FUNCTIONS OF COATED VESICLES DURING PROTEIN ABSORPTION IN THE RAT VAS DEFERENS

The role of coated vesicles during the absorption of horseradish peroxidase was investigated in the epithelium of the rat vas deferens by electron microscopy and cytochemistry. Peroxidase was introduced into the vas lumen in vivo. Tissue was excised at selected intervals, fixed in formaldehyde-glutaraldehyde, sectioned without freezing, incubated in Karnovsky's medium, postfixed in OsO₄, and processed for electron microscopy. Some controls and peroxidase-perfused specimens were incubated with TPP,¹ GP, and CMP. Attention was focused on the Golgi complex, apical multivesicular bodies, and two populations of coated vesicles; large (> 1000 A) ones concentrated in the apical cytoplasm and small (<750 A) ones found primarily in the Golgi region. 10 min after peroxidase injection, the tracer is found adhering to the surface plasmalemma, concentrated in bristle-coated invaginations, and within large coated vesicles. After 20–45 min, it is present in large smooth vesicles, apical multivesicular bodies, and dense bodies. Peroxidase is not seen in small coated vesicles at any interval. Counts of small coated vesicles reveal that during peroxidase absorption they first increase in number in the Golgi region and later, in the apical cytoplasm. In both control and peroxidase-perfused specimens incubated with TPP, reaction product is seen in several Golgi cisternae and in small coated vesicles in the Golgi region. With GP, reaction product is seen in one to two Golgi cisternae, multivesicular bodies, dense bodies, and small coated vesicles present in the Golgi region or near multivesicular bodies. The results demonstrate that (a) this epithelium functions in the absorption of protein from the duct lumen, (b) large coated vesicles serve as heterophagosomes to transport absorbed protein to lysosomes, and (c) some small coated vesicles serve as primary lysosomes to transport hydrolytic enzymes from the Golgi complex to multivesicular bodies.     

Plasma membrane-associated vesicles in retinal capillaries of the rat

The structure and function of abluminal vesicles in endothelial cells of rat retinal capillaries was examined using glutaraldehyde-tannic acid fixation and the hemeproteins--horseradish peroxidase, microperoxidase, and lactoperoxidase--as tracers. Numerous vesicles, delimited by a tannic acid-positive membrane, were distributed along the abluminal front. Other vesicles were arranged in clusters and chains or tubule-like structures. Such vesicles were not found in the vicinity of the capillary lumen. When the retina was exposed to hemeproteins, either in vitro or after intravitreal injection, the abluminal vesicles became labeled with tracer reaction product. Apparently "free" vesicles and tubules seen in tangential sections through the basal lamina were also labeled, suggesting that they were in continuity with the plasma membrane in another plane of section. No enzyme reaction product was present in the capillary lumen. Peroxidase-positive multivesicular bodies were observed, suggesting that some protein was endocytosed and directed to lysosomes where it was presumably degraded. The results suggest that abluminal endothelial vesicles represent pits or invaginations of the plasma membrane and, as such, are not involved in the transendothelial transport of protein from the perivascular space to the capillary lumen. Tannic acid treatment revealed a population of similar vesicles associated with the plasma membrane of pericytes. After exposure to hemeproteins, enzyme reaction product was localized in these vesicles and in a few multivesicular bodies. The results suggest that the majority of these vesicles are in continuity with the plasma membrane and are not involved in endocytosis.     

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.     

Protein transport to the epididymal lumen

Summary Experiments were performed to clarify the debate over the entry of circulating proteins into the epididymal lumen by use of the marker horseradish peroxidase (HRP). Epididymal tubules from the caput epididymidis of the rat were immersed in medium TC 199 containing HRP (3.5 mg/ ml) for 5 min to 3 h at 33° C. Sections were examined for the presence of tracer within the epithelial cells by electron microscopy. From 5 min to 3 h, vesicles containing peroxidase reaction products were found throughout the cytoplasm of the principal cells. Vesicles occurred close to both the basal and apical membranes, and many were found opening into the interstitial space and lumen, depending on the length of incubation. By 5 min labelled vesicles were infrequently found in the apical part of the cells. Reaction product was observed in the epididymal lumen adhering to the microvilli from 30 min of incubation onwards. At all periods of incubation peroxidase was present at the base of the epithelium and between the cells, but it was never found within the tight junctional complexes, and no reaction deposits were found within epithelial cells of tubules incubated in the absence of peroxidase. It is concluded that large molecules leaving the capillaries may enter the epididymal lumen in the caput by means of fluid-phase endocytosis.     

