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The influence of juvenile hormone on the oxygen consumption of the last larval instar of Pyrrhocoris apterus L.
J. Ins. Physiol., 1962, Vol. 8, pp. 145 to 153. Pergamon Press Ltd. Printed in Great Britain
THE INFLUENCE OF JUVENILE HORMONE ON THE OXYGEN CONSUMPTION OF THE LAST LARVAL INSTAROF PYRRHOCORISAPTERUS L. V. J. A. NOVAK Entomological
Institute,
and K. SLhMA
Czechoslovac
Academy
of Sciences,
Praha
(Received 7 October 1961) Abstract-Oxygen consumption during the last larval instar of Pyrrhocoris upterus has been studied under the influence of implantations of active corpora allata. The degree of morphological change effected by tbe juvenile hormone was determined in each specimen after the larval-adult moult. The degree of the induced morphological change is directly proportional to the rate of oxygen consumption. The oxygen consumption curve was practically identical with that of controls until the middle of the intermoult period. The oxygen consumption in the second half of the instar, in which metamorphosis occurs, was less reduced and the period of depression was shorter in specimens that moulted into sixth instar larvae instead of into adults. The intermediates with adultoid characters were intermediate also in this respect. These results are consistent with those of previous work by the authors, accordiig to which the JR-effect on oxygen consumption is an indirect one depending upon increasing the amount of metabolically active tissue.
INTRODUCTION
THE effects of corpus allatum implantation
on the metabolism of insects have been described by many writers since the discovery of the juvenile hormone by WIGGLESWORTHin 1935. With one exception (PFLUGFELDER,1958) they agree that the oxygen consumption of the insect is increased. No agreement has been reached, however, as to whether the observed increase depends upon ‘a direct influence on metabolism, or whether it is an indirect consequence of the increased amount of growing tissue due to the activation of some parts of the body. The first hypothesis has been made by THOMSEN(1949) on the basis of the effects of allatectomy and corpus allatum implantation on oxygen consumption in Calhphora eryth~ocepha~a adults. The other view has been expressed by PFLUGFELDER(1952) for the first time as a result of his work on Dixzppus morosus, but he later (1958) abandoned his earlier view. In a previous paper (NOVAK et al., 1959) the authors attempted another approach to the problem. Oxygen consumption was measured in starving, normal, and castrated Pyrrhocmis female adults after corpus allatum implantation. Specimens in which a piece of muscle of corresponding size was implanted were used as controls. The results revealed that the implantation of active corpora allata increased the oxygen consumption in females with functioning ovaries but did not do so in castrated females (Fig. 1). The authors concluded that the observed increase was due to the activation by the hormone of ovarian follicle cells. From this 145
V. J. A. NOVLKANDK. SLLMA
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it follows that the juvenile hormone has only an indirect effect on the total metabolism, depending on the stimulation of metabolic processes in particular
tissues.
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Days FIG. 1. Oxygen consumption of starved normal (i) and castrated (ii) Pyrrhocoris aptems
female adults following the implantation of active corpus allatum. Solid line: specimens implanted with corpus allatum. Broken line: control specimens implanted with piece of muscle. Arrows indicate the moment of implantation.
Doubts have been thrown upon these results by S~~GESSER(1960), who claims to have found an increase in 0, consumption in Leucophczm after corpus allatum implantation even in castrated females. He agrees, as do THOMSEN and HAMBURGER (1955), that the juvenile hormone directly affects the total metabolism. S&GESSER explains the results of NOVAK et al. (1959) as an effect of starvation. He believes that in starving animals there is no supply of fat reserves necessary for ovarian maturation, so that negative results are not the result of castration but rather of
THE INFLUENCE
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starvation. For reasons mentioned in the discussion, however, this explanation is not adequate. In the present paper a series of experiments were carried out to analyse the influence of corpus allatum implantation on the last larval instar of Pyrrhoco~is apterus, in which the juvenile hormone causes a strong morphogenetical effect. MATERIALS
AND METHODS
All the experiments
were performed on the fifth (i.e. the last) larval instar of Pyrrhoco~i.s apterus L. (Heteroptera, Pyrrhocoridae), Specimens were kept at a constant temperature of 25°C and 16 hr daylight. They were fed on lime-tree seeds and sufficient water was added. They were narcotized by plunging them into water for 10-15 min before the operation. Active corpora allata used for implantation were taken from mating adult females at the beginning of oviposition. Such glands are larger and at least as active in morphogenetical effect as those from young larvae (NOVAK, 1951). Implantations were made into the side of the third abdominal sternite the first day after moulting. Control specimens were operated in the same way except that instead of a gland they received a piece of muscle of similar size. For further detail see Nov& et al. (1959). The method of determining oxygen consumption in Pyrrhocwis apterus has already been described (NOVAKet aZ., 1959; SLAMA, 1960). Warburg manometers have been used. The total volume of the flasks was about 10 ml. The figures for oxygen consumption are based upon averages derived from about ten individuals, the oxygen consumption of each of which represents an average of three or more 1 hr readings on the same day. The total oxygen consumption of forty-nine specimens implanted with c. allatum and forty-one controls were measured daily from the beginning to the end of the last larval instar. RESULTS
