Wilhelm Roux's Archives (1982) 191:257-263 Roux's Archives of Developmental Biology �9 Springer-Verlag 1982

A Correlation between Juvenile Hormone Deficiency and Vitellogenic Oocyte Degeneration in Drosophila melanogaster Thomas G. Wilson Department of Zoology, University of Vermont, Burlington, VT 05405, USA

Summary. It is known from previous work that juvenile hormone (JH) is required to initiate vitellogenin uptake into maturing oocytes of Drosophila rnelanogaster, but addition- al requirements for this hormone during oocyte maturation have not been fully understood. To determine if early vitel- logenic oocytes (stages 8 and 9) require JH for continued development, these oocytes were transplanted to Drosophila female and male hosts which were rendered deficient in JH by three methods. Implanted stage 9 and usually stage 8 oocytes were found to degenerate in JH-deficient hosts unless ZR-515, a JH analogue, was applied to the host shortly after implantation.

These results were confirmed during in situ ovary devel- opment. JH deficiency was produced in gravid females, and ovaries examined at subsequent time intervals were found to be deficient in stage 8-10 oocytes as early as 6 h after treatment. Degenerating oocytes corresponding to these stages were commonly found. ZR-515 prevented oocyte de- generation during at least the first 8 h and continued to support stage 8-10 oocyte development 24 h after applica- tion to these females. The results suggest that JH is required not only for initiation but also for continuation of vitello- genin uptake and oocyte development.

Key words: Juvenile hormone - Precocene - Drosophila mel- anogaster Oocyte degeneration

Introduction

In Drosophila rnelanogaster vitellogenin uptake from hemo- lymph into oocytes begins 8-10 h after eclosion of the adult (reviewed by Mahowald and Kambysellis 1980). Vitellogen uptake characterizes stages 8-10 of oocyte development (staging according to King 1970), and the duration of each of these stages is about 5 h (David and Merle 1968). The first mature oocytes (stage 14) appear within 30-35 h after eclosion. Although stage 10 oocytes complete development to stage 14 in vitro in the absence of hormones (Petri et al. 1979), the hormonal requirements for vitellogen uptake and accompanying stage 8 and 9 oocyte development have not been completely delineated. It is known that 20-hydroxyec- dysone is involved in vitellogenin biosynthesis in the fat body (Jowett and Postlethwait 1980), but this hormone does not appear to function in uptake (Postlethwait and Handler 1979). Juvenile hormone (JH), however, is necessary to initi- ate vitellogen uptake in Drosophila (Postlethwait and Wei- ser 1973; Kambysellis and Heed 1974; Gavin and William- son 1976; Postlethwait et al. 1976) as in other insects (Bell and Barth 1971; Pratt and Davey 1972; Kelly and Daven-

port 1976), since oocytes remain in previtellogenic stages in the absence of JH even though adequate vitellogenin is present in the hemolymph. However, it is not clear in Drosophila whether JH serves only to initiate vitellogen up- take or also is required continously for this process. Horse- radish peroxidase uptake into yolk spheres of vitellogenic oocytes requires JH (Giorgi 1979), suggesting a JH require- ment for continued vitellogenin uptake, but Giorgi's study did not examine vitellogenin uptake or stage 8 and 9 oocyte development to mature oocytes.

In order to assess any JH requirement for development of early vitellogenic oocytes I have transplanted individual Drosophila ovarioles having one stage 8 or 9 oocyte as the most mature, terminal oocyte into hosts deficient in JH. Transplanted into wild-type hosts, ovarioles and even indi- vidual egg chambers develop stage 14 oocytes (Srdic and Jacobs-Lorena 1978), indicating that an intact ovary is not required for oocyte development. I then corroborated the results of transplantation experiments by examining oocyte development in situ in JH-deficient adults. The results in all cases indicated that JH promotes development of stage 8 and 9 oocytes; in the absence of this hormone, degeneration of these oocytes occurs.

