Postembedding staining of Brunner's gland with lectin-ferritin conjugates

Postembedding staining of intracellular carbohydrates of rat Brunner's gland cells embedded in Epon and acrylamide was carried out with Ricinus communis agglutinin-ferritin, concanavalin A-ferritin, and wheat germ agglutinin-ferritin conjugates. Th Golgi vacuoles and mucous granules were stained with these conjugates. In each staining, the tissues embedded in acrylamide were stained more strongly than those embedded in Epon. The staining intensity of wheat germ agglutinin-ferritin was the strongest among the three conjugates and the staining intensity of Ricinus communis agglutinin-ferritin was stronger than that of concanavalin A-ferritin in both embedding methods. Free ferritin showed almost no binding to these structures and staining with the conjugates was inhibited by the addition of appropriate competitive sugars to the staining solutions. Osmium-postfixed tissues were not stained well with the conjugates. Washing of the sections with bovine serum albumin solution after staining was an essential step in the present method to reduce the nonspecific adsorption of the conjugates. The present method was very simple and had good reproducibility.     

Binding and transepithelial transport of immunoglobulins by intestinal M cells: demonstration using monoclonal IgA antibodies against enteric viral proteins

M cells of intestinal epithelia overlying lymphoid follicles endocytose luminal macromolecules and microorganisms and deliver them to underlying lymphoid tissue. The effect of luminal secretory IgA antibodies on adherence and transepithelial transport of antigens and microorganisms by M cells is unknown. We have studied the interaction of monoclonal IgA antibodies directed against specific enteric viruses, or the hapten trinitrophenyl (TNP), with M cells. To produce monospecific IgA antibodies against mouse mammary tumor virus (MMTV) and reovirus type 1, Peyer's patch cells from mucosally immunized mice were fused with myeloma cells, generating hybridomas that secreted virus-specific IgA antibodies in monomeric and polymeric forms. One of two anti-MMTV IgA antibodies specifically bound the viral surface glycoprotein gp52, and 3 of 10 antireovirus IgA antibodies immunoprecipitated sigma 3 and mu lc surface proteins. 35S-labeled IgA antibodies injected intravenously into rats were recovered in bile as higher molecular weight species, suggesting that secretory component had been added on passage through the liver. Radiolabeled or colloidal gold-conjugated mouse IgA was injected into mouse, rat, and rabbit intestinal loops containing Peyer's patches. Light microscopic autoradiography and EM showed that all IgA antibodies (antivirus or anti-TNP) bound to M cell luminal membranes and were transported in vesicles across M cells. IgA-gold binding was inhibited by excess unlabeled IgA, indicating that binding was specific. IgG-gold also adhered to M cells and excess unlabeled IgG inhibited IgA-gold binding; thus binding was not isotype-specific. Immune complexes consisting of monoclonal anti-TNP IgA and TNP-ferritin adhered selectively to M cell membranes, while TNP-ferritin alone did not. These results suggest that selective adherence of luminal antibody to M cells may facilitate delivery of virus-antibody complexes to mucosal lymphoid tissue, enhancing subsequent secretory immune responses or facilitating viral invasion.     

