scholarly journals UNSTIMULATED SECRETION OF PROTEIN FROM RAT EXOCRINE PANCREAS CELLS

1972 ◽  
Vol 52 (1) ◽  
pp. 147-158 ◽  
Author(s):  
M. F. Kramer ◽  
C. Poort
Keyword(s):  
Protein Synthesis ◽  
Cell Membrane ◽  
Secretory Granules ◽  
Apical Cell ◽  
Secretory Protein ◽  
Secretory Process ◽  
Tissue Slices ◽  
Apical Cell Membrane

Our earlier work demonstrated that the rate of protein synthesis in the exocrine cells of the rat pancreas is constant in different physiological states, including prolonged fasting. In this study we have followed the fate of the protein in the pancreatic cells of the fasting animal in vivo as well as in vitro. The data were obtained by quantitative radioautography and by biochemical determinations. In nonanesthesized, fasting rats, without cannulated pancreatic duct, some 80% of the proteins synthesized at a given time leaves the cell within 12 hr by way of secretion, intracellular breakdown not being important. Two mechanisms of fasting secretion exist. The first, starting at a slow rate after 20 min, is inferred to result from fortuitous contacts of young secretory granules with the apical cell membrane. The rate of secretion is the same in vivo as in vitro, at least during the first 4 hr after pulse labeling. Within 7 hr about 20% of the total amount of newly synthesized protein has left the cell. The second mechanism consists of an orderly movement of the mass of secretory granules towards the apical cell membrane as caused by the continuous assembly of new granules. The granules that come into contact with the cell membrane are discharged. It takes about 7–12 hr for secretory protein transported in this way to reach the cell membrane. The addition of new secretory granules to those present is essential for the second mechanism, for the blockade of protein synthesis by cycloheximide decreases the rate of this phase of secretion without interfering with the secretory process proper. Atropin does not inhibit the fasting secretion in vitro, nor does extensive washing of the tissue slices, excluding possible secretagogues as important factors in fasting secretion.

Conductive properties of the rabbit outer medullary collecting duct: inner stripe

1985 ◽  
Vol 248 (4) ◽  
pp. F500-F506 ◽  
Author(s):  
B. M. Koeppen
Keyword(s):  
Cell Membrane ◽  
Collecting Duct ◽  
Basolateral Membrane ◽  
Apical Cell ◽  
Rabbit Kidney ◽  
Apical Cell Membrane ◽  
Basolateral Cell Membrane ◽  
Medullary Collecting Duct ◽  
Conductive Properties

Segments of outer medullary collecting duct were dissected from the inner stripe of the rabbit kidney (OMCDi) and perfused in vitro. The conductive properties of the tubule epithelium and individual cell membranes were determined by means of cable analysis and intracellular voltage-recording microelectrodes. In 35 tubules the transepithelial voltage (VT) and resistance (RT) averaged 17.2 +/- 1.4 mV, lumen positive, and 58.6 +/- 5.3 k omega X cm, respectively. The basolateral membrane voltage, (Vbl) was -29.2 +/- 2.1 mV (n = 23). The apical cell membrane did not contain appreciable ion conductances, as evidenced by the high values of apical cell membrane fractional resistance (fRa = Ra/Ra + Rb), which approached unity (0.99 +/- 0.01; n = 23). Moreover, addition of amiloride or BaCl2 to the tubule lumen was without effect on the electrical characteristics of the cell, as was a twofold reduction in luminal [Cl-]. The conductive properties of the basolateral cell membrane were assessed with bath ion substitutions. A twofold reduction in bath [Cl-] depolarized Vbl by 14.7 +/- 0.4 mV (theoretical, 17 mV), while a 10-fold increase in bath [K+] resulted in only a 0.9 +/- 0.4 mV depolarization (theoretical, 61 mV). Substituting bath Na+ with tetramethylammonium (from 150 to 75 mM) was without effect. Reducing bath [HCO-3] from 25 to 5 mM (constant PCO2) resulted in a steady-state depolarization of Vbl of 8.4 +/- 0.4 mV that could not be attributed to conductive HCO-3 movement. Thus, the basolateral cell membrane is predominantly Cl- selective.(ABSTRACT TRUNCATED AT 250 WORDS)

A reinvestigation of the function of the mammalian urinary bladder

1977 ◽  
Vol 232 (3) ◽  
pp. F187-F195 ◽  
Author(s):  
S. A. Lewis
Keyword(s):  
Urinary Bladder ◽  
Membrane Surface ◽  
Apical Cell ◽  
Membrane Damage ◽  
In Vitro Experiments ◽  
Apical Cell Membrane ◽  
Cell Membrane Damage ◽  
Using Data