Materials and Methods

The Oregon-RC stock was obtained from the Developmen- tal Biology Center, Irvine, California USA. The ap4/SM5 and Canton-S stocks were obtained from the Mid-America Drosophila Stock Center, Bowling Green, Ohio, USA. Flies were raised on a standard brewer's yeast-agar-cornmeal- molasses medium seeded with live baker's yeast at 25 ~ C. Adults eclosing over a 2-h period in the morning were col- lected by direct transfer to unseeded medium at 25 ~ C. In an effort to minimize the circadian rhythm of oocyte matu- ration found for females maintained on a LD 12:12 photo- period (Allemand 1976a, b), both larvae and adults were maintained in total darkness.

Ovaries were dissected free and examined in Drosophila Ringer's solution (Ephrussi and Beadle 1936). Each ovary was gently teased apart, and stage 8-14 oocytes were cen- sused; a good photographic representation of each stage is available (Mahowald and Kambysellis 1980). The term "morphologically normal" refers to oocytes which were judged to be healthy on the basis of their gross morphology. Using standard techniques (Ursprung 1967), ovarioles were transplanted from Oregon-RC females 20-26 h after eclo- sion. Initially, only stage 8 or 9 oocytes were transplanted; however, when it was realized that these oocytes degener-

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Fig. 1. a Ovariole carrying stage 8 oocyte as the most advanced. b Implanted stage 9 oocyte-bearing ovariole recovered after 24 h in an acetone-treated isolated female abdomen host. Note degener- ated stage 9 oocyte (arrow) and development of penultimate oocyte to stage 7. c Implanted stage 8 oocyte-bearing ovariole recovered after 24 h in a ZR-515 treated isolated female abdomen host. Ter- minal oocyte has developed to stage 13, and penultimate oocyte to stage 10. Bar represents 0.1 mm

ated in JH-deficient hosts, entire ovarioles having one stage 8 or 9 oocyte as the most mature, terminal oocyte were transplanted. These ovarioles are usually termed stage 8 or 9 oocytes in this report for simplicity. The implanted ovariole usually could be recovered even if the stage 8 or 9 oocyte degenerated, which in itself could be positively determined since debris from the degenerated oocyte was seen at the posterior end of the ovariole (see Fig. 1).

Degenerating oocytes refer to those egg chambers which had one or usually several of the following morphological characteristics: opacity, an indistinct oocyte-nurse cell in- terface, the presence of one or more vacuoles, or extreme internal disorganization (Fig. 2). When ovaries were placed in a 0.2% solution of trypan blue (Harleco) in Ringer's for 30 min and then rinsed, oocytes designated as degener- ating by the above criteria usually incorporated various amounts of the dye (Fig. 3). Since trypan blue is diagnostic for cell death (Evans and Schulemann 1914), these chambers appeared to be undergoing cell death. However, trypan blue staining did not provide an unambiguous crite- rion for degenerating oocytes, since a fraction ( ~ 15%) of oocytes classified as degenerating failed to stain with trypan blue. Rarely did oocytes judged to be morphologically normal stain with trypan blue, other than the pinocytotic

uptake by vitellogenic oocytes, as expected from the results of Mahowald (1972).

