Structure and possible function(s) of charasomes; complex plasmalemma-cell wall elaborations present in some characean species

Summary Charasomes, complex membrane structures, were found along the longitudinal walls of internodal and lateral branch cells ofChara corallina andC. braunii, but not along their transverse walls or in other cell types. Charasome-complexes were larger and more numerous in the lateral branch cells than in internodal cells. InC. corallina, a dioecious species, especially large elaboration of charasome material occurs in the lateral branch cells of the female plant, sometimes reaching a cross-sectional width which is as great as that of the adjacent cell wall. Chara internodes transport hydroxyl (OH^–) out of the cell and bicarbonate (HCO₃ ^–) into the cell. Spatial distribution of charasomes along the cell was examined with respect to these transport phenomena, which occur at specific identifiable regions along the cell. Charasome-complexes were always found in regions in which HCO₃ ^– transport occurs but were often fewer, reduced in size or absent in areas of OH^– efflux.Nitella flexilis exhibited similar patterns of OH^– and HCO₃ ^– transport along the cell; however, there was a complete absence of charasomes. Ultrastructural examinations onNitella translucens indicated that charasomes were also absent in this species. The observation that charasomes are present in both transport regions ofChara but are totally lacking in the twoNitella spp. indicates that the charasome-complex is not involved in transport of either substance. Other possible functions for the charasomes, including a role in osmoregulation, are discussed.Charasome substructure is the same in bothChara species, consisting of a mass of short (50 nm average length) anastomosing tubules (30 nm average diameter) derived from the plasmalemma. The interior of the tubules is open to the cytoplasm while the area surrounding the tubules is ultimately open to the wall and thus can be considered to be wall space. Charasomes are quite variable in size and shape, but are roughly globular, with the bulk of the structure projecting into the cell cytoplasm. Tubular components of the charasome were sometimes seen to extend into the microfibrillar wall matrix. A three dimensional model of the charasome-complex presented details the great complexity of this membrane system.     

Influence of culture medium pH on charasome development and chloride transport inChara corallina

Summary Internodal cells ofChara, grown in culture either at pH 5.7, 6.5 or 7.5, were studied to determine their chloride influx capability, the quantitative aspects of charasome morphology and the degree to which these two parameters could be correlated. In cells grown at pH 5.7 the charasomes were relatively small, were widely spaced on the plasma membrane, and contributed only a 0.6% increase to the surface area of the plasma membrane in the acid region of the cell. In contrast, the charasome membrane surface area of cells grown at pH 7.5 had increased × 19, the density of charasomes on the cell surface increased × 42, thus producing a × 3.57 increase in the acid region plasma membrane surface area. Chloride influx in cells grown at pH 7.5 was × 8.7–12.7 greater than in cells grown at pH 5.7. Cells that had been starved of chloride exhibited a × 2.4 average increase in the rate of chloride influx. Our observations establish the existence of a positive correlation between the rate of chloride influx and the increase in membrane surface area due to charasomes, although other factors, such as the effect of pH on transport-related enzymes, and the effect of charasome structure on chemical equilibria, may also be of importance.     

The relationship of the charasome to chloride uptake in Chara corallina: physiological and histochemical investigations

A possible role of the charasome in terms of chloride transport into Chara corallina Klein ex. Willd., em. R.D.W. is examined. The branches of Chara contain the most charasome material and are shown to be very effective in acquiring Cl^- to support continued shoot growth. The early maturation of the branches, the rather large Cl^- fluxes into these cells, and their ability of translocate Cl^- to growing cells of the shoot indicate a special role of these branches in Cl^- accumulation. The structure of the charasome, with its extensive periplasmic space, appears especially suited as a site for H⁺–Cl^- cotransport (influx). We show, by histochemical assay, that the charasomes of mature cells contain ATPase activity; such activity is absent in growing charasomes of very young cells. ATPase activity is also associated with the plasmodesmata of C. corallina. Charasome ATPase activity and Cl^- uptake are both inhibited by p-chloromercuribenzenesulfonic acid (1 mM) or diethylstibestrol (40 M; 45 min). The anion transport inhibitor, 4,4-diisothiocyano-2,2-disulfonic acid stilbene (1 mM) had no effect on Cl^- transport and inhibited ATPase activity only when applied after chemical fixation of the cells. Results of an attempt to demonstrate the presence of Cl^- within the cytoplasmic tubules of the charasome, using a silver precipitation technique, proved difficult to interpret because of a reaction between the silver and a cellular substance produced in the light.Abbreviations CPW Chara pond water - DES diethylstilbestrol - DIDS 4,4-diisothiocyano-2,2-disulfonic acid stilbene - Mops 3-(N-morpholino)propanesulfonic acid - PCMBS p-chloromercuribenzenesulfonic acid     

