Historical aspects Directory UMM :Data Elmu:jurnal:I:Insect Biochemistry and Molecular Biology:Vol30.Issue8-9.Sept2000:

Insect Biochemistry and Molecular Biology 30 2000 617–644 www.elsevier.comlocateibmb The juvenile hormones: historical facts and speculations on future research directions Lawrence I. Gilbert a, , Noelle A. Granger b , R. Michael Roe c a Department of Biology, Campus Box 3280 Coker Hall, University of North Carolina at Chapel Hill, Chapel Hill, NC 27599-3280, USA b Department of Cell Biology and Anatomy, Campus Box 7090, University of North Carolina at Chapel Hill, Chapel Hill, NC 27599-7090, USA c Department of Entomology, Campus Box 7647, North Carolina State University, Raleigh, NC 27695-7647, USA Received 31 October 1999; received in revised form 31 December 1999; accepted 25 January 2000

1. Historical aspects

In all of endocrinology there is no more wondrous name for a hormone than the insect juvenile hormone JH. Could V.B. Wigglesworth have predicted some six decades ago that his term “juvenile hormone” would offer promise of immortal youth to the aged, the expec- tation of a bloom of dollars to agrochemical concerns, and the hope of solutions to basic problems by develop- mental biologists and entomologists. The aged have been disappointed and the high expectations of commercial firms have not been met, but hope remains that JH can be used as a probe to ultimately solve basic questions in development. It has been more than two centuries since Lyonet 1762 described granulated vessels in the thorax of lepi- dopteran larvae that proved to be the prothoracic glands. By contrast, the corpora allata were not mentioned in the literature until Mu¨ller 1828 described organs in the cockroach that he called pharyngeal bodies and which he thought innervated the dorsal vessel and esophagus. During the remainder of the 19th century, the corpora allata were described as sympathetic ganglia or other components of the nervous system, as indicated by the various descriptive terms given them, e.g. accessory gan- glia, tracheal ganglia, lateral ganglion, lateral head gang- lion, appendage of the pharyngeal ganglion, etc. In 1899, Heymons dubbed these organs the corpora allata and correctly described their embryological origin, but also believed that they were a pair of sympathetic ganglia concerned with the innervation of the digestive system. Corresponding author. Tel.: + 1-919-966-2055; fax: + 1-919-962- 1344. E-mail address: lgilbertunc.edu L.I. Gilbert. 0965-174800 - see front matter  2000 Elsevier Science Ltd. All rights reserved. PII: S 0 9 6 5 - 1 7 4 8 0 0 0 0 0 3 4 - 5 In 1910, Police suggested that the corpora allata were endocrine organs in phasmids, but that they probably had a nervous function in other insects. It was Nabert in 1913 who, on the basis of studying a variety of insects representing several orders, finally stated that the corpora allata were glandular and exhib- ited periodic internal secretions. This was confirmed by the work of Ito 1918, who concluded that the corpora allata were indeed organs of internal secretion and that they functioned in the adult moth as well. All of this work was anatomical and histological with a bit of microsurgery, with the corpora allata being observed under the light microscope using a variety of stains. Ito was probably the first to use planimetrics to measure the diameter of the corpora allata and to demonstrate variations in size during various stages of metamor- phosis. Burtt 1937, 1938 described the corpora allata in the higher Diptera and found that these organs were a por- tion of the “Weissman’s Ring” or as it is now known, the ring gland Burtt and Hadorn, 1937. He hypothes- ized that certain structures within the ring gland rep- resented modified and fused corpora allata. We now know, of course, that the ring gland is composed of cells with the function of the corpora allata, cells with the function of the prothoracic gland, and an anatomical por- tion that appears to have elements of the corpora car- diaca e.g. Dai and Gilbert, 1991. It was about that time that V.B. Wigglesworth 1935 began his historic studies on many aspects of Rhodnius development and meta- morphosis, making efficient use of surgical techniques such as decapitation and subsequent parabiotic regimens. He established the critical period for molting in this bug, and because the corpus allatum displayed cyclical activity which was correlated with this critical period, he assumed at first that this gland was the source of the 618 L.I. Gilbert et al. Insect Biochemistry and Molecular Biology 30 2000 617–644 molting hormone. Further surgical studies suggested that the insect also contained an “inhibitory factor” which prevented the first four larval stages from molting directly into adults. In 1935 he suggested that both the molting and inhibitory hormones were produced by the corpus allatum, and in 1936 his classic paper on the function of the corpus allatum in the growth of Rhodnius was published. He showed that the corpus allatum was the source of the inhibitory hormone that prevents meta- morphosis in young larvae and that corpora allata from these young larvae when implanted into fifth instars caused them to undergo a supernumerary molt. He per- fected the surgery of allatectomy and showed that such animals could molt, indicating that the corpus allatum was not the source of molting hormone. Wigglesworth concluded that each cell has the potential for larval or adult differentiation and that hormone titers determine which potential is realized. It is the concentration of the corpus allatum hormone that determines the extent of metamorphosis at the next molt, wrote Wigglesworth 1948, and of course, analogous experiments were done on other insects over the next twenty-five years showing that Professor Wigglesworth was correct. It is of interest that some of his work on this subject probably would not be accepted for publication today because of the small sample size used, i.e. only two of the five operated fifth instars molted into intermediates after receiving cor- pus allatum implants from fourth instars. Our own per- sonal bias is that Wigglesworth had the best intuition of any scientist we have met or read about. In the first of many studies of JH function, Piepho 1938a,b, 1950 showed via histological studies