Post-embedding staining of rat gastric mucous cells with lectins

Summary The mucous cells of the rat stomach were stained with lectins by two post-embedding staining methods for electron microscopy. The mucous granules of surface mucous cells and foveolar mucous cells were stained weakly by Ricinus communis agglutinin-ferritin and wheat germ agglutinin-ferritin. The mucous granules of mucous neck cells were stained by concanavalin A-ferritin, Ricinus communis agglutinin-ferritin and wheat germ agglutinin-ferritin. The mucous granules of pyloric gland cells showed an affinity for wheat germ agglutinin-ferritin and concanavalin A-ferritin, while Ricinuscommunis agglutinin-ferritin only slightly stained the granules. The granules of mucous neck cells and pyloric gland cells were also stained by the concanavalin A-horseradish peroxidase-colloidal gold method, but the granules of surface and foveolar mucous cells were not stained by this method. Periodic acid oxidation of the sections before the standard concanavalin A-ferritin procedure enhanced the staining of the granules of mucous neck cells and pyloric gland cells slightly. Reduction of the sections after the periodic acid oxidation weakened the staining. Similar results were obtained using the concanavalin A-horseradish peroxidase-colloidal gold method. Though the staining with Ricinus communis agglutinin-ferritin was inhibited by periodic acid oxidation of the sections before staining, the staining with wheat germ agglutinin-ferritin was not inhibited by the oxidation. It is suggested that the paradoxical staining is closely related to the position of the concanavalin A-binding sugar residues in the carbohydrate chains.This work was supported in part by a grant-in-aid (No. 457008) from the Ministry of Education, Science and Culture, Japan and a grant-in-aid for cancer research (55-21) from the Ministry of Health and Welfare, Japan     

Proteolytic processing of reovirus is required for adherence to intestinal M cells.

Reovirus adheres specifically to apical membranes of mouse intestinal M cells and exploits M-cell transepithelial transport activity to enter Peyer's patch mucosa, where replication occurs. Proteolytic conversion of native reovirus to intermediate subviral particles (ISVPs) occurs in the intestine, but it is not known whether conversion is essential for interaction of virus with M cells. We tested the capacity of native virions, ISVPs, and cores (that lack outer capsid proteins) to bind to intestinal epithelial cells in vivo and found that only ISVPs adhered to M cells. Thus, intraluminal conversion of native reovirus to ISVPs is a prerequisite for M-cell adherence, and outer capsid proteins unique to ISVPs (either sigma 1 or products of mu 1) mediate interaction of virus with M-cell apical membranes.     

Quantitative immunoelectron microscopic localization of (Na+,K+)ATPase in rat parotid gland

Distribution of (Na+,K+)ATPase on the cell membranes of acinar and duct cells of rat parotid gland was investigated quantitatively by immunoelectron microscopy using the post-embedding protein A-gold technique. In acinar cells, ATPase was localized predominantly on the basolateral plasma membranes. A small but significant amount of (Na+,K+)ATPase was, however, detected on the luminal plasma membranes, especially on the microvillar region of the acinar cells; the surface density on the luminal membrane was approximately one third of that on the basolateral membranes. In duct cells, many gold particles were found on the basolateral membrane, especially along the basal infoldings of the plasma membranes, whereas no significant gold particles were found on the luminal plasma membranes, suggesting unilateral distribution of ATPase in duct cells. We suggest that in acinar cells sodium ion is not only transported paracellularly but is also actively transported intracellularly into the luminal space by the (Na+,K+)ATPase located on the luminal plasma membranes, and that water is passively transported to the luminal space to form a plasma-like isotonic primary saliva, while in the duct cells the same ion is selectively re-absorbed intracellularly by (Na+,K+)ATPase found in abundance along the many infoldings of the basal plasma membranes, thus producing the hypotonic saliva.     

Localization of the binding sites for the Ricinus communis, Agaricus bisporus and wheat germ lectins on human erythrocyte membranes.

Freeze-etch electron microscopy has been utilized to localize the binding sites for the Ricinus communis, Agaricus bisporus and wheat germ lectins on human erythrocyte membranes and to determine the relation of these different glycoprotein receptors to the intramembranous particles. A. bisporus lectin, which could be visualized directly on the surface of erythrocyte membranes, and ferritin conjugates of wheat germ agglutinin showed a distribution that correlates exactly with the intramembranous particles at all lectin concentrations tested. The binding sites for both of these lectins are located on the major sialoglycoprotein of the membrane. The R. communis agglutinin-ferritin conjugate which binds to receptors on membrane glycoproteins that are distinct from the major sialoglycoprotein showed a close correlation with the intramembranous particles at low lectin concentrations and a poor correlation at high lectin concentrations. High concentrations resulted in virtually complete coating of the surface of trypsinized ghosts which displayed marked aggregation of the intramembranous particles. We conclude that the intramembranous particles of erythrocyte membranes contain at least two glycoproteins and that some membrane lectin receptors are not associated with the intramembranous particles.     

