Megaspore wall growth inSelaginella (Lycopodiatae)

Different stages of megaspore and megasporangial development inSelaginella argentea (Wallich)Spring,S. bigelowii Unerw., andS. kraussiana (Kze.)A. Br. have been seen and studied. Megaspore wall units give positive reactions for polysaccharides and protein in young megaspores, and become the thick and resistant wall typical of the genus only later.—Units forming the exospore and the spaces between units enlarge from widths of 5–10nm early during development up to over 200 nm at pregermination stages. The spaces enlarge first. Initially they are circular and mostly about 70 nm in diameter. Later, spaces toward the inner part of the exospore enlarge more than those near the outer surface. During pregermination, wall spaces range in size from 4 to 50 times the width of units with the larger spaces located near the inner surface. As a result the exospore would be under tension to spring outward during germination when the laesurae are lysed.—A gap in the exospore, shaped like a half-moon in polar sections, forms in equatorial and distal portions of the spore. This gap becomes enormous, three times the volume of the central space plus the mesospore, and is filled with lipids and other nutrients. Late in development, during the period of tapetal cell degeneration, the gap contents are moved into the central space and the gap is closed.—Late in development the mesospore is degraded. Its products, along with gap contents, seem to be added to the contents of the central cavity and appear as reserve storage globules. A primary wall-like endospore is formed during this period, at the inner surface of the exospore. During germination this endospore develops further at its inner surface.—Changes in the size and shape of megasporangia occur independently of the size of megaspores.Megaspore development inSelaginella. II. For first part seeMorbelli & Rowley (1993).     

Oviposition schedules,survivorship curves,and mortality factors within trees of two cerambycid beetles (Coleoptera: Cerambycidae), the Japanese pine sawyer,Monochamus alternatus hope,and sugi bark borer,Semanotus japonicus lacordaire

Summary Oviposition schedules under laboratory conditions, survivorship curves, and mortality factors within trees of two cerambycid beetles, Japanese pine sawyer (JPS),Monochamus alternatus Hope, and sugi bark borer (SBB),Semanotus japonicus Lacordaire, were investigated. Average longevities of reared adults of JPS were 38.1 days for males and 42.3 days for females; those of SBB were 15.7 days for males and 23.8 days for females. It was confirmed that the JPS must feed on the pine branches for full maturation after emergence, but SBB need not; they are able to lay eggs soon after emergence. The average fecundities of JPS and SBB were 32.9 and 90.5, respectively. Thus, the JPS lay fewer eggs for a long time with continuous maturation feeding, whereas the SBB lay more eggs for short time without maturation feeding. Average survivorship curves of JPS within dead pine trees in 8 pine forests were theDeevey's B type, showing a constant mortality through the pre-imaginal stages in the trees. On the other hand those of SBB in 4 cedar stands approached theDeevey's type, suggesting that the high mortality occurred at an early stage in the trees. Average mortalities of JPS between the appearance of oviposition scars and adult emergence in 8 forests ranged from 62.3% to 95.2%. Intraspecific ompetition of JPS resulting from overcrowding in dead pine trees appeared to be main mortality factor. For SBB, as most larvae were killed by resin flow in living trees, this appeared to be the main mortality factor for this species.     

Seasonal changes in ovarian state in a eusocial halictine bee,Lasioglossum duplex,based on stages of the oldest oocytes in each ovariole (Hymenoptera: Halictidae)

Summary The changes in ovarian activity in the life cycle of the eusocial halictine beeLasioglossum (Evylaeus) duplex (Dalla Torre) was studied in both queens and workers by examining the stages of the terminal follicle in each of the six ovarioles. By this method, ovarian activities of both queens and workers were more quantitatively determined than by observation of gross ovarian features as usually conducted. Queen ovaries clearly exhibited two active periods, corresponding to the spring solitary phase and the summer eusocial phase, with distinctly greater activity in the latter. In ovaries of overwintering queens oosorption of young follicle was observed. Worker ovaries were found more active in orphan than in queenright colonies. The order of ovarian activity obtained from pooled data, summer queens>spring queens>orphan workers>queenright workers, was also recognized by comparison of individual females. Bionomics of the eusocial halictine beeLasioglossum duplex IX. Contribution No. 3107 from the Inst. Low Temp. Sci.     

Somatic embryogenesis from mature leaves of rose (Rosa sp.)

