insect physiology and evolution
mechanical problems of moulting
As the new cuticle cannot harden immediately, all moulting arthropods go through a phase in which their body is only supported by a soft hydrostatic skeleton. This may be particularly problematic for large terrestrial insects, potentially imposing an upper mechanical limit on the insect's body size. The soft cuticle not only leaves insects unprotected and vulnerable to predation, but also makes it more difficult for them to use their powerful muscles. taylor and kier - soft shell (1hr post ecdysis) vs hard shell crab (7 days post ecdysis) soft shell has large fluctuations in blood pressure associated with pinching. hard shell has no fluctuations associated see image
tracheolar supply of flight muscles
Because of the larger dimensions of tracheae and their relatively impermeable lining, virtually all oxygen is taken up through the tracheoles. In the flight muscles of dragonflies (which have fibres ca. 20 μm in diameter), oxygen can reach the mitochondria from outside the fibre. In larger flight muscles, such as those found in flies, the tracheoles form large numbers of indentations in the muscle membrane. This brings the oxygen supply closer to the mitochondria in the muscle so that the distance between a mitochondrion and the nearest tracheole is never more than 2-3μm.
chitin fibrils
Chitin: 1-4B polymer of amino-sugar monomers (N-acetyl-glucosamine). see image crystalline chitin nanofibrils 3nm diameter 19 molecular chains ca. 300nm long
rates of O2 consumption in flying insects
Flying insects can achieve some of the highest known mass-specific rates of O2 consumption in the animal kingdom. How do insects take up oxygen so efficiently and how do they vary respiration with their activity level? In contrast to vertebrates and many other arthropods (such as crustaceans and spiders), the insects' blood (haemolymph) and circulatory system are not essential for oxygen transport. Instead, oxygen is directly transported to the cells via a system of air-filled tubes, the tracheae and tracheoles. Tracheae form by invagination of the epidermis and therefore have a cuticular lining that is continuous with the rest of the cuticle mass specific rate of O2 consumption in flying honeybee =3700 Watts/kg elite cyclist = 30 watts/kg
respiration and body size
If oxygen is taken up only by diffusion, the length of the tracheal tubes (between the spiracles and respiring cells) must be small. Does this impose a limit to the overall body size of insects? There is evidence in both directions: ➢ There are other reasons why insects cannot get very large (see other lectures) ➢ Much larger insects existed during the Carboniferous period ✓ Insect gigantism coincided with higher atmospheric O2 levels during the Carboniferous ✓ Many insects (even flightless ones) grow larger tracheae when kept under low oxygen conditions ✓ Tracheae occupy a greater fraction of the body in larger insects. Tracheal bottlenecks (e.g. at leg orifices) could limit body size eg in beetles ➢ Insect flight is oxygen-sensitive, but some insects can fly even at extremely low oxygen concentrations (5% O2) ➢ Even the largest insects can fly ➢ O2 supply does not only depend on diffusion but can be strongly enhanced by ventilation (at least in larger insects) see image
ecological and evolutionary success of insects
Insects are the most abundant and diverse group of animals on earth. Together with crustaceans, spiders and millipedes, they belong to the Arthropods (jointed-limbed animals), which comprise more than 80% of all animal species. Arthropods are thought to have evolved from soft-bodied segmented worms similar to the extant velvet worms (Onychophora). Why have insects and Arthropods been so successful? see image can quantify success via abundance. estimated 10^8 insects on earth see image
moulting of cuticle
