Chih-Ming Yin

Effect of allatectomy and ventral nerve cord transection on calling, pheromone emission and pheromone production in Lymantria dispar

Abstract The physiological control mechanisms of calling, pheromone production, and pheromone release in female gypsy moths were investigated. No involvement of the corpora allata was found since mean pheromone emission rates from females allatectomized as larvae (14.1 ng/h) were not significantly different from sham-operated (16.4 ng/h), anaesthetized (13.1 ng/h), or untreated controls (13.1 ng/h). Neural input from the brain or other higher centres of the nervous system, however, appeared critical since females which received ventral nerve cord transection anterior to the terminal abdominal ganglion between 0 and 24 h after pupation showed significant reductions in both gland-extractable pheromone ( x = 8.7 ng) and pheromone emission rate ( x = 2.1 ng/h) compared with controls ( x = 25.0 ng and x = 13.1 ng/h, respectively). Differences in calling behaviour were also observed; in control females, the duration of ovipositor protraction was about 16.5 s and retraction about 1.3 s, in ventral nerve cord-transected females the ovipositor was maintained in a full or partially protruded state for periods longer than 300 s. Post-operative inspection of the terminal abdominal ganglion revealed fewer descending nerve branches in ventral nerve cord-transected females compared to controls.

Cold-induced glycerol accumulation by Ostrinia nubilalis larvae is developmentally regulated

Abstract Ostrinia nubilalis larvae reared under both nondiapause and diapause-inducing conditions were chilled at 5°C for various periods and their haemolymph glycerol concentrations were measured enzymatically. The ability of fifth (final) instars to accumulate glycerol was dependent upon cold stress but not the diapause state. Furthermore this response was independent of any cold-induced release of cephalic or thoracic hormones. The capacity of O. nubilalis larvae to express cold-induced glycerol accumulation was found to require ecdysis from the fourth to fifth instar. Eggs as well as second, third and fourth instars were completely incompetent. These results indicate that, at the biochemical level, a specific developmental programme or sequence is required for O. nubilalis to demonstrate this response to cold stress.

Innervation of dromyosuppressin (DMS) immunoreactive processes and effect of DMS and benzethonium chloride on the Phormia regina (Meigen) crop

Antibody to the dipteran myosuppressin peptide, dromyosuppressin, TDVDHVFLRFamide, stained cells and fibers in the brain, optic lobes, subesophageal ganglion, and thoracico‐abdominal ganglion of the blow fly, Phormia regina (Meigen). Dromyosuppressin‐like immunoreactive fibers were detected in the cardiac recurrent nerve, hypocerebral ganglion/corpora cardiaca complex, crop duct, and crop. In order to explore the mechanisms involved in regulating crop movement, we established an in vitro bioassay. The basal rate of crop movement was 50.8 ± 1.5 contractions per minute. Application of 1 μl of saline to the crop did not significantly affect the rate of movement compared to the basal rate (46.1 ± 1.1 contractions per minute, P < 0.05). Application of 1 μl 10‐6 M dromyosuppressin or 1 μl 10‐3 M benzethonium chloride to the crop slowed the rate to 2.2 ± 0.2 and 6.1 ± 0.7 contractions per minute, respectively. Although other data have previously been interpreted to suggest that dipteran crop contractions do not include a neural component, the neuropeptide dromyosuppressin affected P. regina crop motility. Innervation of the crop and crop duct by dromyosuppressin immunoreactive processes that originated in the central nervous system and the effect of dromyosuppressin on crop muscle contractions suggest that dromyosuppressin is released locally to modulate crop contractions and that crop motility is under neural regulation. Myosuppressins isolated from numerous insects have a high degree of structure identity and reduce spontaneous muscle contractions of the hindgut, oviduct, and heart. Benzethonium chloride, previously identified as a myosuppressin agonist on the cockroach hindgut and locust oviduct, mimicked the effect of dromyosuppressin on the crop. This suggests that structural requirements for myosuppressin receptor binding in the cockroach hindgut, locust oviduct, and fruit fly crop are similar. J. Comp. Neurol. 421:136–142, 2000.

Neurological influences on pheromone release and calling behaviour in the gypsy moth, Lymantria dispar

ABSTRACT. Surgical removal of the brain or disconnection of the last abdominal ganglion from the ventral nerve cord prevented sex pheromone release in female Lymantria dispar (L.) (Lymantriidae), as assayed by the male wing‐fanning response. The calling behaviour continued to occur in individuals whose terminal abdominal ganglion had been thus isolated, however, indicating that the neural mechanisms controlling calling function independently in the last abdominal ganglion.

Lack of humoral control in calling and pheromone release by brain, corpora cardiaca, corpora allata and ovaries of the female gypsy moth, Lymantria dispar (L.)

Abstract Investigations into the physiological regulation of calling behaviour and pheromone release by the adult female gypsy moth, Lymantria dispar, showed that the brain, corpora cardiaca, corpora allata and ovaries play no significant endocrine role for either calling behaviour or pheromone release. Removal of corpora allata, corpora cardiaca-corpora allata complex and ovaries from last-instar larvae had no effect on calling behaviour nor on the pheromone release in the resulting moths. Severing the suboesophageal connectives had no effect on calling, but effectively eliminated pheromone release. Since severing nerves should not remove humoral output from the brain, the regulation on pheromone release appears neural.