Several plant growth regulators (0.3–53.3 μm 6-benzyladenine, 2,4-dichlorophenoxyacetic acid, gibberellic acid, 3-indoleacetic acid, p-chlorophenoxyacetic acid, kinetin and α-naphthylacetic acid), alone or in combination, and culture conditions were tested for their capacity to induce somatic embryogenesis from mature leaf and stem explants of rose (Rosa sp.) of four commercial rose cultivars (Baccara, Mercedes, Ronto and Soraya). Somatic embryos were only induced from mature leaf explants derived from Soraya on Murashige and Skoog (MS) medium supplemented with 53.5 μm p-chlorophenoxyacetic acid and 4.6 μm kinetin, although satisfactory callus induction rates were obtained from all cultivars. After subculturing on the same medium, embryos at various developmental stages (globular, heart and torpedo shaped) were transferred for maturation onto a MS medium supplemented with 5.2 μm 6-benzyladenine and 5.7 μm 3-indoleacetic acid. Germination of mature embryos took place after subculturing them onto medium of the same composition. Plantlets regenerated from embryos and bearing three to four leaves were transferred to a greenhouse. Received: 4 February 1997 / Revision received: 28 August 1997 / Accepted: 1 October 1997     

The Drosophilaovarian tumorGene Is Required for the Organization of Actin Filaments during Multiple Stages in Oogenesis

Theovarian tumorgene is required during both early and late stages of oogenesis. Mutations produce a range of phenotypes, including agametic ovarioles, tumorous egg chambers, and late stage oogenic arrest. We demonstrate that each of these phenotypes is associated with specific aberrations in actin distribution. In the earliest case,ovarian tumormutations cause actin filaments to accumulate ectopically in the fusome. This correlates with abnormal fusome morphology and arrested germ cell development in the germaria. Similarly,ovarian tumorfunction is required for the localization of actin that is essential for the maturation of ring canals. This defect gives rise to tumorous egg chambers in which germ cell numbers and morphology are profoundly aberrant. We also confirm thatovarian tumoris required for the formation of the nurse cell cytoplasmic actin array that is essential for the nonspecific transport of cytoplasmic contents to the oocyte during late oogenesis. Our data suggest that at this stageovarian tumorcontrols the site where actin filaments initiate. Taken together, these studies suggest that the diverseovarian tumormutant phenotypes derive from the mislocalization of actin filaments, indicating a role for this gene in organizing the female germline cytoskeleton, and that the misregulation of actin can have profound effects on germ cell division and differentiation.     

Morphology,biology, and systematics ofCryptochlora perforans (Chlorarachniophyta), a phagotrophic marine alga

The life cycle ofCryptochlora perforans is described under culture conditions, including the following stages: Naked and walled amoebae, cysts, coccoid, palmelloid and flagellate stages. Phagotrophy of living diatoms, movement and reproduction are documented. Amoebae ofCryptochlora are chemotactically attracted by damaged algal filaments, perforate and penetrate such filaments, and subsequently engulf part of their contents. A comparison is made with a culture ofChlorarachnion reptans Geitler, according to earlier and own observations. Chloroplast structure as well as zoospore morphology confirm close affinities betweenCryptochlora andChlorarachnion. Generic characters as well as possible phylogenetic affinities with amoeboidXanthophyta andProtozoa are discussed.     

Seasonal variations in the cyanophils of the pituitary gland in correlation with reproductive cycle of Channa striatus (Bloch)

Seasonal changes in the cyanophils of the proximal pars distalis of the pituitary gland of C. striatus are conspicuous, and they closely follow the reproductive cycle. There is a poor concentration of their glycoproteinaceous contents during the resting phase and higher concentration during the maturation phase. The cyanophils also undergo a process of hypertrophy, granulation, followed by degranulation and vacuolisation during the spawning phase. They almost empty their contents during the post-spawning period.     

Ultrastructure and histochemistry of the male reproductive system of the genus Callinectes Stimpson, 1860 (Brachyura: Portunidae)