The function of adult mammalian urinary bladder is evaluated in light of recent in vitro experiments. The discrepancy between in vivo and in vitro experimental results is examined and a possible solution proposed. Techniques for eliminating edge damage and measuring apical membrane surface area are described. A new chamber design for microelectrode studies is illustrated. The possibility of apical cell membrane damage caused by microelectrodes is critically examined and tested using the polyene antibiotic Nystatin. Using data from transepithelial and microelectrode experiments, a model for net Na+ transport across the bladder is proposed and then critically analyzed. The possible clinical implications of the in vitro experiments are briefly discussed.

scholarly journals Neutrophil interaction with influenza-infected epithelial cells

Blood ◽  
1988 ◽  
Vol 72 (1) ◽  
pp. 142-149 ◽  
Author(s):  
DR Ratcliffe ◽  
SL Nolin ◽  
EB Cramer
Keyword(s):  
Influenza Virus ◽  
Epithelial Cells ◽  
Cell Membrane ◽  
Mdck Cells ◽  
Apical Cell ◽  
Apical Cell Membrane ◽  
Protein Insertion ◽  
Neutrophil Adherence ◽  
Vitro Model

Abstract An in vitro model system was used to study the early neutrophil response to influenza-infected epithelia. In the absence of serum, neutrophil adherence to influenza-infected confluent monolayers of Madin-Darby canine kidney epithelial cells (MDCK) was approximately 590 times greater than neutrophil binding to control cultures. The leukocytes bound specifically to virus-infected cells. Neutrophil adherence to influenza-infected MDCK cells was monitored during the course of one replication cycle, and binding began at a time (4.5 hours) that coincided with viral protein insertion in the apical cell membrane. Ultrastructural examination at 4.5 hours showed that greater than 90% of the neutrophils adhered to the epithelial cell membrane in the absence of budding virus and, at 6.5 hours, 100% of the neutrophils adhered to the epithelium with emerging virions. The number of neutrophils bound to influenza-infected MDCK cells was not affected by the presence or absence of calcium or magnesium but did depend on the amount of viral inoculum and on the temperature of the culture. In direct contrast to hemadsorption of RBCs, neutrophil binding to influenza-infected MDCK cells was 100% greater at 37 degrees C than at 4 degrees C. The neutrophil surface molecules that bound influenza virus appeared to become functionally polarized because the adherence of neutrophils to budding influenza virus or to a virus-coated surface inhibited the neutrophils from binding additional influenza virus to their nonadherent surface.

Neuroendocrine secretory protein 7B2: structure, expression and functions

2001 ◽  
Vol 357 (2) ◽  
pp. 329-342 ◽  
Author(s):  
Majambu MBIKAY ◽  
Nabil G. SEIDAH ◽  
Michel CHRÉTIEN
Keyword(s):  
Secretory Pathway ◽  
Secretory Granules ◽  
Secretory Protein ◽  
Neuroendocrine Cells ◽  
High Similarity ◽  
Granule Formation ◽  
Proteolytic Activation ◽  
Polyproline Motif

7B2 is an acidic protein residing in the secretory granules of neuroendocrine cells. Its sequence has been elucidated in many phyla and species. It shows high similarity among mammals. A Pro-Pro-Asn-Pro-Cys-Pro polyproline motif is its most conserved feature, being carried by both vertebrate and invertebrate sequences. It is biosynthesized as a precursor protein that is cleaved into an N-terminal fragment and a C-terminal peptide. In neuroendocrine cells, 7B2 functions as a specific chaperone for the proprotein convertase (PC) 2. Through the sequence around its Pro-Pro-Asn-Pro-Cys-Pro motif, it binds to an inactive proPC2 and facilitates its transport from the endoplasmic reticulum to later compartments of the secretory pathway where the zymogen is proteolytically matured and activated. Its C-terminal peptide can inhibit PC2 in vitro and may contribute to keep the enzyme transiently inactive in vivo. The PC2–7B2 model defines a new neuroendocrine paradigm whereby proteolytic activation of prohormones and proneuropeptides in the secretory pathway is spatially and temporally regulated by the dynamics of interactions between converting enzymes and their binding proteins. Interestingly, unlike PC2-null mice, which are viable, 7B2-null mutants die early in life from Cushing's disease due to corticotropin (‘ACTH’) hypersecretion by the neurointermediate lobe, suggesting a possible involvement of 7B2 in secretory granule formation and in secretion regulation. The mechanism of this regulation is yet to be elucidated. 7B2has been shown to be a good marker of several neuroendocrine cell dysfunctions in humans. The possibility that anomalies in its structure and expression could be aetiological causes of some of these dysfunctions warrants investigation.

scholarly journals Abnormal apical cell membrane in cystic fibrosis respiratory epithelium. An in vitro electrophysiologic analysis.

