Physiological control of pheromone production in Choristoneura fumiferana and C. rosaceana

The diel periodicity of calling behavior and pheromone production are synchronous in virgin females of both Choristoneura fumiferana and C. rosaceana (Lepidoptera: Tortricidae). Newly emerged females decapitated prior to scotophase produced no or very little pheromone 24 h later. However, injection of PBAN or Br-SEG homogenates, obtained from donors of the same or the other species, stimulated pheromone production to normal levels. Transection of the ventral nerve cord (VNC) or extirpation of the terminal abdominal ganglion (TAG) did not affect pheromone production in control females. Similarly, injections of PBAN or Br-SEG homogenates into decapitated females reactivated pheromone production to normal levels, whether or not the VNC was intact or the TAG present. Furthermore, octopamine was not effective in stimulating pheromone production in decapitated females. Taken together, these results indicate that the regulation of pheromone production is not neurally mediated in either Choristoneura species. However, there was no evidence that hemolymph collected from pheromone-producing females contained pheromonotropic activity. Similarly, isolated glands incubated with PBAN did not produce pheromone. The presence of the bursa copulatrix was required to produce pheromone in both tortricids as production was not restored in decapitated bursa-less females injected with PBAN or a Br-SEG homogenate. However, an extract of the bursa copulatrix did not elicit pheromonotropic activity in decapitated females or incubated glands of either species. The bursa copulatrix is only involved in pheromone production of some species of tortricids but our results do not support the current explanation for such interspecific differences. We postulate that the relative importance of a bursa factor may be related to the evolution of different desaturation systems used for pheromone biosynthesis in the Tortricidae. Arch.     

Regulation of pheromone production by female pink bollworm moths Pectinophora gossypiella (Sanders) (Lepidoptera: Gelechiidae)

Abstract. We present in this study data which indicate that there is a diel periodicity in the pheromone production of the pink bollworm moth Pectinophora gossypiella (Sanders) (Lepidoptera: Gelechiidae) but that it is not well defined. Moreover the control mechanism of pheromone production differs somewhat from that reported for other moths. No pheromonotropic response was obtained when photophase females were injected with synthetic Helicoverpa zea pheromone biosynthesis activating neuropeptide (Hez-PBAN). After decapitation for 24 h, Hez-PBAN did not induce pheromonotropic activity above control levels, which themselves remained relatively high. No effect on pheromone production was observed after treatment with the non-steroidal ecdysone agonist (RH5999). Decapitation for 72 h resulted in a significant drop in the control levels of pheromone titres. After decapitation for 72 h, stimulation by injections of Hez-PBAN and pink bollworm head extracts was observed. In addition, an enhancement of the PBAN stimulation was observed when combined with severance of the ventral nerve cord before injection. On the other hand, pink bollworm head extracts did not cross-react with Hez-PBAN antiserum in a radioimmunoassay, indicating that the pheromonotropic factor present is sufficiently different from Hez-PBAN and does not recognize the antigenic binding sites. In studies using isolated abdomen and pheromone gland cultures in vitro , no stimulation of de novo pheromone biosynthesis was observed but a 3-fold increase in the de novo fatty acid biosynthesis was detected in pheromone gland cultures.     

PBAN selective antagonists: inhibition of PBAN induced cuticular melanization and sex pheromone biosynthesis in moths

A D-Phe scan (sequential D-Phe replacement) library of linear peptides, synthesized on the basis of a slightly modified active sequence of PBAN (YFSPRL-amide) was employed to detect potential inhibitors of cuticular melanization in Spodoptera littoralis larvae and to compare their stimulatory and inhibitory melanization activity with their pheromonotropic agonistic and antagonistic activities. A quantitative melanotropic assay was used to monitor the extent of cuticular melanization elicited by Hez-PBAN1-33NH2 in S. littoralis larvae in the presence and absence of the D-Phe peptides. The data revealed the presence of two partial melanotropic antagonists, and disclosed the presence of selective pure melanotropic agonists and pure pheromonotropic antagonists indicating differences in the inhibitory and stimulatory patterns of the library with respect to both activities. The differences between the pheromonotropic and melanotropic inhibitory patterns of the peptides hints at the possibility that sex pheromone biosynthesis in the pheromone gland of Heliothis peltigera females and induction of cuticular melanization in S. littoralis may be mediated by different receptors (that may result either from presence of different receptor sub-types or may reflect species differences in receptor structure and/or properties) despite the fact that they are induced by the same peptide (PBAN1-33NH2).     