Endocytosis in the epithelium of the mouse oviduct

We studied the pathway of serum protein transport into the lumen of the mouse oviduct by localizing several tracer proteins in the oviduct after intravenous injection on days 1, 5, and 11 of pregnancy. Fluorescent proteins were observed in the lamina propria and in vesicles in the lumenal epithelial cells mainly in the preampulla segment on days 5 and 11 of pregnancy. In the isthmus, there was much less fluorescence in the lamina propria and no fluorescent vesicles in lumenal epithelial cells. This is similar to previous observations on day 1 and indicates that the uptake of serum proteins into lumenal epithelial cells in the preampulla is not limited to the time when embryos are present in the oviductal lumen. Horseradish peroxidase (HRP) was present in the lamina propria of the preampulla on days 1 and 5, but direct tracer movement into the oviductal lumen was blocked by the epithelial junctional complexes. Within the epithelial cells, HRP was localized in endocytic vesicles along the basolateral membrane, multivesicular bodies (mvb), elongated dense bodies below the nucleus (bdb), and many small vesicles near the apical surface of the cells. Ferritin was also used as a tracer and was observed in the same locations as HRP. Acid phosphatase in the epithelial cells of the preampulla on day 1 was localized in mvb and bdb, indicating that these structures are lysosomes. It appeared that HRP and ferritin followed two pathways after basolateral endocytosis by the epithelial cells in the preampulla: 1) they were transported to apical vesicles that may release their contents into the oviductal lumen, or 2) they were transported to lysosomes.(ABSTRACT TRUNCATED AT 250 WORDS)     

ELECTRON MICROSCOPY OF ABSORPTION OF TRACER MATERIALS BY TOAD URINARY BLADDER EPITHELIUM

The absorption of Thorotrast and saccharated iron oxide by the epithelium of the toad urinary bladder was studied by electron microscopy. Whether the toads were hydrated, dehydrated, or given Pitressin, no significant differences in transport of colloidal particles by epithelial cells were observed. This implies that these physiological factors had little effect on the transport of the tracer particles. Tracer particles were encountered in three types of epithelial cells which line the bladder lumen, but most frequently in the mitochondria-rich cells. Tracer materials were incorporated into the cytoplasm of epithelial cells after being adsorbed to the coating layer covering the luminal surface of the cells. In the intermediate stage (1 to 3 hours after introducing tracer) particles were present in small vesicles, tubules, and multivesicular bodies. In the later stages (up to 65 hours), the particles were more commonly seen to be densely packed within large membrane-bounded bodies which were often found near the Golgi region. These large bodies probably were formed by the fusion of small vesicles. Irrespective of the stages of absorption, no particles were found in the intercellular spaces or in the submucosa. Particles apparently did not penetrate the intercellular spaces of the epithelium beyond the level of the tight junction.     

Ultrastructure of the perfused rat epididymis: Effect of luminal sodium ion concentration

Summary The appearance of the rat epididymal epithelium changed when it was perfused in vivo through the lumen with unphysiologically high sodium ion concentrations; dilatation of intercellular spaces (ICS) at threshold concentrations of 30mM-Na⁺ in the cauda and about 55mM-Na⁺ in the corpus was associated with absorption of water from the lumen. Despite the distended ICS, junctional complexes appeared intact, and their integrity was confirmed by the exclusion of luminal horseradish peroxidase (HRP) from the ICS, and by demonstrating that circulating [³H]inulin did not enter the lumen. Smooth ER and lipid droplets in the principal cells of the corpus epididymidis were well maintained, and the preservation of granular ER in principal cells of the cauda epididymidis lent morphological support to the continued secretion of protein in this segment. However, occasional distension or involution of inner Golgi cisternae was evident in principal cells after 3–6 h perfusion. In contrast to multivesicular bodies of principal cells, the apical and basal vacuoles characteristic of clear cells changed in size with different perfusing solutions. When low Na⁺ concentrations were perfused large translucent vacuoles were frequently found in the apical cytoplasm of clear cells in the corpus and cauda epididymidis, and filled vacuoles became larger and showed a decrease in content density in the cauda epididymidis. These large vacuoles were absent from tissue perfused with high Na⁺ concentrations. Normal pinocytotic activity of both cell types was demonstrated by perfusing HRP which was taken up by the normal route in principal cells, with some transfer to the Golgi cisternae. By far the most HRP was accumulated in clear cell vacuoles irrespective of the composition of the perfusing solution.     

