticle to be deposited; it is formed from plaques on the top of microvilli and it appears to be stabilized and made impermeable by an oxidative tanning process. Beneath the envelope the proper epicuticle is deposited by exocyto-sis from the epidermal cells, it is typically 2-4 urn thick and consists of proteins and lipids and is characterized by the absence of chitin. The combined envelope and epicuticle remain poorly characterized because they are difficult to purify, dissolve, and degrade.
The envelope is covered by a waterproofing wax layer, containing complex mixtures of extractable lipids, secreted during the molting process from integumental oenocytes and epidermal cells. This layer is again covered by a protective cement layer, secreted immediately after ecdysis from glands in the integument.
The extractable lipids in the wax layer have been characterized for several insect species. They appear to be species-specific mixtures of a wide range of lipids, including normal and branched, saturated and unsaturated hydrocarbons, fatty acids, alcohols, esters, sterols, and aldehydes. Differences in lipid composition have been used to discern closely related insect species. The epicuticular lipid composition can also vary between instars and sex of the same species, and the lipids often play an essential role in recognition and communication between insects.
PROCUTICLE
The region of the cuticle, located between epicuticle and the epidermal cell layer, is called procuticle; it constitutes the main part of the total cuticle. Histologically, the sclerotized regions (sclerites) are often subdivided into layers with different staining properties: (1) the outermost layer, the exocuticle, may be dark colored because of sclerotiza-tion, but is refractory to staining; (2) the innermost, uncolored layer, the endocuticle, stains blue; and (3) in between these two layers one often observes a layer of mesocuticle, staining red with Mallory triple stain. The flexible cuticle (arthrodial membranes), which connects the sclerites, stains blue with Mallory throughout most of its thickness. Exocuticle may correspond to the part of the procuticle deposited before ecdysis, stabilized by sclerotization. Mesocuticle plus endocuti-cle often correspond to the post-ecdysially deposited procuticle, and if these layers are sclerotized at all, it is only slightly.
The procuticle consists mainly of chitin and proteins; water is an essential component, and other materials, such as lipids, phenolic compounds, salts, pigments, and uric acid may be present. Chitin (poly 1,4-p-N-acetylglucosamine) is a polysaccharide, present as long and nearly straight microfibrils, usually about 2.8 nm in diameter and of indeterminate length. The filaments tend to run parallel to the cuticular surface, but columns of chitin filaments running perpendicular to the surface have been described for some types of cuticle (lepidopteran larval cuticle). The function of such chitinous columns remains uncertain.
The chitin microfibrils are organized in various patterns, and the organization seems to be important for the mechanical properties of the cuticle. The most commonly observed patterns are the heliocoidal pattern, where the microfibril direction changes by a small, constant angle between neighboring layers; the preferred, unidirectional orientation, where the fibrils run in the same direction in all layers; and the pseudo-orthogonal orientation, where unidirectional layers of chitin microfibrils alternate with layers running at nearly right angles to each other. In certain cuticles the pattern of chitin microfibrils depends on a daily rhythm: in locust tibiae, heliocoidal cuticle is deposited during the night and unidirectional cuticle is deposited during the day, making it possible to determine the number of days since ecdysis.
The chitin microfibrils are embedded in a protein matrix; the protein content tends to equal the chitin content in flexible cuticles and is usually three to four times higher than the chitin content in hard cuticles. The number of different proteins present in a given type of cuticle can vary from about 10 to more than 100. A database describing known arthropod cuticular proteins is available at http://www. bioinformatics.biol.uoa.gr/cuticleDB. Different types of protein are present in flexible and hard cuticles; the proteins are species specific, and some of them are also specific for certain cuticular regions. A characteristic amino acid sequence region, common to a large number of cuticular proteins, links these proteins to the chitin microfibrils. The proteins are often extractable immediately after deposition, but in many cuticular regions they are later rendered inextractable by scle-rotization, whereby low molecular weight phenolic compounds (N-acetyldopamine and N- (3-alanyldopamine) are oxidatively incorporated into the cuticular matrix, cross-linking the proteins, making the cuticle harder and stiffer and more difficult to digest with enzymes.
Sclerotization may occur soon after a molt when the insect has expanded its new cuticle to a larger size, but the regions that are not enlarged may have been sclerotized in the pharate stage, which is the stage that is present before emergence from the exuvium, or old cuticle. The elastic protein, resilin, present in rubberlike cuticu-lar regions, is cross-linked as soon as it is deposited extracellularly. The cross-linking process is different from that in sclerotized cuticle because no low molecular weight compounds are involved, but tyro-sine residues in the protein chains are oxidatively coupled to each other, forming di- and trityrosine residues.
SUBCUTICLE
A narrow, histochemically distinct layer, called subcuticle, is situated between the procuticle and the epidermal cells. It stains positively for muco- and glycoproteins. It has been suggested that it serves to bind cuticle and epidermis together and that this layer is the deposition zone, where new cuticular material is assembled and added to the already existing cuticle.