Pheromonotropic and pheromonostatic activity in moths

Pheromone biosynthesis in many species of moths requires a pheromonotropic neurosecretion, the pheromone biosynthesis activating neuropeptide (PBAN), from the brain-subesophageal ganglion-corpora cardiaca complex. Some investigators suggest that PBAN is released into the hemolymph and acts directly on sex pheromone glands (SPG) via a Ca⁺⁺/calmodulin-dependent adenylate cyclase. Others suggest, however, that PBAN acts via octopamine that is released by nerves from the terminal abdominal ganglion innervating the SPG. These findings suggest that there are controversies on the mode of action of PBAN and other pheromonotropic factors, sometimes even within the same species. Mating in many insects results in temporary or permanent suppression of pheromone production and/or receptivity. Such a suppression may result from physical blockage of the gonopore or deposition of pheromonostatic factor(s) by the male during copulation that result in suppressed pheromone production and/or receptivity in females either directly or by a primer effect. In several species of insects, including moths, a pheromonostatic factor is transferred in the seminal fluid of males. Similar to the controversies associated with the pheromonotropic activity of PBAN, sometimes even within the same species, there appear to be controversies in pheromonostasis in heliothines as well. This paper reviews these conflicting findings and presents some data on pheromonostatic and pheromonotropic activity in Heliothis virescens that support and conflict with current information, raising further questions. Answers to some of the questions are partly available; however, they remain to be answered unequivocally. © 1994 Wiley-Liss, Inc.     

Age-related changes in the competency of the pheromone gland and the pheromonotropic activity of the brain of both virgin and mated females of two Choristoneura species

Nine-day-old decapitated females injected with different doses of Hez-PBAN produced significantly less pheromone than 1-day-old individuals, suggesting that the age-related decline in the pheromone titre of Choristoneura fumiferana and C. rosaceana virgin females was primarily the result of a reduced ability of the glands to produce pheromone. In C. fumiferana, lower pheromonotropic activity of the Br-SEG may also contribute to the pheromone decline with age but not in C. rosaceana, as the pheromonotropic activity of the Br-SEG remained constant throughout the females' life. In both Choristoneura species, mating also suppressed pheromone production (pheromonostasis) after 24 h. The Br-SEG of mated females contained PBAN but there was no indication that its concentration changed with time post-mating since Br-SEG homogenates obtained from different-aged mated females showed the same level of pheromonotropic activity in both Choristoneura species. However, as observed in virgins, pheromone glands of older mated females were less sensitive to PBAN than those of younger ones. This suggests that the probability of Choristoneura females to attract a second mate may decrease with an increase in the refractory period following the first mating.     

Pheromone-producing cells in the silkmoth, Bombyx mori: identification and their morphological changes in response to pheromonotropic stimuli

A method to isolate functional clusters of viable pheromone gland cells of Bombyx mori was developed. The 8th-9th intersegmental invaginated membrane corresponding to the pheromone gland was dissected, trimmed and separated into two distinct layers, the outer and inner layers, by enzymatic digestion with papain. The outer layer mainly consists of cuticle, while the inner layer consists of homogeneous cells with many refractile granules. The solubilized microsome fraction prepared from the inner layer retained the ability to produce bombykol in vitro, whereas the outer layer fraction did not produce bombykol. Moreover, in tissue incubations, the inner layer - but not the outer layer - produced bombykol in response to the pheromonotropic peptide TKYFSPRLamide, ionomycin and calcium ionophore A23187. These results indicate that the inner-layer cells are indeed the pheromone-producing cells, which retain their functional integrity after separation with papain. These cells could be cultured successfully in Grace's medium for at least 5days.The presence or absence of pheromonotropic stimuli prior to dissection greatly influenced the size, number and distribution of refractile granules in the cytoplasm of the pheromone-producing cells. Staining with Nile Red proved that these refractile granules were lipid droplets. When pheromone production was studied under normal conditions or stimulated in decapitated females with pheromone-biosynthesis-activating neuorpeptide (PBAN) charge, the size of lipid droplets observed in the pheromone-producing cells reduced prominently and their number increased dramatically with time. By contrast, when pheromone production was suppressed by decapitation, the size and number of the lipid droplets remained constant. Lipid droplets observed in the pheromone-producing cells could be carriers of pheromone precursors and/or the pheromone bombykol. The present results suggest that the isolated cell preparation can be used for quantitative visualization of the cellular dynamics during pheromone production in B. mori.     