Uptake of horseradish peroxidase from CSF into the choroid plexus of the rat,with special reference to transepithelial transport

Summary Protein uptake from cerebral ventricles into the epithelium of the choroid plexus, and transport across the epithelium were studied ultrastructurally in rats. Horseradish peroxidase (HRP, MW 40,000) was used as protein tracer. Steady-state ventriculo-cisternal perfusion with subatmospheric pressure (-10cm of water) in the ventricular system was applied. HRP dissolved in artificial CSF was perfused from the lateral ventricles to cisterna magna for various times, and ventriculo-cisternal perfusion, vascular perfusion or immersion fixation with a formaldehyde-glutaraldehyde solution was performed.Coated micropinocytic vesicles containing HRP were seen both connected with the apical, lateral and basal epithelial surface and within the cells. Heavily HRP-labeled vesicles were often fused with the lining membrane of slightly labeled or unlabeled intercellular spaces. Since the apical tight junctions of the epithelium never appeared open or never contained HRP in the spaces between the fusion points, and since the intercellular spaces between adjacent epithelial cells below the junctions only infrequently contained tracer after 5 min, by increasing amounts after 15–60 min of HRP perfusion, a vesicular transport of HRP from the apical epithelial surface to the intercellular spaces, bypassing the tight junctions, is suggested.In addition to the transepithelial transport, micropinocytic vesicles also transported HRP to the lysosomal apparatus of the epithelial cells. With increasing length of exposure to HRP, a sequence of HRP-labeled structures could be evaluated, from slightly labeled apical vacuoles and multivesicular bodies to very heavily labeled dense bodies.     

Role of epithelial clear cells of the rat epididymis in the disposal of the contents of cytoplasmic droplets detached from spermatozoa

Upon release from the seminiferous epithelium, spermatoza show a small droplet of cytoplasm attached to the neck region. During transit of spermatozoa in the caput epididymidis, this cytoplasmic droplet migrates along the middle piece of the flagellum. In the corpus epididymidis, the droplet shows a lateral displacement, while in the cauda epididymidis it detaches from the spermatozoon. In the electron microscope, cytoplasmic droplets attached to spermatozoa were seen to contain numerous, short, straight or C-shaped, flattened membranous elements referred to as lamellae, small vesicles, and small particles (35-nm diameter) with a diffuse wall showing no apparent unit membrane. The lamellae were stacked closely on one another or arranged in a loose array. Structurally as well as cytochemically, with different cytochemical markers, the lamellae and vesicular elements failed to show any evidence of being components of the Golgi apparatus or elements of the endoplasmic reticulum. The lamellae, vesicular elements, and 35-nm particles were also seen free in the lumen of the corpus epididymidis but were especially prominent in the cauda epididymidis at a time when droplets were being released from spermatozoa. The lumen of the epididymis, as spermatozoa passed from the caput to the cauda epididymidis, was also noted to acquire progressively a flocculent background material. The epididymal epithelium is composed predominantly of principal and clear cells. The endocytic activity of clear cells was examined in rats at different time intervals after a single injection of cationic ferritin into the lumen of the cauda epididymidis. At 2 min the tracer was bound to the microvilli of these cells and was also observed within large coated and uncoated pits, subsurface coated vesicles, and numerous subsurface small uncoated vesicular membranous elements (150-200-nm diameter). At 5 min, in addition to the above structures, the tracer was present in endosomes, while at 15 and 30 min, pale and dense multivesicular bodies appeared labeled, respectively. At 1 and 2 hr, but more so at 6 hr large dense membrane-bound bodies identified cytochemically as secondary lysosomes became labeled. All of the above endocytic structures were also seen to contain the 35-nm particles, flattened or vesicular membranous profiles, and a fine flocculent background material reminiscent of those seen free in the lumen or found in cytoplasmic droplets attached to spermatozoa. (ABSTRACT TRUNCATED AT 400 WORDS)     

