AJP - GI Ad Instruments
HOME HELP FEEDBACK SUBSCRIPTIONS ARCHIVE SEARCH TABLE OF CONTENTS
QUICK SEARCH:   [advanced]


     

Am J Physiol Gastrointest Liver Physiol 293: G1333-G1338, 2007. First published August 23, 2007; doi:10.1152/ajpgi.00275.2007
0193-1857/07 $8.00
This Article
Right arrow Abstract Freely available
Right arrow Full Text (PDF)
Right arrow Supplemental Figure and Movie
Right arrow All Versions of this Article:
293/6/G1333    most recent
00275.2007v1
Right arrow Alert me when this article is cited
Right arrow Alert me if a correction is posted
Right arrow Citation Map
Services
Right arrow Email this article to a friend
Right arrow Similar articles in this journal
Right arrow Similar articles in Web of Science
Right arrow Similar articles in PubMed
Right arrow Alert me to new issues of the journal
Right arrow Download to citation manager
Citing Articles
Right arrow Citing Articles via Web of Science (1)
Right arrow Citing Articles via Google Scholar
Google Scholar
Right arrow Articles by Voronina, S. G.
Right arrow Articles by Tepikin, A. V.
Right arrow Search for Related Content
PubMed
Right arrow PubMed Citation
Right arrow Articles by Voronina, S. G.
Right arrow Articles by Tepikin, A. V.

INNOVATIVE METHODOLOGY

Visualizing formation and dynamics of vacuoles in living cells using contrasting dextran-bound indicator: endocytic and nonendocytic vacuoles

Svetlana G. Voronina, Mark W. Sherwood, Oleg V. Gerasimenko, Ole H. Petersen, and Alexei V. Tepikin

The Physiological Laboratory, The University of Liverpool, Liverpool, United Kingdom

Submitted 18 June 2007 ; accepted in final form 21 July 2007


   ABSTRACT
TOP
 ABSTRACT
MATERIALS AND METHODS
 RESULTS
 DISCUSSION
 GRANTS
 REFERENCES
 
Here we describe a technique that allows us to visualize in real time the formation and dynamics (fusion, changes of shape, and translocation) of vacuoles in living cells. The technique involves infusion of a dextran-bound fluorescent probe into the cytosol of the cell via a patch pipette, using the whole-cell patch-clamp configuration. Experiments were conducted on pancreatic acinar cells stimulated with supramaximal concentrations of cholecystokinin (CCK). The vacuoles, forming in the cytoplasm of the cell, were revealed as dark imprints on a bright fluorescence background, produced by the probe and visualized by confocal microscopy. A combination of two dextran-bound probes, one infused into the cytosol and the second added to the extracellular solution, was used to identify endocytic and nonendocytic vacuoles. The cytosolic dextran-bound probe was also used together with a Golgi indicator to illustrate the possibility of combining the probes and identifying the localization of vacuoles with respect to other cellular organelles in pancreatic acinar cells. Combinations of cytosolic dextran-bound probes with endoplasmic reticulum (ER) or mitochondrial probes were also used to simultaneously visualize vacuoles and corresponding organelles. We expect that the new technique will also be applicable and useful for studies of vacuole dynamics in other cell types.

pancreatic acinar cells; confocal microscopy; patch clamp; cholecystokinin

THE FORMATION OF INTRACELLULAR vacuoles has been observed by using fixed tissues in a large number of cell types (e.g., hepatocytes, cardiomyocytes, neurons, and different exocrine secretory cell); it reflects wide-ranging pathological conditions and is used as an important indicator of cell damage (5, 10, 23, 24). In particular, the appearance of intracellular vacuoles is hallmark of the morphological changes in pancreatic acinar cells in conditions of acute pancreatitis. These structures were observed in different models of pancreatitis including pancreatic duct obstruction, overstimulation with calcium-releasing secretagogues, and stimulation with bile acids (8, 9, 12, 14–16, 23). In our recent study we have identified aberrant endocytosis as one mechanism for the formation of large intracellular vacuoles in pancreatic acinar cells (18). These vacuoles were visualized by using fluid-phase tracer for endocytosis added to the extracellular solution. The limitation of this technique is that it labels only one specific type of vacuoles. In the present study we decided to utilize contrasting indicator added to the cytosol to visualize, in real time, the formation and dynamics of all large membrane-bounded vacuoles in living cells. We hope that the technique described below will be useful for studies of other cell types in which the formation of intracellular vacuoles reflects important physiological or pathological processes.