We described the ultrastructure and histochemistry of the reproductive system of five Callinectes species, and evaluate the seasonal variation in weight of the reproductive system and hepatopancreas by comparing annual changes of somatic indices. The somatic indices changed little throughout the year. In Callinectes, spermatogenesis occurs inside the lobular testes and, within each lobule, the cells are at the same developmental stage. Spermatogenesis and spermiogenesis follow the same development pattern in all Callinectes studied. Mature spermatozoa are released into the seminiferous ducts through the collecting ducts. Cells of the vas deferens are secretory as evidenced by rough endoplasmic reticulum, Golgi complex, and secretory vesicles that produce the seminal fluid. The anterior vas deferens shows two portions: proximal and distal. In proximal portion (AVDp), spermatozoa are clustered and embedded in an electron-dense, basophilic glycoproteinaceous secretion Type I. In the distal portion (AVDd), the spermatophore wall is formed by incorporation of a less electron-dense glycoproteinaceous secretion Type II. The secretion Type I change to an acid polysaccharide-rich matrix that separates the spermatophores from each other. The median vas deferens (MVD) stores the spermatophores and produces the granular glycoproteinaceous seminal fluid. The posterior vas deferens (PVD) has few spermatophores. Its epithelium has many mitochondria and the PVD seminal fluid changes into a liquid and homogeneous glycoprotein. Many outpocketings in the PVD and MVD help to increase the fluid production. Overall, the reproductive pattern of Callinectes is similar to other species that produce sperm plugs. The secretions of AVD, MVD, and PVD are responsible for the polymerization that forms the solid, waxy plug in the seminal receptacle. The traits identified here are common to all Portunidae species studied so far.     

Ovarian and fat body protein concentrations in Ips sexdentatus (Coleoptera: Scolytidae) parasitized by nematodes

The protein concentrations of fat body and ovaries in Ips sexdentatus either uninfected or infected by Parasitaphelenchus sp., P. sexdentati, or Contortylenchus diplogaster were measured at various stages of insect development, from preswarming maturation to the first oviposition (24 hr after mating). Weight variations of the fat body and ovaries in insects infected by C. diplogaster show the same evolution as those observed in uninfected insects, but at a much lower level. Fat body proteins in uninfected insects reach their minimum level during swarming, but they remain fairly constant throughout the maturation of the first egg. After dropping shortly after swarming, the ovarian protein level in such insects increases in two stages during ovarian maturation. The first stage, which corresponds to a slow protein incorporation, takes place during the first 12 hr after mating. During the second stage, i.e., beyond 12 hr, a significant level of proteins is rapidly incorporated into the ovaries. In insects infected by Parasitaphelenchus fat body proteins are reduced and protein incorporation into the ovaries is reduced; Parasitaphelenchus would thus affect at least some proteins required for ovarian maturation in their host. Fat body protein levels are even more affected by C. diplogaster than by Parasitaphelenchus, while incorporation into ovaries seems to be less affected in spite of slower ovarian growth. C. diplogaster might thus essentially act both upon proteins which are not required for the ovarian maturation of their host and upon nonproteinaceous substances that are required for such maturation. Results are discussed in relation to the possible mode of action of parasitic nematodes.     

Spawning behaviour of barred spiny eel,Macrognathus pancalus,in the River Gomti,India

The present study investigated the spawning behaviour of the spiny eel, Macrognathus pancalus, and its relationship with the environment (photoperiod and temperature). The results show that M. pancalus breeds during February to September with peak spawning during February/March and July/August, indicating two main breeding periods. Initiation of breeding occurs during increasing day lengths in early spring (February/March), and regression during decreasing day lengths in autumn (September/October). The breeding status confirmed by gonadosomatic index (GSI) and ovarian histology showed six stages of ovarian maturation (viz. stage I: chromatin nucleaolar; II: perinucleolar; III: cortical alveolar; IV: vitellogenesis; V: ripe; and VI: partially spent), with predominance of one or more stages at a particular time of the year. Males also followed a similar seasonal pattern in gonadal maturation as reflected by their GSI. Results show that M. pancalus is a seasonal breeder with bimodal spawning activity. The breeding pattern suggests an influence of prevailing day length conditions at this latitude.     

Ovarian maturation and oogenesis in the blue swimmer crab,Portunus pelagicus (Decapoda: Portunidae)

The study was aimed at understanding the process of reproduction and the changes happening in the ovary of Portunus pelagicus during maturation, which would be useful for its broodstock development for hatchery purposes. For that, tissue samples from different regions of the ovary at various stages of maturation were subjected to light and electron microscopy, and based on the changes revealed and the differences in ovarian morphology, the ovary was divided into five stages such as immature (previtellogenic oocytes), early maturing (early vitellogenic oocytes), late maturing (late vitellogenic oocytes), mature (vitellogenic oocytes), and spent (resorbing oocytes). The ovarian wall comprised of an outermost thin pavement epithelium, a middle layer of connective tissue, and an innermost layer of germinal epithelium. The oocytes matured as they moved from the centrally placed germinal zone toward the ovarian wall. The peripheral arrangement of nucleolar materials and the high incidence of cell organelles during the initial stages indicated vitellogenesis I. Movement of follicle cells toward oocytes in the early maturing stage and low incidence of mitochondria and endoplasmic reticulum in the ooplasm during late vitellogenic stage marked the commencement and end of vitellogenesis II, respectively. Yolk granules at various stages of development were seen in the ooplasm from late vitellogenic stage onwards. The spent ovary had an area with resorbing oocytes and empty follicle cells denoting the end of one reproductive cycle and another area with oogonial cells and previtellogenic oocytes indicating the beginning of the next.     

