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K Hiruma

Publications and source records attributed to K Hiruma.

80 records · Page 5Linked to original sources

Regulation of prothoracicotropic hormone release by juvenile hormone in the penultimate and last instar larvae of Mamestra brassicae.

The penultimate instar larvae of Mamestra brassicae have one ecdysteroid peak 2 days after ecdysis which induces larval ecdysis. Allatectomy deleted this peak and caused a larger peak 7 days later just before precocious pupation, in a pattern similar to the ecdysteroid titer of the last instar. Application of juvenile hormone (JH) immediately after allatectomy restored the normal larval pattern and larval ecdysis. The brain of the normal Day 1 or 2 penultimate instar larva had higher prothoracicotropic hormone (PTTH) activity than that of the allatectomized larva. Therefore, JH in the penultimate instar activates the brain to secrete and/or produce PTTH. Allatectomy of last instar larvae prolonged the period from gut purge to pupation by causing a delay of the ecdysteroid peak necessary for pupation. Application of JH restored the normal timing of ecdysteroid release. Thus, JH is important for the occurrence of the ecdysteroid surge on Day 7, since its absence delayed this surge by a day.

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Roles of dopa decarboxylase and phenoloxidase in the melanization of the tobacco hornworm and their control by 20-hydroxyecdysone.

When Manduca sexta larvae are allatectomized 5 h before head capsule slippage (HCS) in the final larval molt, the new larval cuticle contains granules that melanize 3 h before ecdysis when the ecdysteroid titer falls (Curtis et al. 1984). In both the epidermis and hemolymph of these allatectomized larvae dopamine was higher than dopa prior to and at the time of melanization. Dopamine also increased in the new cuticle as melanization began. Dopa decarboxylase (DDC) activity increased in the epidermis, cuticle, and fat body beginning 16 h after HCS, with a two-fold greater increase in the epidermis of allatectomized larvae. Both alpha-MDH and alpha-fluoromethyl-dopa inhibited epidermal DDC activity and inhibited melanization in vitro when dopa was used as a precursor. Addition of dopamine to the medium allowed melanization in the presence of the inhibitors. All these results indicate that dopamine is likely the primary precursor of cuticular melanin. The diphenoloxidase in the premelanin granules was activated in vivo between 19 and 21 h after HCS and was found to prefer dopamine to dopa and not to convert tyrosine to melanin. The activation of the prophenoloxidase was inhibited by 20-hydroxyecdysone (20-HE), both in vivo and in vitro, if hormone was given by 16 h after HCS. Infusion of 1.2 microgram/ml 20-HE into allatectomized larvae for 24 h from HCS prevented both the increase in DDC activity and the activation of the premelanin granules. Although the larvae ecdysed after a 15 h delay, melanization never occurred.

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Hormonal regulation of dopa decarboxylase during a larval molt.

Cuticular sclerotization in insects requires dopamine derivatives and thus the presence of dopa decarboxylase (DDC), the enzyme which converts dopa to dopamine. During the last half of the larval molt of the tobacco hornworm, Manduca sexta, beginning at 16 hr after head capsule slippage, the epidermal DDC activity increased fourfold. By contrast, allatectomized larvae which were destined to produce a melanized cuticle showed a sevenfold increase. This increase in DDC activity was prevented by infusion of 20-hydroxyecdysone (20HE) into the larva, indicating that the fall of the ecdysteroid titer is necessary for the increase. In vitro 20HE also prevented the increase in a dose-dependent manner when the epidermis was explanted at 16 hr after head capsule slippage but had less effect on epidermis explanted 3 hr later. Both 5 micrograms/ml alpha-amanitin and 100 micrograms/ml cycloheximide also prevented the increase. Application of juvenile hormone I showed that the critical period for determination of the level of the later increase in DDC activity was about 4 hr after head capsule slippage at the peak of the ecdysteroid titer. Apparently then the rise and fall of ecdysteroid regulate different aspects of DDC synthesis, the rise determining its later appearance and the fall timing this appearance.

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Regulation of melanization of tobacco hornworm larval cuticle in vitro.

When tobacco hornworm larvae (Manduca sexta) are allatectomized 5-6 hr before head capsule slippage in the molt to the fifth (final) larval instar, the new cuticle melanizes 3 hr before ecdysis. After explantation between 7 and 3 hr before the onset of melanization, the new cuticle was found to melanize in vitro in Grace's medium only if beta-alanine was removed. When explanted at the onset of melanization, the presence of beta-alanine had no effect on melanization. The addition of either dopa or dopamine was found to be necessary for complete melanization of pieces explanted before the onset of melanization with 0.3 mM of either dopa or dopamine being optimal. Both of these compounds were incorporated into the cuticular melanin. In this optimal medium, melanization occurred over about a 9-hr period after a 5- to 6-hr lag period presumably required for adjustment to the medium. Fifty ng/ml 20-hydroxyecdysone was found to inhibit melanization of pieces explanted 7 hr but not 3 hr before melanization. The hormone neither inhibited uptake of dopa into the epidermis nor prevented melanization in the cuticle once the prophenoloxidase in the premelanin granules was activated. Therefore, 20-hydroxyecdysone may inhibit the activation of the phenoloxidase in the pre-melanin granules, or may inhibit the incorporation of dopa into the granules.

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