   MATERIALS AND METHODS
TOP
 ABSTRACT
 MATERIALS AND METHODS
RESULTS
 DISCUSSION
 GRANTS
 REFERENCES
 
Reagents. Dextran Alexa Fluor 488 10,000 mW, Dextran Texas Red, 3,000 mW, Dextran Cascade Blue 10,000 mW, ER-Tracker Blue-White DPX, MitoTracker Deep Red, and NBD-C6-ceramide were from Molecular Probes/Invitrogen (Eugene, OR). Collagenase was from Worthington Biochemical (Lakewood, NJ). All other reagents were from Sigma-Aldrich (Gillingham, UK).

Cell isolation and solutions. Mouse pancreatic acinar cells were isolated by collagenase digestion as described previously (22). Pancreata were obtained from adult male mice (CD1) that had been killed by cervical dislocation. In accordance with the Animals (Scientific Procedures) Act of 1986 (UK), training and oversight of procedures were conducted by competent personnel from the University of Liverpool (in compliance with national requirements). The standard extracellular solution used for cell preparation and for perfusion of cells during experiments contained 140 mM NaCl, 4.7 mM KCl, 1.3 mM MgCl2, 1 mM CaCl2, 10 mM D-glucose, 10 mM HEPES (pH 7.4). CCK was added to this solution at concentrations indicated for the specific experiments. Fluorescent probe Dextran Alexa Fluor 488 10,000 mW was added to the extracellular solution to attain a final concentration of 100 µM. The intracellular patch pipette solution contained 120 mM KCl, 10 mM NaCl, 1.5 mM MgCl2, 2 mM MgATP, 100 µM EGTA, and 40 mM HEPES (pH 7.2). NBD-C6-ceramide was loaded into the cells by incubation with 5 µM of the probe for 15 min at 4°C. The procedure was similar to that described in our previous publication (3). ER Tracker Blue-White DPX was loaded into the cells by incubation with 100 nM of the probe for 20–25 min at 37°C. MitoTracker Deep Red was loaded into the cells by incubation with 100 nM of the probe for 15 min at 37°C. All probes for cellular organelles were removed (by centrifugation) from the extracellular solution prior to the beginning of experiments. Dextran Texas Red 3,000 mW (1 mg/ml) or Dextran Cascade Blue 10,000 mW (1 mg/ml) was added to the patch pipette solution.

Experiments were conducted at room temperature (20–23°C).

Electrophysiological and optical recordings. The whole-cell configuration of the patch-clamp technique was used in this study. Patch pipettes were pulled from borosilicate glass capillaries (Harvard Apparatus, Edenbridge, Kent, UK) and fire polished. The pipettes had a resistance of 3–5 m{Omega} when filled with the intracellular solution (see Cell isolation and solutions). Further details on patch-clamp protocols used in our laboratory can be found in MATERIALS AND METHODS section of Ref. 2.

Experiments were performed on Zeiss 510 confocal microscope with water immersion objective x63/numerical aperture 1.2; axial resolution was selected at ~1 µm. For excitation of Dextran Texas Red we used a 543-nm laser line; emission was collected in the band 585–615 nm. Dextran Alexa Fluor 488 was excited with a 488-nm laser line and emission collected in the range 505–550 nm. Experiments were conducted in Multi Track configuration (in this configuration the combinations of excitation wavelength and emission filter for specific probes are applied sequentially). NBD-C6-ceramide was excited with a 488-nm laser line and emission collected between 505 and 550 nm. ER Tracker Blue-White DPX was excited with a 364-nm laser line and emission collected between 475 and 525 nm. MitoTracker Deep Red was excited with a laser line of 633 nm and emission collected with a long-pass filter (650 nm). Dextran Cascade Blue was excited with 357 nm and emission collected between 380 and 430 nm. Four-dimensional imaging of cells was used in some of our experiment to find the region of vacuole formation. In these experiments Z stacks of 10–20 two-dimensional images were collected every 60 s. Two-dimensional images from a specific confocal section were later selected to illustrate the process of vacuole formation.