Ideally, the most direct assessment of the hormonal re- quirements for maintenance and advancement of stage 8 and 9 oocytes could be made during in vitro culture; howev- er, the culture conditions have not been determined for these stages, although short-term culture has been achieved (Tedesco et al 1981). Therefore, in vivo culture was carried out using host preparations rendered deficient in JH by surgical, genetical, or chemical methods. Surgically, female abdomens were isolated within 2-4 h after eclosion prior to JH secretion from the corpus allatum located in the thorax (Handler and Postlethwait 1977), and these prepara- tions remained deficient in JH as judged by previtellogenic ovaries (Handler and Postlethwait 1977; Postlethwait and Handler 1978) and delayed larval fat body histolysis (Post- lethwait and Jones 1978). JH-deficient adults can also be produced genetically by use of the mutation ap 4, character- ized by nonvitellogenic oocyte development, which is non- autonomous (King and Bodenstein 1965) and can be rescued by application of ZR-515 (Postlethwait and Weiser 1973). Although ap 4 adults die precociously (Butterworth and King 1965; Wilson 1980), their adult lifespan is suffi- cient for utilization of this mutation in these experiments. Results from an examination of ap 4 mosaics suggested that the precocious death might be due to abnormal hemolymph (Wilson 1981). I f this is the case, ap 4 hemolymph does not seem to be toxic to ovary tissue, since ap 4 ovaries undergo apparently normal development in wild-type hosts (King and Bodenstein 1965) or in response to ZR-515 application (Postlethwait and Weiser 1973; Tedesco et al. 1981). Final- ly, JH-deficient adults can be produced by treatment of wild-type adults with the compound precocene I (Bowers and Martinez-Pardo 1977; Landers and Happ 1980). Newly eclosed adults are rendered deficient in JH for 72-96 h after treatment (Landers and Happ 1980) and were useful in the present study since a 24-48 h period of JH deficiency was required. Landers (1980) has clearly demonstrated a strong probability of JH deficiency as judged by failure of larval fat body histolysis within 30 h after eclosion (Postlethwait and Jones 1978), failure of induction of acid phosphatase in ovary tissue (Postlethwait and Gray 1975), and nonvitel- logenic oocyte development.

The JH analog ZR-515 (isopropyl ll-methoxy-3,7,11- trimethyl dodeca-2,4-dienoate) was a gift from Zoecon Cor- poration, Palo Alto, California, USA. It was dissolved in acetone at a concentration of 0.2 gg/jal and applied in a volume of 0.25 pl or 0.20 gl with a I gl microcapillary tube (Drummond Scientific) to the ventral half of the abdomen of females or males, respectively. Control host flies were treated with 0.25 gl acetone. 20-hydroxyecdysone (Sigma Biochemicals) was dissolved in Drosophila Ringer's contain- ing 5% ethanol, and 0.1-0.3 gl of a 5 x 1 0 - * M solution was injected into each fly. A dose of 10 gg/fly of precocene I was topically applied in 0.25 gl of acetone solution to the ventral abdomen in initial experiments (Table 1). Due to the significant mortality resulting from topically applied precocene as well as the time required, later experiments (Tables 2-4) utilized a method devised to treat flies with precocene incorporated into the food. In this method 5 ~tl of precocene I was dissolved in 0.1 ml of 95% ethanol and suspended in 1.8 ml H20. To this was added 0.6 g of instant Drosophila food (Carolina Biological), and the mixture was placed in an 8 dram shell vial. Since precocene-treated

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Fig. 2a-f. Ovarioles bearing normal-appearing oocytes isolated from 24-h post-eclosion Oregon-RC females, a Stage 8, b stage9, e stage 10. Ovarioles bearing degenerating oocytes which were recovered from abdomens of Oregon-RC females isolated 24 h after eclosion and maintained for 6 h after abdomen isolation, d Stage 8, e stage 9, f stage 10. Bar represents 0Amm

adults were sensitive to ether vapors, they were immobilized with ice, if possible. Also, adults did not feed on the preco- cene medium, presumably because of an unacceptable taste, but fed readily on regular medium smeared on the styro- foam plug capping the vial.

In most of the experiments Oregon-RC females were used. Highly inbred wild-type stocks have been implicated as having reduced egg maturation, presumably due to ho- mozygosity of recessive gene variants impairing oogenesis, and several investigators have used F1 hybrid females in their studies (David and Merle 1968; Allemand 1976b). In one experiment I used F 1 hybrid females issuing from a cross between Oregon-RC females and Canton-S males, two wild-type Drosophila stocks. Although the average number of stage 8, 9 and 10 oocytes were significantly dif- ferent 24 h after eclosion between these females and

Oregon-RC females (Tables 2 and 3), the difference was not striking, and no difference in their response to preco- cene could be detected.

