1. During normal flight of the desert locust, auxiliary ventilating mechanisms do not appear, and dorso-ventral abdominal pumping continues at increased frequency and amplitude. When flight stops hyperventilation together with auxiliary forms appear briefly. Removal of the abdomen has shown that pterothoracic and neck ventilation are adequate for sustained flight.

  2. Spiracles 2 and 3 open wide during flight: when flight is weaker they make incipient closing movements. A central inhibitory reflex controls their activity, in addition to the peripheral action of carbon dioxide on spiracle 2. The incipient closing movements are shown not to have a functional significance; they are probably the expression of two competing mechanisms, and may arise by negative induction.

  3. Spiracles 1 and 4-10 remain synchronized with ventilation, and thereby permit adequate ventilation of the central nervous system.

  4. The isolation of the pterothoracic tracheal system is enhanced by the occlusion of two pairs of cross-links. The occlusion of a further three pairs in the prothorax and head ensures that the head has priority on the inspired air.

  5. The occlusion of all the cross-links takes place after the first instar, at which time spiracle synchronization first regularly appears and a directed airstream becomes possible.

  6. In flight there are two largely independent ventilating systems. The first, a two-way system, ventilates the flight muscles through the open spiracles 2 and 3 and is pumped by the flight movements. The second, a one-way system, ventilates primarily the central nervous system and is pumped by the abdomen, in through the dorsal orifice of spiracle 1, and out through spiracles 5-10.

The possibility that the thoracic tracheae might be ventilated by flight movements has been realized for a long time (du Buisson, 1924), and Krogh & Zeuthen (1941) have stated that ‘abdominal pumping movements and increased ventilation are insufficient during flight and serve mainly to raise the body temperature’, and again Krogh (1941) that ’the conclusion appears inevitable that the flight movements must themselves provide the necessary ventilation of the flight muscles’. Not until the work of Krogh & Weis-Fogh (1951), however, was thoracic ventilation more than an inference. These authors were able to show that the resting oxygen consumption of Schistocerca gregaria is about 0·63 l./kg./hr., and that during flight it rises to 15 l. The volume of air ventilated by the abdomen at rest is about 40 l./kg./hr. and with very vigorous pumping, induced by carbon dioxide and oxygen lack, a maximum of 300 l. is attained (Weis-Fogh, 1960). However, Weis-Fogh has shown that abdominal ventilation is not pumping more than 180 l. during the first 5 min. of flight and subsequently about 150 l. Consequently, while the oxygen consumption is increased twenty-four times, the volume pumped by the abdomen is no more than four or five times greater.

Thoracic movements during flight account for 600 l./kg./hr., of which 250 1. supply the large anterior abdominal air-sacs and 350 l. the pterothorax. Only the latter is available for pterothoracic respiration, but it is more than adequate (Weis-Fogh, 1953).

Ventilation induced by wing movements can be divided into two steps:

  • Ventilation of the intramuscular air-sacs by contraction of the flight muscles.

  • Ventilation of the large extramuscular air-sacs by alteration of the volume of the pterothorax due to vertical movements of the nota and lateral movements of the walls, with each wing-stroke (Weis-Fogh, 1953).

Weis-Fogh has inferred from his measurements of the changes in thoracic volume and pressure that the thoracic spiracles must remain open in flight, but Fraenkel (1932) reported that while in Libellula, Vespa and Tipula they opened immediately flight started and stayed open, in Schistocerca they continued to be synchronized with abdominal ventilation. In this paper a re-examination of the behaviour of the spiracles and ventilation during flight is described. It has shown that spiracles 2 and 3 are more or less wide open during flight, while the remainder continue to be synchronized with abdominal ventilation. Some suggestions are made as to how this uncoupling is achieved, and the significance of the continued synchronization of the remainder is discussed with reference to modifications of the thoracic tracheal system.

Material

Adult Schistocerca gregaria were used. Arrangements for keeping them have been described elsewhere (Miller, 1960 a).

Mature males are the best fliers. Even among these, however, it was necessary to conduct preliminary flight trials on a multiple stand, on which seven locusts could be flown simultaneously in front of the wind tunnel.