Spatial and temporal distribution of pheromone biosynthesis-activating neuropeptide in Helicoverpa (Heliothis) armigera using RIA and in vitro bioassay.

A [3H]-PBAN (pheromone biosynthesis-activating neuropeptide) analog was synthesized, and binding of the radioligand to a specific PBAN-antiserum was achieved. The inhibition of binding of the radioligand by unlabeled PBAN, several PBAN analogs, and other competitors was studied and a specific radio-immunoassay was developed. Using this radioimmunoassay we found PBAN-like immunoreactivity in methanol extracts of hemolymph and neural tissues from females. Higher levels of PBAN-like immunoreactivity in extracts of brain-suboesophageal ganglion complexes, corpora cardiaca, thoracic ganglia, and abdominal ganglia were observed during the 4-5th h scotophase when compared to the PBAN-like immunoactivity levels during the 6-11th h photophase. On the other hand, the concentrations of PBAN-like immunoreactivity, in the terminal abdominal ganglion were higher during the photophase relative to minimal levels observed during the scotophase, indicating an accumulation before the onset of pheromone production. These differences in concentrations of PBAN were also reflected in the stimulation of in vitro pheromone glands, whereby significant stimulations were obtained by scotophase and photophase brain extracts, scotophase thoracic ganglia extracts, and photophase terminal abdominal ganglia extracts. No detectable levels of PBAN were found in hemolymph extracts during the sampling periods.     

A pheromonotropic peptide of Helicoverpa zea,with melanizing activity,interaction with PBAN,and distribution of immunoreactivity

The sequence of an 18-amino acid residue peptide was deduced from the gene encoding PBAN and other peptides with common C-termini in Helicoverpa zea. The peptide caused melanization in larvae and pheromone production in females of H. zea, and was designated pheromonotropic melanizing peptide (Hez-PMP). The peptide has a 83% sequence homology with a pheromonotropic peptide isolated from Pseudaletia separata. PMP caused melanization and mortality when injected into larvae just before molting. Whereas intense melanization was caused with a dose of 1,000 pmol, peak mortality occurred at 100 pmol, with 50% of larvae dying within 48 h after injection. Pheromonotropic activity of PMP was dose dependent. Co-injection of Hez-PMP and Hez-PBAN into a female resulted in suppression of the pheromonotropic effect of PBAN. Whole-mount immunocytochemical studies revealed PMP-like immunoreactivity in frontal ganglion, subesophageal, thoracic, and abdominal ganglia as well as the esophageal nerve.     

昆虫神经肽PBAN的细胞内信号转导

昆虫性信息素多数为长链的不饱和醇、醋酸酯、醛或酮类,链长一般为10-20碳,主要在性信息素腺体内由乙酰辅酶A经过脂肪酸合成、碳链缩短、去饱和以及碳酰基的还原修饰等步骤合成的;而性信息素合成激活肽(pheromone biosynthesis activating neuropeptide,PBAN)是由昆虫食管下神经节中的部分神经细胞合成和分泌的神经肽,通常由33个氨基酸组成,在C-末端有一个相同的五肽序列,主要调控性信息素的生物合成。有关PBAN的细胞内信号转导是近几年的研究热点,研究显示 PBAN首先与性信息素腺体细胞表面的G蛋白偶联受体结合,随后依据昆虫种类的不同,其细胞内信号转导方式主要有三种:(1)以cAMP信号传导途径进行信号转导;(2)以cAMP和磷脂酰肌醇信号传导途径共同进行信号转导;(3)主要以Ca2 为第二信使进行信号传导。     

The effect of the juvenile hormone analog, fenoxycarb on the PBAN-receptor and pheromone production in adults of the moth Helicoverpa armigera: an "aging" hormone in adult females?