Insects grow by a series of moults from the egg to the adult. Although some insects can increase in size without moulting (by cuticle expansion), moulting is generally necessary to grow and replace tanned exocuticle and to renew the cuticle's water-proofing properties. Moulting also allows fundamental changes in anatomy (metamorphosis - allows occupying of different ecological niches) to occur. Based on the way that adult insects (and their wings) develop, three major groups can be distinguished: a) Apterygotes: wingless insects b) Exopterygotes: insects with young stages similar to the adult forms (which gradually develop wings). hemimetabolous c) Endopterygotes: insects with juvenile and adult forms that look very different and are separated by a pupal stage. holometabolous Moulting is triggered by hormones, most importantly 20-hydroxyecdysone (20-HE) and juvenile hormone (JH). This ensures appropriate timing of the moults, depending on the insect body size and state of nutrition (20-HE. initiates moult) and also determines the character of the moult (JH). Moulting of the insect cuticle includes a series of specific steps. First, the epidermal cells detach from the cuticle, followed by rapid cell divisions. Digestive enzymes (chitinases, proteases and lipases which are initially inactive) are then secreted into the gap. Only once the new epicuticle has been formed, the digestion of the old cuticle begins by activation of the enzymes. Most insects reabsorb the moulting fluid before breaking open the cuticle (ecdysis). see image ecdysial line = line of weakness
morphology of the exoskeleton
Insects possess external skeletons made of cuticle. A cuticular exoskeleton with jointed appendages is the distinctive feature of all arthropods. In comparison with the hydrostatic skeletons of ancestral forms, the arthropod exoskeleton conveys a number of advantages, including: • Mechanical stability • protection • makes very complex structures possible - bizarre shapes possible • allows new muscle attachment sites see image rigid cuticle meets soft cuticle -> joints. allows for locomotion systems.
material properties of the cuticle
Many biological functions of the cuticle depend on its material properties. Cuticle is an extremely versatile material; it can be as hard as steel and as soft as rubber, as resilient as a spring and as dampening as a shock absorber. The material properties of cuticle depend on its ultrastructure and chemical composition see image Insect cuticle has highly versatile material properties. A measure of its stiffness is the Young's modulus, defined as the ratio of stress (force per unit area) to strain (=extension per initial length).
discontinuous gas exchange
Many insects open and close their spiracles in a cyclic pattern. Spiracles may open only once in several minutes or hours, depending on how active the insect is (called 'discontinuous gasexchange cycles' or DGC). When the spiracles are closed, respiration reduces the O2 concentrations in the tracheae to a low level of ca. 5 kPa. As a result, the pressure in the tracheal system falls. This triggers the spiracles to "flutter", i.e. they open slightly, causing air to flow in rapidly. As a consequence, neither CO2 nor water is released from the tracheae. The intermittent fluttering of the spiracles goes on until the concentration of CO2 (despite being partly dissolved in the haemolymph) increases so much that the spiracles then open fully, allowing CO2 to be released.
Elastic cuticle - resilin
One specialised form of procuticle contains no chitin fibres but a special, rubbery protein called resilin. Like rubber, resilin has infrequent crosslinks allowing it to be stretched, thereby losing entropy and tending to return elastically to its unordered state. see image changes happen with little energy loss. doesnt produce a lot of heat. there is a tendency to return to the unordered state. unstrained = infrequent crosslinking by di- and trityrosins. cross linking -> UV autoflourescence so can identify them as blue patches under UV Like plasticised cuticle, resilin is highly extensible, but resilin is special in that it can return up to 97% of the elastic energy that is put into it by stretching or compressing. Resilin has a variety of special biological functions. It is used for rapid or repeated movements where heat and material fatigue must be avoided (e.g. in flight muscle tendon or wing folds), as a light-weight replacement for muscles, and as a power amplifier in jumping insects (in combination with stiffer cuticle).
structure of insect cuticle
The cuticle is an extracellular material, produced by a single layer of epidermal cells. It not only forms an outer shell, but also structures within the insect body (e.g. tendons, head endoskeleton and air-breathing tubes); this occurs through invagination of the epidermis.
why perform DGC?