Transcytosis in thyroid follicle cells

Inside-out follicles prepared from pig thyroid glands were used for studies on endocytosis. endocytosis. In this in vitro system, only the apical plasma membranes of follicle cells were exposed to tracers added to the culture medium. Cationized ferritin (CF) bound to the apical plasma membrane and was transferred first to endosomes and to lysosomes (within 5 min). Later, after approximately 30 min, CF was also found in stacked Golgi cisternae. In addition, a small fraction of endocytic vesicles carrying CF particles became inserted into the lateral (at approximately 11 min) and the basal (at approximately 16 min) plasma membranes. Morphometric evaluation of CF adhering to the basolateral cell surfaces showed that the vesicular transport across thyroid follicle cells (transcytosis) was temperature-sensitive; it ceased at 15 degrees C but increased about ninefold in follicles stimulated with thyrotropin (TSH). Thyroglobulin-gold conjugates and [3H]thyroglobulin (synthesized in separate follicle preparations in the presence of [3H]leucine) were absorbed to the apical plasma membrane and detected mainly in lysosomes. A small fraction was also transported to the basolateral cell surfaces where the thyroglobulin preparations detached and accumulated in the newly formed central cavity. As in the case of CF, transcytosis of thyroglobulin depended on the stimulation of follicles with TSH. The observations showed that a transepithelial vesicular transport operates in thyroid follicle cells. This transport is regulated by TSH and includes the transfer of thyroglobulin from the apical to the basolateral plasma membranes. Transcytosis of thyroglobulin could explain the occurrence of intact thyroglobulin in the circulation of man and several mammalian species.     

Ultrastructural and cytoarchitectural features of lymphoreticular organs in the colon and rectum of adult BALB/c mice

The structure and function of colonic mucosal lymphoid organs remain largely unexplored, especially in the rectum hidden within the pelvic vault. Two-month-old female BALB/c mice were anesthetized, and the entire colon was removed from cecum to anus. Distal colonic patches were then prepared for electron microscopy or were quick-frozen and sectioned for immunoperoxidase localization of B cells and T cell subsets. Aggregated lymphoid follicles were distributed irregularly along the entire colon with an average of 1.4 patches per centimeter of colon length. There were large collections of follicles opposite the ileocecal valve (cecal patches), variable numbers of patches throughout the colon, and at least one patch within 10 mm of the anus (rectal patch). Follicles were adjacent to branching crypts lined by epithelium infiltrated by lymphoid cells and containing few goblet cells. In electron micrographs, M cells were identified by their short, irregular microvilli; intraepithelial lymphoid cells; reduced lysosomal dense bodies; and an expanded tubulovesicular network. Small germinal centers were seen. Cytoarchitectural components of colonic lymphoid follicles and Peyer's patch follicles were remarkably similar, despite differences in surrounding mucosa and luminal microbial exposure. The presence of organized lymphoid tissue with M cells and germinal centers suggests that transepithelial particle transport and antigen recognition can take place in the rectum. Whether such tissue has the capacity for uptake of luminal microorganisms is of particular interest, not only because colonic follicles may be sites for local initiation of immune responses but also because they may be important entry points for systemic infection.     