In a previous study we showed that juvenile hormone (JH) or its analog, fenoxycarb (FX), is involved in the up-regulation of pheromone biosynthesis-activating neuropeptide (PBAN) competence. JH causes induction of binding to a putative PBAN-receptor (PBAN-R) and the subsequent pheromone production by pheromone glands of pharate females. The present study demonstrates that pheromone production by the adult female is age-dependent. The pheromonotropic response increased to reach a maximum at 4 days, after which a decreased response was observed. Binding of the PBAN-R was also age-dependent. Treatment with FX inhibited both binding of PBAN to the PBAN-R and the pheromonotropic response as reflected by the production of the main pheromone component, Z-11-hexadecenal. Thus, in contrast to its up-regulatory role in pharate females, FX treatment of adult females causes down-regulation of both pheromone production and specific binding to the PBAN-R. In addition, behavioural observations showed that calling behaviour, mating success and subsequent egg-fertility are affected by treating females with FX.     

Pheromonotropic stimulation of moth pheromone gland cultures in vitro

The direct neurohormonal control of pheromone biosynthesis by pheromone biosynthesis activating neuropeptide (PBAN) was demonstrated in Helicoverpa (Heliothis) spp. using pheromone gland cultures in vitro. Pheromone gland activation involved the de novo production of the main pheromone component (Z)-11-hexadecenal as revealed by radio-TLC, radio-HPLC, and radio-GC. Activation was found to be a specific response attributed to pheromone gland cultures alone. Specificity of pheromonotropic activation was demonstrated to be limited to nervous tissue extracts. A sensitive and specific radioimmunoassay was developed using [³H]-PBAN, and the spatial and temporal distribution of PBAN-immunore-activity was studied. PBAN-immunoreactivity in brain complexes was found throughout the photoperiod and in all ages. From the distribution of PBAN-immunoreactivity it appears that PBAN release is affected by photoperiod. Pheromone gland cultures were found to be competent to pheromone production irrespective of age and photoperiod. Therefore, the neuroendocrine control of pheromone production operates at the level of neuropeptide synthesis and/or release and not at the level of the target tissue itself. The involvement of cyclic-AMP as a second messenger system was demonstrated. Brain extracts and PBAN were shown to stimulate dose- and time-dependent changes in intracellular cyclic-AMP levels. The role of cyclic-AMP in this mechanism was further verified by the ability of cyclic-AMP mimetics to mimic the pheromonotropic effect of brain extracts and PBAN. However, dose-response studies using PBAN and a hexapeptide C-terminal fragment of PBAN suggested that PBAN induces a two mechanism response, one occurring at low PBAN concentrations (high affinity receptor) and another at higher PBAN concentrations (low affinity receptor). Further evidence indicating a dual receptor system was obtained with the observation that the active phorbol ester (phorbol-12-myristate 13-acetate), the diacyl-glycerol analog (1,2-dioleolyl-sn-glycerol), and the intracellular calcium ionophore (ionomycin) mimicked the physiological action of PBAN and that lithium chloride had a pheromonostatic effect. The results indicate that pheromone glands also possess receptors that are linked to inositol phosphate hydolysis. © 1994 Wiley-Liss, Inc.     

Mating Effect on Sex Pheromone Production of the Oriental tobacco budworm,Helicoverpa assulta

This study was undertaken to clarify the suppression phenomenon of sex pheromone production after mating and its relationship to the physiological mechanism in adult females of Helicoverpa assulta, and determine the mating factor from males causing depletion of sex pheromonc production. Sex pheromone production of H. assulta females was mostly terminated in 3 hours after mating. Mated females maintained with a low titer of sex pheromone until 3 days when it started to increase again, which showed a characteristic of species mating more than once. The mated female again produced pheromone upon injection of pheromone biosynthesis activating neuropeptide (PBAN) or extracts of brain-suboesophageal ganglion complexes (Br-Sg) of mated female, which were shown similar pheromonotropic activities as compared with virgin females. These results indicated that the mating did not inhibit the receptivity of pheromone gland itself and PBAN biosynthesis in suboesophageal ganglion of the mated females. And it seems to support that the depletion of sex pheromone production is responsible for blocking of PBAN release from head. To investigate the mating factor from adult males, when extracts of reproductive organs of male were injected into hemocoel of virgin females evoking depletion of sex pheromone production as shown in mated female. The results suggest that a chemical substance(s) from the male reproductive organs could be responsible for the loss of sex pheromone biosynthesis in H. assulta.     