Methods

Standard conditions for flying tethered locusts have been described by Weis-Fogh (1956a) and these were followed closely, the wind tunnel used being that described in his paper. The suspension was a simple version of Weis-Fogh’s type 3, the locust being fixed by the sterna to a rigid post with a mixture of wax and resin.

24 hr. before flying, the metathoracic and certain other legs, according to which spiracles were to be observed, were removed under carbon-dioxide narcotization and the wounds sealed with wax.

Locusts were flown for up to 2 hr. in a constant wind speed of 3·5 m./sec., at a temperature of 28° C., and with a body angle of 10° to the horizontal. Wing-beat frequency and the flight performance were observed by means of a stroboscope (Stroboflash—Dawe Instrum. Co.).

Mirrors were fixed to the spiracles, and the movements of a reflected beam of light were photographed as described elsewhere (Miller, 1960b). Ventilation frequency was recorded manually with a signal marker on a revolving smoked drum ; ventilation amplitude was estimated with the help of a binocular and a scale fixed behind the abdomen.

Weis-Fogh (1956b) has briefly described a method of calculating the metabolic rate of a flying locust from measurements of the steady-state temperature within the thorax, the surface index of the locust (Weis-Fogh, 1952), and the wind speed. To measure the thoracic temperature in flight, a thermistor was mounted in a glass capillary, surrounded by narrow gauge polythene tubing and inserted between the crop and muscle 113. It was waxed in position several hours before flight.

To record impulses from the spiracle nerves during flight, the same electrodes and recording apparatus were used as are described elsewhere (Miller, 1960b). The locust was mounted on the flight stand, which was then turned through 90°so that the spiracle under investigation was uppermost. Impulses were recorded from the nerve under oil. It was not always possible to avoid recording muscle potentials as well, but they are clearly distinguishable on the trace, and give some indication of the wing-beat frequency. When recording from the transverse nerve to spiracle 3, it was necessary to remove most of the hind wing on that side.

During the initial 5 min. of flight the wing-beat frequency is high. After about 10 min. the locust settles down to a steady performance which may continue for several hours with little change. Considerable attention was paid to this initial period as well as to the subsequent steady flight.

Ventilation

For the first few seconds of flight abdominal ventilation ceases entirely. Subsequently, it is resumed at a high frequency and a slightly greater amplitude. None of the three auxiliary ventilating mechanisms (Miller, 1960a) appears during normal flight. If spiracles 1, 2 or 3 are blocked, however, longitudinal abdominal and neck ventilation may continue indefinitely in flight Blocking more than one pair causes vigorous hyperventilation and a rapid decline of the flight performance. Injection of 0·1 ml. 5% carbon dioxide into the mandibular air-sac causes hyperventilation and the momentary appearance of all auxiliary forms. Injection of larger amounts of 10% carbon dioxide causes excessive hyperventilation and flight soon ceases.

If flight is forcibly stopped during the first 5 min. considerable hyperventilation follows, often involving all three forms of auxiliary ventilation. After longer flights, the ensuing hyperventilation is less marked. These transient increases do not last for more than a few seconds, and must not be confused with the prolonged rise in oxygen consumption lasting for more than an hour after flight (Krogh & Weis-Fogh, 1951).

Locusts were flown for at least an hour after section of the nerve cord posterior to the metathoracic ganglion, and also after complete removal of the abdomen, the wound being sealed with wax. In neither case did the lack of abdominal ventilation appear to affect the flight performance, although neck and prothoracic ventilation continued during and after flight. Apparently abdominal ventilation is not essential for flight.

Spiracle 1

In order to attach a mirror to this spiracle, part of the overlying cuticle was removed and the prothorax rigidly waxed to the mesothorax. This did not reduce the flight performance or impede the spiracle movements.

Dining the initial few seconds of flight, when ventilation ceases, the valve remains closed but the opener contracts fully (Miller, 1960b). As soon as ventilation starts, the spiracle resumes normal synchronized movements, and the amount of opening is reduced after a minute (Fig. 1). When flight is stopped, the spiracle again opens wide during the first few inspirations, although this is less marked after longer flights.

Fig. 1.