The function of the DGC is controversial. The following explanations have been proposed: • DGC minimises water loss through the spiracles • DGC is an adaptation to hypoxic and hypercapnic conditions (for example in underground burrows) • DGC helps to maintain a low oxygen concentration in the tracheal system, thereby protecting the tissues against oxidative damage see image
layers of the cuticle
The layers of the cuticle include: • Procuticle (= Exocuticle + Endocuticle): a thick layer between the epicuticle and epidermis, made up of chitin microfibrils in a matrix of protein, water and lipids. • Epicuticle: •Wax (lipid) layer: this is secreted through the pore and wax canals and is essential for waterproofing (see lecture 3) •Cuticulin (envelope): a thin but dense layer (15-20nm). This is the first layer to be formed when a new cuticle is secreted. •Inner epicuticle: a thicker layer (0.1-10 μm) which does not contain chitin. Exocuticle: tanned and therefore often darkly coloured and hard Endocuticle: untanned and mainly secreted after the insect moults.
fibrous structure of procuticle
The procuticle is made of stiff fibres in a soft matrix. This has the advantage that small cracks cannot propagate, so the material is very fracture-resistant, a property found in many manmade fibrous composites such as plywood and fibre glass. The orientation of the cuticle fibrils determines the mechanical properties and is adapted for different functions (e.g. tendons, elytra, adhesive pads). Cuticle fibril orientation can probably be controlled by microvilli on the surface of the epidermal cells.
structure of the tracheal system
The tracheal system of insects is a complex network of air-filled tubes and includes: • Spiracles: openings to the air. These are formed in several pairs along the body and are controllable via opening and closing muscles. They protect the tracheal system and control respiration. act as doors to control access to tracheal system •Air sacs: collapsible structures within the breathing system. •Tracheae: 2μm-1mm diameter tubes. These are stabilized by spiral thickenings (taenidia) and have an impermeable cuticular lining. •Tracheoles: 0.1-1 μm intracellular permeable tubes. These directly supply tissues with oxygen. see image
mechanisms of respiration control
There are several mechanisms that insects use to increase and control their uptake of oxygen. These include: •Ventilation: enhancing O2 uptake by compressing or expanding parts of the tracheal system, thereby setting up a flow of air through the tracheae - Autoventilation of thorax during flight - Abdominal pumping: compresses abdominal air sacs see image Effect of atmospheric oxygen partial pressure (PO2) on the ventilation rate in grasshoppers (Schistocerca americana). Breathing frequency increases as oxygen pressure decreases. • Spiracle movements •Changing the tracheolar fluid level. As O2 diffuses much faster in air than in water, fluid movement into the endings of the tracheoles reduces the transport of O2 from the tracheoles into the tissues, whereas removal of fluid increases it. The fluid level is probably controlled by active ion pumping by the tracheolar cells. see image
plasticisation of cuticle
Untanned endocuticle can be stretched considerably, allowing body expansion after a moult. Stretching without moulting is also important in other contexts, such as in blood-sucking insects, queens of social insects and honeypot ants that store food for the colony. Before expansion, the endocuticle needs to be prepared for stretching. It is plasticised (softened) considerably by only slightly increasing its water content. This may be achieved by changing the intracuticular pH to reduce hydrogen bonding between proteins. water content increases -> cuticle swells hydration achieved by reducing the intracuticular pH, away from the isoelectric point of the proteins cuticle softens dramatically. honey pot ants - expand abdomen. blood feeding insects need to stretch very fast. 12x increase in body size in 10 mins in kissing bugs. unfed = not stretchable, indicating there needs to be preparation for stretching
respiration in aquatic insects