Uptake and fate of luminally administered horseradish peroxidase in resting and isoproterenol-stimulated rat parotid acinar cells

The formation and fate of apical endocytic vesicles in resting and isoproterenol-stimulated rat parotid acinar cells were studied using luminally administered horseradish peroxidase (HRP) to mark the vesicles. The tracer was taken up from the lumen by endocytosis in small, smooth-surfaces "c"- or ring-shaped vesicles. About 1 h after HRP administration the vesicles could be found adjacent to the Golgi apparatus. At later times HRP reaction product was localized in multivesicular bodies and lysosomes; in isoproterenol-stimulated cells it was also present in autophagic vacuoles. HRP reaction product was never localized in any structure associated with secretory granule formation. These results suggest that the apical endocytic vesicles play a role in membrane recovery, but that they are degraded and not reutilized directly in secretory granule formation. Additionally, it was found that when isoproterenol was injected before HRP administration, the apical junctional complexes became permeable to the tracer, allowing it to gain access to the lateral and basal intercellular spaces. This permeability may provide an additional route whereby substances in the extracellular fluid could reach the saliva.     

Receptor-mediated and adsorptive endocytosis by male germ cells of different mammalian species

Summary The routes for adsorptive and receptor-mediated endocytosis were studied in vivo after microinjection of tracers into the lumen of the seminiferous tubules, and in vitro in isolated germ cells of different mammals. Cationic ferritin was located on the plasma membrane, in vesicles, in tubules, in multivesicular bodies and in lysosome-like granules of mouse spermatocytes. In these cells the number of multivesicular bodies varied during spermatogenesis. Spermatids and to a lesser extent residual bodies also performed adsorptive endocytosis. In the rat and monkey (Macaca fascicularis) diferric transferrin was specifically taken up by germ cells via receptor-mediated endocytosis. The labelling was observed subsequently in membrane pits, vesicles, endosome-like bodies and pale multivesicular bodies. A progressive decrease in the frequency of the labelling of the germ cells by transferrin-gold particles was observed from spermatogonia to spermatocytes and to early spermatids, which could indicate that iron is particularly required by germ cells during the mitotic and meiotic processes. Adsorptive and receptor-mediated endocytosis therefore occurs in all classes of germ cells. These endocytic processes are most probably required for germ cell division, differentiation and metabolism.     

Ultrastructure of langur monkey epididymidis prior to and following vasectomy and vasovasostomy.

The ultrastructural changes in langur monkey epididymis prior to and following vasectomy or vasovasostomy were studied. The epididymal epithelium of the intact langur monkey was found to consist mainly of principal cells and basal cells and frequently apical or mitochondria rich cells were found. Besides these cells intraepithelial lymphocytes were also a consistent feature of the epididymal epithelium. Principal cells identified by means of the tuft of the stereocilia on their apical surface, bear well developed Golgi bodies, endoplasmic reticulum, vesicles, vacuoles and multivesicular bodies. This suggests their active involvement in absorption and secretion. Basal cells present at the base of the lamina bear a few cellular organelles and strong interdigitations with the adjacent cells. Apical or mitochondria rich cells were characterized by clusters of mitochondria in the apical region of the cell and few microvilli on their apical surface. Lymphocytes with a large nucleus and a pale rim of cytoplasm were also found at the base of the epithelium. Secretory and absorptive functions of principal cells of the epididymal epithelium were found to be increased after vasectomy, as indicated by bulging of the apical portion of the principal cells and membrane bound structure in the lumen. An extensive increase in the number of lysosomes, vesicles and vacuoles was also observed. An increase in the number of macrophages with spermatozoa remnants in the lumen of epididymis suggests that the principal mechanism for spermatozoa disposal following vasectomy is intraluminal endocytosis by macrophages. Changes following vasectomy persisted in vasovasostomized animals even after 12 months of recanalization, which may contribute to the failure of functional reanastomosis.     