Summary of the behaviour of the spiracles and of ventilation during flight. A, the start of flight. B, about 30 min. after the start of flight; incipient closing movements in spiracles 2 and 3. C, the end of flight. Cl, closed; O, open; insp., inspiration; exp., expiration.

Fig. 1.

Summary of the behaviour of the spiracles and of ventilation during flight. A, the start of flight. B, about 30 min. after the start of flight; incipient closing movements in spiracles 2 and 3. C, the end of flight. Cl, closed; O, open; insp., inspiration; exp., expiration.

Spiracle 2

Immediately flight starts, spiracle 2 opens wide and remains open without movement (Fig. 1). The wide-opening mechanism is fully effective. In most locusts incipient closing movements, in phase with abdominal expiration, appear after 5 or 10 min. They normally result in a 10–20% reduction in opening, although when flight is weak and the wing-beat frequency low, they may more than half close the valve. On the cessation of flight the valve immediately closes fully, and then resumes normal synchronized movements. Fig. 2 shows a tracing of a mirror recording from this spiracle.

Fig. 2.

Tracing from an actual mirror recording of spiracle 2 during flight. A, before flight. B, 5 sec. after the start of flight. C, 5 min. after the start of flight. D, 30 min. after the start of flight. E, 90 min. after the start of flight.

Fig. 2.

Tracing from an actual mirror recording of spiracle 2 during flight. A, before flight. B, 5 sec. after the start of flight. C, 5 min. after the start of flight. D, 30 min. after the start of flight. E, 90 min. after the start of flight.

Appropriate cuts in the cuticle close to the spiracle have been made to put the wide-opening mechanism out of action. During flight the spiracle remains 30-40% open and no movement occurs in the valve for the first few minutes. This shows that the closer muscle is not prevented from closing the spiracle by the wide-opening mechanism, but initially makes no contraction, and subsequently only very weak contractions. It has been confirmed by lightly pressing the cuticle close to the spiracle of a non-flying locust and so causing wide-opening; after a few seconds the muscle is able to close the spiracle completely against the mechanism.

Since the spiracle opens the instant flight starts and closes immediately it ceases, the reaction is unlikely to be mediated entirely by chemical stimulus. That it is not wholly a result of the action of carbon dioxide on the muscle membrane (Hoyle, 1960) has been shown by recording the impulses in the transverse nerve under oil during flight (Fig. 3). Initially the nerve is silent; later, as the incipient movements commence, bursts of impulses are recorded, but each burst comprises no more than about 6–10 impulses as compared with 50–80 at rest.

Fig. 3.

Oscilloscope records from the transverse nerves of spiracles 2 (A–E) and 3 (F and G) during flight. A and B, flight starts at the arrow; bursts of motor impulses to the spiracle closer cease. C and D, records from the transverse nerve when the spiracles are making incipient closing movements. E, flight is very poor and the valve is nearly full closing. F and G, flight starts at the arrow but is weak Opener impulses start at a high frequency, the spiracle opens wide, but irregular bursts of closer impulses continue. Time markers: 50 cyc./sec. (trace) and 1·0 sec. (dots).

Fig. 3.

Oscilloscope records from the transverse nerves of spiracles 2 (A–E) and 3 (F and G) during flight. A and B, flight starts at the arrow; bursts of motor impulses to the spiracle closer cease. C and D, records from the transverse nerve when the spiracles are making incipient closing movements. E, flight is very poor and the valve is nearly full closing. F and G, flight starts at the arrow but is weak Opener impulses start at a high frequency, the spiracle opens wide, but irregular bursts of closer impulses continue. Time markers: 50 cyc./sec. (trace) and 1·0 sec. (dots).

Hoyle (1959) describes an inhibitory reflex causing opening of spiracle 2, which is initiated by strong contractions in the abdomen. Since the behaviour of spiracle 2 was unchanged when the locust was flown after the complete removal of the abdomen, this cannot be responsible for spiracle opening in flight. It would appear that a central inhibitory mechanism initiated by flight is responsible for the maintained opening of the spiracle.

