The fine structure of Crithidia fasciculata with special reference to the organelles involved in the ingestion and digestion of protein

Summary As in other trypanosomatids, the cell membrane of Crithidia fasciculata overlies a single layer of microtubules. Each microtubule possesses a large number of periodically arranged drumstick-like appendages and adjacent microtubules are joined by fibrillar connectives. Anteriorly, the microtubules gradually taper to terminate just before or just after entering the reservoir. An attempt is made to correlate microtubule tapering with maintenance of form of the truncated anterior end of the cell. Smooth and coated vesicles are proliferated from the Golgi saccules and the prominent contractile vacuole lies nearby. The single mitochondrion is extensive and expanded at one point to form a capsule for the kinetoplast. The cristae are predominantly plate-like but other configurations do occur. The cytostome, a shallow invagination of the reservoir membrane, is found between two constrictions in the reservoir wall. Supporting the cytostome are several microtubules which penetrate deeply into the cytoplasm. Ingestion of ferritin occurs by pinocytosis from the cytostome and by coated vesicle formation from the reservoir membrane. Digestion probably occurs in multivesicular bodies which contain acid phosphatase activity.     

Multivesicular bodies in the transitional epithelium of the neonatal mouse urinary border.

The origin of late endosomes - multivesicular bodies (MVBs) in the superficial cells of 16 and 17 embryonic old transitional epithelium of mouse urinary bladder was studied by electron microscopy, lectin labelling and HRP tracing. Analysis of hexagonally structured membrane particles, WGA, and RCA I binding sites revealed structural similarity between plasmalemma, fusiform vesicles and multivesicular bodies. Early endosomes are lined by symmetric unit membrane as well as by asymmetric thickened membrane regions. Multivesicular bodies and fusiform vesicles have asymmetric unit membranes. MVBs may be derived from primary endosomes as well as from fusiform vesicles in the cytoplasm.     

Endocytotic pathways at the ruffled borders of rat maturation ameloblasts

Summary Using horseradish peroxidase (HRP) as a soluble protein tracer, electron microscopic studies were carried out in order to analyze endocytosis in the ruffle-ended ameloblasts of rat incisors. Accumulated HRP was initially incorporated from the ruffled border into the cytoplasm by means of pinocytotic vacuoles (pinosomes) and pinocytotic coated vesicles. The majority of the HRP was taken up by the large number of pinosomes, which then formed large endocytotic vacuoles by fusing either with each other or with preexisting endocytotic vacuoles. As time passed HRP accumulated, not in the pinosomes and ruffled border but in the endocytotic vacuoles and multivesicular bodies. Frequent connections between HRP-labeled coated vesicles and these cytoplasmic bodies indicate that these vesicles serve as an HRP carrier. These findings strongly suggest that ruffle-ended ameloblasts actively absorb soluble proteins from the enamel matrix during enamel maturation.     

Structural modulation of the surface and cytoplasm of oocytes during vitellogenesis in the lobster,Homarus americanus. An electron microscope-protein tracer study

The present investigation describes the ultrastructural changes which occur at the surface and in the cytoplasm of developing oocytes of the lobster, Homarus americanus, during vitellogenesis. The immature oocytes showed no surface specializations of the oolemma and no pinocytotic activity was observed. Horseradish peroxidase (HRP) tracer studies showed penetration of the tracer into the perivitelline space, but no uptake by the oocytes. The surfaces of oocytes examined during vitellogenesis, when yolk protein accumulation was maximal, exhibited numerous microvilli that projected into the perivitelline space, often appearing to be embedded in the follicular cell mass. In addition, the plasma membrane of vitellogenic oocytes contained many pinocytotic pits frequently situated at the bases of microvilli. The perivitelline space was engorged with electrondense material which appeared similar to that contained in pinocytotic structures of the oocytes. Vitellogenic oocytes incubated in HRP showed uptake of tracer reaction product by the coated pits and vesicles of the oolemma. Aggregation and subsequent fusion of these vesicles into large multivesicular bodies of ingested material were also observed in vitellogenic oocytes. Animals artificially induced to undergo vitellogenesis exhibited modulations of oocyte ultrastructure similar to those of normal vitellogenesis, notably, pinocytotic incorporation of extra-oocytic material and hypertrophy of oocyte surface microvilli. This study supports the hypothesis for a dual source of yolk protein in the American lobster.     