The course of oxygen consumption in Py~~hocoris during the intermoult period is characterized by a rise of 0, consumption in the beginning of each instar and by its subsequent decrease in the second half of the instar. A new rise in consumption begins immediately before the next ecdysis. The course of the 0, consumption in the last larval instar is similar, but the depression in the second half of the instar is distinctly deeper (SL&EA, 1960). The 0, consumption of the fifth instar larvae implanted with a piece of muscle tissue used as a control is on the average identical with that of the normal specimens (cf. SUMA (1960) with Fig. 2). The very small decrease in oxygen consumption following the operation shows that narcotizing and implantation of themselves have no significant influence on oxygen consumption. The quantitatively different effects of corpus allatum implantation have shown themselves not only by morphogenetical changes but also by differences in the length of the intermoult period. The most affected specimens which moulted into giant sixth instar larvae had the shortest intermoult period (on an average 25 per
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FIG. 2. Weight and oxygen consumption of Pyrrhocoris aptwus f&lx instar larvae moulted on sixth, seventh, and implanted with active corpus allatum. A, B, C-specimens Each point represents an average of thirteen individuals eighth days respectively, D-controls. in group (a), eleven individuals in group (b), ten in group (c), and fourteen in group (d). Arrows indicate the moment of implantation. For further explanation see text.
cent shorter), whereas the adultoid larvae moulted later. All the specimens str, -d therefore fall into the following four groups according to the amount of morphulogical change and the length of the instar :
THE INFLUENCE
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_rlost affected specimens, the intermoult period of which was reduced to 6 days and which moulted into complete sixth instar larvae; (b) specimens moulting into slightly adultoid sixth instar larvae the seventh day after the preceding ecdysis; (c) adultoid larvae moulting on the eighth day; and (d) control specimens implanted with a piece of muscle moulting into normal adults on the eighth day. The weight of the body increases steadily from the beginning of the instar. However, its growth stops 1 or 2 days before the next ecdysis. The specimens of all three groups moulting into sixth instar larvae reached nearly the same average weight (45-48 mg) at the end of the instar. The controls were about 15 per cent heavier (average, 54 mg). The last larval instar in Pynhocoti may be subdivided into two parts on the basis of the growth of the body and the oxygen consumption. In the first of these, at the beginning of the instar, the weight of the body increases steadily and the oxygen consumption per individual increases. In the second part, the growth of the body slows down and the oxygen consumption decreases in cormexion with the intensive morphogenetical processes. The interdependence between growth and an increase in 0, consumption is striking, as previously noted (SW, 1960). A close resemblance between the changes in body weight and 0, consumption is evident from Fig. 2, where both curves rise similarly. In the affected specimens, as in the controls, no additional increase in oxygen consumption has been observed in the first part of the instar: the Os-consumption curves follow the curve of the body weight. It means that the presence of the juvenile hormone, does not produce any change in metabolism during this period. As might be supposed, c. allatum implantation influences the metabolism in the second part of the instar where the bulk of metamorphosis occurs. In the controls a steep fall of 0, consumption occurs in this period. This decrease occurs independently from the increase in the weight of body, being most probably the consequence of the loss of metabolic activity in the larval parts of the body which are partly replaced by the imaginal parts during the metamorphosis. This depression of the U-shaped C&-consumption curve does not reach a similar depth in the holometabolous pupa. The amount of tissue undergoing metamorphosis in Pyrrhocoris is smaller, as a larger propbrtion of tissue survives to the adult stage in the Heterometabola. The corpus allatum implantation results, as has been generally agreed, in activation of the larval tissues which therefore continue their growth at the same rate as the imaginal ones, so that the growth of the body as a whole is an isometric one and no metamorphosis occurs (Nov.&, 1951, 1960). It might thus be expected that the fall in oxygen consumption in these specimens will be less intensive than in the normal metamorphosis controls; and this is what is actually observed, as shown in Fig. 2. Thus the more an individual is affected by a corpus allatum implantation, the less the 0, consumption falls in the second part of the instar. Hence the 0, consumption of the most affected specimens in group (a) falls to 660 $/g/hr, and in the less affected groups (b) and (c) to 650 and 620 ,ul/g/hr respectively, whereas IO