   RESULTS
TOP
 ABSTRACT
 MATERIALS AND METHODS
 RESULTS
DISCUSSION
 GRANTS
 REFERENCES
 
Visualizing vacuoles using contrasting dextran-bound indicator. The procedure for visualizing the formation of intracellular vacuoles is illustrated in Fig. 1. The vacuole formation is seen as the appearance of a dark area on the background of brightly fluorescent (red color) contrasting indicator (Fig. 1, A and B). The indicator Dextran Texas Red was delivered into the cell via patch pipette (Fig. 1C) in whole cell patch-clamp configuration. Dextran-bound indicator was used in this study to make it more difficult for the probe to cross the membrane of vacuoles. Addition of dextran-bound indicator into the patch pipette does not preclude recording the electrophysiological parameters from the cell; an example of this is the calcium-depended Cl– current shown on Fig. 1D. The formation of vacuoles was initiated by supramaximal concentration of CCK (10 or 20 nM). A second indicator, Dextran Alexa Fluor 488, was added together with CCK to the extracellular solution as a probe for fluid endocytosis; note the appearance of fluorescence (green) in the extracellular solution during CCK stimulation (Fig. 1, A and B). Images on Fig. 1A and the profile of fluorescence on Fig. 1B indicate that the large vacuole, formed as a result of CCK stimulation in this specific experiment, does not show fluorescence of Dextran Alexa Fluor 488 and it is not therefore of endocytic origin. Figure 1E shows changes of shape of two neighboring nonendocytic vacuoles that could be interpreted as fusion of these structures (Fig. 1E).


Figure 1
View larger version (53K):
[in this window]
[in a new window]

  		Fig. 1. Imaging formation of nonendocytic vacuole. A: images of a cell infused with Dextran Texas Red, before (top row) and after (bottom row) stimulation with 10 nM CCK. Recording started (time 0) ~5 min after establishment of whole cell patch-clamp conditions. Fluorescence of Dextran Texas Red is shown in red. Fluorescence of Dextran Alexa Fluor 488 is shown in green. 3D images of the cell were collected over 60 s. Projection containing large intracellular vacuole, formed following CCK application, is shown. The 360-s image (top right) is the last image collected in this confocal section before CCK application; the 420-s image (bottom left) is the first image collected after CCK application. Profile of fluorescence, plotted in B, was recorded along the white arrow in A (bottom right image). Note the dip in the Dextran Texas Red fluorescence profile (red trace), which corresponds to the vacuole. The profile of fluorescence (here and in other figures) also serves to indicate the size of cells and vacuoles. C: transmitted light image. The patch pipette is visible in the top part of the image. D: calcium-dependent Cl– current recorded at –30 mV. E: fragment of the cell image (only Dextran Texas Red fluorescence) showing the fusion of a large dark vacuole with a smaller structure.

 
Combining intracellular and extracellular probes. The extracellular probes were utilized recently by two groups to study exocytosis in living pancreatic acinar cells (11, 20). These investigations observed the formation of large postexocytic {Omega} shapes as a consequence of compound exocytosis. In our previous study we extended this technique to measure endocytosis in the acinar cells and observed the disconnection of postexocytic structures producing large vacuoles (18). Here we combine this method with a new technique for monitoring vacuole formation by using contrasting dextran-bound indicator delivered into the cells via patch pipette.