Termination of sex pheromone production in mated females of the silkworm moth

A mating duration of more than 6 h was necessary to permanently terminate the production of the sex pheromone (bombykol) in the silkworm moth, Bombyx mori L. (Lepidoptera: Bombycidae), although the female formed a bursa copulatrix including a spermatophore and laid fertilized eggs even after mating for only 0.5 h. The 6-h mated female again produced bombykol if given an injection of synthetic pheromonotropic neuropeptide (PBAN), which is known to activate pheromone biosynthesis in a virgin female. Extracts of brain-suboesophageal ganglion (SG) complexes, which were removed from 6- and 24-h mated females, showed strong pheromonotropic activities. These results indicated that the pheromone gland of the mated female maintained its ability to biosynthesize bombykol; however, it could not produce pheromone due to a suppression of PBAN secretion from the SG. Furthermore, bombykol titers did not decrease after mating in females with a transected ventral nerve cord, even after the injection of a spermatophore extract, suggesting that the suppression of PBAN secretion was mediated by a neural signal and not by a substance in the spermatophore. The mated females accumulated (10E, 12Z)-10,12-hexadecadienoic acid, a precursor of bombykol biosynthesis, in their pheromone glands as did decapitated females. © 1996 Wiley-Liss, Inc.     

Hormonal control of female sex pheromone biosynthesis in the redbanded leafroller moth,Argyrotaenia velutinana

Pheromone biosynthesis in female redbanded leafroller moths (RBLR) is under control of a neuropeptide produced in the brain. A bioassay consisting of isolated abdomens was developed to test the mode of action of the pheromone biosynthesis activating neuropetide (PBAN). Pheromone titer and incorporation of radiolabeled acetate into pheromone could be monitored with this bioassay. Synthetic PBAN with sequences identical to PBAN isolated from Heliothis zea and Bombyx mori were active in inducing synthesis of pheromone in RBLR. Removal of the ventral nerve cord in isolated abdomens did not inhibit the action of PBAN. Small amounts of PBAN-like activity was found in hemolymph collected from normal females but not from decapitated females. Severing the VNC in vivo in normal females did not lower the pheromone titer. These data indicate that PBAN is released into the hemolymph and then travels to its site of action. A two-fold increase in both pheromone titer and radiolabeled acetate incorporation upon incubation with PBAN was shown with isolated pheromone glands. However, the differences between control and PBAN-induced values were smaller than those obtained with the isolated abdomen culture bioassay where a seven-fold increase was observed. A decrease in pheromone titer was seen upon the in vivo removal of the corpus bursae from normal females. Removal of the corpus bursae in the isolated abdomen cultures also abolished the activity of PBAN. However, cutting the cervix bursae and leaving the corpus bursae in the abdomen culture increased both titer and radiolabeled acetate incorporation into pheromone without the presence of PBAN. An aqueous extract made from the corpus bursae of 5-day-old females was also active by itself in inducing pheromone biosynthesis in the isolated abdomen cultures. Experiments performed using newly emerged females confirmed that the corpus bursae extracts will induce pheromone biosynthesis. These results indicate that both PBAN and the corpus bursae are involved in controlling pheromone biosynthesis in RBLR.     

Octopamine and tyramine modulate pheromone-sensitive olfactory sensilla of the hawkmoth <Emphasis Type="Italic">Manduca sexta</Emphasis> in a time-dependent manner

In moths octopamine improved pheromone-dependent mate search time dependently. In the nocturnal hawkmoth Manduca sexta long-term tip recordings of trichoid sensilla were performed to investigate whether biogenic amines modulate pheromone transduction time dependently. At three Zeitgebertimes octopamine, tyramine and the octopamine antagonist epinastine were applied during non-adapting pheromone-stimulation. At ZT 8-11, during the photophase, when sensilla were adapted, octopamine and to a lesser extent tyramine increased the bombykal-dependent sensillar potential amplitude and initial action potential (AP) frequency. In addition, during the photophase, when sensilla are less able to resolve pheromone pulses, octopamine rendered pheromone responses more phasic and sensitive, and raised the spontaneous AP frequency. During the late scotophase, at ZT 22-1, when the antenna appeared maximally sensitized for pheromone pulse detection and endogenous octopamine levels are high, exogenously applied octopamine was ineffective. Epinastine blocked the pheromone-dependent AP response at ZT 8-11 and slightly affected it at ZT 22-1, while it had no effect on the sensillar potential amplitude. Epinastine decreased the spontaneous AP activity during photophase and scotophase and rendered pheromone responses more tonic in the scotophase. We hypothesize that the presence of octopamine in the antenna is obligatory for the detection of intermittent pheromone pulses at all Zeitgebertimes.     