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in the control group it falls to a 560 $/g/hr. A similar decrease of the O,-consumption curve as in group (a) occurs also in the preceding larval instars when their own c. allatum functions (SLA~VIA,1960). Similarly, the observed shortening of the intermoult period in the most affected animals may be explained as due to the reduction of the second part of the instar in which the fall occurs. Thus the depression of the O,-consumption curve in group (a) occurs in the last day before moulting, whereas it takes 2 days in (b) and (c) and 3 days in the controls. The period of the increase in 0, consumption is thus equally long, taking 4-5 days in all groups. The terminal increase in 0, consumption immediately before ure end ot the in-star may be explained by the influence of ecdysis, as suggested by SW (1960). No decline in 0, consumption has been observed in comparison with that of the controls in specimens in which there was only a local effect of the implanted gland. The results of the experiments show clearly that the observed positive effect of the juvenile hormone on 0, consumption is not a direct and general one; it is just a consequence of the increase in total metabolism due to the share of the larval parts of the body activated by the hormone, which would have otherwise lost their activity. If the effect of the implanted gland were a direct one, it would have affected the Q-consumption curve during the first part of the instar in the same way. DISCUSSION The results of this paper thus agree fully with those of NOVAK et aZ. (1959) on the female adult of Pywhacoris. As shown in Fig. 1, implantation of c. allata induces a distinct rise of 0, consumption in normal starving females in comparison with controls implanted with a piece of muscle of similar size. This agrees with JONAN~~ON(1958), who found that corpus allatum implantation induces egg-laying even in starving (i.e. physiologically allatectomized) females of Oncopeltusfmciatus. Implantation of active c. allata into females which have been castrated at the beginning of the last larval instar does not affect metabolism. From this it may be concluded that the rise in 0, consumption in normal females following c. allatum implantation is due to the activation of the ovarian follicle cells. According to S~~GESSER (1960), the lack of juvenile hormone effect in castrated females in the experiments of NOVAK et aZ. (1959) may be due to the lack of available fat reserves in starving specimens. This is, however, scarcely likely as the same treatment in the similarly starved normal females retaining their ovaries has been effective. In addition, the fat reserves in the castrated specimens are on the average higher than in the normal ones, where the reserves are utilized by the growing ovaries. In our view the results of SAGESSERon Leucophaeu may be interpreted in the following way. %GESSER implanted nine castrated females of Leucophau (five of them deprived of both ovaries and accessory glands, the others of ovaries only) with active corpora allata. The 0, consumption of the experimental animals was about one-third higher than that of the controls with ovaries implanted with active
THEINFLUFNCEOFJWENILEHORh3ONE
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c. allata. Even if we take as significantthe difference found (O-1172 0.02 ml O,/g/hr) within the individual variability of normal specimens (not less than O-3ml O,/g/hr, according to Fig. 2 of this paper), the following note by the author makes more probable a different explanation: ‘Es ist zu beachten, dass fiir die Implantation aktiver c. allata immer das Tier benutzt wurde, das vor der Operation den geringsten Verbrauch angewiesen hatte’ (UGESSER, 1960, p. 277). The results show that the bulk of the individual difference in 0, consumption per unit weight depends upon the stage of digestion of the animals. Those which have accepted the food immediately before the measurement have a higher body weight and lower 0, consumption per g per hr due to the stage of consumption of the food from the last meal. .Thus their respiration in relation to body weight is lower. On the other hand, those specimens which are at the peak of digestion of the previous meal have a low body weight, their 0, consumption per unit of weight being relatively high. However, the stage of digestion changes and may be quite different from one measurement of 0, consumption to another. There is another reason to prefer our explanation The specimens ‘with inactive corpora allata’ used as controls received the inactive corpora allata from females with oiithecae. However, the 5 days following the implantation spent in the new ‘active’ medium (females which had deposited their oiithecae 4 days before the operation were used) had necessarily to bring the implanted gland to the stage of full activation at the time of the measurement. These are therefore the true experimental animals comparable with ‘Ringer controls’. Our interpretation is also supported by the observation of SAGESSER that there is no qualitative difference in the Oa-consumption curve between the one but last and last larval instar of Leucophaea ,m.aderae, so long as we do not refuse the conception of only one corpus allatum hormone. As a matter of fact, there is no experimental evidence that could be better explained by the supposition of two hormones, as postulated by SBGESSER: one of them the morphogenetically active juvenile hormone and the other with metabolic activity said to be identical with the supposed adult ovarian follicle activating hormone. The fallacy of SAGESSER’S conclusion that the larval corpus allatumwould differ in its influence on metabolism from that of the adult female may be clearly shown by measuring the 0, consumption of specimens implanted with larval corpora allata if the concentration level of the hormone produced is taken into account. Similarly all other evidence in favour of two corpora allata hormones by other authors (L&cm and SPRINGBTTI, 1960.; ENGEL-, 1957; SCHNEIDERMAN, 1961) is inconclusive and may be more simply explained by the supposition of one corpus allatum hormone, as shown by NOVAK(1960). Th is is not the case with the extract of cecropia male abdomens described by WILLIAMS(1956) and SCHNEIDERMAN and GILBERT(1959), which seems to be a composite mixture of lipoid materials which can cause a regressive metathetely with progressive prothetely effects, and is probably not identical with the juvenile hormone of the corpus allatum (SLAMA, 1961)., The same is true of the findings by DE WILDE (1959) on corpus allatum and cecropia extract effects in Leptinotarsa akxmlineata and the homogenates of its tissues.