Spiracle 3

The behaviour of this spiracle in flight is very similar to that of spiracle 2 (Fig. 1). Wide-opening is followed by incipient closing movements which start slightly earlier than those in spiracle 2. At the end of flight the spiracle closes fully, and subsequently opens with inspiration no more than 10–20%. Oscilloscope recordings during flight (Fig. 3) show that wide-opening of the spiracle is accompanied by a high frequency (up to 200/sec.) of small (opener) impulses in the transverse nerve. When incipient closing starts, short bursts of larger impulses correspond to the weak contractions of the closer; as in spiracle 2 each burst may comprise no more than 6 impulses. They are superimposed on the opener impulses so that the closer operates against the opener—the latter acting like a rubber band.

This behaviour is again unlikely to be the result of chemical stimulation, and probably originates as a flight reflex in the central nervous system inhibiting closer and exciting opener impulses.

Spiracles 4–10

The behaviour of these spiracles is similar to that of spiracle 1. During flight they remain synchronized with abdominal ventilation.

The possible significance of incipient closing

Nearly simultaneous recordings were made of the wing-beat frequency and the amount of incipient closing of spiracles 2 and 3. Fig. 4, the results from several flights made by a male weighing 2·0 g., shows that there is a fairly close relation between the two, with spiracle 3 always slightly more closed than spiracle 2.

Fig. 4.

The relation between the amount of incipient closing of spiracles 2 and 3 in flight (i.e. the percentage opening during expiration) and the wing-beat frequency.

Fig. 4.

The relation between the amount of incipient closing of spiracles 2 and 3 in flight (i.e. the percentage opening during expiration) and the wing-beat frequency.

Wing-beat frequency is only one aspect of the total flight effort of the locust. If the incipient closing has any functional significance it would be expected to be more closely related to the metabolic rate.

From thoracic temperature measurements made in flight the metabolic rate has been calculated (Weis-Fogh, 1956b) and correlated with the amount of incipient closing. Readings were commenced 10 min. after the start of flight and were made only when flight was steady and when no change in the wing-beat frequency had occurred for at least 2 min. The results (Fig. 5) show that the amount of incipient closing is more nearly related to the wing-beat frequency than to the metabolic rate. Weis-Fogh (1956a) mentions that there is a close relationship between lift and power output, but not between wing-beat frequency and power output. Since the size of the incipient closing movements appears to be more nearly elated to the wing-beat frequency, it presumably has little functional significance.

Fig. 5.

The relation between the amount of incipient closing of spiracles 2 and 3 in flight and the excess temperature of the thorax over that of the air. (This is directly related to the metabolic rate—see text.)

Fig. 5.

The relation between the amount of incipient closing of spiracles 2 and 3 in flight and the excess temperature of the thorax over that of the air. (This is directly related to the metabolic rate—see text.)

This conclusion has been supported by the results of measuring the air flow through spiracle 2. The spiracle was cut out, waxed to the end of a long narrow glass tube, and air was driven through under a constant pressure of 3 cm. water. The time for the meniscus to pass between two fixed points was measured. The valves were set in various positions with wax and their maximum separation noted. This distance was then plotted against the air flow in ml./min. (Fig. 6). Point 1 on the graph indicates the separation of the valves due to muscle relaxation alone, and point 2 that due to the wide-opening mechanism. While the separation of the valves increases with wide-opening by 70%, the air flow increases by only 27%. In subsequent experiments in which the pressure was increased to 20 cm. water, wide-opening doubled the air flow. The pressure changes synchronous with wing movements do not exceed 1–3 cm., while those resulting from abdominal ventilation are as much as 20 cm. water (Weis-Fogh, 1960). Most of the incipient movements take place between 80 and 100% open where they can have hardly any effect on the air flow. On the other hand, very slight adjustment of the position of the valves, when they are almost closed, may be of great importance.

Fig. 6.

The air flow through spiracle 2 at a pressure of 3 cm. water, with the valves set in different positions. Point 1, maximum opening at rest. Point 2, maximum opening in flight

Fig. 6.