We found that both nonendocytic and endocytic vacuoles formed in pancreatic acinar cells following supramaximal stimulation with CCK. Combination of intracellular and extracellular fluorescence probes helped us to identify the nature of the vacuoles. For example, in the experiment shown on Fig. 2A a nonendocytic vacuole was formed in the top cell following CCK stimulation, whereas the bottom cell responded by producing an endocytic vacuole of similar size (revealed by internalized fluorescence probe; see also Fig. 2B).


Figure 2
View larger version (52K):
[in this window]
[in a new window]

  		Fig. 2. Visualizing endocytic and nonendocytic vacuoles. A, left: transmitted light image of 2 acinar cells (patch pipette is visible in the top part of the image). A, middle: nonendocytic vacuole formed in the top cells, visualized with the help of infused Dextran Texas Red (fluorescence of this probe is shown by red color). A, right: endocytic vacuole in the bottom cell; green color (here and in C) shows fluorescence of Dextran Alexa Fluor 488, which was added to the extracellular solution together with CCK (20 nM). The fluorescence images were recorded following 580 s of CCK stimulation. B: overlay and profile of fluorescence. The left image shows overlay of fluorescence of the 2 dextrans (from A). Right: profile of fluorescence recorded along the arrow shown in the image (left). The peak of the fluorescence of Dextran Alexa Fluor 488 (green trace) corresponds to the endocytic vacuole. The dip in the fluorescence of Dextran Texas Red (red trace) corresponds to the nonendocytic vacuole. C: 2 types of vacuoles formed in the same cell. Scale bar corresponds to 5 µm. Left: transmitted light image of a doublet of pancreatic acinar cells. Top cell was patched and Dextran Texas Red (red color) infused into the cell. Dextran Alexa Fluor 488 (green color) was added into the extracellular solution (together with CCK) and shows endocytic vacuole formed as a result of CCK stimulation (20 nM for 890 s). Overlay of 2 images (right) shows the larger endocytic vacuole and 2 nonendocytic vacuoles above it formed in the same cell.

 
Both nonendocytic and endocytic vacuoles can form in the same cells (Fig. 2C). In this case both types of vacuoles form dark imprints on the brightly fluorescent image of the cell loaded with Dextran Texas Red. The endocytic vacuoles were revealed by the presence of Dextran Alexa Fluor 488. It is interesting to note that the boundary of the image of nonendocytic vacuoles (imprint on Dextran Texas Red fluorescence) was usually sharper than the boundary of endocytic vacuoles.

Observation of vacuole formation following patch-pipette withdrawal. Experiments were also conducted in conditions when the patch pipette was withdrawn following the delivery of the contrasting indicator into the cytosol and the plasma membrane resealed (Fig. 3). In these conditions intracellular pressure changes, which could potentially be induced by changes in the concentration of electrolytes and water, following the stimulation of the cell, cannot be relieved by the patch pipette. The relatively rapid termination of contact between the pipette and the cell interior should also minimize the washout of proteins and other macromolecules from the cytosol. In our experience the probability of successful (without destroying the patched cell) pipette withdrawal increases when the pipette is removed soon (<5 min) after establishing the whole cell configuration. In conditions of our experiments a few minutes of pipette-cytosol contact was sufficient to load the dextran-bound indicator. These experiments were conducted on larger clusters. It is known that the morphology of individual pancreatic acinar cells is better maintained in clusters than in isolated cells (13). In these experiments we were also able to observe the formation of intracellular vacuoles (Fig. 3).


Figure 3
View larger version (37K):
[in this window]
[in a new window]

  		Fig. 3. Imaging vacuole formation in a cell following patch pipette withdrawal. A: images of cells before stimulation. The transmitted light image is overlaid with fluorescence image of a cell loaded with Dextran Texas Red. Images were acquired after the patch pipette, used to deliver the indicator into the cell, was withdrawn. B: this image shows the fluorescence of this cell prior to stimulation. C: the same confocal section as in B but following stimulation with CCK (10 nM for 1,480 s). The dark imprint on the red fluorescence image reveals the formation of a nonendocytic vacuole. The fluorescence of Dextran Alexa Fluor 488 (added together with CCK) is shown in green. Note the green area in the right cell, which suggests the formation of an endocytic structure.