V. J. A. NoviiK ANDK. S&
152
All other evidence by earlier authors (PFEIFFER-WEED, 1945; THOMSEN, 1949; PFLUGFELDER, 1952, 1958; L’HBLIAs, 1954), as shown in Nov.& et al. (1959), does not affect the question of whether the observed juvenile hormone effect on the 0, consumption is a direct or indirect one; and it may be completely explained as here suggested. It may be thus concluded that there is only one corpus allatum hormone responsible for all the observed effects of corpus allatum transplantations. Its influence on oxygen consumption depends upon increasing the total metabolism by the share taken by the activated larval tissues in the last larval instar or the share taken by the ovarian follicles in adults. SUMMARY 1. The rate of oxygen consumption in the last instar larvae of Pyvhoco~i~ is directly proportional to the degree of the induced morphological changes. 2. The oxygen-consumption curve of the implanted specimens is practically identical with that of the controls until the middle of the intermoult period. 3. The oxygen consumption in the second half of the instar is less reduced and the period of depression is shorter in specimens that moulted into sixth instar larvae instead of into adults, the intermediates with adultoid characters being intermediate also in this respect; 4. The results are consistent with those of a previous paper by the authors on the oxygen consumption of starving normal and castrated Pyrrhocoris female adults. 5, There,is only one c. allatum secretion, the juvenile hormone, responsible for all the observed effects of corpus allatum transplantations. 6. The JH-influence on oxygen consumption depends upon its increasing the amount of tissue that participates in the active metabolism. 7. The opposite conclusions by different authors based partly on the two corpus allatum hormones conception are inconclusive for several reasons on the basis of the experimental evidence available.
apterus
REFERENCES (1957) Die Steuerung der Ovarfunktion bei der ovoviviparen Schabe Leucophaea maderaz (Fabr.). J. Ins. Physiol. 1,257-278.
ENGELMANN
F.
JOHANSSON A. S. 11958) Relation of nutrition to endocrine-reproductive functions in the milkweed bug Oncopeltusfasciatus (Dal.). Nytt Msg. Zool. 7, l-132. L’H~LIAS C. (1954) I%ude du metabolisme basal chez le phasme Dixipplls morosus apres ablation des corpora allata. C.R. Acad. Sci., Paris 239, 778-780. L~~SCHER M. and SPRINGHETTIA. (1960) Untersuchungen iiber die Bedeutung der Corpora allata fiir die DifIerenzierung der Kasten bei der Termite Kalotermes flavivollis F. J. Ins. Physiol. 5, 190-212. Novk V. J. A. (1951) The metamorphosis hormones and morphogenesis in Oncopeltus fast. Dal. Acta Sot. zool. Bohem. 15, l-48. NoviiK V. J. A. (1960) Insektenhomzone (2nd ed.). NCSAV, Prague. NOVAKV. J. A., %.&MA K., and WENIG K. (1959) Influence of implantation of corpus allatum on the oxygen consumption of Pywhocoti apte-rus. Acta Symp. Evol. Ins. pp. 147-151. Prague.
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PFEIFFER-WBED I. (1945) Effect of the corpora ahata on the metabolism of adult female grasshoppers. J. exp. Zool. 99, 183-233. PFLUGFELDER0. (1952) Entwicklungsphysiologie der Insekten (I. AL@. Leipzig. PFLUGPELDER0. (1958) EntwickZungsphysioZogie der Insekten (II. Au&). Leipzig. S~~GESSERH. (1960) aber die Wirkung der Corpora ahata auf den SauerstofIverbrauch bei der Schabe Leucophaea made-rue (F.). 3. Inns. Physiol. 5, 264-285. SCwiEIDERMAN H. A. (1961) The juvenile hormone and other insect growth hormones. Acta Sot. ent. &l. 58 (l), 1-12. ScDNEIDERMANH. A. and GILBERT L. I. (1959) The chemistry and physiology of insect growth hormones. Cell, Organism and Milieu 6,157-187. SL.&A K. (1960) Oxygen consumption during the postembryonic development of Pyrrhoco& aptems and its comparison with that of Holometabola. Ann. ent. Sot. Amer. 53, 606-610. S&A K. (1961) Pseudo-juvenihxing effect in insects. Acta Sot. ent. &Z. 58, 1X7-120. THOMSEN E. (1949) InfIuence of the corpus allatum on the oxygen consumption of adult Calliphora erythrocepkala Meig. J. exp. Biol. 26, 137-149. THOMSEN E. and HAMBURGERK. (1955) Oxygen consumption of castrated females of the blow-fly, Calliphora erythrocephala Meig. 3. e@. Biol. 32, 692-699. WIGGLESWORTHV. B. (1935) Function of the corpus allatum in insects. N&me, Land. 136, 338. WILDE J. DE (1959) Diapause in the Colorado beetle Leptinotarsa decenzlineata Say as an endocrine deficiency syndrome of the corpora ahata. Acta Symp. Evol. Ins. pp. 226-230. Prague. WILLIAMS C. M. (1956) The juvenile hormone of insects. Nature, Lond. 178,212-213.
