Frank Dittmann

The first gonocycle of the parasitic mite (Varroa juobsoni) in relation to preimaginal development of its host, the honey bee (Apis mellifra carnicar)

Summary Initiation of oocyte development in Varroa jacobsoni depends on whether the female enters the brood cell of Apis mellifra before operculation and subsequently sucks hemolymph from a late fifth instar bee larva. Females inserted into the brood 24 h after cell capping, during the spinning phase of the bee larva, do not initiate oocyte development. All stages of mite vitellogenesis and embryogenesis which are described here develop in a close temporal relation to the host stages. A timetable of the first gonocycle is given showing its duration in worker and drone host cells. About 70 h after capping of the bee brood cell, the first egg is laid containing a nearly completed protonymph that hatches within the next day.

Juvenile hormone-controlled vitellogenin cycles in Dysdercus intermedius (Heteroptera)

Abstract Two vitellogenic female-specific haemolymph proteins and one yolk-specific protein were demonstrated in Dysdercus intermedius. The yolk-specific protein includes all female-specific protein subunits. A cyclical change in the temporal pattern of the female specific polypeptides occurs during the first gonadotropic cycle. Female specific polypeptides do not occur in the haemolymph during the pre-vitellogenic stages of oocyte development and during formation of the chorion. Volume changes of the corpus allatum are correlated with changes in yolk precursors in the haemolymph. Allatectomy by decapitation of the female during the early pre-vitellogenic stage suppress the formation of the major female-specific polypeptides. Applications of juvenile hormone-III or corpus allatum transplantation restores the ability to produce these polypeptides.

Voltage gradients and microtubules both involved in intercellular protein and mitochondria transport in the telotrophic ovariole of Dysdercus intermedius

SummaryIn the telotrophic ovariole of Dysdercus intermedius the intercellular transport consists of different subsystems. Microinjection of FITC-labeled slowly diffusing proteins with opposite electrical net charges and of mitochondria was used to study the translocation of macromolecules and organelles. a) By intracellular measurements a voltage gradient of about 4 mV between the tropharium as the more negative side and the previtellogenic oocytes could be demonstrated. b) After injection into the tropharium negatively charged proteins migrated according to the electropotential gradient via the trophic cords into the oocytes. Positively charged proteins, however, were retained in the tropharium. c) After injection into previtellogenic oocytes both negatively and positively charged proteins moved into the trophic cords. Thus, the effectiveness of the electropotential gradient on the distribution of charged proteins is more pronounced from the tropharium side. d) Mitochondria microinjected into the trophic core were probably aligned along microtubules and translocated towards the trophic cords. — These results suggest that in the telotrophic bug ovariole a number of intercellular transport subsystems contribute to provide previtellogenic oocytes with nurse cells products. An electrophoretic transport mechanism for soluble proteins acting especially within the tropharium and a microtubule-associated transport for mitochondria could be evidenced.

Movement of mitochondria in the ovarian trophic cord of Dysdercus intermedius (Heteroptera) resembles nerve axonal transport

SummaryThe motile behaviour of mitochondria in the ovarian trophic cord of the red cotton bug, Dysdercus intermedius, was observed optically using video-enhanced differential interference contrast (AVEC-DIC) microscopy. The motion of 258 video-recorded mitochondria was analysed of which 10%–30% were found to move during the observation periods. Of the moving mitochondria 76% travelled towards the oocyte with an average velocity of 3.37 μm/ min, and 24% towards the tropharium with 2.84 μm/min. The movement was found to be basically of the saltatory type I as known from nerve axons characterized by the absence of directional reversal. In some cases short periods of interrupted motion of type II, i.e. with local oscillations, were observed. Individual mitochondria often showed velocity variations during the excursions. The hemipteran trophic cords are known to contain numerous parallel microtubules. As the observed type of mitochondrial motility resembles axonal transport, a modified transport hypothesis is presented for the microtubule-based motility of organelles in the nurse strands of telotrophic insect ovarioles.