The Fine Structure of Trophozoites and Gametocytes in Plasmodium coatneyi*

SYNOPSIS. An electron microscope study of Plasmodium coatneyi in the rhesus monkey supplied information on the fine structure of trophozoites, gametocytes and of the host cell. The trophozoites resemble other mammalian malaria parasites. They do not have typical protozoan mitochondria, but instead a concentric double-membraned organelle, which, it is assumed, performs mitochondrial functions. They feed on the host cell by pinocytosis, engulfing droplets of erythrocytes thru invaginations of the plasma membranes at any region of the cell or thru the cytostome. Digestion of hemoglobin takes place in small vesicles pinched off from the food vacuole proper. Gametocytes can be clearly distinguished into macro- and microgametocytes. Macrogametocytes are covered by 2 plasma membranes, the inner one appearing thicker in some places. The cytoplasm is filled with Palade's particles and has numerous vesicles of endoplasmic reticulum and toxonemes. In microgametocytes most of the inner membrane is thickened, the cytoplasm has few Palade's particles and vesicles of the endoplasmic reticulum and does not have toxonemes. Erythrocytes with trophozoites are irregularly scallop-shaped and have elevated points with knob-like protrusions covered by a double membrane. If these protrusions are sticky they might be in part responsible for clumping and arresting the schizonts and segmenters in the capillaries. The host cell contains numerous Maurer's clefts which in some instances are continuous with the membranes of the parasite suggesting that they might originate from them.     

The Fine Structure of the Cytostome of the Apostomatous Ciliate Hyalophysa chattoni*

SYNOPSIS. The fine structure of the organelles concerned with the ingestion of exuvial fluid by the trophont of the apostome ciliate, Hyalophysa chattoni, has been examined. One of the taxonomic characteristics of the order Apostomatida is that cytostomes of ciliates within the taxon are reduced and evolving toward astomy. When examined by electron microscopy the cytostome of H. chattoni appears as a small region of active pinocytosis which is continuous with a very large cortical area, the extended cytostome. The fine structure of the extended cytostome resembles that of the cytostomes of ciliates from other orders in that it is covered by a single membrane underlain with microtubular ribs. Beneath the extended cytostome are accumulations of peculiar organelles that may represent stored membrane for recycling during food vacuole formation. Associated with the site of pinocytosis is a complex fiber that may be contractile.     

Pinocytosis in the root cells of a salt-accumulating halophyte <Emphasis Type="Italic">Suaeda altissima</Emphasis> and its possible involvement in chloride transport

Ultrastructure of root cells in salt-accumulating halophyte Suaeda altissima (L.) Pall. was examined with transmission electron microscopy. Plants were grown hydroponically on nutrient media containing 3, 50, 250, and 500 mM NaCl. Some plants were exposed to hypersomotic salt shock by an abrupt increase in NaCl concentration from 50 to 400 mM. Growing S. altissima plants at high NaCl concentrations induced the formation of type 1 pinocytotic structures in root cells. Type 1 structures appeared as pinocytotic invaginations of two membranes, the plasmalemma and tonoplast. These invaginations into vacuoles gave rise to freely ‘floating’ multivesicular bodies (MVB) enclosed by a double membrane layer. The pinocytotic invaginations and MVB contained the plasmalemma-derived vesicles and membranes of endosome origin. The hyperosmotic salt shock led to formation of type 2 and type 3 pinocytotic structures. The type 2 structures were formed as pinocytotic invaginations of the tonoplast and gave rise to MVB in vacuoles. Unlike type 1 MVB, the type 2 MVB had only one enclosing membrane, the tonoplast. The type 3 structures appeared as the plasmalemma-derived vesicles located in the periplasmic space. The cytochemical electron-microscopy method was applied to determine the intracellular Cl^? localization. This method, based on sedimentation of electron-dense AgCl granules in tissues treated with silver nitrate, showed that the pinocytotic structures of all types contain Cl^? ions. The presence of Cl^? in pinocytotic structures implies the involvement of these structures in Cl^? transport between the apoplast, cytoplasm, and the vacuole.     