Results

Development of Stage ~10 Oocytes Transplanted into JH-deficient Hosts

When an ovariole containing one stage 8 or 9 oocyte as the most mature, terminal oocyte was transplanted into JH-deficient females, in nearly all cases the vitellogenic oocyte failed to develop (Table 1). In many cases, the vitel- logenic oocyte was found to be degenerating 24 h after im- plantation (Fig. 1). Stage 9 implicants were always found to be degenerating, whereas stage 8 implants frequently ap- peared morphologically normal. Previtellogenic oocytes in

260

Fig. 3 a, b. Ovarioles bearing trypan blue-stained oocytes dissected from an isolated abdomen of a 24 h post-eclosion Oregon-RC female maintained for 6 h after abdomen isolation. The arrows denote the staining of degenerating oocytes of a probable stage 8 and b stage 9. Note complete exclusion of dye by previtellogenic oocytes in each ovariole

these ovarioles generally appeared healthy. Stage 10 oocytes transplanted into JH-deficient hosts developed into mor- phologically normal stage 14 oocytes (Table 1), thus corro- borating the in vitro results of Petri et al. (1979).

When JH-deficient hosts implanted with a stage 8 or 9 oocyte were treated with ZR-515, a high percentage of the implants developed at least to stage 10 and usually to stage 14 within 24 h (Table 1). This result indicates that ZR-515 can substitute either directly or indirectly for the

Table 1. Development of terminal stage 8 or 9 oocytes in ovarioles implanted into normal 24 h

missing factor(s) needed for development. Since JH is known to be involved in fat body vJtellogenin biosynthesis (Postlethwait and Handler 1978; Jowett and Postlethwait 1980, 1981 ; Postlethwait and Shirk 1981), there was a possi- bility that hemolymph vitellogenin titers were deficient in these JH-deficient hosts, resulting in the oocyte develop- mental failure. To test this possibility, isolated abdomens 4-6 h prior to implantation were injected with a concentra- tion of 20-hydroxyecdysone known to induce vitellogenin biosynthesis within several hours in fat body (Postlethwait and Handler 1978). As shown in Table 1, 20-hydroxyecdy- sone treatment failed to stimulate development of the im- planted oocytes, suggesting that neither ecdysone nor vitel- logenin was the missing factor.

In situ Development of Vitellogenic Oocytes in JH-deficient Females If stage 8 or 9 oocytes failed to develop when transplanted to JH-deficient hosts, would these oocytes develop in situ in JH-deficient females? To answer this question, I pro- duced JH deficiency in females 24 h after eclosion by abdo- men isolation or precocene treatment and followed the fate of the vitellogenic oocytes present at that time. Although the half-life of endogenous JH is unknown in Drosophila hemolymph or tissues, it seemed reasonable to assume that a 24-h waiting period after abdomen isolation or precocene treatment would allow for catabolism of endogenous hormone, based on metablic rates of exogenous JH in Dip- teran insects (Slade and Zibitt 1972; Wilson and Gilbert 1978). Table 2 shows the distribution of vitellogenic oocytes in JH-deficient females at 48 or 72 h after eclosion. The ovaries of most of these females contained stages 8 and 14 as the only vitellogenic oocytes present. Degenerating stage 8 10 (based on size) oocytes were common in these ovaries. Degenerating stage 11-14 oocytes were never ob- served, and these stage oocytes were grouped together in Tables 2 and 3 to simplify data presentation. Control females were provided not only by untreated females but also by females decapitated at 24 h after eclosion. Many of the decapitated females were alive and healthy-appearing after 48h; they remained upright and jumped when prodded. Since decapitated females neither eat nor oviposit, the presence of stages 9 13 oocytes in their ovaries at 24 and 48 h after decapitation shows that isolated abdomens fail to maintain vitellogenic oocyte development for reasons

or juvenile hormone-deficient hosts for

Host Ovarioles Ovarioles implanted recovered

Terminal ooeyte Terminal oocyte arrested in developed to at development or least stage 10 degenerated