Charasomes are not essential for photosynthetic utilization of exogenous HCO3 − inChara corallina

Summary Cells ofChara corallina grown under high CO₂ culture conditions were able to utilize exogenous HCO₃ ^– to give appreciable rates of net photosynthesis. Since these rates of photosynthesis could be detected within 10 min of being transferred from high-CO₂ to normal HCO₃ ^– (pH 8.2) culture conditions, it would appear that the HCO₃ ^–-accumulating system ofChara is not fully repressed under these high CO₂ culture conditions. The membrane potential of these cells also responded to light/dark treatments in a manner consistent with the operation of a HCO₃ ^– acquisition system. With prolonged exposure (2–6 days) to CPW/B, net photosynthesis continued to increase towards the expected control rate and, in parallel, the electrical responses elicited by light/dark treatments converged towards those obtained on control (CPW/B-grown)Chara cells. Charasomes were absent in CPW/CO₂-grownChara, but redeveloped in mature cells once the culture was returned to CPW/B conditions; a minimum period of 7 days in CPW/B was required before charasomes were detected in tissue examined in the transmission electron microscope. As the above-detailed physiological and electrophysiological features were observed with both axial and whorl cells ofChara in which charasomes were completely absent, we conclude that this specialized organelle is not an essential component for photosynthetic utilization of exogenous HCO₃ ^– in this species.Abbreviations CPW/B Chara pond water containing 1.0 mM NaHCO₃, pH8.2 - CPW/CO₂ Chara pond water containing dissolved CO₂, pH 5.5 - DIC dissolved in organic carbon - D.H. dark-induced membrane hyperpolarization - L.H. light-induced membrane hyperpolarization - TEM transmission electron microscopy     

Inversion of extracellular current and axial voltage profile inChara andNitella

Summary Reducing the pH of the bathing solution from 8.2 to pH 6 can induce an inversion of the extracellular current pattern that develops at the surface ofChara corallina internodal cells. A similar result can be obtained on some cells by changing the medium to a pH value of 10. In noninvertingChara cells the currents were strongly reduced when the pH value of the medium was changed between 3 and 11. Simultaneous measurements of theChara transmembrane potential in the acid and alkaline regions revealed that a light-induced electrical potential gradient of approximately 24 mV was present in the axial (or longitudinal) direction. Correlated to the external current pattern inversion was an inversion of this internal longitudinal voltage gradient. Reillumination ofNitella cells, after a period of darkness, often resulted in a complete inversion of the extracellular current pattern. These results are discussed in terms of spatial and temporal control of membrane transport processes, and in particular the control of current loops that pass through these cells.     

Electron tomographic characterization of a vacuolar reticulum and of six vesicle types that occupy different cytoplasmic domains in the apex of tip-growing <Emphasis Type="Italic">Chara</Emphasis> rhizoids

We provide a 3D ultrastructural analysis of the membrane systems involved in tip growth of rhizoids of the green alga Chara. Electron tomography of cells preserved by high-pressure freeze fixation has enabled us to distinguish six different types of vesicles in the apical cytoplasm where the tip growth machinery is accommodated. The vesicle types are: dark and light secretory vesicles, plasma membrane-associated clathrin-coated vesicles (PM-CCVs), Spitzenkoerper-associated clathrin-coated vesicles (Sp-CCVs) and coated vesicles (Sp-CVs), and microvesicles. Each of these vesicle types exhibits a distinct distribution pattern, which provides insights into their possible function for tip growth. The PM-CCVs are confined to the cytoplasm adjacent to the apical plasma membrane. Within this space they are arranged in clusters often surrounding tubular plasma membrane invaginations from which CCVs bud. This suggests that endocytosis and membrane recycling are locally confined to specialized apical endocytosis sites. In contrast, exocytosis of secretory vesicles occurs over the entire membrane area of the apical dome. The Sp-CCVs and the Sp-CVs are associated with the aggregate of endoplasmic reticulum membranes in the center of the growth-organizing Spitzenkoerper complex. Here, Sp-CCVs are seen to bud from undefined tubular membranes. The subapical region of rhizoids contains a vacuolar reticulum that extends along the longitudinal cell axis and consists of large, vesicle-like segments interconnected by thin tubular domains. The tubular domains are encompassed by thin filamentous structures resembling dynamin spirals which could drive peristaltic movements of the vacuolar reticulum similar to those observed in fungal hyphae. The vacuolar reticulum appears to serve as a lytic compartment into which multivesicular bodies deliver their internal vesicles for molecular recycling and degradation. Electronic supplementary material The online version of this article (doi:) contains supplementary material, which is available to authorized users.     