THE INFLUENCE OF JUVENILE HORMONE ON THE OXYGEN CONSUMPTION OF THE LAST LARVAL INSTAROF PYRRHOCORISAPTERUS L. V. J. A. NOVAK Entomological
Institute,
and K. SLhMA
Czechoslovac
Academy
of Sciences,
Praha
(Received 7 October 1961) Abstract-Oxygen consumption during the last larval instar of Pyrrhocoris upterus has been studied under the influence of implantations of active corpora allata. The degree of morphological change effected by tbe juvenile hormone was determined in each specimen after the larval-adult moult. The degree of the induced morphological change is directly proportional to the rate of oxygen consumption. The oxygen consumption curve was practically identical with that of controls until the middle of the intermoult period. The oxygen consumption in the second half of the instar, in which metamorphosis occurs, was less reduced and the period of depression was shorter in specimens that moulted into sixth instar larvae instead of into adults. The intermediates with adultoid characters were intermediate also in this respect. These results are consistent with those of previous work by the authors, accordiig to which the JR-effect on oxygen consumption is an indirect one depending upon increasing the amount of metabolically active tissue.
INTRODUCTION
THE effects of corpus allatum implantation
on the metabolism of insects have been described by many writers since the discovery of the juvenile hormone by WIGGLESWORTHin 1935. With one exception (PFLUGFELDER,1958) they agree that the oxygen consumption of the insect is increased. No agreement has been reached, however, as to whether the observed increase depends upon ‘a direct influence on metabolism, or whether it is an indirect consequence of the increased amount of growing tissue due to the activation of some parts of the body. The first hypothesis has been made by THOMSEN(1949) on the basis of the effects of allatectomy and corpus allatum implantation on oxygen consumption in Calhphora eryth~ocepha~a adults. The other view has been expressed by PFLUGFELDER(1952) for the first time as a result of his work on Dixzppus morosus, but he later (1958) abandoned his earlier view. In a previous paper (NOVAK et al., 1959) the authors attempted another approach to the problem. Oxygen consumption was measured in starving, normal, and castrated Pyrrhocmis female adults after corpus allatum implantation. Specimens in which a piece of muscle of corresponding size was implanted were used as controls. The results revealed that the implantation of active corpora allata increased the oxygen consumption in females with functioning ovaries but did not do so in castrated females (Fig. 1). The authors concluded that the observed increase was due to the activation by the hormone of ovarian follicle cells. From this 145
V. J. A. NOVLKANDK. SLLMA
146
it follows that the juvenile hormone has only an indirect effect on the total metabolism, depending on the stimulation of metabolic processes in particular
tissues.
700
-
600
-
500 -
400
I1 [i)
I
I
2
I
I
I
I
3
4
5
6
Days
L f
\”
0”
“E
600
E
500
Days FIG. 1. Oxygen consumption of starved normal (i) and castrated (ii) Pyrrhocoris aptems
female adults following the implantation of active corpus allatum. Solid line: specimens implanted with corpus allatum. Broken line: control specimens implanted with piece of muscle. Arrows indicate the moment of implantation.
Doubts have been thrown upon these results by S~~GESSER(1960), who claims to have found an increase in 0, consumption in Leucophczm after corpus allatum implantation even in castrated females. He agrees, as do THOMSEN and HAMBURGER (1955), that the juvenile hormone directly affects the total metabolism. S&GESSER explains the results of NOVAK et al. (1959) as an effect of starvation. He believes that in starving animals there is no supply of fat reserves necessary for ovarian maturation, so that negative results are not the result of castration but rather of
THE INFLUENCE
OF JUVENILE HORMONE
147
starvation. For reasons mentioned in the discussion, however, this explanation is not adequate. In the present paper a series of experiments were carried out to analyse the influence of corpus allatum implantation on the last larval instar of Pyrrhoco~is apterus, in which the juvenile hormone causes a strong morphogenetical effect. MATERIALS
AND METHODS
All the experiments
were performed on the fifth (i.e. the last) larval instar of Pyrrhoco~i.s apterus L. (Heteroptera, Pyrrhocoridae), Specimens were kept at a constant temperature of 25°C and 16 hr daylight. They were fed on lime-tree seeds and sufficient water was added. They were narcotized by plunging them into water for 10-15 min before the operation. Active corpora allata used for implantation were taken from mating adult females at the beginning of oviposition. Such glands are larger and at least as active in morphogenetical effect as those from young larvae (NOVAK, 1951). Implantations were made into the side of the third abdominal sternite the first day after moulting. Control specimens were operated in the same way except that instead of a gland they received a piece of muscle of similar size. For further detail see Nov& et al. (1959). The method of determining oxygen consumption in Pyrrhocwis apterus has already been described (NOVAKet aZ., 1959; SLAMA, 1960). Warburg manometers have been used. The total volume of the flasks was about 10 ml. The figures for oxygen consumption are based upon averages derived from about ten individuals, the oxygen consumption of each of which represents an average of three or more 1 hr readings on the same day. The total oxygen consumption of forty-nine specimens implanted with c. allatum and forty-one controls were measured daily from the beginning to the end of the last larval instar. RESULTS