Lepidocoleus cf.ulrichi (Machaeridia) from the Trenton Group (Ordovician) of Ontario (Canada)

A specimen of the machaeridianLepidocoleus cf.ulrichi described herein focuses attention on taxonomical difficulties with extremely dorso-ventrally elongated sclerites found in some lepidocoleid taxa.Lepidocoleus ulrichi Withers, 1926 andLepidocoleus sigmoideus Withers, 1926 exhibit very similar appearances and prove indistinguishable based onWithers 1926 criteria. The key to separating taxa as similar as these may instead lie in the most juvenile stages of growth.     

A new mobergellan (small shelly fossils) from the early middle cambrian of morocco and its significance

A new mobergellan genus and species,Tateltella ranoculata, is described from the early Middle Cambrian (Agdzian Stage) of Morocco. The new taxon is characterized by only four pairs of muscle scars and is furthermore distinguished from other mobergellans by its strongly concave shape and its distally rising muscle scars. The individual specimens ofTateltella ranoculata distinctly vary in size and display different ontogenetic stages. Juvenile, intermediate, and adult stages can be distinguished by means of the development of the muscle scars that differ in Position relative to the apex, size, and distinctness between individual stages. The shell ofT. ranoculata is composed of a succession of thin phosphatic lamellae separated by interlamellar gaps, presumably originally filled by organic material. The interlamellar gaps may be divided by septum-like structures producing discrete cavities. The specimens are the youngest mobergellans known so far and correlation of their stratigraphic position suggests a correspondence with the lower part of the Amgan stage of the Siberian Platform. In addition, they are the first reported mobergellans from the present day continent Africa. Other mobergellan taxa and mobergellan-like species are briefly reviewed and the genusHippoklosma Missarzhevsky, previously assigned to the Mobergellidae, is excluded from the family due to its different shell structure. An evolutionary trend of reduction in the number of muscle scars from 14 in the early Early Cambrian to only eight in the early Middle Cambrian is apparent among mobergellans.      

Pericarp histogenesis and histochemistry during fruit development in Butia capitata (Arecaceae)

Palm fruits show great structural complexity, and in-depth studies of their development are still scarce. This work aimed to define the developmental stages of the fruit of the neotropical palm Butia capitata and to characterize the ontogenesis of its pericarp. Biometric, anatomical, and histochemical evaluations were performed on pistillate flowers and developing fruits. The whole fruit develops in three phases: (I) histogenesis (up to 42 days after anthesis – DAA), when the topographic regions of the pericarp are defined; (II) pyrene maturation (42 to 70 DAA), when the sclerified zone of the pericarp is established; and (III) mesocarp maturation (70 to 84 DAA), when reserve deposition is completed. During pericarp ontogenesis (i) the outer epidermis and the outer mesophyll of the ovary give origin to the exocarp (secretory epidermis, collenchyma, parenchyma, sclerenchyma, and vascular bundles); (ii) the median ovarian mesophyll develops into the mesocarp, with two distinct topographical regions; (iii) the inner ovarian epidermis originates the endocarp; and in the micropylar region, it differentiates into the germination pore plate, a structure that protects the embryo and controls germination. (iv) Most of the inner region of the mesocarp fuses with the endocarp and, both lignified, give rise to the stony pyrene; (v) in the other regions of the mesocarp, carbohydrates and lipids are accumulated in a parenchyma permeated with fiber and vascular bundles. The development of the B. capitata pericarp presents high complexity and a pattern not yet reported for Arecaceae, which supports the adoption of the Butia-type pyrenarium fruit class.