Broad-range effects of ionophore X-537A on pollen tubes of Lilium longiflorum

The effects of the broad-range cationophore X-537A on pollen tubes of Lilium longiflorum were investigated, using both light and electron microscopy. Pollen tube growth is completely inhibited within 30 min after the application of 5·10^-5 M ionophore X-537A; cytoplasmic streaming is stopped only after 60 min of ionophore treatment. Ultrastructurally, X-537A effects are a vacuolation of Golgi cisternae and a general vacuolation. The wall is thickened at the very tip. Coated vesicles and coated regions are enriched close to and at the plasma membrane. The results indicate that pollen tube tip growth needs a specific ion distribution.Abbreviations CTC chlorotetracycline - DMSO dimethylsulfoxide     

The ultrastructural characteristics of the apical cell in the neuroepithelial bodies of the toad lung (Bufo marinus)

Summary The cytological features and membrane specialisations of neuroepithelial cells (apical cells) in direct contact with the lumen of the lung were studied with transmission and scanning electron microscopy. The luminal surface of the apical cell is characterised by microvilli, a cilium with an 8+1 microtubular pattern and numerous coated vesicles. The cytoplasmic region immediately beneath the luminal plasma membrane contains numerous smooth-walled vesicles, tubules and microtubules, a few microfilaments and dense granules (15–20 nm in diameter). The luminal pole of the cell is marked off from the basal or vascular pole by a well-defined terminal web associated with junctional complexes. Protrusion of the luminal pole occurs as a transient phenomenon and is accompanied by a pinching in of the cell at the terminal web. It is proposed that the distinctive features of the luminal pole of the apical cell are comparable to those of recognised chemoreceptor cells. It is also proposed that in view of the common features of apical and basal cells the apical cell functions as a receptor/transducer and the basal cells serve as an accessory source of peptides/5-hydroxytryptamine to be released on stimulation of the apical cell. Furthermore, we have drawn attention to the structural heterogeneity of the neuroepithelial bodies in various vertebrate classes.     

The plasma membrane of young Chara internodal cells revealed by rapid freezing

Young elongating internodal cells of Chara globularis var. capillacea (Thuill.) Zanev. were rapidly frozen and freze-fractured in order to observed transient events occurring within the plasma membrane. Several structures have been observed. Relatively small depressions, varying in depth, are prolific and scattered at random over the plasma membrane. Charasomes and clusters of particle rosettes are common. Arrays of intramembrane particle lines are a characteristic feature of the internodal cell plasma membrane. The charasomes and the arrays of particle lines occupy a considerable proportion of the plasma membrane. In these young cells, substantial movement must take place across this membrane and its basic structure must fluctuate accordingly. The innumerable small depressions may represent pinocytotic and secretory processes. The array of intramembrane particle lines may represent stages in fusion between the membranes of vesicles within the cytoplasm and the plasma membrane. The technique of ultra-rapid freezing allows these events and their intermediate stages to be visualised; some features of the membrane may only be seen by this method.     

Plasma membrane coat and a coated vesicle-associated reticulum of membranes: their structure and possible interrelationship in Chara corallina

Primary fixation with buffered glutaraldehyde plus 2.0 mM CaCl2 and 0.1% tannic acid results in the preservation of certain portions of the plasma membrane coat of Chara when seen with the electron microscope. Such a coat is not observable after fixation with glutaraldehyde alone. The coat appears to be present on all the above ground, vegetative cells of the male plant. Within complex invaginations of the plasma membrane, which are known as charasomes, the coat has two structural components, a central core that is either tubular or solid and a fibrous or granular peripheral region that surrounds the core. The coat material appears to be at least partially derived, via exocytosis, from the contents of single membrane-bound organelles known as glycosomes. Glycosomes seem to originate from within an assemblage of membranes and coated vesicles that can be described, in purely structural terms, as a partially coated reticulum. Such a reticulum is distinguishable from Golgi stacks because the reticulum (a) is not composed of stacked membranes, (b) is extensively involved with large, clearly detailed coated vesicles and coated invaginations, (c) is closely associated with glycosomes, and (d) is only slightly stained by the zinc-iodide- osmium tetraoxide reagent.     