Oregon-RC female Oregon-RC isolated abdomen Oregon-RC isolated abdomen plus ZR-5t 5 Oregon-RC female plus precocene ap4/ap 4 female ap4/ap 4 female plus ZR-515 Oregon-RC isolated abdomen plus 20-hydroxyecdysone Oregon-RC female plus 5% ethanol in Ringer's solution Oregon-RC isolated abdomen

20 18 4 14 95 78 78 0 67 60 16 44 43 31 29 2 22 17 13 4 2O 18 3 15 35 26 26 0 15 11 1 10 20 a 16 4 12 b

Ovarioles having stage 10 oocytes implanted b Development to stage 13-14

Table 2. Appearance of ovaries 24 and 48 h after various treatments of 24-h post-eclosion Oregon-RC females

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Treatment N Time Females Average number oocytes after with per female (Standard error of mean) treatment stage 9 13 Stage (h) oocytes

8 9 10 11 14

None 43 0 43 9 (0.6) 3.7 (0.4) 4.0 (0.4) 1.0 (0.2) Precocene 25 24 5 7 (0.8)* 0.4 (0.2)* 0.1 (0.1)* 5.2 (0.5) Abdomen isolated 31 24 0 3 (0.6)* 0* 0* 5.4 (0.6) Decapitated 27 24 22 10 (0.7) 3.1 (1.0) 2.6 (0.4) 7.0 (1.0) None 10 24 10 11 (1.3) 3.5 (0.7) 3.9 (0.6) 5.8 (1.0) Precocene; 23 24 16 8 (0.7) 3.0 (0.7)* 2.6 (0.7)* 5.0 (0.8)

ZR-515 at 31 h Abdomen isolated; 33 24 25 6 (0.7)* 5.3 (0.8)* 4.0 (0.7)* 7.2 (0.6)

ZR-515 at 30 h Precocene 29 48 4 4 (0.8)* 0.3 (0.2)* <0.1 (<0.i)* 5.4 (0.5) Decapitated 11 48 7 8 (0.6) 1.8 (0.6) 0.3 (0.1) 14.5 (1.2) None 10 48 10 8 (0.8) 2.5 (0.6) 2.9 (0.5) 8.1 (1.0)

* Significant difference (P<0.05) for stage 8, 9, or 10 oocytes between experimental and same-time control or between hormone treatment group and similar JH-deficient group

Table 3. Appearance of ovaries at various times after initiation of precocene treatment of 24-h post-eclosion Oregon-RC/Canton-S hybrid females

Treatment N Time Females having after > 2 degenerating treatment vitellogenic (h) oocytes/ovary

Average number oocytes per female (Standard error of mean) Stage

8 9 10 11-14

None 25 0 0 None 10 3 0 None 10 6 5 None 15 9 6 Precocene 10 3 3 Precocene 20 6 20 Precocene 20 9 20 Acetone, 10 8 t0

then precocene ZR-515, 27 8 9

then precocene ZR-515 2 h 20 7 20

> precocene

13 (0.8) 6.6 (0.6) 5.8 (0.6) 1.4 (0.3) 12 (0.9) 4.5 (0.6) 7.9 (0.7) 4.0 (0.4) /5 (1.0) 1.7 (0.8) 6.9 (0.6) 5.6 (0.9) 16 (1.3) 2.1 (0.6) 0.8 (0.3) 4.8 (0.6) 11 (l . t) 3.5 (0.8) 4.7 (/.0)* 3.9 (0.6) 10 (0.6)* 0* 1.9 (0.4)* 5.2 (0.7) 12 (1.1)* 0.1 (0.1)* 0.1 (0.1)* 6.8 (0.7) 10 (1.0) 0.1 (0.1) 0 6.5 (1.1)

17 (0.8)* 5.4 (0.5)* 2.9 (0.5)* 8.9 (0.9)

15 (0.9)* 0.5 (0.3) 0.1 (0.1) 8.3 (1.0)

* See Table 2 footnote

other than an inadequate food intake or any type of feed- back inhibi t ion of nonoviposi ted stage 14 oocytes.