Regulation of sex pheromone biosynthesis in the oriental tobacco budworm,Helicoverpa assulta (Lepidoptera: Noctuidae)

The five components, Z9-16:Ald, 16:Ald, Z11-16:Ald, Z9-16:Ac and Z11-16:Ac, of the sex pheromone in Helicoverpa assulta were mostly detected during the scotophase, with their titer peaking at the 4th hour during the scotophase under a 15L/9D regime. They were not detected during the photophase, but were produced during the photophase when decapitated females were injected with extracts of virgin female (FHE), male heads (MHE), homogenates of the brain-suboesophageal ganglion complex (Br-SOG), or synthetic Hez-PBAN. Production of Z9-16:Ald increased during the first 45min after FHE injection and then declined to a very low level after 2h during the photophase. Synthetic Hez-PBAN stimulated the sex pheromone glands for at least 2h and the effect was more or less proportional to the concentration of the peptide. From the present results, we suggest the following: PBAN is released continuously into the haemolymph to stimulate pheromone biosynthesis at least during the first half of the scotophase, PBAN is synthesized and accumulated independent of photoperiod or sex, and the release starts just prior (about 1h) to the beginning of the scotophase.     

Discovery of a linear lead antagonist to the insect pheromone biosynthesis activating neuropeptide (PBAN)

We report the discovery of a linear lead antagonist for the insect pheromone biosynthesis activating neuropeptide (PBAN) which inhibits sex pheromone biosynthesis in the female moth Heliothis peltigera. Two approaches have been used in attempting to convert PBAN agonists into antagonists. The first involved omission of the C-terminal amide and reduction of the sequence from the N-terminus in a linear library based on PBAN 1-33NH(2.) The second involved replacement of L amino-acids by the D hydrophobic amino acid D-Phe in a linear library based on PBAN28-33NH(2.) Screening of the two libraries for pheromonotropic antagonists resulted in the disclosure of one compound out of the D-Phe library (Arg-Tyr-Phe-D-Phe-Pro-Arg-Leu-NH(2)) which inhibited sex pheromone production by 79 and 64% at 100 pmol in two moth colonies and exhibited low agonistic activity. Omission of the C-terminal amide in PBAN 1-33NH(2) and its shorter analogs did not lead to the discovery of an antagonistic compound.     

Pyrokinin/PBAN radio-receptor assay: development and application for the characterization of a putative receptor from the pheromone gland of Heliothis peltigera

A radio-receptor assay (RRA) for the insect pyrokinin/PBAN family has been developed. The development involved examination of the ligand (3H-tyrosyl-PBAN28-33NH2)-receptor interaction under various incubation conditions and variations on sex pheromone gland membrane preparation. Application of the RRA for a partial characterization of the putative pyrokinin/PBAN receptor in the pheromone gland of H. peltigera revealed age-dependence of its expression. Pharmacological characterization revealed a high correlation between the binding-affinity to the receptor of various PBAN-derived peptides and their in vivo pheromonotropic bioactivity, and shed light on the interaction of backbone cyclic and linear ([Arg27,D-Phe30]PBAN28-33NH2) PBAN antagonists with the receptor.     

Stimulation of sex pheromone production by proteinaceous extracts of the bursa copulatrix in the redbanded leafroller moth

Pheromone biosynthesis in the redbanded leafroller moth, Argyrotaenia velutinana, was stimulated by homogenates of the bursa copulatrix. Although pheromonotropic activity was also extractable from the ovary, the activity of pheromone biosynthesis activating neuropeptide (PBAN) or bursa extracts was not impaired in isolated abdomens by removal of the ovary. Response to the bursa extracts was dependent on the dose administered and the time of incubation. Amounts of pheromone present in adult females of different ages appeared to be correlated with the extractable amount of pheromonotropic activity from their bursa copulatrix. Decapitation did not result in the suppression of burse factor production. Homogenates of the bursa elicited similar effects in both isolated gland and isolated abdomen incubations, but the brain neuropeptide, PBAN, was less active in the former than in the latter. Bursa extracts stimulated pheromone production in isolated abdomen incubations deprived of the bursa copulatrix, but PBAN did not. Loss of activity of bursa homogenates after treatment with either pronase E or carboxypeptidase Y indicated that the pheromonotropic factor is a proteinaceous substance. The mechanism through which pheromone production is regulated in redbanded leafroller moths is discussed. © 1992 Wiley-Liss, Inc.