The course of oxygen consumption in Py~~hocoris during the intermoult period is characterized by a rise of 0, consumption in the beginning of each instar and by its subsequent decrease in the second half of the instar. A new rise in consumption begins immediately before the next ecdysis. The course of the 0, consumption in the last larval instar is similar, but the depression in the second half of the instar is distinctly deeper (SL&EA, 1960). The 0, consumption of the fifth instar larvae implanted with a piece of muscle tissue used as a control is on the average identical with that of the normal specimens (cf. SUMA (1960) with Fig. 2). The very small decrease in oxygen consumption following the operation shows that narcotizing and implantation of themselves have no significant influence on oxygen consumption. The quantitatively different effects of corpus allatum implantation have shown themselves not only by morphogenetical changes but also by differences in the length of the intermoult period. The most affected specimens which moulted into giant sixth instar larvae had the shortest intermoult period (on an average 25 per
148
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K. %kMA
AND
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900
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800 -
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FIG. 2. Weight and oxygen consumption of Pyrrhocoris aptwus f&lx instar larvae moulted on sixth, seventh, and implanted with active corpus allatum. A, B, C-specimens Each point represents an average of thirteen individuals eighth days respectively, D-controls. in group (a), eleven individuals in group (b), ten in group (c), and fourteen in group (d). Arrows indicate the moment of implantation. For further explanation see text.
cent shorter), whereas the adultoid larvae moulted later. All the specimens str, -d therefore fall into the following four groups according to the amount of morphulogical change and the length of the instar :
THE INFLUENCE
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_rlost affected specimens, the intermoult period of which was reduced to 6 days and which moulted into complete sixth instar larvae; (b) specimens moulting into slightly adultoid sixth instar larvae the seventh day after the preceding ecdysis; (c) adultoid larvae moulting on the eighth day; and (d) control specimens implanted with a piece of muscle moulting into normal adults on the eighth day. The weight of the body increases steadily from the beginning of the instar. However, its growth stops 1 or 2 days before the next ecdysis. The specimens of all three groups moulting into sixth instar larvae reached nearly the same average weight (45-48 mg) at the end of the instar. The controls were about 15 per cent heavier (average, 54 mg). The last larval instar in Pynhocoti may be subdivided into two parts on the basis of the growth of the body and the oxygen consumption. In the first of these, at the beginning of the instar, the weight of the body increases steadily and the oxygen consumption per individual increases. In the second part, the growth of the body slows down and the oxygen consumption decreases in cormexion with the intensive morphogenetical processes. The interdependence between growth and an increase in 0, consumption is striking, as previously noted (SW, 1960). A close resemblance between the changes in body weight and 0, consumption is evident from Fig. 2, where both curves rise similarly. In the affected specimens, as in the controls, no additional increase in oxygen consumption has been observed in the first part of the instar: the Os-consumption curves follow the curve of the body weight. It means that the presence of the juvenile hormone, does not produce any change in metabolism during this period. As might be supposed, c. allatum implantation influences the metabolism in the second part of the instar where the bulk of metamorphosis occurs. In the controls a steep fall of 0, consumption occurs in this period. This decrease occurs independently from the increase in the weight of body, being most probably the consequence of the loss of metabolic activity in the larval parts of the body which are partly replaced by the imaginal parts during the metamorphosis. This depression of the U-shaped C&-consumption curve does not reach a similar depth in the holometabolous pupa. The amount of tissue undergoing metamorphosis in Pyrrhocoris is smaller, as a larger propbrtion of tissue survives to the adult stage in the Heterometabola. The corpus allatum implantation results, as has been generally agreed, in activation of the larval tissues which therefore continue their growth at the same rate as the imaginal ones, so that the growth of the body as a whole is an isometric one and no metamorphosis occurs (Nov.&, 1951, 1960). It might thus be expected that the fall in oxygen consumption in these specimens will be less intensive than in the normal metamorphosis controls; and this is what is actually observed, as shown in Fig. 2. Thus the more an individual is affected by a corpus allatum implantation, the less the 0, consumption falls in the second part of the instar. Hence the 0, consumption of the most affected specimens in group (a) falls to 660 $/g/hr, and in the less affected groups (b) and (c) to 650 and 620 ,ul/g/hr respectively, whereas IO