The air flow through spiracle 2 at a pressure of 3 cm. water, with the valves set in different positions. Point 1, maximum opening at rest. Point 2, maximum opening in flight

Modifications of the tracheal system and the continued synchronization of spiracle 1 during flight

The flight tracheal system

Weis-Fogh (1960) has pointed out the considerable isolation of the tracheae which supply the flight muscles from the remainder. This pterothoracic tracheal system comprises on each side (Fig. 7) the supra-ventral trunk running posteriorly from the ventral orifice of spiracle 1, with branches to the second and third legs, to the flight muscles and to the plexus of spiracle 2 ; a dorsally running trachea from the ventral orifice of spiracle 1, supplying the flight muscles and the main thoracic air-sacs; and finally tracheae from spiracles 2 and 3 to the flight muscles and the air-sacs.

Fig. 7.

Diagram of the pterothoracic tracheal system and its junctions with the other main tracheae. GI, GII and GIII, pro-, meso-and metathoracic ganglia; LI, LII and LIII, pro-, meso-and metathoracic legs; SP, spiracle; CL 4 and 5, degenerate cross-linking tracheae.

Fig. 7.

Diagram of the pterothoracic tracheal system and its junctions with the other main tracheae. GI, GII and GIII, pro-, meso-and metathoracic ganglia; LI, LII and LIII, pro-, meso-and metathoracic legs; SP, spiracle; CL 4 and 5, degenerate cross-linking tracheae.

The flight tracheal system joins the tracheal system of the remainder of the insect at the following points (Fig. 7).

  • Branches from the anterior end of the main thoracic air-sacs which run into the dorsal cephalic tracheae. They are small ; the ratio of their cross-sectional area to that of the cephalic tracheae is 1:14.

  • At spiracle 1 across the atrium from the ventral to the dorsal orifice. This route is only effective when the spiracle is closed, and then only when the opener is relaxed and the ventral orifice open (Miller, 1960b).

  • Across the ventral plexus of spiracle 1 into the leg trachea, and thence to the dorsal cephalic trachea via a small loop, the ratio of whose cross-sectional area to that of the dorsal cephalic trachea is 1:20.

  • From the plexus of spiracle 3 through a long thin air-sac to the longitudinal ventral trunk.

  • From the plexus of spiracle 3 through branches which run posteriorly and anastomose with the alimentary tracheae.

Routes (iv) and (v) will be effective only when the spiracle is closed.

The trachea from the large anterior abdominal air-sac runs down close to spiracle 4, where it bends sharply to the posterior and then joins the longitudinal ventral trunk. At the bend the occluded remains of a branch into the tracheal plexus can be seen. In the more posterior spiracles the corresponding branch is normal. There is no connexion between the anterior abdominal air-sacs and the main thoracic air-sacs in spite of their contiguity. The former may have little respiratory significance and be more important in reducing the mechanical damping of the wing muscles (Weis-Fogh, 1953).

Two pairs of non-functional cross-linking tracheae run between the supraventral trunks and the longitudinal ventral trunks (Fig. 7). In the more anterior pair (CL 4), situated just in front of the mesothoracic ganglion, each trachea is flattened and, near its junction with the ventral trunk, filled with liquid. The ratio of its cross-sectional area to that of the ventral trunk is 1:47. The posterior pair (CL 5), level with the metathoracic ganglion, comprises longer tracheae entirely flattened and usually liquid-filled near the ventral trunk. The corresponding ratio is 1:25. In Locusta migratoria the equivalent cross-links are thinner and entirely liquid-filled.

In the first instar Schistocerca both pairs of cross-links are relatively larger (the corresponding ratios are for the anterior 1:2, and for the posterior 1:3) and appear as normally functional tracheae. In the second instar they are flattened, and by the third they become liquid-filled.

A number of tests on the adult has demonstrated that these links never conduct air. Adults of all ages and both sexes have been inspected after long flights. The links have been observed during hyperventilation through windows glued to the sterna. Air has been blown into spiracles 1 and 2 under pressures of up to 20 cm. water. During some of these tests the flattened portions of the links were inflated, but the liquid was never expelled.

The large trachea from spiracle 4 supplies almost entirely the metathoracic leg. Since spiracles 2 and 3 supply principally the pterothoracic system, the main source of inspired air for the central nervous system and the rest of the locust is the dorsal orifice of spiracle 1.