Cytochemical and ultrastructural characteristics of the lining epithelium of the distal part of the epididymis in hamsters (Mesocricetus auratus)

In the terminal segment of the hamster epididymidis there was some evidence of micro-merocrine protein secretion a the level of the principal cells and clear evidence of granular secretion in the light cells, presumable of glycoproteins. The PAS and protein cytochemistry reactivities observed in both these cells, of the ductus epithelial lining, but especially in the light cells, are suggestive of mucopolysaccharides and protein complexes synthesis and secretion. This secretion is carried out to the epididymal epithelium from the lumen and luminal content. A complex of small vacuoles and vesicles appeared to form from the Golgi complex is showed in the principal cells. It was suggested that this complex may represented merocrine secretory vacuoles and vesicles in these cells. Dense granules, at the TEM level, are observed in all the cytoplasm of the light cells, with correspondence to similar PAS-positive granules observed in these cells, at the light microscope level. These granules, at the TEM level, are actually secreted to the epididymal duct lumen, by the apical cytoplasms of the light cells. Signs of absorption were suggested to the principal and light columnar cells through the ultrastructural observations of micropinocytosis, apical multivesicular bodies or great membrane-bounded vacuoles in the adluminal cytoplasms.     

ENDOCYTOSIS DURING POSTNATAL DIFFERENTIATION IN SUPERFICIAL CELLS OF THE MOUSE URINARY BLADDER EPITHELIUM

This electron microscopical study was performed in order to follow the endocytic pathway of horseradish peroxidase and colloidal gold tracers and to determine the involvement of endocytosis in postnatal differentiation in superficial cells of the mouse urinary bladder epithelium. Morphometric analyses of late endosomes/multivesicular bodies from day of birth to day 25 were performed. The internalisation and intracellular transport of luminal plasmalemma to multivesicular bodies via endocytic vesicles, early endosomes and pleomorphic compartments was established. Dynamic changes in endocytic activity took place within the first few days of postnatal differentiation. During this period the number of multivesicular bodies changed in an inverse ratio to their size. After the third day endocytic activity gradually approached the low rate of adult urothelium.     

Vascular permeability (problem of the blood-brain barrier) in the pineal organ of the rainbow trout,Salmo gairdneri

Summary The problem of the blood-brain barrier in the pineal organ of the rainbow trout, Salmo gairdneri, was investigated following intraperitoneal or intracardial injections of several tracers and dyes with different molecular weights. As demonstrated at the light-microscopic level, repeated injections of trypan blue or horseradish peroxidase (HRP) resulted in an accumulation of these substances in the pineal epithelium (parenchyma). By use of the electron microscope, HRP was found in electron-dense bodies, probably lysosomes, in (i) the endothelial cells and perivascular macrophages 4 h after intraperitoneal injection, (ii) the supporting cells and intrapineal or luminal macrophages 8 h after injection, and (iii) the receptor cells 24 h after injection of the tracer. Ferritin particles penetrated the fenestrated endothelium of pineal capillaries. They were confined to vesicles, vacuoles and the smooth endoplasmic reticulum of the supporting cells as well as to the synaptic vesicles and the smooth endoplasmic reticulum of the pineal photoreceptors. The intercellular passage of tannic acid mixed with the fixative was blocked at the luminal junctional complex separating the pineal lumen from the basal portion of the pineal epithelium. The passive intercellular transport of substances with high molecular weight from the bloodstream to the cerebrospinal-fluid compartment is thus prevented. However, no blood-brain barrier exists for exogenously administered proteins, which are rapidly taken up by pineal cells and actively transported in a transcellular manner.The findings on the blood-brain barrier of the pineal organ of the rainbow trout are discussed with particular reference to the endocrine capacity of pineal sensory organs.Fellow of the Alexander von Humboldt Foundation, Federal Republic of Germany.     

Lamellar bodies are the intracellular site of membrane turnover in insect fat body

Analysis of the time course of highly cationic ferritin uptake by fat body cells has shown that the tracer bound to the plasma membrane and was pinocytosed by coated vesicles. The first sites of intracellular accumulation were multivesicular bodies which became filled with ferritin between 30-60 min after cells were exposed to the tracer. At no time during the experiments were any parts of the Golgi complex labeled by the tracer. By 60 min, the ferritin was increasingly found in lamellar bodies. The different types of 'light' and 'dark' multivesicular bodies suggest that lamellar bodies form from multivesicular bodies as they fill with tracer. The occurrence of lamellar bodies in many different cell types suggests an important role in membrane dynamics.     