 
Using contrasting indicator for vacuole formation in combination with Golgi, ER, or mitochondrial probes. Nonendocytic vacuoles are frequently formed in parts of the cell that are a few micrometers away from the apical region. One of the organelles present in this region is the Golgi apparatus (3, 19). We found that staining of the NBD-C6-ceramide is compatible with the fluorescence of the contrasting indicator Dextran Texas Red (Fig. 4). We can therefore simultaneously visualize Golgi structures and the formation of vacuoles (note the disruption of Golgi by the growing vacuole; see Fig. 4 and the Supplementary Movie). Previous studies, utilizing electron microscopy, suggested that cytosolic vacuoles form in the vicinity of the Golgi (15, 23); we therefore initially developed a protocol specifically for combined staining of vacuoles and Golgi structures. We later also found combinations of fluorescence-labeled dextran and organellar probes suitable for visualizing vacuoles and the endoplasmic reticulum (ER) (combination of Dextran Texas Red and ER Tracker Blue-White DPX; see Supplementary Fig. 1A) and also vacuoles and mitochondria (combination of Dextran Cascade Blue and MitoTracker Deep Red, Supplementary Fig. 1B).


Figure 4
View larger version (54K):
[in this window]
[in a new window]

  		Fig. 4. Visualization of a vacuole formation and partitioning of the Golgi apparatus. A: top row of images was taken before CCK application, middle row following 50 s of continuous CCK (20 nM) stimulation, and bottom row following 200 s of the CCK stimulation. The vacuole formation is seen as an imprint on the Dextran Texas Red fluorescence (orange). NBD-C6-ceramide fluorescence (green) highlights Golgi apparatus. Transmitted image of the cells is shown in B. C shows the profiles of Dextran Texas Red fluorescence (orange) and NBD-C6-ceramide fluorescence (green), recorded along the arrow shown in the image (just above the fluorescence profile).

 

   DISCUSSION
TOP
 ABSTRACT
 MATERIALS AND METHODS
 RESULTS
 DISCUSSION
GRANTS
 REFERENCES
 
Here we have outlined technology for monitoring the formation and dynamics of vacuoles in acinar cells. In the configuration of our experiments, contrasting dextran-bound indicator allowed us to resolve vacuoles that are 0.7 µm in diameter. However, we had difficulty obtaining sufficient contrast for individual secretory granules that are known to be ~0.5 µm (4). The resolution of the new technique, as used in this study, is therefore somewhere between these two values. It is possible that the technique could be further optimized by increasing the concentration of the contrasting indicator and by increasing the intensity of excitation light. Brownian movement of vacuoles/granules (or other organelles) in the cytosol of living cells could be a factor limiting the resolution of this method. Another factor is the nonuniform light-scattering environment of the cytoplasm. For example, the apical part of acinar cells is known to cause strong scattering of visible light (11, 18). Therefore the resolution of the technique could be different in different parts of the cell. Pancreatic acinar cells have well-developed cellular organelles. In the acinar cells of guinea pig pancreas the rough surfaced ER occupies 22% of the acinar cell volume, the mitochondria take up 8.1%, and the Golgi occupies 9.9% of the surface area of cellular membranes (1). It is interesting to note that these organelles do not make a clear imprint on the images of Dextran Texas Red fluorescence. The reason for this is that although the total volume occupied by these organelles is very substantial the individual components are very thin: less than 0.2 µm for ER strands and most of the Golgi cisternae (1, 17). Mitochondria are also less than 0.5 µm in cross section (1, 7). It is possible that small variations of brightness exist over the regions occupied by the organelles but it certainly does not prevent the recordings of vacuole formation (e.g., Fig. 4). Previously, the majority of the studies of vacuole formation in pancreatic acinar cells and other cell types were conducted using fixed tissue. Electron microscopy, visible light microscopy, and fluorescence microscopy were used in these investigations. Although these studies provided remarkable details of the vacuole structure and suggested a plethora of the mechanisms responsible for vacuole formation (9, 12, 15, 23), they lacked the capability to resolve the process of vacuole formation (which can be very fast: <60 s following stimulation with CCK), the ability to visualize translocation of individual vacuoles, and the dynamics of vacuole interaction with other cellular organelles. The new approach should allow one to visualize transport of individual vacuoles, to study effects of second messengers on the rate of transport and could help to identify molecular motors responsible for this transport. Further refinement of this experimental approach will be necessary to combine all of these with the ability to characterize the origin of vacuoles in live cells. The nonendocytic (dark) vacuoles usually appear in the vicinity of the Golgi apparatus, which could suggest involvement of Golgi components or nearby condensing vacuoles in the formation of these structures. Indeed the notion that large vacuoles, formed as result of caerulein hyperstimulation, look like swollen condensing vacuoles was discusses in a study from Meldolesi's laboratory (23). The other type of large membrane bounded organelle that drastically increases in number as a result of stimulation with calcium-releasing agonists (6, 23) and is found in the Golgi area (23) is an autophagic vacuole. Both the size and the location of these organelles are similar to that of the dark vacuoles observed in our study. The dark vacuoles could be forming as a result of fusion between different types of organelles; for example autophagic vacuoles have been shown to fuse with late endosomes and lysosomes (21). One cannot also exclude the possibility that the vacuoles are formed by fusion of zymogen granules and lysosomes (15). Combination of optical microscopy, utilizing infused dextran in live cells, followed up by fixation and electron microscopy investigation of the same samples, could be necessary to characterize both the dynamics and the histological nature of the dark vacuoles.