Selective adherence of IgA to murine Peyer's patch M cells: evidence for a novel IgA receptor

M cells represent the primary route by which mucosal Ags are transported across the intestinal epithelium and delivered to underlying gut-associated lymphoid tissues. In rodents and rabbits, Peyer's patch M cells selectively bind and endocytose secretory IgA (SIgA) Abs. Neither the nature of the M cell IgR nor the domains of SIgA involved in this interaction are known. Using a mouse ligated ileal loop assay, we found that monoclonal IgA Abs with or without secretory component, but not IgG or IgM Abs, bound to the apical surfaces of Peyer's patch M cells, indicating that the receptor is specific for the IgA isotype. Human serum IgA and colostral SIgA also bound to mouse M cells. The asialoglycoprotein receptor or other lectin-like receptors were not detected on the apical surfaces of M cells. We used recombinant human IgA1 and human IgA2 Abs and domain swapped IgA/IgG chimeras to determine that both domains Calpha1 and Calpha2 are required for IgA adherence to mouse Peyer's patch M cells. This distinguishes the M cell IgA receptor from CD89 (FcalphaI), which binds domains Calpha2-Calpha3. Finally, we observed by immunofluorescence microscopy that some M cells in the human ileum are coated with IgA. Together these data suggest that mouse, and possibly human, M cells express an IgA-specific receptor on their apical surfaces that mediates the transepithelial transport of SIgA from the intestinal lumen to underlying gut-associated organized lymphoid tissues.     

Epithelial M cells in the rabbit caecal lymphoid patch display distinctive surface characteristics

The follicle-associated epithelium (FAE) in the rabbit caecal lymphoid patch is characterised by the presence of membranous (M) cells, which are believed to be functionally equivalent to those present at other sites of gut-associated lymphoid tissue (GALT). Caecal patch M cells display distinctive features compared with those of other GALT sites, despite similar general morphology and expression of the M cell marker vimentin, suggesting marked heterogeneity in the apical surface of M cells at discrete GALT sites. Electron microscopy reveals that rabbit caecal patch M cells differ from those in the small intestinal Peyer's patch FAE: the former have a prominent aspect within the epithelium and possess microvilli which are longer than those of adjacent enterocytes. Many of the M cells in peripheral regions of the caecal patch FAE are not associated with leucocytes and may thus represent an immature M cell population. The M cells are also histochemically distinct from adjacent enterocytes and from Peyer's patch M cells, showing greater expression of brush-border alkaline phosphatase activity and affinity for certain lectins (peanut and wheat germ agglutinins, Bandeiraea simplicifolia agglutinin II). The differences in the brush-border morphology and glycocalyx structure between M cells at different GALT sites may affect their function at these sites by influencing the interaction of luminal antigens and microorganisms with the M cell surface. The present data also support the hypothesis that M cells arise directly from differentiation of crypt stem cells and not from the transformation of existing fully differentiated enterocytes.     

Localization of the binding sites for the Ricinus communis, Agaricus bisporus and wheat germ lectins on human erythrocyte membranes

Freeze-etch electron microscopy has been utilized to localize the binding sites for the Ricinus communis, Agaricus bisporus and wheat germ lectins on human erythrocyte membranes and to determine the relation of these different glycoprotein receptors to the intramembranous particles. A. bisporus lectin, which could be visualized directly on the surface of erythrocyte membranes, and ferritin conjugates of wheat germ agglutinin showed a distribution that correlates exactly with the intramembranous particles at all lectin concentrations tested. The binding sites for both of these lectins are located on the major sialoglycoprotein of the membrane. The R. communis agglutinin-ferritin conjugate which binds to receptors on membrane glycoproteins that are distinct from the major sialoglycoprotein showed a close correlation with the intramembranous particles at low lectin concentrations and a poor correlation at high lectin concentrations. High concentrations resulted in virtually complete coating of the surface of trypsinized ghosts which displayed marked aggregation of the intramembranous particles. We conclude that the intramembranous particles of erythrocyte membranes contain at least two glycoproteins and that some membrane lectin receptors are not associated with the intramembranous particles.     