Membrane cycling and macrovacuolation under the influence of cytochalasin: kinetic and morphometric studies

Fibroblasts exposed to higher doses of cytochalasin accumulate very big discrete endoplasmic vacuoles, the membrane of which is derived by internalization of plasmalemma. Morphometry confirms that the amount of surface interiorized is equal to the difference between the original cell surface area (before CD) and the reduced surface area measurable after CD-induced rounding. Correspondingly, there is a nearly two-fold increase in the activity of the ectoenzyme 5'-nucleotidase (a marker for plasma membrane) internally within the cytoplasm, after treatment with CD. Macrovacuolation increases cell volume by approximately 30%. Surface membrane is internalized as micropinocytotic vesicles at a rate measurable by the accumulation of HRP, a marker of fluid-phase pinocytosis. Uptake of HRP is shown to be enhanced at all times during exposure to CD, and is balanced by accelerated exocytic recycling of membrane except during a phase (approximately 4-8 hr) in which pinocytic uptake exceeds exocytosis. Vesicular membrane accumulated intracellularly in this period is retained in the endoplasm, and by successive fusions forms vacuoles in close approximation to microfilament aggregates. Once established, this new macrovacuolar membrane compartment is in dynamic equilibrium with the cell surface, and its membrane is cycled like the plasma membrane, in a mutual exchange of pinosomes between the several vacuoles and the cell surface. In drug-free medium vacuole membrane apparently reverts to the surface by pinocytotic recycling, and the cells recover normal characteristics 4-6 hr after withdrawal of cytochalasin.     

Protein Uptake and Digestion in Bloodstream and Culture Forms of Trypanosoma brucei*

SYNOPSIS. The mechanisms of ferritin uptake and digestion differ in bloodstream and culture forms of Trypanosoma brucei. Ferritin enters bloodstream forms from the flagellar pocket by pinocytosis in large spiny-coated vesicles. These vesicles become continuous with straight tubular extensions of a complex, mostly tubular, collecting membrane system where ferritin is concentrated. From the collecting membrane system the tracer enters large digestive vacuoles. Small spiny-coated vesicles, which never contain ferritin, are found in the Golgi region, fusing with the collecting membrane system, and around the flagellar pocket. Acid phosphatase activity is present in some small spiny-coated vesicles which may represent primary lysosomes. This enzymic activity is also found in the flagellar pocket, pinocytotic vesicles, the collecting membrane system, the Golgi (mature face), and digestive vacuoles of bloodstream forms. About 50% of the acid phosphatase activity of blood forms is latent. The remaining nonlatent activity is firmly cell-associated and probably represents activity in the flagellar pocket. The structures involved in ferritin uptake and digestion are larger and more active in the short stumpy than in the long slender bloodstream forms. The short stumpy forms also have more autophagic vacuoles. No pinocytotic large, spiny-coated vesicles or Golgi-derived, small spiny-coated vesicles are seen in culture forms. Ferritin leaves the flagellar pocket of these forms and enters small smooth cisternae located just beneath bulges in the pocket membrane. The tracer then passes through a cisternal collecting membrane network, where it is concentrated, and then into multivesicular bodies. In the culture forms, acid phosphatase activity is localized in the cisternal system, multivesicular bodies, the Golgi (mature face), and small vesicles in the Golgi and cisternal regions. The flagellar pocket has no acid phosphatase activity, and almost all the activity is latent in these forms. The culture forms do not release acid phosphatase into culture medium during 4 days growth. Uptake of ferritin by all forms is almost completely inhibited by low temperature. These differences among the long slender and short stumpy bloodstream forms and culture forms are undoubtedly adaptive and reflect different needs of the parasite in different life cycle stages.     