Directional flow of sensillum liquor in blowfly (Phormia regina) labellar chemoreceptors.

Utilizing horseradish peroxidase as a tracer protein, it is shown that trichogen and tormogen cells have a secretory function. Protein tracer from the haemolymph enters these cells by endocytosis and is transported to the sensillum liquor cavity by transport vacuoles and multivesicular bodies. It is also suggested that closely associated pigment cells may be involved in macromolecular transport.     

Morphology of the epithelium of the extratesticular rete testis, ductuli efferentes and ductus epididymidis of the adult male rabbit

The fine structure of the epithelium lining the extratesticular rete testis, ductuli efferentes and ductus epididymidis of the rabbit has been investigated. In the ductuli efferentes the epithelium is composed of two cell types, principal cells and ciliated cells. The latter type is distinguished from principal cells by the presence of cilia projecting into the lumen and the position of the nucleus in the apical half of the cell. Principal cells in this segment are characterized by micropinocytotic vesicles on the surface plasma membrane and a variety of small dense bodies scattered throughout the cytoplasm. In the ductus epididymidis basal cells replace ciliated cells as the second cell type, but differences between various segments of the epididymis are related to the fine structure of the principal cells. In the proximal caput epididymidis (Nicander's region 1) the principal cells are tall with long microvilli. They typically contain a small Golgi apparatus and a cluster of dense bodies adjacent to the nucleus. In the distal caput epididymidis (Nicander's regions 2-5) the apical cytoplasm of principal cells is filled with numerous micropinocytotic vesicles and large multivesicular bodies; these features are interpreted as signs of absorptive activity. The multivesicular bodies are absent from the cytoplasm of principal cells in the corpus epididymidis (Nicander's region 6) and, instead, numerous elements of smooth endoplasmic reticulum, a large Golgi apparatus, lipid droplets and dense bodies characterize principal cells in this segment. Towards the proximal cauda epididymidis (Nicander's region 7), the number of dense bodies (lysosomes) in the cytoplasm increases considerably. In the globose cauda (Nicander's region 8), the principal cells are reduced in height, and in addition to the features described in region 7, are characterized by a concentric array of rough endoplasmic reticulum in the basal cytoplasm. These observations are discussed in relation to the role of the epididymis in promoting the maturation and survival of spermatozoa.     

Endocytotic activity in the endometrium during conceptus attachment in the cow

The uptake of horseradish peroxidase tracer injected into the uterine lumen of the cow was studied during the period of conceptus attachment (Days 18-21; Day 0 = oestrus) and also in cyclic animals. Endocytosis occurred in pregnant and non-pregnant cows but was especially marked when circulating progesterone concentrations were high. By 20 min after injection, the tracer was located in apical endocytotic vesicles and in organelles of the lysosomal system. In addition, some of the horseradish peroxidase-containing vesicles were associated with the lateral membranes of the cells and the tracer was also present in the intercellular spaces and beneath the basal membrane, especially in pregnant animals by the time of conceptus attachment. There was no evidence that pinopod-like functions could be attributed to large cytoplasmic protrusions from endometrial cells. Rather, the protrusions seemed to be involved in secretory processes. The presence of clear vesicles among the endocytotic vesicles suggested a coupled secretory-endocytotic activity of the cells, the significance of which remains to be determined.     

SECRETION AND ENDOCYTOSIS IN INSULIN-STIMULATED RAT ADRENAL MEDULLA CELLS

Insulin was used to deplete the adrenalin stores of rat adrenal medulla cells. Release of secretion was observed to occur by exocytosis. In addition, during the stages of massive release of secretory granules, the insulin-treated preparations showed greatly enhanced endocytic uptake of horseradish peroxidase. The tracer was taken up within vesicles, tubules, multivesicular bodies, and dense bodies. From acid phosphatase studies and from previous work it appears that many of the structures in which peroxidase accumulates are lysosomes or are destined to fuse with lysosomes. Subsequent to the period of intense exocytosis and endocytosis, there is a transient accumulation of lipid droplets in the adrenalin cells. The cells then regranulate, with new granules forming near the Golgi region. These results suggest that under the conditions used, much of the membrane that initially surrounds secretory granules is degraded after release of the granules.