An ultrastructural search for lectin-binding sites on surfaces of spinach leaf organelles

Organelles isolated from leaves of spinach (Spinacia oleracea L.) were prefixed in glutaraldehyde and then incubated with ferritin conjugates of four lectins — Concanavalin A (Con A), Ricinus communis L. agglutinin, MW 120,000 (RCA), soybean agglutinin (SBA), and wheat germ agglutinin (WGA) — in order to probe their cytoplasmic surfaces for saccharide residues. In each case the major leaf organelles, including microbodies, mitochondria and chloroplast derivatives, failed to exhibit labeling when examined with the electron microscope. Tobacco (Nicotiana tabacum L.) leaf protoplasts, incubated simultaneously with and under identical conditions to the spinach organelles, showed specific labeling of their plasma membranes with all four lectin conjugates, thus establishing the efficacy of the procedure for demonstrating the presence of binding sites when they exist. Further attempts to show binding of one of the lectins, Con A, by labeling with fluorescein-Con A and by organelle agglutination, yielded results consistent with the absence of ultrastructural labeling. It is concluded that no saccharide residues recognized by the four lectins are present on the cytoplasmic surfaces of organelles and that those residues reported to be constituents of intracellular membranes, therefore, are most likely exposed on the luminal (extracytoplasmic) surfaces.Abbreviations Con A Concanavalin A - RCA Ricinus communis agglutinin, MW 120,000 - SBA soybean agglutinin - WGA wheat germ agglutinin     

Transfer of carbonic anhydrase-positive particles from the follicle-associated epithelium to lymphocytes of Peyer's patches in foetal sheep and lambs

Summary Carbonic anhydrase cytochemistry of the ileal Peyer's patch in foetal and neonatal lambs has indicated secretion from the follicle-associated epithelium to the follicles. Reaction for carbonic anhydrase in the follicle-associated epithelium was found in the luminal plasma membrane, in cytoplasmic vesicles, and in vacuoles containing 50-nm membrane-bounded particles that seemed to be shed to the intercellular space. The lateral plasma membrane was negative for carbonic anhydrase, indicating that formation of carbonic anhydrase-positive particles was restricted to vacuoles. Administration of ferritin to ileal loops of sheep foetuses showed ferritin localized in vesicles and vacuoles of the follicle-associated epithelium followed by exocytosis, together with carbonic anhydrase-positive particles, into the indentations of the lateral cell border. The carbonic anhydrase-positive particles seemed to be transported to the centres of lymphoid follicles where many were attached to the plasma membrane of lymphocytes. Carbonic anhydrase-positive particles were also seen in vesicles and sometimes free in the cytoplasm of the lymphocytes or attached to their nuclear envelope. Light microscopically, carbonic anhydrase reactivity of the follicle-associated epithelium was associated with the early formation of the ileal Peyer's patch at about 100 days gestation. At this time the follicle-associated epithelium showed a strong luminal but at most a week lateral staining. With further foetal development there was a progressive increase in the amount of carbonic anhydrase-positive reaction product in extracellular particles, both along the lateral cell borders of the follicle-associated epithelium and among the lymphocytes of the follicle centres.     

Transtubular transport of proteins in rabbit proximal tubules

The purpose of the present experiments was to study possible different pathways of intracellular transport of proteins after luminal and basolateral uptake in isolated rabbit proximal tubules. Tubules were exposed to cationized ferritin (CF) in the perfusion fluid and horseradish peroxidase (HRP) in the bath simultaneously or to HRP in the bath alone for 30 min. The peritubular fluid (bath) and perfusion fluid were then exchanged and the tubules either fixed immediately or allowed to function during chase-periods for 10, 20, 30, or 60 min before fixation to follow the migration of the proteins through the cells. The proteins were to a large extent found separated in different vacuoles and lysosomes at all time periods studied, indicating separate pathways after uptake via the luminal and basolateral membranes respectively. About 0.5% of the CF taken up by the cells was transported through the cells and became located in the intercellular spaces. HRP was transported from the peritubular fluid to the apical cytoplasm of the tubules indicated by a gradual accumulation of small HRP-containing vesicles, first in the basal part of the cells and then in the apical cytoplasm. In tubules perfused with both CF and HRP in the perfusate, the CF and HRP were found together in apical vacuoles and lysosomes. After perfusion with HRP alone, this tracer was found in similar large vacuoles and lysosomes in the apical cytoplasm, in contrast to the small HRP-filled vacuoles seen after uptake from the bath.     