Water-living insects have a problem taking up enough O2 (getting rid of CO2 is easier, as it is more soluble in water, see table at the end of handout). Aquatic insects have developed a variety of solutions allowing them to acquire oxygen underwater: a) Breathing tubes Many fly larvae breathe air through long tubes with a pair of spiracles at the rear of the insect. The opening is usually surrounded by hydrophobic hairs and thus strongly water-repellent. b) Gills Very small insects can survive under water without any spiracles, by simple diffusion of oxygen across the cuticle into the closed tracheal system. Larger insects such as mayfly larvae have evolved gill-like structures on their abdomen with tracheal branches inside them. c) Air bubbles ('physical gill') Many diving insects take bubbles of air underwater or have films of air attached to their body. Backswimmers (Notonecta spp.) can survive with a bubble of air in cool water for several hours, longer than they could if the bubble consisted of pure oxygen. When the insect takes up O2, the partial pressure of O2 in the bubble decreases, while that of N2 increases. Therefore, O2 diffuses from the water into the air bubble, down this concentration gradient, providing the insects with additional oxygen. At the same time, however, N2 diffuses out of the bubble, so that the bubble shrinks. As N2 diffuses more slowly than O2, the bubble is stable for long enough to act as a "gill". You will investigate this mode of respiration in the practical. d) Plastron respiration A plastron is a permanent thin layer of air around the body that is stabilized by a dense cover of water-repellent hairs. As for air bubbles, N2 has a tendency to diffuse out, but the air layer cannot shrink much, because this would only be possible if water could enter the space beneath the hairs. Water can only penetrate these hydrophobic hairs under very high pressures. This is because the air/water interface would have to curve strongly between two adjacent hairs, requiring high pressures to overcome surface tension according to Laplace's law. The denser the hairs, the higher the required pressure. Therefore, the air layer is stable and can act as a gill down to considerable water depths see image
diffusive transport of oxygen - requirements
a) Diffusion of oxygen in air is 300,000 times faster than in water, because of a 30x higher O2 concentration and a 10,000x higher diffusion coefficient in air (see table at the end of this handout). This means that the diffusion of oxygen to the cells will be fastest if the transport pathway through water is as short as possible. see image Calculations have predicted that for the breathing system to cope with the high respiration rates of insect flight muscles, the distance between tracheoles and respiring cells should never be more than ca. 8-10 μm. b) If oxygen is taken up only via diffusion through air from the spiracles to the tissues, the length of the tracheal tubes should be limited
hardening of the cuticle
aka sclerotisation The hardening of cuticle involves the formation of bonds between the protein molecules or between the proteins and the chitin fibrils. Two alternative mechanisms for have been proposed for how this is achieved: phenolic tanning and dehydration. see image Phenolic tanning leads to protein-protein or protein-chitin cross-links, and it occurs by the action of a tanning agent (mainly N-acetyldopamine) that is released into the cuticle, and activated locally by enzymes (incorporated into the cuticle beforehand). The high stiffness of exocuticle may be based both on phenolic tanning and cuticle dehydration (possibly achieved by the secretion of N-acetyldopamine or pH changes). Cuticle parts exposed to high stresses and wear (such as the tips of claws or cutting edges of herbivore mandibles) often contain high concentrations of metal as a further reinforcement. 3d network of crosslinks and chitin. sclerotised usually brown colour -> melanin = poylmer formed. zinc = 16% of dry weight in cuticle of leaf cutting ant. manganese other common metal for reinforcement.
cuticle deposition zone
chitin microfibrils aligned with microvilli of the underlying epidermal cells see images
uses of resilin
flight muscle tendon wing folding cockroach tarsus -> lightweight construction, saving of material catapults in jumping insects -> used in combination with stiff chitinous cuticle, like a composite archery bow
moulting size dependent?
not just size dependent small insects with higher no of moults than some larger insects. wouldnt expect to see this if just moult for growth small stone fly - 20-30 moults to reach adult size huge atlas moth - 2-4 moults macrotermes queen - can grow abdomen to huge size without moulting
orientation of fibres in function
parallel arrangement - where pull/push in one direction eg cartilage helicoidal - more ability to move in different directions eg wings orthogonal - protective and fracture resistant eg hindlimbs of beetles
decline in insect numbers
worldwide declines in insects -habitat loss from agriculture and deforestation -chemical pollution (mostly from agriculture) -climate change