Electron microscopic studies of coated membranes in two types of gill epithelial cells of lamprey

Summary Coated membranes in two types of gill epithelial cell of adult lamprey, Lampetra japonica, were studied by electron microscopy. The type 3 gill epithelial cells possess well-developed microvilli or microfolds, apical vesicles and abundant mitochondria. The cytoplasmic surface of the microvillous plasma membrane is covered by a coat of regularly spaced particles with a center-to-center distance of about 15 nm. Each particle consists of a bulbous free end, about 10 nm in diameter, and a connecting piece, about 5 nm long. Apical vesicles are covered by a surface coat which consists of fine filamentous material but lack any special coating on their cytoplasmic surface.The type 4 cells (chloride cells) are characterized by apical vesicles, abundant mitochondria and cytoplasmic tubules. These tubules possess a coat on their luminal surface which consists of spirally wound parallel rows of electron-dense materials. The rows are about 16 nm apart and wound at a pitch of about 45°. The cytoplasmic surface of these tubules does not display a special coat. These coated membranes are assumed to be the sites of active ion transport across the plasma membrane. In particular, particles in type 3 cells and linear coat materials in chloride cells may be either loci of transport enzymes or energy generating systems. Apical vesicles lack any coating on their cytoplasmic surface but a fine filamentous coat is present on their luminal surface. They contain intraluminal vesicles and are continuous with apical ends of cytoplasmic tubules.     

Plasma membrane domains participate in pH banding of Chara internodal cells

We investigated the identity and distribution of cortical domains, stained by the endocytic marker FM 1-43, in branchlet internodal cells of the characean green algae Chara corallina and Chara braunii. Co-labeling with NBD C(6)-sphingomyelin, a plasma membrane dye, which is not internalized, confirmed their location in the plasma membrane, and co-labelling with the fluorescent pH indicator Lysotracker red indicated an acidic environment. The plasma membrane domains co-localized with the distribution of an antibody against a proton-translocating ATPase, and electron microscopic data confirmed their identity with elaborate plasma membrane invaginations known as charasomes. The average size and the distribution pattern of charasomes correlated with the pH banding pattern of the cell. Charasomes were larger and more frequent at the acidic regions than at the alkaline bands, indicating that they are involved in outward-directed proton transport. Inhibition of photosynthesis by DCMU prevented charasome formation, and incubation in pH buffers resulted in smaller, homogenously distributed charasomes irrespective of whether the pH was clamped at 5.5 or 8.5. These data indicate that the differential size and distribution of charasomes is not due to differences in external pH but reflects active, photosynthesis-dependent pH banding. The fact that pH banding recovered within several minutes in unbuffered medium, however, confirms that pH banding is also possible in cells with evenly distributed charasomes or without charasomes. Cortical mitochondria were also larger and more abundant at the acid bands, and their intimate association with charasomes and chloroplasts suggests an involvement in carbon uptake and photorespiration.     

Aspects ultrastructuraux des relations entre les couches folliculaires et l'ovocyte depuis la formation du follicule jusqu'au début de la vitellogenèse chez le Lézard Lacerta vivipara Jacquin

Résumé Des contacts étroits s'établissent d'abord entre la granulosa et la membrane plasmique de l'ovocyte. L'aire pellucide les sépare ensuite. L'ovocyte émet de nombreuses microvillosités digitiformes et des processus intenses de micropinocytose se produisent au niveau des invaginations ovocytaires. Des granules provenant de la thèque externe, passent dans les espaces intercellulaires de la granulosa, se répandent dans l'aire pellucide et envahissent les invaginations de l'ovocyte. Des vésicules tapissées, différenciées à partir de la membrane plasmique de ces invaginations, captent des granules et s'isolent dans le cytoplasme; elles perdent leur revêtement externe et donnent naissance à des sphères vitellines corticales. Le rôle attribué jusqu'alors aux cellules piriformes n'est pas confirmé. Le grand développement de l'appareil de Golgi dans ces cellules peut indiquer une fonction sécrétoire qui serait à l'origine du matériel amorphe périovocytaire.