Examination of the cephalic tracheal system, which is supplied by the dorsal orifice, has revealed three further pairs of degenerate cross-links (Fig. 8), the degeneration occurring in each case after the first instar. They all run between the ventral cephalic trachea and the longitudinal ventral trunk. The third (CL3) is near the front legs and comprises on each side a broad flattened trachea, liquid-filled at its dorsal end. The second (CL2) is a long, collapsed and partially liquid-filled air-sac close to the third, and the first (CL1), inside the head, is a short trachea entirely liquid-filled. Tests, similar to those already described, suggest that none of these conducts air in the adult.

Fig. 8.

Diagram of the main tracheae to the head and their relation to the dorsal orifice of spiracle 1. The arrows indicate the probable direction of the movement of air resulting from abdominal ventilation. LI, prothoracic leg base; CL 1, 2 and 3, degenerate cross-linking tracheae; SP1, spiracle 1.

Fig. 8.

Diagram of the main tracheae to the head and their relation to the dorsal orifice of spiracle 1. The arrows indicate the probable direction of the movement of air resulting from abdominal ventilation. LI, prothoracic leg base; CL 1, 2 and 3, degenerate cross-linking tracheae; SP1, spiracle 1.

Their occlusion means that air can pass from the cephalic tracheae to the longitudinal trunks only through anastomosing air-sacs and tracheal loops close to the cephalic ganglia. Thus inspired air from the dorsal orifice of spiracle 1 goes straight to the brain and then down the ventral trunks to the thoracic ganglia. The continued synchronization of spiracle 1 during flight will ensure the maintenance of this stream of air, thereby adequately ventilating the central nervous system.

Only after the first instar do regular synchronized movements of the spiracles commence, and a directed stream of air through the insect then becomes possible. Since all five pairs of cross-links become non-functional after the first instar, their occlusion is probably associated with this air-stream. Tracheae which have apparently outlived their usefulness are notorious for their persistence (Hamilton, 1931 ; Kramer, 1937; Smith, 1958). It is probable, therefore, that the occlusion of these cross-links enhances the air flow through the insect, directing it close to the cephalic and thoracic ganglia.

It has been shown that the opening of spiracle 2 with flight is controlled by a reflex mechanism. It was argued (Miller, 1960b) that the peripheral action of carbon dioxide on the muscle membrane is much more effective when the frequency of motor impulses is low; it seems most probable that the small number of motor impulses which reaches the muscle during incipient closing fails to cause more than a very weak contraction because of the presence of 5 % carbon dioxide near the spiracle. The full closing immediately after flight is due to a greatly increased frequency of motor impulses. In this way the ganglion is effectively altering the sensitivity of the local reaction of the spiracle to carbon dioxide.

To summarize, spiracle 2 stays open for the first 5-10 min. of flight because it receives no motor impulses: subsequently it makes only very weak closing movements due to the combination of a small number of motor impulses and the peripheral action of carbon dioxide.

Locust flight muscle is non-fibrillar and the ratio of nerve impulses to muscle contractions is 1:1. This means that the size of the incipient closing movement is related to the frequency of flight motor impulses originating from the meso-and metathoracic ganglia. There would appear to be competition between an hypothetical flight centre, which inhibits impulses in the motor nerves to the spiracle closers, and a ventilation centre which rhythmically excites them. The higher the wing-beat frequency, the greater the frequency of impulses in the nerves to the flight muscles and the more complete the inhibition of the closer neurones. The rhythmical centre of ventilation is in the metathoracic ganglion, and a spiracleinhibiting centre exists in the same ganglion, at least in the dragonfly (Miller, unpublished). This behaviour suggests the concept of negative induction (Pavlov, 1927), which may be described as ‘a concentrated state of excitement which produces an inhibition around itself’ (Konorski, 1948).

During the flight of the locust the ‘concentrated state of excitement’ of the flight motor neurones may be adequate to induce a field of inhibition around themselves, and possibly internuncial fibres, which supply the closer motor neurones of spiracles 2 and 3, have synapses within this field. More simply, the flight centre may supply inhibitory nerves to these synapses: such a scheme is represented in Fig. 9.

Fig. 9.