1987 ◽  
Vol 79 (1) ◽  
pp. 80-85 ◽  
Author(s):  
C U Cotton ◽  
M J Stutts ◽  
M R Knowles ◽  
J T Gatzy ◽  
R C Boucher
Keyword(s):  
Cystic Fibrosis ◽  
Cell Membrane ◽  
Apical Cell ◽  
Respiratory Epithelium ◽  
Apical Cell Membrane

CYTOTOXIC EFFECTS OF PUROMYCIN ON THE GOLGI APPARATUS OF PANCREATIC ACINAR CELLS, HEPATOCYTES AND AMELOBLASTS

1970 ◽  
Vol 18 (12) ◽  
pp. 875-886 ◽  
Author(s):  
ALFRED WEINSTOCK
Keyword(s):  
Protein Synthesis ◽  
Golgi Apparatus ◽  
Single Injection ◽  
Secretory Granules ◽  
Acinar Cells ◽  
Secretory Protein ◽  
Pancreatic Acinar Cells ◽  
Cytotoxic Effects ◽  
Total Inhibition

A single injection of puromycin was given to rats in a dose sufficient to produce almost total inhibition of protein synthesis in pancreas and liver. By 10 min after injection, Golgi saccules in pancreatic acinar cells, hepatocytes and ameloblasts appeared grossly and irregularly distended, and almost devoid of content. In pancreas the condensing vacuoles near the inner face of the Golgi were often altered, and those normally present in ameloblasts were lacking. Between 2 and 3 hr after injection, protein synthesis had started anew. At this time, granules without limiting membranes appeared within cisternae of the rough endoplasmic reticulum in acinar cells and ameloblasts. These intracisternal granules are believed to consist of newly synthesized secretory protein which could not be transported through the disrupted Golgi apparatus to be packaged into secretory granules. Indeed, by 3 hr postinjection the secretory granules which normally abound in the apical processes of ameloblasts were sparce or absent. Thus, while biochemical evidence indicates that puromycin blocks protein synthesis on the ribosomes, the use of this antibiotic in vivo results in alterations in the Golgi apparatus and interruption of the packaging of protein into secretory granules.

scholarly journals CELL MEMBRANE RESORPTION IN THE RAT EXOCRINE PANCREAS CELL AFTER IN VIVO STIMULATION OF THE SECRETION, AS STUDIED BY IN VITRO INCUBATION WITH EXTRACELLULAR SPACE MARKERS

1973 ◽  
Vol 57 (1) ◽  
pp. 159-174 ◽  
Author(s):  
J. J. Geuze ◽  
C. Poort
Keyword(s):  
Cell Membrane ◽  
Extracellular Space ◽  
Protein Transport ◽  
Secretory Protein ◽  
Zymogen Granule ◽  
Basal Face ◽  
Rat Exocrine Pancreas ◽  
Pancreas Cell

Pancreatic secretion in the rat was stimulated in vivo by pilocarpine injection causing 90% of the storage granules to be discharged within 2 h. Incubation in vitro with [14C]sorbitol indicated that maximal ingestion of this extracellular space marker occurred 3 h after secretogogue injection. Morphological cell membrane measurements on cells with stimulated secretion revealed a simultaneous decrease in amount of membrane bordering the microvilli at the cell apex, lamellar processes, and infoldings present at the latero-basal face of these cells. In 3-h stimulated cells, having the average zymogen granule content characteristic for that phase of secretion, ferritin treatment in vitro showed that the infoldings and related fragmentation vesicles had ingested ferritin and could consequently be considered as being transport vehicles for redundant cell membrane. During stimulated secretion numerous vesicles and vacuoles appeared in the apical cytoplasm. Part of these structures were postulated to be related to the Golgi complex and were discussed in relation to secretory protein transport. Another part of these structures was assumed to have an endocytotic nature, although they never contained ferritin.

Intracellular microelectrode characterization of the rabbit cortical collecting duct

1983 ◽  
Vol 244 (1) ◽  
pp. F35-F47 ◽  
Author(s):  
B. M. Koeppen ◽  
B. A. Biagi ◽  
G. H. Giebisch
Keyword(s):  
Cell Membrane ◽  
Collecting Duct ◽  
Basolateral Membrane ◽  
Apical Cell ◽  
Fold Increase ◽  
K Channel ◽  
Cortical Collecting Duct ◽  
Apical Cell Membrane ◽  
Basolateral Cell Membrane