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Ultrastructural aspects of the relationships between follicular layers and oocyte from the formation of the follicle to the beginning of vitellogenesis in the lizard Lacerta vivipara Jacquin

Summary First the granulosa cells establish close contact with the plasma membrane of the oocyte. Then, the zona pellucida separates them. The oocyte acquires numerous microvillosities, and an intensely active micropinocytosis is observed in the ovocytic invaginations. Granules originating in the theca externa pass through the intercellular spaces of the granulosa and invade the zona pellucida and the invaginations of the oocyte. At the level of these invaginations, coated vesicles incorporate granules and form granular vesicles without coating. These granular vesicles are transformed into cortical vitelline spheres. The nutritive function, generally assigned to the pyriform cells, is not confirmed in this study. The great development of the Golgi apparatus in these cells suggests a secretory function. Perhaps the pyriform cells secrete the amorphous component of the periovocytic space.

Avec la collaboration technique de Madame M. Hubert.     

Immunolocalization of myosin in rhizoids ofChara globularis Thuill

Summary Myosin-related proteins have been localized immunocytochemically in gravity-sensing rhizoids of the green algaChara globularis using a monoclonal antibody against the heavy chain of myosin from mouse 3T3 cells and a polyclonal antibody to bovine skeletal and smooth muscle myosin. In the basal zone of the rhizoids which contain a large vacuole, streaming endoplasm and stationary cortical cytoplasm, the monoclonal antibody stained myosin-related proteins as diffusely fluorescing endoplasmic strands. This pattern is similar to the arrangement of subcortical actin filament bundles. In the apical zone which contains an aggregation of ER membranes and secretory vesicles for tip growth, diffuse immunofluorescence was detected; the intensity of the signal increasing towards the apical cell wall. The most prominent myosin-staining was associated with the surface of statoliths in the apical zone. The polyclonal antibody produced a punctate staining pattern in the basal zone, caused by myosin-related proteins associated with the surface of drganelles in the streaming endoplasm and the periphery of the nucleus. In the apical zone, this antibody revealed myosin-immunofluorescence on the surface of statoliths in methacrylate-embedded rhizoids. Neither antibody revealed myosin-immunofluorescence on the surface of organelles and vesicles in the relatively stationary cytoplasm of the subapical zone. These results indicate (i) that different classes of myosin are involved in the various transport processes inChara rhizoids; (ii) that cytoplasmic streaming in rhizoids is driven by actomyosin, corresponding to the findings onChara internodal cells; (iii) that actindependent control of statolith position and active movement is mediated by myosin-related proteins associated with the statolith surfaces; and (iv) that myosin-related proteins are involved in the process of tip growth.     

Secondarily-forming Coated Vesicles in Ray Parenchyma Cells of Pinus radiata Needle Traces

Coated vesicles in ray parenchyma cells of Pinus radiata needletraces were observed to form secondarily from smooth vesicles.The nascent coated vesicles were seen as small protrusions ofthe bounding membrane of smooth vesicles. Initially they wereellipsoidal or dome shaped, later becoming spheroidal. Althougha coat was observable at the spheroidal stage the coat was mostpronounced in vesicles that had separated from parent smoothvesicles. Coated vesicles were most common near dictyosomeswhere they were often seen in stages of formation from smoothvesicles and in the cortical cytoplasm near the plasmalemma.The plasmalemma developed small protrusions some of which appearedcoated at the cytoplasmic face. Coated vesicles, Pinus radiata, needle trace, ray parenchyma     

Distribution of isoasparagine among different characean species

Thirteen species of Characeae were analyzed for their free amino acid contents. Large amounts of isoasparagine, accounting for 10 to 50% of the total free amino acids, were found in extracts fromChara (5 species including one unidentified),Nitellopsis (1 species), andLamprothamnium (1 species). In contrast, no isoasparagine was detected inNitella (5 species) andTolypella (1 species), except forN. flexilis in which as much as 40% of the free amino acids was isoasparagine. Other major amino acids found in the tested materials were Ala, Asp, Glu and Gln.     