Conditions for estimation of corpus allatum activity in the blowfly, Phormia regina, in vitro

ABSTRACT. Incubation conditions have been established for the corpus cardiacum‐corpus allatum (CC‐CA) complex of female Phormia regina (Meigen), which will support CC‐CA biosynthetic activities in vitro as measured by the incorporation of a labelled methyl group with L‐[methyl‐3H]methionine as the methyl donor. After incubation, radioactivity in the organic extract of the medium was determined by scintillation counting. Analysis of the organic extract with reverse phase high‐performance liquid chromatography (HPLC) revealed that a compound which has similar retention time (UV absorbance) with synthetic JH III was synthesized by the CC‐CA complexes of liver‐fed females. By using this short‐term, in vitro, radiochemical assay for CA activity, it was shown that a protein diet significantly increases the activity of the CA compared with females fed only a sugar‐water diet. Furthermore, use of HPLC separation, in conjunction with scintillation counting of time‐collected fractions, demonstrated the existence of a moiety containing incorporated radiolabeled methyl group (from the methionine) which did not co‐elute with JH I or JH III standards. These results suggest that in P. regina use of the incorporation of a radiolabeled methyl group to measure JH biosynthesis (CA activity) can be misleading if the compounds which do not co‐elute with JHs are not considered.

Subunit composition of vitellin, and concentration profiles of vitellogenin, and vitellin in Phormia regina (Meig.) following a protein meal

1. n1. The vitellin (Vt) of Phormia regina eggs was isolated by extraction at low ionic strength and purified by TEAE-cellulose chromatography. SDS polyacrylamide gel electrophoresis and staining with horseradish peroxidase-concanavalin A showed that the Vt contains four subunits of Mr 42,000, 43,000, 44,000 and 45,500, all of which are glycosylated. n n2. n2. Murine polyclonal antibodies to the Vt, employed in conjunction with radioimmunoblotting, recognized all four subunits, and revealed that Vt and/or vitellogenin (Vg) are present exclusively in the hemolymph, fat bodies and ovaries of liver-fed females but not those fed sugar. n n3. n3. Using these antibodies in an ELISA, concentrations of Vg in the hemolymph and Vt in the ovaries were determined periodically following liver feeding. At 28°C, the hemolyph Vg titer reaches a maximum 28 hr post-feeding and then decreases rapidly as ovaries begin to sequester Vt, whose concentration in that organ peaks about 48 hr post-feeding.

Ultrastructure of honey bee, Apis mellifera, sperm with special emphasis on the acrosomal complex following high‐pressure freezing fixation

Abstract. The ultrastructure of ejaculated sperm of the honey bee, Apis mellifera L., was studied after sperm were processed by using a high‐pressure freezing fixation and freezing substitution method. The electron micrographs of samples processed by this method clearly revealed previously unobserved or unresolved fine details, particularly those in the acrosomal complex. The present study demonstrates that the acrosomal complex of honey bee sperm consists of an anterior tubular acicular apex, an enlarged spherical region, and the elongated acrosomal proper. Internally, the structures of acrosomal complex confirm the general description of the tri‐layer model; having an extra‐acrosomal layer, an acrosome vesicle, and a central acrosomal rod located in a subacrosomal cavity. The acrosomal rod terminates anteriorly with an electron dense corpuscle in an ensheathing cap at the spherical region. The electron micrographs also show the presence of a centriolar adjunct and a structure possibly of a centriolar or basal body origin. The chemical and functional aspects of these ultrastructures remain unknown.

Effect of rapid freezing and thawing on cellular integrity of honey bee sperm

Abstract. Direct observations on the effect of rapid freezing and thawing on honey bee (Apis melliferaL.) sperm were made by light, scanning and transmission electron microscopy. Rapid freezing of honey bee ejaculated sperm, suspended in freezing diluent, in liquid nitrogen followed by rapid thawing can cause cellular injuries which lead to the death of the sperm. The frozen‐thawed sperm, supravitally stained, showed a significant decrease in cell viability compared with that of the control fresh sperm (P<0.001). Significant uptake of the stain in the dead sperm resulted from damage in the cell membrane. The scanning electron micrographs of frozen‐thawed sperm further demonstrated that the injury of cell membrane can lead to the splitting of mitochondrial derivatives from the flagellar axoneme. More cellular injuries including the release of acrosomal content and membrane damage at the acrosome, nucleus and the tail regions were further revealed by transmission electron microscopy. The impact of cellular injuries on the quality of honey bee sperm cryopreserved for artificial insemination of honey bee queens is discussed.

Influence of age on the electroantennogram response of the female blowfly (Phormia regina) (Diptera: Calliphoridae)

Abstract By means of electroantennogram recording techniques, we have monitored the antennal olfactory sensitivity, from the time of eclosion until complete egg maturation, of female blowflies (Phormia regina) fed either a protein diet or a protein free one. The tested stimuli were swormlure-4 (SL-4) which is a potent lure for calliphorid flies, 1-hexanol as a reference stimulus and air as a control. After taking the electroantennograms, egg and ovarian development were evaluated. Stimulation with SL-4 and 1-hexanol evoked electroantennograms increasing in amplitude with age regardless of whether or not flies were fed protein during the first 5 days of adult life. In the protein-fed flies eggs were fully developed whereas those of the no-protein group remained undeveloped. The peak olfactory sensitivity occurs at a time when the female fly begins to search for an oviposition substrate.