Scheme to account for the inhibition by negative induction of the closer motor neurones of spiracles 2 and 3 in flight. F, flight centre; V, ventilation centre; —, inhibitory nerves; +, excitatory nerves ; other abbreviations as in Fig. 7.

Fig. 9.

Scheme to account for the inhibition by negative induction of the closer motor neurones of spiracles 2 and 3 in flight. F, flight centre; V, ventilation centre; —, inhibitory nerves; +, excitatory nerves ; other abbreviations as in Fig. 7.

Several possible advantages might arise from keeping the pterothoracic tracheal system isolated. Volume changes induced by flight movements might be able to ventilate the pterothorax more efficiently through the open spiracles 2 and 3, since the system forms an isolated unit. High carbon-dioxide tension and oxygen lack might to some extent be limited to the pterothorax, while the abdomen continues to ventilate the central nervous system through spiracle 1, and does not draw ‘used air’ into the longitudinal trunks. The ability of carbon dioxide to diffuse through animal tissues faster than oxygen (Krogh, 1919) probably means that pterothoracic isolation is more important in preventing the withdrawal of oxygen from other tissues (in particular the central nervous system), than in limiting high concentrations of carbon dioxide to the pterothorax.

That oxygen lack, and possibly carbon dioxide accumulation, are partially limited to the pterothorax is suggested by a number of observations. Auxiliary ventilating mechanisms seldom function in flight ; nevertheless, when a resting locust is placed in an atmosphere of 5 % carbon dioxide, they continue for at least an hour without diminution. 5% carbon dioxide and 15% oxygen are concentrations commonly occurring in the main thoracic air-sacs in flight (Weis-Fogh, 1960), so that either the ventilatory centres are depressed by flight or they remain unstimulated. The results of injecting 5 % carbon dioxide into the mandibular air-sac of a flying locust, giving rise to considerable hyperventilation, show that the ventilatory regulation centres are not depressed but remain unstimulated. The considerable increase in ventilation and the wider opening of spiracle 1, which occur momentarily after flight, may be explained by the closing of spiracles 2 and 3 and the flooding of the ganglionic regulatory centres with pterothoracic gases which are driven anteriorly by abdominal expiration.

At the beginning of flight, and subsequently if there is a sudden increase in wing-beat frequency, spiracle 1 opens fully with inspiration as a result of a strong and maintained contraction of the opener. When the spiracle is closed, this contraction constricts the ventral orifice completely, but when the closer relaxes the orifice opens a limited amount (Miller, 1960b). The constriction of the ventral orifice prevents gases in the pterothorax from being blown by abdominal expiration across the atrium, into the cephalic tracheae and so to the central nervous system. Opener contractions are initiated by carbon dioxide in the head and gases will still be able to reach the head from the thoracic air-sacs through the cephalic tracheae (Fig. 8). However, the sudden initial contraction of the opener at the start of flight suggests it may then be controlled by a nervous reflex, independent of chemical stimulation.

Hoyle (1959) has drawn attention to the rich tracheation of the closer muscle of spiracle 2. The small tracheae to the muscle arise from an air-sac which is itself in direct communication with flight muscle air-sacs. Moreover, a ‘through trachea’, arising from this air-sac, passes across the muscle sending small twigs into it, and then joins a larger trachea which leads straight into the spiracle plexus. This may enable the muscle to sample gases on their way to the exterior during flight, and seems to refute the suggestion that to have a carbon-dioxide sensing mechanism in the spiracle is to have it ‘in the worst possible situation, that is, almost outside the insect’ (Case, 1957). ‘Through tracheae’ have been observed in association with the dragonfly spiracles, which appear to be controlled by a comparable local reaction to carbon dioxide (Miller, unpublished).

My thanks are due to Prof. V. B. Wigglesworth for much encouragement and supervision of this work. I am most grateful to Prof. T. Weis-Fogh for help and advice, for the generous loan of apparatus and for permission to quote his un-published results. I would like to thank Prof. Wigglesworth, Prof. Weis-Fogh, Mr F. Darwin and Mr J. S. Edwards for reading and criticizing the manuscript, and the Agricultural Research Council for financial support.

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