Effect of lectins on the transport of food ingredients in Caco-2 cell cultures

We investigated the effect of several lectins, such as soy bean lectin (SBA), concanavalin A (Con A), and wheat germ agglutinin (WGA), on the transport of some food ingredients (isoflavones, quercetin glycosides, carnosine/anserine) across Caco-2 cell monolayers. After incubation of food ingredients (0.03 approximately 2 mmol/L) in the presence or absence of lectins (1 approximately 180 microg/ml) on the apical side, aliquots were taken from the apical and basolateral solution, and were subjected to HPLC analysis. We also examined the effect of lectins on the permeability of the tight junction by measuring the transepithelial electrical resistance (TER) value of the Caco-2 cell monolayer. Isoflavones, which was not transported to the basolateral solution without lectins, could be transported in the presence of lectins, whereas their aglycones were detected at the same levels with or without the lectin treatment. The transport of quercetin glycosides also increased in the presence of lectins, however, that of peptides was not affected by the lectins. Con A and WGA, but SBA, decreased the TER value, indicating that Con A and WGA increased the transport via paracellular pathway, whereas SBA did via a different pathway.     

Phosphate transport by reconstructed monolayers from cultured mouse kidney cells

A reconstructed monolayer was formed using epithelial cells from normal mouse kidney to investigate the hormonal effect on phosphate transport by the renal cells. The cells, when cultured on a Millipore filter, formed a monolayer with an apical negative transepithelial potential of 8.4 +/- 0.4 mV. When radioactive phosphate was added onto the apical surface of the monolayer (corresponding to the luminal surface of a renal tubule), the phosphate was transported through the cell layer to the basolateral surface (corresponding to the peritubular surface of a renal tubule). This transport process was saturable, energy-dependent, and inhibited by 2,4-dinitrophenol or ouabain. Dose-dependent parathyroid hormone-induced inhibition (73% of the control) was also evident in this system. Similar inhibition (69% of the control) was observed with DBcAMP. Thus, monolayers reconstructed from cultured mouse kidney cells show characteristics similar to those of renal tubules.     

Differences in neutral amino acid and glucose transport between brush border and basolateral plasma membrane of intestinal epithelial cells.

A comparison of L-valine and D-glucose transport was carried out with vesicles of plasma membrane isolated either from the luminal (brush border) or from the contra-luminal (basolateral) region of small intestinal epithelial cells. The existence of transport systems for both non-electrolytes was demonstrated by stereospecificity and saturability of uptake, as well as tracer coupling. Transport of L-valine and D-glucose differs markedly in the two types of plasma membrane with respect to stimulation by Na+. The presence of Na+ stimulated initial L-valine and D-glucose uptake in brush border, but not in basolateral membrane. Moreover, an electro-chemical Na+ gradient, oriented with the lower potential on the inside, supported accumulation of the non-electrolytes above medium concentration only in the brush border membrane. L-Valine and D-glucose transport also were saturated at lower concentrations in brush border (10-20 mM) than in basolateral plasma membranes (30-50 mM). A third difference between the two membranes was found in the effectiveness of known inhibitors of D-glucose transport. In brush border membranes phlorizin was more potent than phloretin and 2', 3', 4'-trihydroxy-4-methoxy chalcone and cytochalasin B did not inhibit at all. In contrast, with the basolateral plasma membranes the order of potency was changed to phloretin = 2',3',4'-trihydroxy-4-methoxy chalcone greater than cytochalasin B greater than phlorizin. These results indicate the presence of different types of transport systems for monosaccharides and neutral amino acids in the luminal and contra-luminal region of the plasma membrane. Active transepithelial transport can be explained on the basis of the different properties of the non-electrolyte transport systems in the two cellular regions and an electro-chemical Na+ gradient that is dependent on cellular metabolism.     