An examinat ion of the stage dis tr ibut ion of vitellogenic oocytes in either isolated abdomens or precocene-treated females gave an indicat ion of the fate of stage 8-10 oocytes (Table 2). First , it appears that most of the stage 10-13 oocytes present at the time of isolat ion or precocene treat- ment developed to stage 14, as expected. Since very few eggs were laid by females exposed to precocene, and none by isolated abdomens, the number of stage 11-14 oocytes represents accumulated stage 14 oocytes at 24 and 48 h after JH treatment. Since the numbers o f accumulated mature oocytes are similar at 24 and 48 h after treatment, apparent- ly few oocytes were matured from the larger pool of stage 8 and 9 oocytes after treatment. Stage 9 oocytes were not found in JH-deficient females and, as will be subsequently shown, quickly degenerate after JH withdrawal. Stage 8 oocytes were more resistant to degeneration, a finding con- sistent with the frequent detection of morphological ly

normal stage 8 oocytes t ransplanted to JH-deficient hosts. It cannot be ruled out that stage 7 to 8 development oc- curred in these females; if this is the case, then init iat ion of vitellogen uptake required less JH than continued devel- opment of stage 8 and 9 oocytes.

When ZR-515 was applied to either isolated abdomens or precocene-treated females (~7 h later, the numbers of stage 9 and 10 oocytes found 17 18 h after hormone appli- cat ion increased significantly (P<0 .05 ) over acetone- treated controls (which produced no increase in vitellogenic oocytes over those from 24-h isolated abdomens or preco- cene-treated females), indicating that vitellogenic oocytes can develop in JH-deficient adults if ZR-515 is supplied.

In order to further characterize the degenerat ion phe- nomenon as well as understand in more detail the precocene effect, I exposed 24-h post-eclosion Oregon-RC/Canton-S hybrid females to precocene and examined their ovaries at 3-h time intervals thereafter (Table 3). A deficiency of morphological ly normal stage 8-10 oocytes was apparent

262

Table 4. Development of terminal stage 8 or 9 oocytes in ovarioles implanted into normal or juvenile hormone-deficient male hosts for 24 h

Host Ovarioles Ovarioles Terminal Terminal treatment impianted recovered oocyte oocyte

arrested in developed development to stage 10 or degenerated

None 17 15 1 14

Abdomen 22 15 15 0 isolated

Abdomen 34 29 2 27 isolated; ZR-5t5 treated

Precocene 19 18 13 5 treated

ZR-5t5, 26 24 4 20 precocene treated

at 6 h after the initial exposure, and this loss corresponded with the appearance of degenerating oocytes of stage 8, 9, and often 10 oocytes. Degenerating oocytes of these stages were also apparent in untreated control females to a lesser extent.

I f degeneration of stage 8-10 oocytes is due to JH defi- ciency, the application of ZR-515 might biock the preco- cene-induced oocyte degeneration. Table 3 shows that ZR- 515 applied immediately before precocene exposure dramat- ically blocked degeneration, either by supplying an exoge- nous source of hormone, by inhibiting the effect of preco- cene on the endogenous JH titer, or by both mechanisms. If applied 2 h after the initiation of precocene exposure, however, ZR-515 was much less effective, suggesting that precocene results in a rapid decrease in the endogenous JH titer. Stage 8 and 9 oocytes are sensitive to this decrease and begin to degenerate, which cannot be reversed with subsequent ZR-515 application.

JH Requirement for Ovary-autonomous Vitellogenesis

In Drosophila females vitellogenin is derived from both fat body and ovary, the latter due to follicle cell contribution (Brennan et al. 1982). I f previtellogenic ovaries are trans- planted into males or into females of certain other Drosophi- la species, however, the vitellogenin is derived exclusively from the implanted ovary (Srdic et al. 1979; Bownes 1980). To determine if vitellogenic oocytes engaged solely in ovary- autonomous vitellogenesis require JH for development, I transplanted stage 8 and 9 oocytes into 0-2 h post-eclosion males, then either isolated the host abdomes or exposed the hosts to either precocene or acetone within 2 h after implantation. When examined after 24 h, the implanted oocytes generally failed to develop in precocene-treated males and consistently failed in isolated abdomens (Ta- ble 4). Stage 8 and 9 oocytes transplanted into acetone- treated males underwent morphologically normal develop- ment, corroborating previous results with transplanted ovaries (Bodenstein 1947; Srdic and Jacobs-Lorena 1978). ZR-515 applied immediately before precocene exposure or after abdomen isolation increased the number of implanted