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in the control group it falls to a 560 $/g/hr. A similar decrease of the O,-consumption curve as in group (a) occurs also in the preceding larval instars when their own c. allatum functions (SLA~VIA,1960). Similarly, the observed shortening of the intermoult period in the most affected animals may be explained as due to the reduction of the second part of the instar in which the fall occurs. Thus the depression of the O,-consumption curve in group (a) occurs in the last day before moulting, whereas it takes 2 days in (b) and (c) and 3 days in the controls. The period of the increase in 0, consumption is thus equally long, taking 4-5 days in all groups. The terminal increase in 0, consumption immediately before ure end ot the in-star may be explained by the influence of ecdysis, as suggested by SW (1960). No decline in 0, consumption has been observed in comparison with that of the controls in specimens in which there was only a local effect of the implanted gland. The results of the experiments show clearly that the observed positive effect of the juvenile hormone on 0, consumption is not a direct and general one; it is just a consequence of the increase in total metabolism due to the share of the larval parts of the body activated by the hormone, which would have otherwise lost their activity. If the effect of the implanted gland were a direct one, it would have affected the Q-consumption curve during the first part of the instar in the same way. DISCUSSION The results of this paper thus agree fully with those of NOVAK et aZ. (1959) on the female adult of Pywhacoris. As shown in Fig. 1, implantation of c. allata induces a distinct rise of 0, consumption in normal starving females in comparison with controls implanted with a piece of muscle of similar size. This agrees with JONAN~~ON(1958), who found that corpus allatum implantation induces egg-laying even in starving (i.e. physiologically allatectomized) females of Oncopeltusfmciatus. Implantation of active c. allata into females which have been castrated at the beginning of the last larval instar does not affect metabolism. From this it may be concluded that the rise in 0, consumption in normal females following c. allatum implantation is due to the activation of the ovarian follicle cells. According to S~~GESSER (1960), the lack of juvenile hormone effect in castrated females in the experiments of NOVAK et aZ. (1959) may be due to the lack of available fat reserves in starving specimens. This is, however, scarcely likely as the same treatment in the similarly starved normal females retaining their ovaries has been effective. In addition, the fat reserves in the castrated specimens are on the average higher than in the normal ones, where the reserves are utilized by the growing ovaries. In our view the results of SAGESSERon Leucophaeu may be interpreted in the following way. %GESSER implanted nine castrated females of Leucophau (five of them deprived of both ovaries and accessory glands, the others of ovaries only) with active corpora allata. The 0, consumption of the experimental animals was about one-third higher than that of the controls with ovaries implanted with active
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c. allata. Even if we take as significantthe difference found (O-1172 0.02 ml O,/g/hr) within the individual variability of normal specimens (not less than O-3ml O,/g/hr, according to Fig. 2 of this paper), the following note by the author makes more probable a different explanation: ‘Es ist zu beachten, dass fiir die Implantation aktiver c. allata immer das Tier benutzt wurde, das vor der Operation den geringsten Verbrauch angewiesen hatte’ (UGESSER, 1960, p. 277). The results show that the bulk of the individual difference in 0, consumption per unit weight depends upon the stage of digestion of the animals. Those which have accepted the food immediately before the measurement have a higher body weight and lower 0, consumption per g per hr due to the stage of consumption of the food from the last meal. .Thus their respiration in relation to body weight is lower. On the other hand, those specimens which are at the peak of digestion of the previous meal have a low body weight, their 0, consumption per unit of weight being relatively high. However, the stage of digestion changes and may be quite different from one measurement of 0, consumption to another. There is another reason to prefer our explanation The specimens ‘with inactive corpora allata’ used as controls received the inactive corpora allata from females with oiithecae. However, the 5 days following the implantation spent in the new ‘active’ medium (females which had deposited their oiithecae 4 days before the operation were used) had necessarily to bring the implanted gland to the stage of full activation at the time of the measurement. These are therefore the true experimental animals comparable with ‘Ringer controls’. Our interpretation is also supported by the observation of SAGESSER that there is no qualitative difference in the Oa-consumption curve between the one but last and last larval instar of Leucophaea ,m.aderae, so long as we do not refuse the conception of only one corpus allatum hormone. As a matter of fact, there is no experimental evidence that could be better explained by the supposition of two hormones, as postulated by SBGESSER: one of them the morphogenetically active juvenile hormone and the other with metabolic activity said to be identical with the supposed adult ovarian follicle activating hormone. The fallacy of SAGESSER’S conclusion that the larval corpus allatumwould differ in its influence on metabolism from that of the adult female may be clearly shown by measuring the 0, consumption of specimens implanted with larval corpora allata if the concentration level of the hormone produced is taken into account. Similarly all other evidence in favour of two corpora allata hormones by other authors (L&cm and SPRINGBTTI, 1960.; ENGEL-, 1957; SCHNEIDERMAN, 1961) is inconclusive and may be more simply explained by the supposition of one corpus allatum hormone, as shown by NOVAK(1960). Th is is not the case with the extract of cecropia male abdomens described by WILLIAMS(1956) and SCHNEIDERMAN and GILBERT(1959), which seems to be a composite mixture of lipoid materials which can cause a regressive metathetely with progressive prothetely effects, and is probably not identical with the juvenile hormone of the corpus allatum (SLAMA, 1961)., The same is true of the findings by DE WILDE (1959) on corpus allatum and cecropia extract effects in Leptinotarsa akxmlineata and the homogenates of its tissues.