that the formation and nature of the cuticle of Galleria was under hormonal control. In the initial experiments, the integu- ment from one larva was grafted to another, and the graft integument molted with the host. This led to a series of experiments in which a fragment of integument from one larva was implanted into the abdominal cavity of another. The epidermis of the fragment regenerated around the cuticle of the implant to give rise to an epi- dermal vesicle with cuticle at its interior. Piepho’s lab- oratory showed that when the host molted, the vesicle molted in concert. His studies very beautifully demon- strated that molting was under hormonal control, that the epidermis reacts to JH by laying down larval structures cuticle and that the epidermis does not “count the molts”. The classic study of Piepho 1942, 1950 revealed important phenomena concerning tissue com- petence and tissue responsiveness. First, it appeared that the epidermal cells remained responsive to JH until very late in the last stadium. When implants were done very late in the stadium, there was a patch of larval cuticle at the wound site in the resulting pupa. Second, the results showed that wounded epidermis is much more sensitive to the action of the JH than intact regions. This was a vital finding for the development of the Galleria wax test that was used successfully for some twenty years before analytical analyses of JH came to the fore Gilbert and Schneiderman, 1960; Schneiderman and Gilbert, 1964, and allowed the first semiquantitative titer of JH Gilbert and Schneiderman, 1961b; Table 1. During that same era, the classic studies of Bounhiol 1936–1938 showed the effects of corpora allata extir- pation in Lepidoptera Bombyx. Bounhiol’s results were confirmed and extended by those of Fukuda 1942; Fukuda 1944. It is of interest that despite the use of HPLC, mass spectrometry, etc., to measure JH, our con- cept of how the changing titer affects developmental pro- cesses has not advanced a great deal over the past fifty years. The “modern” era of JH research began with the criti- cal finding by Carroll Williams 1956 that the male Hyalophora cecropia moth contained a store of a lip- oidal “golden oil” with JH activity when assayed on lepi- dopteran pupae. The original intent of the experiment was to determine if the life of the male saturniid moth could be extended if it were parabiosed to a pupa. As is so common in biology, serendipity ruled, and the pupa molted into a second pupa, indicating that the moth had furnished the pupa with JH. Furthermore, Williams’ dis- covery elicited a frenzy of studies of JH in a variety of insects — for the first time, an active extract of this amazing hormone was available and it worked on most insect species. Indeed, from one male H. cecropia abdo- men enough JH could be extracted and diluted in peanut oil or mineral oil to conduct hundreds or even thousands of experiments. If you could find one male moth, you were in business for life If Williams had used the other saturniid studied in his laboratory in his original experi- ments, the parabiotic pupal partner would have molted into an adult since the adult Antheraea polyphemus is almost devoid of JH. In retrospect, it is clear from these early experiments that Williams, Wigglesworth, Piepho, etc. all had an almost uncanny ability to interpret cor- rectly from unexpected, incomplete and seemingly bizarre results. It was the patience and persistence of a young German scientist, Herbert Ro¨ller, that allowed his group to finally deduce the structure of JH I Ro¨ller et al., 1967. At the time, there was competition between the Ro¨ller labora- tory and that of Howard Schneiderman, the latter con- firming the structure of JH I and identifying JH II several years later Meyer et al., 1970. The difference between the approach of the two laboratories was that Ro¨ller et al. used glass columns in their gas chromatographic analyses while the Schneiderman group used all metal columns, which enhanced the lability of the JH mol- ecule. In any event, pure JHs were available about thirty years ago. It should be noted that we have chosen only three top- ics for our review and speculations. These are: control of the corpora allata; control of JH titer; and JH action 619 L.I. Gilbert et al. Insect Biochemistry and Molecular Biology 30 2000 617–644 Table 1 Juvenile hormone content during the life history of the Cecropia silkworm a Stage Approximate juvenile Approximate juvenile Approximate juvenile hormone concentration of hormone concentraion per hormone activity per gram extract: Cecropia insect or fragment: fresh weight: Cecropia unitsgram extract Cecropia unitsanimal or unitsgram animal or fragment fragment fresh weight Unfertilized eggs Minimum of: 165 0.042 8.151 Unfertilized eggs from alatectomized “0” “0” “0” 7-day embryos Minimum of: 165 0.031 7.095 7-day embryos from allatectomized Minimum of: 70 0.012 3.108 1st instar larvae newly hatched Minimum of: 165 0.031 7.475 5th instar larvae mixed sex 35 4.162 0.571 Diapausing pupae 1 month old 20 5.780 1.468 Diapausing pupae 1 month old 20 6.300 1.114 Chilled pupae 6 months old “0” “0” “0” Chilled pupae 6 months old “0” “0” “0” 2-day old developing adults “0” “0” “0” 8-day old developing adults “0” “0” “0” 11-day old developing adults “0” “0” “0” 14-day old developing adults “0” “0” “0” 17-day old developing adults “0” “0” “0” 20-day old developing adults 125 25 13.70 22-day old developing adults 400 122.68 201.64 Adult 4 days 675 151.88 298.75 Adult 7 days 1000 178.30 418.00 Adult 7 days 125 17.50 11.63 a Activity of extracts is expressed in Cecropia units. One Cecropia unit is equivalent to the juvenile hormone activity found in one milligram of extract obtained from the abdomens of seven-day old male Cecropia moths. Extractions of developing adults and adult moths were conducted on the abdomens only and the activity noted is for the abdominal extract. From Gilbert and Schneiderman 1961b. receptors. We simply could not summarize or do jus- tice to all the presentations made during the symposium and chose topics we felt were important and with which we were personally involved. We apologize to those individuals we did not cite since it is possible that their contributions may be even more important than those summarized here.

2. Control of the corpora allata