vitellogenic oocytes which developed to at least stage 10; in fact, most of the terminal oocytes developed to stage 14 and appeared morphologically normal, although par- tially collapsed.

Discussion

The experiments described here showing a requirement, ei- ther direct or indirect, of JH for continued development of stage 8 and 9 oocytes support the results of Giorgi (1979) for a JH requirement for pinocytotic uptake and incorpora- tion of horseradish peroxidase into yolk spheres. I found that stage 10 oocytes usually developed in JH-deficient hosts, consistent with other results demonstrating that stage 10 oocytes develop in vitro in the absence of JH (Petri et al. 1979). The in vitro and in vivo demonstration of stage 10 oocyte emancipation from a JH requirement for contin- ued development can be explained in several ways: a de- creased JH requirement for the waning phase of vitellogen uptake, an ability of stage 10 oocytes to complete morpho- logical development even if vitellogen uptake is prematurely terminated, or a storage of sufficient JH within stage 10 oocytes for completion of development.

At present there is conflicting evidence defining the hor- monal requirements for ovary-autonomous vitellogen up- take in Drosophila. Although it is clear that ecdysone is not involved in vitellogenin synthesis in ovaries (Jowett and Postlethwait 1980), JH has been shown to be required for ovary-autonomous vitellogenin synthesis (Jowett and Post- lethwait 1980), yet application of ZR-515 to various species of Drosophila males implanted with an immature melano- gaster ovary failed to induce oocyte maturation (Bownes 1980). The present results support a JH requirement for ovary autonomous vitellogenesis. Perhaps a factor neces- sary for ovary maturation to stage 8 was missing from the heterospecific male hosts. If this is the case, applied ZR-515 might encourage continued development of stage 8 oocytes transplanted to these hosts.

Degeneration of oocytes and subsequent oosorption has been reported in ovaries of a wide variety of other insects and can be triggered by a variety of stimuli, such as nutri- tional deficiency, virginity, or unsuitable ovipositional sites (reviewed by Bell and Bohm 1975). In several insects oocyte degeneration has been correlated with JH deficiency (High- nam etal. 1963; DeLoof and DeWilde 1970). Although it seems clear that in Drosophila degeneration of stage 8 and 9 oocytes occurs in the absence of JH, whether this phenomenon is physiologically significant is open to ques- tion. Yet there are subtle indications that Drosophila females may in part control the rate of oocyte maturation by manipulating the JH titer: 1) The finding of occasional degenerating stage 8-10 oocytes in control females (Table 3) coupled with a previous report of cell death of stage 7-8 oocytes in ovaries of wild-type females (Giorgi and Deri 1976) documents the occurrence of these oocytes, 2) it is known that maturation of stage 8-10 oocytes follows a circadian rhythm when females are maintained on a 12:12 LD cycle (Allemand 1976a), and 3) Drosophila cor- pus allatal cell nuclear volume, presumably reflecting JH secretory activity, also follows a circadian rhythm, although it is bimodal (Rensing 1964). Although oocyte degeneration as a response to a lowered JH titer seems somewhat dramat- ic as a means of lowering oocyte production, it does have advantages: a tight control over oocyte maturation and

263

a presumed recycling of materials during resorption of the degenerated oocytes.

Acknowledgements. I wish to thank Timothy Fain for the baby hair used for ligatures. Support for this research was provided by grants from the Monsanto Company and the National Science Foundation (PCM 81-12221).

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Received December 12, 1981

Accepted in revised form April 16, 1982