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All other evidence by earlier authors (PFEIFFER-WEED, 1945; THOMSEN, 1949; PFLUGFELDER, 1952, 1958; L’HBLIAs, 1954), as shown in Nov.& et al. (1959), does not affect the question of whether the observed juvenile hormone effect on the 0, consumption is a direct or indirect one; and it may be completely explained as here suggested. It may be thus concluded that there is only one corpus allatum hormone responsible for all the observed effects of corpus allatum transplantations. Its influence on oxygen consumption depends upon increasing the total metabolism by the share taken by the activated larval tissues in the last larval instar or the share taken by the ovarian follicles in adults. SUMMARY 1. The rate of oxygen consumption in the last instar larvae of Pyvhoco~i~ is directly proportional to the degree of the induced morphological changes. 2. The oxygen-consumption curve of the implanted specimens is practically identical with that of the controls until the middle of the intermoult period. 3. The oxygen consumption in the second half of the instar is less reduced and the period of depression is shorter in specimens that moulted into sixth instar larvae instead of into adults, the intermediates with adultoid characters being intermediate also in this respect; 4. The results are consistent with those of a previous paper by the authors on the oxygen consumption of starving normal and castrated Pyrrhocoris female adults. 5, There,is only one c. allatum secretion, the juvenile hormone, responsible for all the observed effects of corpus allatum transplantations. 6. The JH-influence on oxygen consumption depends upon its increasing the amount of tissue that participates in the active metabolism. 7. The opposite conclusions by different authors based partly on the two corpus allatum hormones conception are inconclusive for several reasons on the basis of the experimental evidence available.
apterus
REFERENCES (1957) Die Steuerung der Ovarfunktion bei der ovoviviparen Schabe Leucophaea maderaz (Fabr.). J. Ins. Physiol. 1,257-278.
ENGELMANN
F.
JOHANSSON A. S. 11958) Relation of nutrition to endocrine-reproductive functions in the milkweed bug Oncopeltusfasciatus (Dal.). Nytt Msg. Zool. 7, l-132. L’H~LIAS C. (1954) I%ude du metabolisme basal chez le phasme Dixipplls morosus apres ablation des corpora allata. C.R. Acad. Sci., Paris 239, 778-780. L~~SCHER M. and SPRINGHETTIA. (1960) Untersuchungen iiber die Bedeutung der Corpora allata fiir die DifIerenzierung der Kasten bei der Termite Kalotermes flavivollis F. J. Ins. Physiol. 5, 190-212. Novk V. J. A. (1951) The metamorphosis hormones and morphogenesis in Oncopeltus fast. Dal. Acta Sot. zool. Bohem. 15, l-48. NoviiK V. J. A. (1960) Insektenhomzone (2nd ed.). NCSAV, Prague. NOVAKV. J. A., %.&MA K., and WENIG K. (1959) Influence of implantation of corpus allatum on the oxygen consumption of Pywhocoti apte-rus. Acta Symp. Evol. Ins. pp. 147-151. Prague.
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PFEIFFER-WBED I. (1945) Effect of the corpora ahata on the metabolism of adult female grasshoppers. J. exp. Zool. 99, 183-233. PFLUGFELDER0. (1952) Entwicklungsphysiologie der Insekten (I. AL@. Leipzig. PFLUGPELDER0. (1958) EntwickZungsphysioZogie der Insekten (II. Au&). Leipzig. S~~GESSERH. (1960) aber die Wirkung der Corpora ahata auf den SauerstofIverbrauch bei der Schabe Leucophaea made-rue (F.). 3. Inns. Physiol. 5, 264-285. SCwiEIDERMAN H. A. (1961) The juvenile hormone and other insect growth hormones. Acta Sot. ent. &l. 58 (l), 1-12. ScDNEIDERMANH. A. and GILBERT L. I. (1959) The chemistry and physiology of insect growth hormones. Cell, Organism and Milieu 6,157-187. SL.&A K. (1960) Oxygen consumption during the postembryonic development of Pyrrhoco& aptems and its comparison with that of Holometabola. Ann. ent. Sot. Amer. 53, 606-610. S&A K. (1961) Pseudo-juvenihxing effect in insects. Acta Sot. ent. &Z. 58, 1X7-120. THOMSEN E. (1949) InfIuence of the corpus allatum on the oxygen consumption of adult Calliphora erythrocepkala Meig. J. exp. Biol. 26, 137-149. THOMSEN E. and HAMBURGERK. (1955) Oxygen consumption of castrated females of the blow-fly, Calliphora erythrocephala Meig. 3. e@. Biol. 32, 692-699. WIGGLESWORTHV. B. (1935) Function of the corpus allatum in insects. N&me, Land. 136, 338. WILDE J. DE (1959) Diapause in the Colorado beetle Leptinotarsa decenzlineata Say as an endocrine deficiency syndrome of the corpora ahata. Acta Symp. Evol. Ins. pp. 226-230. Prague. WILLIAMS C. M. (1956) The juvenile hormone of insects. Nature, Lond. 178,212-213.