In this study we had to work on a much shorter time scale than in our previous investigation of endocytic vacuoles (18). The reason for this is that cells only rarely survive in patch-clamp experiments beyond 20–30 min. By that time we could indeed observe formation of both nonendocytic vacuoles (which we could not resolve in our previous work because of the absence of a cytosolic marker) and endocytic vacuoles. But the endocytic vacuoles are only in the early stage of their formation and transport and are still clustered in the apical part of the cell. The technology of monitoring the dynamics of both types of vacuoles could be improved by further refinement of the procedure of patch pipette withdrawal; at the moment these experiments are still rather difficult.

In some cells we observed vacuoles immediately following infusion of the Dextran Texas Red (before any stimulation). This prehistory of the vacuole formation revealed by the indicator could identify prior episodes of cell damage that are undetectable by other techniques. The technique therefore allows one to visualize both already formed vacuoles and de novo generation of vacuoles.

Combination of Dextran Texas Red as a contrasting indicator to visualize vacuoles with NBD-C6-ceramide to label the Golgi apparatus is an example of colabeling of vacuoles and other cellular organelles. We also tested combinations of probes for visualizing vacuoles and ER or vacuoles and mitochondria. The broad range of indicators now available in dextran-conjugated form together with substantial and rapidly increasing range of probes for cellular organelles (including targeted fluorescent proteins) should provide an experimenter with the possibility of visualizing relationships between vacuoles and cellular organelles without major overlap between the spectra of the probes.


   GRANTS
TOP
 ABSTRACT
 MATERIALS AND METHODS
 RESULTS
 DISCUSSION
 GRANTS
REFERENCES
 
The study was supported by Grant G8801575 from the Medical Research Council (UK).


   ACKNOWLEDGMENTS
 
We are grateful to Mark W. Houghton for excellent technical assistance.


   FOOTNOTES
 

Address for reprint requests and other correspondence: A. Tepikin, The Physiological Laboratory, The Univ. of Liverpool, Crown St., Liverpool L69 3BX UK (e-mail: a.tepikin{at}liv.ac.uk)

The costs of publication of this article were defrayed in part by the payment of page charges. The article must therefore be hereby marked "advertisement" in accordance with 18 U.S.C. Section 1734 solely to indicate this fact.