Antidiuretic hormone response in the amphibian urinary bladder: time course of cytochalasin-induced vacuole formation, an ultrastructural study employing ruthenium red

Cytochalasin is known to inhibit the antidiuretic hormone-induced hydro-osmotic response (bulk water flow) in the amphibian urinary bladder without altering hormone-stimulated diffusional water permeability or short-circuit current. In addition, histological studies have shown that the mold metabolite induces the formation of large intracellular vacuoles or lakes in the epithelial cells. We report here a transmission electron microscopic time-course study which indicates that during the early phases of the ADH response cytochalasin causes the formation of numerous multivesicular bodies or aggregates derived from individual basolateral pinocytotic vesicles. Because of their apparent hypertonic nature, the vesicles, as well as the vesicular aggregates, accumulate water during hormone-stimulated hydro-osmotic flow. As a result, the multivesicular bodies dilate and fuse to form the large intracellular lakes characteristic of cytochalasin treatment in the presence of both an applied osmotic gradient and vasopressin. In the presence of mucosal ruthenium red, the luminal glycocalyx was heavily stained with this tracer. At no time, however, even in the presence of hormone, was there any evidence for the uptake of this dye at the apical epithelial border. In the presence of serosal ruthenium red, the lateral intercellular spaces, basolateral pinocytotic vesicles, basal lamina, and collagen, as well as other subepithelial structures, were ruthenium positive. With cytochalasin D, vasopressin, and serosal ruthenium red, both the pinocytotic vesicles and the multivesicular bodies demonstrated an apparent membrane associated ruthenium positive coat. The tracer data indicates that the basolateral pinocytotic vesicles, increased by the presence of hormone, are indeed endocytotic in nature. The mucopolysaccharide coat associated with these structures may be involved in ionic and/or fluid transport.     

THE FEEDING MECHANISM OF AVIAN MALARIAL PARASITES

Electron microscope studies of the erythrocytic forms, including gametocytes and asexual schizonts, of the protozoa Plasmodium fallax, P. lophurae, and P. cathemerium, have revealed a "cytostome," a specialized organelle of the pellicular membrane which is active in the ingestion of host cell cytoplasm. In material fixed in glutaraldehyde and postfixed in OsO₄, the cytostome appears in face view as a pore limited by two dense circular membranes and having an inside diameter of approximately 190 mµ. In cross-section, the cytostome is a cavity bounded on each side by two dense segments corresponding to the two dense circles observed in face view; its base consists of a single unit membrane. In the process of feeding, the cytostome cavity enlarges by expansion of its membrane, permitting a large quantity of red cell cytoplasm to come into contact with the cytostome wall. Subsequent digestion of erythrocyte cytoplasm occurs exclusively in food vacuoles which emanate from the cytostome invagination. As digestion progresses, the food vacuoles initially stain more densely and there is a marked build-up of hemozoin granules. In the final stage of digestion, a single membrane surrounds a cluster of residual pigment particles and very little of the original host cell cytoplasm remains. The cytostome in exoerythrocytic stages of P. fallax has been observed only in merozoites and does not seem to play the same role in the feeding mechanism.     

Endocytotic pathways at the ruffled borders of rat maturation ameloblasts

Using horseradish peroxidase (HRP) as a soluble protein tracer, electron microscopic studies were carried out in order to analyze endocytosis in the ruffle-ended ameloblasts of rat incisors. Accumulated HRP was initially incorporated from the ruffled border into the cytoplasm by means of pinocytic vacuoles ( pinosomes ) and pinocytotic coated vesicles. The majority of the HRP was taken up by the large number of pinosomes , which then formed large endocytotic vacuoles by fusing either with each other or with preexisting endocytotic vacuoles. As time passed HRP accumulated, not in the pinosomes and ruffled border but in the endocytotic vacuoles and multivesicular bodies. Frequent connections between HRP-labeled coated vesicles and these cytoplasmic bodies indicate that these vesicles serve as an HRP carrier. These findings strongly suggest that ruffle-ended ameloblasts actively absorb soluble proteins from the enamel matrix during enamel maturation.     