Renal organic anion transport: a comparative and cellular perspective

A major system for net transepithelial secretion of a wide range of hydrophobic organic anions (OAs) exists in the proximal renal tubules of almost all vertebrates. This process involves transport into the cells against an electrochemical gradient at the basolateral membrane and movement from the cells into the lumen down an electrochemical gradient. Transport into the cells at the basolateral membrane, which is the dominant, rate-limiting step, is a tertiary active transport process, the final step which involves countertransport of the OA into the cells against its electrochemical gradient in exchange for alpha-ketoglutarate moving out of the cells down its electrochemical gradient. The outwardly directed gradient for alpha-ketoglutarate is maintained by metabolism ( approximately 40%) and by transport into the cells across both the basolateral and luminal membranes by separate sodium-dicarboxylate cotransporters ( approximately 60%). The inwardly directed sodium gradient driving alpha-ketoglutarate uptake is maintained by the basolateral Na(+)-K(+)-ATPase, the primary energy-requiring transport step in the total tertiary process. The basolateral OA/alpha-ketoglutarate exchange process now appears to be physiologically regulated by several factors in mammalian tubules, including peptide hormones (e.g., bradykinin) and the autonomic nervous system acting via protein kinase C (PKC) pathways and epidermal growth factor (EGF) working via the mitogen-activated protein kinase (MAPK) pathway.     

Energetics of potassium ion transport in Aplysia gut

Basolateral membranes of Aplysia californica foregut epithelia contain an ATP-dependent Na(+)/K(+) transporter (Na(+)/K(+) pump or Na(+)/K (+) -ATPase). This Na(+)/K(+) pump accounts for both the intracellular Na(+) electrochemical potential (micro) being less than the extracelluar Na(+) micro and the intracellular K(+) micro being more than the extracellular K(+ ) micro. Also, K(+) channel activity resides in both luminal and basolateral membranes of the Aplysia foregut epithelial cells. Increased activity of the Na(+)/K(+) pump, coupled to luminal and basolateral membrane depolarization altered the K(+) transport energetics across the basolateral membrane to a greater extent than the alteration in K(+) transport energetics across the luminal membrane. These results suggest that K(+) transport, either into or out of the Aplysia foregut epithelial cells, is rate-limiting at the basolateral membrane.     

Prolactin is transported across the epithelium of the jejunum and ileum of the suckling rat

Milk prolactin is transferred from the gastrointestinal tract to the circulation of the suckling rat. To identify the site of prolactin penetration and to determine the mechanism by which the hormone traverses the mucosal barrier, we followed the uptake of prolactin from ligated loops of jejunum or ileum in vivo by three methods: autoradiography, transport of prolactin-gold conjugates, and immunocytochemistry. Autoradiographic studies demonstrated specific binding sites for 125I-prolactin on apical membranes of the jejunum and ileum. Excess cold prolactin reduced radiolabel in apical and basal compartments. Gel autoradiography of portal sera showed the presence of intact prolactin and a prolactin fragment following jejunal transport but only a prolactin fragment following ileal transport. Uptake of prolactin-gold conjugates demonstrated that, in the jejunum, label was present at the luminal surface, within endosomal compartments and lysosomes, in basal coated and smooth vesicles, within basal coated pits, and beyond the basolateral surface. In the ileum, label was found at the luminal surface; within the tubulocisternae, endosomal vesicles, lysosomes, and basal smooth vesicles; and beyond the basolateral surface. Immunoreactive prolactin was present throughout the transepithelial pathways. This study demonstrates that prolactin is selectively and nonselectively absorbed in the jejunum and ileum and that the hormone is directed either to the lysosome for degradation or across the epithelium by means of a transcellular pathway.