Induction of yolk formation in hemipteran previtellogenic oocytes (Dysdercus intermedius)

Summary Yolk formation has been studied in previtellogenic oocytes of the telotrophic-meroistic ovariole of the red cotton bug Dysdercus intermedius (Heteroptera: Pyrrhocoridae) in the absence of the follicular epithelium (“skinned oocytes”). Early preparation for endocytosis was seen by urea gel electrophoresis and immunoblotting, which showed that cytosolic clathrin (light chain) is already present in the previtellogenic trophocyte-oocyte syncytium. The ability of these previtellogenic skinned oocytes to form yolk was studied by incubating them in physiological saline to which rhodarnine-labelled hemolymph proteins were added. These oocytes formed a peripheral band of fluorescent yolk spheres when incubated in vitellogenin-containing hemolymph proteins obtained from 6-day-old adult females but not when in hemolymph proteins from 3-day-old females, which lack vitellogenin. AVEC-DIC microscopy was used to record fluorescent protein uptake as it occurred in living, previtellogenic oocytes. Adsorption to th...

Developmental patterns of DNA-accumulation and nuclear division in the follicle epithelium of the red cotton bug, Dysdercus intermedius (Heteroptera)

Abstract We demonstrate the spatial and the developmental pattern of DNA content in the follicular epithelium nuclei in the red cotton bug, Dysdercus intermedius, during the first gonadotropic cycle. The prefollicular tissue is composed of nuclei of different DNA material formed by underreplication of DNA. This disposition of DNA material does not change in the sequel of development of previtellogenic follicles. As yolk is incorporated, the ploidy changes, establishing a new batch of polyploid nuclei by a final underreplication of DNA. These nuclei form the basis for the development of nuclei in the epithelium of vitellogenic follicles. Either they duplicate their genome, forming mononucleate cells or divide into binucleate cells. Yolk incorporation is not dependent on the composition of polyploid nuclei in the follicular epithelium. Precocene-II treatment and juvenile hormone-III application cause no apparent alteration in the ploidy pattern of the previtellogenic follicles. While juvenile hormone exerts no influence upon the epithelium of previtellogenic follicles, it stimulates ploidy changes in vitellogenic follicles. The developmental pattern of juvenile hormone-induced ploidizations indicates a direct mode of action.

The effect of ooplasmic pH regulation on the formation of yolk spheres in the telotrophic ovariole of Dysdercus intermedius

Vitellogenic oocytes of Dysdercus intermedius (Heteroptera: Pyrrhocoridae) were treated with the proton ionophore monensin in order to load the ooplasm with protons along the electrochemical gradient. Additionally, changes in the ooplasmic pH (DeltapH(OOC)) were recorded during exposure the oocytes to potassium-free medium (K(+)(MED)=0mM; choline for K(+)) or sodium-free medium (Na(+)(MED)=0mM; 40mM of choline for 40mM of Na(+)). The following observations were made: 1) The average ooplasmic pH (pH(OOC)) recorded during immersion in physiological saline solution (PSS) was pH(OOC(PSS))=7.40. 2) K(+)(MED) had no effect on pH(OOC) (pH(OOC(K-FREE)) congruent with pH(OOC(PSS))). 3) In sodium-free medium the pH(OOC) decreased by H(+) influx in the magnitude of DeltapH(OOC(Na-FREE))=pH(OOC(Na-FREE))-pH(OOC(PSS))=-0.05 pH units. 4) The decreased pH(OOC) observed in sodium-free medium returned to initial values (7.40) by pumping out H(+) when 40mM of choline were replaced by 40mM of Na(+). 5) Addition of monensin (10&mgr;M; under the condition of Na(+)(MED)=0mM) reduced pH(OOC) in the magnitude of DeltapH(OOC(MON))=pH(OOC(MON))-pH(OOC(PSS))=-0.14. 6) Monensin induced ooplasmic proton loading was reversible when 40mM choline were replaced by 40mM Na(+).VITELLOGENESIS WAS DEMONSTRATED BY THE ACCUMULATION OF FLUORESCENCE LABELLED HEMOLYMPH PROTEINS IN YOLK SPHERES IN THE CORTEX OF THE OOCYTE: 1) Yolk formation continued in potassium-free medium. 2) The formation of yolk spheres came to a halt in sodium-free medium and, additionally, in the presence of monensin (10&mgr;M; Na(+)(MED)=0mM). 3) Breaks in yolk formation under the condition of Na(+)(MED)=0mM or during monensin treatment were stopped by replacing 40mM of choline with 40mM of Na(+). The results obtained using proton-specific microelectrodes and the in vitro assay to detect the formation of yolk spheres indicate that both the ooplasmic pH regulation and the acidification of vesicles during vitellogenesis are under control of a H(+)/Na(+) antiporter.