Membrane production and yolk degradation in the early fly embryo (Calliphora erythrocephala meig.): An ultrastructural analysis

Formation of nuclear envelopes during the last cleavage mitosis and the formation of the cell membranes during the cellularization of the blastoderm have been studied ultrastructurally in the blowfly egg. Dense bodies arising from yolk granules by budding could contain membrane material destined to be incorporated into the new membranes of the blastoderm. The presence of transitional structures indicates that these bodies can be converted into dark multivesicular bodies. Large amounts of endoplasmic reticulum are found around the mitotic nuclei. Clusters or branched chains of vesicles associated with this are interpreted as evidence for the formation of endoplasmic reticulum by the breakdown of dark multivesicular bodies. Nuclear envelopes of mitotic daughter nuclei probably originate from endoplasmic reticulum. The egg contains both intranuclear and extranuclear annulate lamellae. The main events of cytokinesis are furrow initiation and cell membrane growth during the slow first phase, but probably only cytokinetic movement during the rapid second phase. On the assumption that cell membrane growth occurs by incorporation of complete membrane pieces, the addition of coated vesicles and/or light multivesicular bodies is definitely most probable. Some intermediate profiles indicate that light and dark multivesicular bodies are related. The membrane needed for second phase cytokinesis could well be provided by the unfolding of surface microvilli and protuberances of the furrow canal.     

THE DISTRIBUTION OF EXOGENOUS FERRITIN IN TOAD SPINAL GANGLIA AND THE MECHANISM OF ITS UPTAKE BY NEURONS

Toad spinal ganglion cells are individually enclosed in sheaths consisting of one or more attenuated layers of satellite cell cytoplasm surrounded externally by a basement membrane. Narrow (~150 A) extracellular channels separate these layers from one another and from the underlying neuron. In both in vivo and in vitro experiments it was found that molecules of ferritin, a water-soluble protein, are to some extent able to pass across the basement membrane and through these channels to reach the neuronal plasma membrane. Ferritin particles arriving at the neuronal surface are engulfed by the neuron in 0.1 to 0.2 µ "coated" vesicles. The concentration of ferritin in these vesicles is higher than in the perineuronal space. The ferritin incorporated into the neuron is segregated, apparently intact, in multivesicular bodies. It is inferred that the 150A channels in the satellite cell sheath are patent, aqueous spaces through which molecules with a diameter as large as 95 A are able to pass, and that these neurons are capable of taking up whole protein from their immediate environment by the process of pinocytosis.     

Evidence for both prelysosomal and lysosomal intermediates in endocytic pathways

Horseradish peroxidase (HRP), an enzyme internalized by fluid phase pinocytosis, has been used to study the process by which pinosome contents are delivered to lysosomes in Chinese hamster ovary cells. Pinosome contents were labeled by allowing cells to internalize HRP for 3-5 min. Following various chase times, cells were either processed for HRP and acid phosphatase (AcPase) cytochemistry or homogenized and fractionated in Percoll gradients. In Percoll gradients, pinosomes labeled by a 3-5 min HRP pulse behaved as a vesicle population more dense than plasma membrane and less dense than lysosomes. In pulse- chase experiments, internalized HRP was chased rapidly (3-6 min chase) to a density position intermediate between the "initial" pinocytic vesicle population and lysosomes. With longer chase periods, a progressive accumulation of HRP in more dense vesicles was observed. Correspondence between the HRP distribution and lysosomal marker distribution was reached after a approximately 1-h chase. By electron microscope cytochemistry of intact cells, the predominant class of HRP- positive vesicles after pulse uptakes or a 3-min chase period was characterized by a peripheral rim of reaction product and was AcPase negative. After 10-120-min chase periods, the predominant class of HRP- positive vesicles was characterized by luminal deposits and HRP activity was frequently observed in multivesicular bodies. HRP-positive vesicles after a 10- or 30-min chase were AcPase-positive. No HRP activity was detected in Golgi apparatus. Together these observations indicate that progressive processing of vesicular components of the vacuolar apparatus occurs at both a prelysosomal and lysosomal stage.     