     

Paleolimnological reconstruction of the trophic state in Lake Balaton (Hungary) using Cladocera remains

The sediment of Lake Balaton (Hungary) provides important information about the lake’s history, particularly with regard to eutrophication. In this study, we used fossil pigment analysis and subfossil Cladocera remains preserved in a dated sediment core to identify trophic stages from ~250 bc to present. Dates of the most recent eutrophic events are in good agreement with previously published data. In general, the abundance and diversity of the Cladocera community increased with eutrophication and decreased with oligotrophication. The sediments of Lake Balaton were characterised by Chydoridae remains, of which Alona species were the most abundant. Of these, Alona quadrangularis and Alona affinis accounted for 40 and 20% of the total Cladocera remains, respectively. The trophic state of Lake Balaton varied between mesotrophic and eutrophic regimes. Seven different trophic periods were identified in Lake Balaton on the basis of Sedimentary Pigment Degradation Unit (SPDU) content of the sediment. Eutrophic states were (1) from ~250 to ~30 bc, (3) between ~300 and ~590 ad, (5) between 1834 and 1944 and (7) from the 1960s until present. Mesotrophic states were (2) ~30 bc to ~300 ad, (4) 590–1834, (6) 1944–1960s. Discriminant analysis of the cladoceran data confirmed these historic events, except for the short mesotrophic episode between 1944 and 1960. The first stage of eutrophication of Lake Balaton (~250 to ~30 bc) was characterised by extensive macrophyte vegetation, as indicated by the increasing abundance of vegetation-associated Cladocera species (Eurycercus lamellatus, Sida crystallina, Pleuroxus sp.). Intensification of eutrophication was identified since the 1980s, reflected by a high abundance of Bosmina species. The most significant planktivorous fish of Lake Balaton was the Sabre carp (Pelecus cultratus), and when its number decreased, the abundance of Bosmina species increased. This study shows that Cladocera are responsive to trophic state changes, underlining their importance as a tool for the assessment of lake eutrophication.     

Ovarian maturation of Penaeus subtilis (Decapoda: Penaeidae): A new insight to describe oocyte development and somatic structures

We observed the presence of follicular cells (FC) in the ovaries of Penaeus subtilis (n = 1198), which led us to classify the development of germ cells into six phases: oogonia, previtellogenic oocytes, primary and secondary vitellogenic oocytes, mature oocytes and atretic oocytes. The FC changes their shape according to the development of germ cells and showed a different distribution along the ovary, which allowed differentiating vitellogenic oocytes into primary and secondary. We also observed that the postovulatory follicles (POF) are composed of follicular cells. The presence of POF in penaeids ovaries is rarely reported, but allows the differentiation between spent and resting stages, commonly grouped in reproductive biology research. Furthermore, observation of ovarian lining was useful to differentiate immature females from females that had spawned at least once. Thus, ovarian development was classified into six stages: immature, early developing, advanced developing, ripe, spent and resting. The distribution and shape variations of FC, ovarian lining features and presence of POF were considered crucial for the classification of ovarian maturation stages. The methods developed here may improve estimates of their reproductive cycle, size at first maturity and spawning season, which are important variables in future studies of the reproductive dynamics.     

Reproductive dynamics of the white shrimp Litopenaeus schmitti (Burkenroad 1936) in a beach seine fishery in northeastern Brazil

The aim of the present study was to investigate the reproductive dynamics of the Litopenaeus schmitti through macroscopic and microscopic analyses. Specimens were collected monthly from December 2016 to November 2017 through artisanal beach seine fishery in the municipality of Lucena, state of Paraíba, Brazil. A total of 1710 shrimp were caught and the proportion of males was significantly lower than that of females (43.51% and 56.49%). The ovarian maturation stages were classified as immature, initial development, advanced development, mature and spent. These classification criteria added an additional stage than those previously reported for this species on the Brazilian coast. The proportion of ovarian maturation stages per month suggest a seasonal reproductive cycle that extends from September to June, with no reproductive activity occurring in July and August. The reproductive activity was directly related to the precipitation and salinity in the area of the present study. Mean cephalothorax length at first gonad maturity for females (2.20 cm) was larger than values reported 13 years ago for this species in the same area. This finding may indicate that the prohibition of motorized trawler fishing in the area since 1990 is contributing to the more sustainable exploitation of this resource.     