Cortical collecting ducts of the rabbit were perfused in vitro and the intracellular potential (Vbl) was measured with KCl-filled microelectrodes. The ratio of apical to basolateral membrane resistance (Ra/Rbl) was estimated from the voltage divider ratio using cable analysis. In control tubules Vbl averaged--84.0 +/- 2.5 mV and Ra/Rbl was 0.83 +/- 0.11. Pretreatment of the rabbits with mineralocorticoid caused Vbl to hyperpolarize to--105.8 +/- 3.1 mV and Ra/Rbl to decrease slightly to 0.62 +/- 0.10. A 10-fold increase of the luminal [K+] caused a 40.6 +/- 3.1 mV depolarization of Vbl in control tubules and a 33.0 +/- 4.2 mV depolarization in tubules from DOCA-pretreated rabbits. Concurrently, Ra/Rbl decreased in both groups, consistent with the existence of a conductive K+ channel at the apical cell membrane. This apical K+ channel was not sensitive to amiloride but was blocked by Ba2+. Conductive movement of Na+ across the apical membrane was also apparent in that Ra/Rbl increased with amiloride from 0.61 +/- 0.10 to 1.45 +/- 0.28. A 10-fold increase in the bath [K+] caused a 28.6 +/- 3.8 and a 49.4 +/- 4.4 mV depolarization of Vbl in tubules obtained from control and DOCA-pretreated rabbits, respectively. In both groups Ra/Rbl increased, suggesting that the basolateral cell membrane also contains a conductive K+ channel. Taken together the results support a model in which the transepithelial reabsorption of Na+ and the transepithelial secretion of K+ are driven by the Na+-K+-ATPase located in the basolateral cell membrane, with passive movement of these ions occurring through separate conductive pathways in the apical cell membrane.

Further studies of organic base secretion by rabbit proximal tubules

1984 ◽  
Vol 246 (3) ◽  
pp. F282-F289 ◽  
Author(s):  
T. D. McKinney
Keyword(s):  
Cell Membrane ◽  
Apical Cell ◽  
Proton Exchange ◽  
Proximal Tubules ◽  
Organic Base ◽  
Apical Cell Membrane ◽  
Major Mechanism ◽  
Collecting Tubules

The aim of the present studies was to further characterize the mechanisms responsible for the secretion of the organic base procainamide by measuring [3H] procainamide transport in superficial (SF) and juxtamedullary (JM) S2 segments of rabbit proximal tubules perfused in vitro. When perfusate pH was changed from 7.4 to 6.2, procainamide secretion increased by 227% in SF and by 159% in JM tubules. In the presence of ouabain the acidic perfusate caused only a minimal increase in procainamide secretion. When the perfusate pH was increased, procainamide secretion was reduced. Acetazolamide, 10(-5) M in the bath, reduced procainamide secretion in both SF and JM tubules. In SF tubules amiloride, 10(-4) M in the perfusate, inhibited procainamide secretion, whereas nonradioactive procainamide, quinidine, and cimetidine did not. When ambient sodium was reduced to 10 mM, the rate of procainamide secretion was low and was not affected by amiloride in the perfusate or acetazolamide in the bath. Amiloride in the bath caused a dose-related inhibition of procainamide secretion. The rate of procainamide secretion by cortical collecting tubules was very low and was not affected by acetazolamide. The mechanisms of renal organic base secretion are complex. Some of the present data are consistent with a mechanism involving organic base-proton exchange across the apical cell membrane. However, other data are not consistent with this being the major mechanism involved.

Electrophysiological properties of amphibian late distal tubule in vivo

1988 ◽  
Vol 255 (1) ◽  
pp. F158-F166
Author(s):  
G. Planelles ◽  
T. Anagnostopoulos
Keyword(s):  
Cell Membrane ◽  
Apical Cell ◽  
Electrical Coupling ◽  
Distal Tubule ◽  
Ion Concentration ◽  
Transference Numbers ◽  
Apical Cell Membrane ◽  
Electrophysiological Properties ◽  
Concentration Changes

This study was undertaken to determine the passive electrophysiological properties of the diffusive barriers of the late distal tubule (LDT) in Necturus. The transepithelial resistance (RT) determined by cable analysis was 1,130 omega.cm2, which puts the LDT in the class of "tight" epithelia. Using two different methods, we did not find significant cell-to-cell electrical coupling. The fractional apical resistance was 0.93, and it did not vary with distance from the current-injecting electrode. Relative permeabilities of K+, Na+, and Cl- during peritubular ion concentration changes were assessed by circuit analysis. The conclusions are as follows. The basolateral cell membrane is highly permeable to K+; its apparent K+ transference number is 0.78. Basolateral chloride transference was very small. Sodium removal from peritubular fluid produced depolarization, suggesting carrier-mediated electrogenic Na+ transport. The high fractional resistance of the apical cell membrane prevented assessment of apical transference numbers. However, Cl- removal from luminal fluid produced cell hyperpolarization; the underlying mechanism has not been established with certainty. The paracellular pathway does not discriminate between Na+, Cl-, and some of their substitutes; it is poorly permeable to gluconate and prefers K+ to Na+.

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