   REFERENCES
TOP
 ABSTRACT
 MATERIALS AND METHODS
 RESULTS
 DISCUSSION
 GRANTS
 REFERENCES
 

  1. Bolender RP. Stereological analysis of the guinea pig pancreas. I. Analytical model and quantitative description of nonstimulated pancreatic exocrine cells. J Cell Biol 61: 269–287, 1974.[Abstract/Free Full Text]
  2. Chvanov M, Gerasimenko OV, Petersen OH, Tepikin AV. Calcium-dependent release of NO from intracellular S-nitrosothiols. EMBO J 25: 3024–3032, 2006.[CrossRef][Web of Science][Medline]
  3. Dolman NJ, Gerasimenko JV, Gerasimenko OV, Voronina SG, Petersen OH, Tepikin AV. Stable Golgi-mitochondria complexes and formation of Golgi Ca(2+) gradients in pancreatic acinar cells. J Biol Chem 280: 15794–15799, 2005.[Abstract/Free Full Text]
  4. Ermak TH, Rothman SS. Large decrease in zymogen granule size in the postnatal rat pancreas. J Ultrastruct Res 70: 242–256, 1980.[CrossRef][Web of Science][Medline]
  5. Foo NC, Ahn BY, Ma X, Hyun W, Yen TS. Cellular vacuolization and apoptosis induced by hepatitis B virus large surface protein. Hepatology 36: 1400–1407, 2002.[CrossRef][Web of Science]
  6. Hammel I, Shor-Hazan O, Eldar T, Amihai D, Lew S. Morphometric studies of secretory granule formation in mouse pancreatic acinar cells. Dissecting the early structural changes following pilocarpine injection. J Anat 194: 51–60, 1999.[CrossRef][Web of Science][Medline]
  7. Johnson PR, Dolman NJ, Pope M, Vaillant C, Petersen OH, Tepikin AV, Erdemli G. Non-uniform distribution of mitochondria in pancreatic acinar cells. Cell Tissue Res 313: 37–45, 2003.[CrossRef][Web of Science][Medline]
  8. Kim JY, Kim KH, Lee JA, Namkung W, Sun AQ, Ananthanarayanan M, Suchy FJ, Shin DM, Muallem S, Lee MG. Transporter-mediated bile acid uptake causes Ca2+-dependent cell death in rat pancreatic acinar cells. Gastroenterology 122: 1941–1953, 2002.[CrossRef][Web of Science][Medline]
  9. Lerch MM, Saluja AK, Runzi M, Dawra R, Steer ML. Luminal endocytosis and intracellular targeting by acinar cells during early biliary pancreatitis in the opossum. J Clin Invest 95: 2222–2231, 1995.[Web of Science][Medline]
  10. Miyata S, Takemura G, Kawase Y, Li Y, Okada H, Maruyama R, Ushikoshi H, Esaki M, Kanamori H, Li L, Misao Y, Tezuka A, Toyo-Oka T, Minatoguchi S, Fujiwara T, Fujiwara H. Autophagic cardiomyocyte death in cardiomyopathic hamsters and its prevention by granulocyte colony-stimulating factor. Am J Pathol 168: 386–397, 2006.[Abstract/Free Full Text]
  11. Nemoto T, Kojima T, Oshima A, Bito H, Kasai H. Stabilization of exocytosis by dynamic F-actin coating of zymogen granules in pancreatic acini. J Biol Chem 279: 37544–37550, 2004.[Abstract/Free Full Text]
  12. Otani T, Chepilko SM, Grendell JH, Gorelick FS. Codistribution of TAP and the granule membrane protein GRAMP-92 in rat caerulein-induced pancreatitis. Am J Physiol Gastrointest Liver Physiol 275: G999–G1009, 1998.[Abstract/Free Full Text]
  13. Park MK, Lee M, Petersen OH. Morphological and functional changes of dissociated single pancreatic acinar cells: testing the suitability of the single cell as a model for exocytosis and calcium signaling. Cell Calcium 35: 367–379, 2004.[CrossRef][Web of Science][Medline]
  14. Raraty M, Ward J, Erdemli G, Vaillant C, Neoptolemos JP, Sutton R, Petersen OH. Calcium-dependent enzyme activation and vacuole formation in the apical granular region of pancreatic acinar cells. Proc Natl Acad Sci USA 97: 13126–13131, 2000.[Abstract/Free Full Text]