CA2+ CURRENT FROM OOCYTE TO NURSE CELLS AND SUPPRESSION OF URIDINE INCORPORATION IN THE GERMINAL VESICLE OF HYALOPHORA CECROPIA

Summary In ovarian follicles of Hyalophora cecropia, [3H]uridine incorporation by germinal vesicles is normally arrested at the start of vitellogenesis when a Ca2+ current arises through the intercellular bridges connecting the oocyte to its nurse cells. We show here that the germinal vesicle resumes uridine incorporation following inhibition or reversal of this current. Inhibition/reversal was achieved by treating follicle with any one of three inhibitors that block the Ca2+ current by very different mechanisms. Vanadate was used to block Ca2+ extrusion by the nurse cells, which drives the current. BAPTA was used to buffer Ca2+ activities in the nurse cero, and A23187 was used to increase the membrane conductance of Ca2+. Measurements with ion selective microelectrodes indicated that in all these cases [Ca2+]i in nurse cells equaled or exceeded that in the oocyte; and in all these cases uridine incorporation resumed in the germinal vesicle. Resumption correlated not with the level of [Ca2+] i in the oocy...

Yolk formation in the telotrophic-meroistic ovariole of Dysdercus intermedius depends on extracellular sodium

Abstract With the present study we attempted to integrate electrophysiological results with effects of ion substitution in the vitellogenic oocyte of Dysdercus intermedius . The permeability of the oocyte membrane to ions in the in vitro medium was determined by equimolar substitutions of K + or Na + by choline; Ca ++ by Mg ++ Mg ++ by Sr ++ , and by sulfate for Cl − . The impermeability of the oocyte membrane to Na + , Ca ++ , and Mg ++ , as opposed to the permeability of K + and Cl − , renders the latter two as the major ions that contribute to the membrane potential ( E m ) of −34 mV. Intracellular activities of K + , Na + , Ca ++ , Mg ++ and Cl − were measured with ion selective microelectrodes. Liquid Ion Exchange (LIX) pH microelectrode determination revealed an ooplasmic pH of 7.4 which is 0.6 pH units higher than the pH (6.8) of the in vitro medium. Variations of pH med was of no influence on pH ooc . We visualised the effects of ion substitution on yolk formation by in vitro uptake of rhodamine-labeled hemolymph proteins derived from vitellogenic females which results in the formation of large fluorescent yolk spheres. Cl − replacement did not affect the protein uptake. However, in the absence of Ca ++ yolk formation was restrained. Replacement of external K + by choline did not influence protein uptake when [Na + ] med was present, but resulted in a marked inhibition of yolk formation when in vitro medium was Na + -free. Thus this process appears to depend on extracellular Na + rather than intracellular or extracellular K + .

Intercellular connection between the lyrate organ and the growing oocyte in Varroa jacobsoni as revealed by Lucifer Yellow dye-coupling

SUMMARYMicro-injected Lucifer Yellow CH (LY) was used to study cytoplasmic continuity and transport between the lyrate organ and the developing oocyte in the ovary of the mite, Varroa jacobsoni, an ectoparasite of honey bees. When LY was micro-injected into the lyrate organ of young female mites in their phoretic phase, movement of the dye towards the late stage 2 oocyte did not occur. Further oocyte development was initiated by transfer of young female mites into brood cells of Apis mellifera shortly after operculation (‘reproductive phase’ of mites). Uptake of haemolymph from an L5 (feeding phase) bee larva coincided with the migration of LY both from one lobe of the lyrate organ to the other as well as from the lyrate organ towards the oocyte. Since translocation of LY in vitro reflects the alimentation of oocytes (nutrimentary oogenesis), we highlight the temporal pattern of intercellular transport during oogenesis.