Ovarian and immunological responses to alternating exogenous gonadotropin regimens in the ocelot (Leopardus pardalis) and tigrina (Leopardus tigrinus)

Exogenous gonadotropins frequently are used to stimulate ovarian follicular growth and ovulation in mammalian species, including felids. However, repeated exogenous gonadotropin treatment can result in decreased ovarian responsiveness due to antibody formation. In this study, our objectives were to assess the effectiveness of alternating gonadotropin regimens on ovarian responses in ocelots and tigrinas, and investigate the humoral immune responses to these gonadotropins in each species. Females were treated four to six times with alternating equine chorionic gonadotropin (eCG)/human chorionic gonadotropin (hCG) and porcine follicle stimulating hormone (pFSH)/luteinizing hormone (pLH) regimens at 4‐month intervals. With each treatment, the females were evaluated laparoscopically to assess ovarian follicular development and recover oocytes from mature follicles. Blood was collected before each treatment and at laparoscopy. Overall, the ocelots averaged more (P<0.05) follicles and corpus luteum (CL) (6.8±0.8; mean±SEM) per stimulation than the tigrinas (2.3±0.4), but the percentage of mature oocytes (mean range=54–55%) did not differ (P<0.05). Within species, both gonadotropin regimens were equally effective (P>0.05) in inducing follicular growth and oocyte maturation. The total number of ovarian structures and oocyte maturation percentages did not decrease (P<0.05) in either species with sequential stimulations. Although the percentage of blood samples containing anti‐gonadotropin immunoglobulins increased (P<0.05) with sequential treatment, the presence of positive titers did not cause a decrease (P<0.05) in ovarian responsiveness. In summary, the female ocelots and tigrinas continued to respond to these alternating ovarian stimulation protocols after repeated use, despite the formation of anti‐gonadotropin antibodies in some of the females. These findings suggest that the use of alternating gonadotropin regimens may permit more intensive reproductive management of these endangered cat species for conservation. Zoo Biol 00:1–14, 2005. © 2005 Wiley‐Liss, Inc.     

Oogenesis and the ovarian cycle in Salamandra salamandra infraimmaculata Mertens (Amphibia; Urodela; Salamandridae) in fringe areas of the taxon's distribution

Reproductive cycle and oogenesis were studied in specimens of Salamandra salamandra infraimmaculata Mertens that inhabit fringe areas of the taxon's distribution in the Mediterranean region. Both ovarian mass and length are correlated significantly with body mass and length. Ovarian length is also correlated with the number of oocytes. During the oogenetic cycle six stages in oocyte development were recognized. Three occur during previtellogenesis: stage 1, in which oogonia divide and form cell nests; stage 2 in which oogonia differentiate into oocytes; and stage 3, in which the oocyte cytoplasm increases in volume. In the vitellogenic phase two additional stages, 4 and 5, were recognized: stage 4, in which lipid accumulates in vacuoles in the periphery followed by the appearance of yolk platelets near the cytoplasmic margin; and stage 5, in which oocyte volume increases rapidly due to increased number of yolk platelets until it reaches its maximal size. During postvitellogenesis one stage was recognized: stage 6, in which the beginning of maturation is characterized by movement of the nucleus toward the animal pole. Oogenesis continues year-round. The first four stages were seen in all ovaries examined. The ovarian cycle is independent of season and reproductive stage apart from the number of mature, postvitellogenic oocytes that increases following gestation toward the beginning of spring (March-April). J. Morphol 231:149–160, 1997. © 1997 Wiley-Liss, Inc.     

A cytochemical study on the effects of energy deprivation on autophagocytosis in Ehrlich ascites tumor cells

Summary The effect of energy deprivation on autophagocytosis in Ehrlich ascites tumor cells was studied using cytochemical techniques. Autophagocytosis was induced with vinblastine incubation (0.1 mM) and the cellular ATP-level was lowered with 2-deoxy-d-glucose (0.35 mM). Acid phosphatase was used as a marker for lysosomal enzymes and imidazole-buffered osmium tetroxide impregnation in order to study the effects of energy deprivation on the maturation of autophagic vacuole (AV) membranes.Control and vinblastine treated cells maintained their ATP-levels throughout the incubation period tested (120 min). 2-Deoxy-d-glucose alone and with vinblastine decreased the intracellular ATP-level significantly after only 3 min incubation. Most of the AV's in control and vinblastine treated cells contained degraded material and acid phosphatase activity. Their membranes were stained only slightly or not at all with imidazole-buffered osmium tetroxide. 2-Deoxy-d-glucose alone as well as with vinblastine induced in particular an accumulation of early stages of AV's. These vacuoles contained undegraded cytoplasmic material and no acid phosphatase activity and their membranes were stained usually partly with imidazole-buffered osmium tetroxide. The membranes of some early AV's resembled endoplasmic reticulum and still had attached ribosomes.It was concluded that the inhibition of cellular energy production used in the present study did not inhibit autophagic sequestration but retarded the maturation of AV membranes and impaired the functioning of lysosomal hydrolases.