The cellular uptake of horseradish peroxidase and its poly(lysine) conjugate by cultured fibroblasts is qualitatively similar despite a 900-fold difference in rate

When horseradish peroxidase (HRP) is conjugated to poly(L-lysine) of molecular weight (MW) 13,000, its transport into cultured L929 fibroblasts in 1 hour at 37 degrees C is increased 918-fold. The kinetics of uptake are linear with time and concentration, reflecting a process of nonreceptor-mediated adsorptive endocytosis. Neither HRP-poly(Lys) conjugate nor free poly(Lys) of any size cause any increase in fluid phase endocytosis. All evidence indicates that the covalently bound poly(Lys) increases the binding of HRP to the cell surface and that this adequately accounts for an increased internalization occurring at a steady rate. In the absence of Ca++, both surface binding and uptake of the conjugate, but not of free HRP, are decreased. Cytochalasin B does not significantly inhibit the transport of either form of HRP. The half-lives of HRP and HRP-poly(Lys) are 7.6 and 5.6 hours, respectively, when measured over a period of 12 hours. Electron microscopic analysis of cells that have ingested comparable amounts of HRP shows that the intracellular localization of free and conjugated HRP are comparable and that both are found inside the same array of structures. Thus HRP, which binds very poorly to the cell surface and is considered a fluid-phase marker for the "contents" of pinocytotic vesicles, and HRP-poly(Lys), which binds very strongly to the cell surface and is considered a membrane marker for adsorptive endocytosis, are taken up along the same endocytotic pathway. Moreover, despite the fact that neither markers are transported by receptor-mediated endocytosis, both are seen in structures that were described as receptosomes. It appears, therefore, that the pathways utilized in receptor-mediated transport are available to all other forms of pinocytosis.     

Effect of Metabolic Inhibitors on Pinocytosis of Uranyl lons by Radish Root Cells: Probable Mechanisms of Pinocytosis

Pinocytotic uptake of uranyl ions by root cells of the radishRaphanus sativus L. cv. Red Giant and effects of monoiodacetate(MIA), 2,4-dinitrophenol (DNP), 2-mercaptoethanol (ME) and cytochalasinB (CB) on this process were studied. The number of invaginationsand pinocytotic vesicles in cells incubated with 0.15 mM uranylacctate (UA) is 15–20 times greater than in control cells,as estimated by counting the structures (plasmalemma derivatives)in serial sections. Monoiodacetate (0.05 mM) slightly stimulatedand ME (1.5 mM) completely inhibited pinocytosis. DNP (0.1 mM)inhibits UA pinocytosis by 45 per cent: DNP combined with MIAdiminishes pinocytotic activity by nearly 70 per cent. In thepresence of CB (4 µM) and UA quite large invaginations(exceeding 1 µm) have been observed. Cells treated withUA and UA $ MIA contain considerably more secretory vesicles.The results provide evidence for two types of pinocytosis inradish root cells; one independent of metabolic energy (I) andthe other essentially dependent on energy from respiration (II).Experiments with liposomes which show a pinocytosis-like behaviourin the presence of the pinocytosis inductors (including UA)indicate that type I is due solely to the effect exerted bythe inductor on the membrane. Type II is probably directly dependenton metabolism and may be regarded as a combination of uptakeand of sui generis removal of any excess plasmalemma increasedby exocytotic secretion. Raphanus sativus, pinocytosis, exocytosis, metabolic inhibitors, liposomes, electrostatic interaction, lipid peroxidation     