Artificial control of cytoplasmic pH and its bearing on cytoplasmic streaming,electrogenesis and excitability of characeae cells

Effects of changing the cytoplasmic pH on the cytoplasmic streaming, membrane potential and membrane excitability were studied in tonoplast-free cells ofChara australis andNitellopsis obtusa. The cytoplasmic pH was varied by internal perfusion of pH-buffered media.Nitellopsis cells were perfused only once, whileChara cells were perfused twice to control the pH more accurately. In both materials the rate of cytoplasmic streaming was maximum at about pH 7, low at pH 8.5–9 and almost zero at pH 5–5.5. The membrane potential was most negative at about pH 7. InChara the membrane potential supported by Mg·ATP was strongly inhibited at pH 5.5, and almost zero at pH 9, supporting the results obtained by Fujiiet al. (1979) on cells ofChara australis which were perfused once. The action potential could be induced by electrical stimulation inChara at pH 6.0–9.0 and inNitellopsis at pH 6.6–7.9. The membrane resistance ofNitellopsis was high at acidic and neutral pH values and low at alkaline pH, while that ofChara was low at both acidic and alkaline pH values.     

Ultrastructure of chytridiomycete and oomycete zoospores using spray-freeze fixation

Summary The ultrastructure of zoospores of several zoosporic fungi was examined using a modified cryofixation technique. An atomizer was used to spray a zoospore suspension into the cold propane reservoir of a conventional plunge freeze-substitution apparatus. Spray-freeze fixation and freeze-substitution of zoospores porvided better fixation of vacuolar structures, membranes and the extracellular coat than that obtained with chemical fixation. The overall shape of cryofixed spores was closer to that seen in living zoospores. Two types of vacuoles were seen in cryofixed zoospores ofMonoblepharella andChytridium. One type of vacuole contained electron-opaque material within the lumen while the other type had no visible internal material in the lumen and appeared to be part of the water expulsion vacuole complex. Coated pits and coated vesicles were observed associated with both the water expulsion vacuoles and the plasma membrane inMonoblepharella andPhytophthora, suggesting that endocytosis of the plasma membrane and expulsion vacuoles is part of membrane recycling during osmoregulatory events. An extracellular coat was seen on the outer surface of cryofixed zoospores ofMonoblepharella sp.,Chytridium confervae andPhytophthora palmivora without the use of carbohydrate-specific stains. The spray-freeze method gave good and reproducible fixation of the wall-less spores in quantities greater than those obtained in previously described zoospore cryofixation studies. The technique is potentially useful for cell suspensions in that freeze damage from excess water is limited.Abbreviations ddH₂O deionized distilled water - PME Pipes/MgCl2/EGTA buffer - WEV water expulsion vacuole     

Presence of a partially-coated reticulum in angiosperms

Summary The form and distribution of partially-coated membrane systems and coated vesicles in the leaf glands ofPhaseolus and the root cortical cells ofZea were investigated. Partially-coated membranes exist as a reticulum which is either sparsely branched or extensively anastomosed. Coated vesicles and coated membrane regions may occur at all points that are in the vicinity of such a reticulum. Golgi stack membranes, though associated with the partially-coated reticulum in many cases, show no consistent orientation to it. Occasionally, the reticulum and the associated coated vesicles are located away from any Golgi stack. We examined other plant tissues from such species asAllium cepa, Beta vulgaris andNicotiana tabacum and the partially-coated reticulum was observed in all material. We suggest that membrane flow may be occurring from the partially-coated reticulum to either the Golgi stack or to other intracellular compartments.     

The fine structure of the carapace integument of Daphnia magna straus (Crustacea branchiopoda)

Summary The structure of the two integumental layers comprising the carapace of female D. magna was examined at several points through the molt cycle. The epicuticle and procuticle are simple in organisation; pore canals are absent but intracuticular fibres are present, forming complexes with invaginations of the epidermal plasma membrane similar to such complexes described in the literature for other arthropods. The epidermis consists almost entirely of cuticle-secreting cells. Secretion of the new cuticle begins when 50–67% of the instar has elapsed by which time the epidermal cells have increased in height and their nuclei have become more rounded. However, other presumed secretory phenomena observed viz. the formation of dense core vesicles by Golgi bodies, and the occurrence of these and coated vesicles near the apical plasma membrane are not restricted to any particular period during the molt cycle. This suggests that the mechanisms of cuticle secretion do not undergo marked changes in activity as they do in decapods; presumably this relative continuity is related to the much shorter molt cycle of cladocerans.The technical assistance of G.A. Bance, and the financial support provided by the National Research Council of Canada are gratefully acknowledged