PINOCYTOSIS IN ACANTHAMOEBA CASTELLANII : Kinetics and Morphology

The uptake of radioactively labeled albumin, inulin, leucine, and glucose by Acanthamoeba castellanii (Neff strain) was measured. The uptake is linear with time and appears to be continuous under the conditions of these experiments. Uptake is abolished at 0°C. No evidence for saturation of the uptake mechanism was obtained with either albumin or leucine. Each of the four tracer molecules enters the ameba at a similar rate when the uptake is calculated as volume of fluid ingested per unit time. The data suggest that each of these molecules enters the cell by pinocytosis. The highest rate of uptake was obtained with cells in their usual culture medium containing proteose peptone, glucose, and salts but pinocytosis also continued at a reduced rate in a simple salt solution. The calculated volume of fluid taken in during pinocytosis in culture medium was about 2 µl/hr per 10⁶ cells. The route of uptake was examined in the electron microscope using horseradish peroxidase (HRP) as a tracer. HRP activity was found exclusively within membrane profiles within the cytoplasm, confirming the pinocytotic mode of uptake. An estimate of the rate of surface membrane turnover due to pinocytosis was made using the biochemical and morphological data obtained. This estimate suggests that the plasma membrane turnover of one cell is on the order of several times an hour.     

AN ELECTRON MICROSCOPIC STUDY OF PINOCYTOSIS IN AMEBA : II. The Cytoplasmic Uptake Phase

This report is devoted principally to a consideration of the fate of the pinocytotic vacuole and its content in the ameba Pelomyxa carolinensis (Chaos chaos). High resolution micrographs of the plasmalemma have shown it to consist of three layers, i.e., an outermost filamentiferous zone, a middle homogeneous zone, and an inner zone which appears to be a unit membrane. The three zones can be identified in the membranes lining the pinoyctotic tunnels and vacuoles of amebas fixed shortly after pinocytosis occurred. The first apparent change in the pinocytotic vacuole is an increase in the surface-to-volume ratio which occurs during the 1st hour of its existence. Within 24 hours the marker substance commonly collects in defecation vacuoles which can be identified by the profiles of bacteria usually found in the lumen. Occasionally, however, thorotrast can be seen in the lumen of the contractile vacuole. The thorotrast appears to enter the two excretory organelles by the coalescence of vesicular fragments of the pinocytotic vacuoles with the limiting membranes of the excretory organelles.     

Endocytosis in the uterine luminal and glandular epithelial cells of mice during early pregnancy

We have localized horseradish peroxidase (HRP) in the mouse uterus after intravenous administration on days 1 and 5 of pregnancy in an effort to understand how serum proteins reach the uterine lumen. Direct movement of HRP into uterine and glandular lumina was blocked by the epithelial tight junctions on both days. In luminal and glandular epithelial cells at both times, HRP was localized in endocytic vesicles along the basolateral membranes, multivesicular bodies (mvb), elongated dense bodies below the nucleus (bdb), and many small vesicles near the apical surface of the cells. The uptake of HRP was most extensive in the luminal epithelium on day 1: the number of tracer-containing apical vesicles and bdb was largest, and there were also clusters of vesicles containing the tracer above the nucleus. Acid phosphatase was localized on day 1 in mvb and bdb in both cell types, indicating that these structures are lysosomes. It appeared that HRP followed two pathways after basolateral endocytosis by the epithelial cells: it was transported to the apical region of the cells, where it was present in small vesicles that may release their contents into the uterine or glandular lumina, or it was transported to lysosomes. To investigate whether macromolecules may be transported from the uterine lumen to the stroma, we also studied endocytosis at the apical pole of luminal epithelial cells after intraluminal injection of HRP. There was no detectable uptake of HRP from the lumen on day 1, and no tracer was detected in the intercellular spaces or basement membrane region. On day 5, a large amount of HRP was taken up from the lumen into apical endocytic vesicles, mvb, and dense bodies, but tracer was not present in the Golgi apparatus, lateral intercellular spaces, or the basement membrane region at the times studied. These observations indicate that there was no transport of luminal macromolecules to the uterine stroma on day 1, while the possibility of transport on day 5 requires further study.