  15. Saito I, Hashimoto S, Saluja A, Steer ML, Meldolesi J. Intracellular transport of pancreatic zymogens during caerulein supramaximal stimulation. Am J Physiol Gastrointest Liver Physiol 253: G517–G526, 1987.[Abstract/Free Full Text]
  16. Saluja A, Saito I, Saluja M, Houlihan MJ, Powers RE, Meldolesi J, Steer M. In vivo rat pancreatic acinar cell function during supramaximal stimulation with caerulein. Am J Physiol Gastrointest Liver Physiol 249: G702–G710, 1985.[Abstract/Free Full Text]
  17. Sesso A, de Faria FP, Iwamura ES, Correa H. A three-dimensional reconstruction study of the rough ER-Golgi interface in serial thin sections of the pancreatic acinar cell of the rat. J Cell Sci 107: 517–528, 1994.[Abstract]
  18. Sherwood MW, Prior IA, Voronina SG, Barrow SL, Woodsmith JD, Gerasimenko OV, Petersen OH, Tepikin AV. Activation of trypsinogen in large endocytic vacuoles of pancreatic acinar cells. Proc Natl Acad Sci USA 104: 5674–5679, 2007.[Abstract/Free Full Text]
  19. Thomas DD, Weng N, Groblewski GE. Secretagogue-induced translocation of CRHSP-28 within an early apical endosomal compartment in acinar cells. Am J Physiol Gastrointest Liver Physiol 287: G253–G263, 2004.[Abstract/Free Full Text]
  20. Thorn P, Fogarty KE, Parker I. Zymogen granule exocytosis is characterized by long fusion pore openings and preservation of vesicle lipid identity. Proc Natl Acad Sci USA 101: 6774–6779, 2004.[Abstract/Free Full Text]
  21. Tooze J, Hollinshead M, Ludwig T, Howell K, Hoflack B, Kern H. In exocrine pancreas, the basolateral endocytic pathway converges with the autophagic pathway immediately after the early endosome. J Cell Biol 111: 329–345, 1990.[Abstract/Free Full Text]
  22. Voronina SG, Barrow SL, Gerasimenko OV, Petersen OH, Tepikin AV. Effects of secretagogues and bile acids on mitochondrial membrane potential of pancreatic acinar cells: comparison of different modes of evaluating DeltaPsim. J Biol Chem 279: 27327–27338, 2004.[Abstract/Free Full Text]
  23. Watanabe O, Baccino FM, Steer ML, Meldolesi J. Supramaximal caerulein stimulation and ultrastructure of rat pancreatic acinar cell: early morphological changes during development of experimental pancreatitis. Am J Physiol Gastrointest Liver Physiol 246: G457–G467, 1984.[Abstract/Free Full Text]
  24. Zhang JH, Sampogna S, Morales FR, Chase MH. Age-related changes in cholinergic neurons in the laterodorsal and the pedunculo-pontine tegmental nuclei of cats: a combined light and electron microscopic study. Brain Res 1052: 47–55, 2005.[CrossRef][Web of Science][Medline]




This Article
Right arrow Abstract Freely available
Right arrow Full Text (PDF)
Right arrow Supplemental Figure and Movie
Right arrow All Versions of this Article:
293/6/G1333    most recent
00275.2007v1
Right arrow Alert me when this article is cited
Right arrow Alert me if a correction is posted
Right arrow Citation Map
Services
Right arrow Email this article to a friend
Right arrow Similar articles in this journal
Right arrow Similar articles in Web of Science
Right arrow Similar articles in PubMed
Right arrow Alert me to new issues of the journal
Right arrow Download to citation manager
Citing Articles
Right arrow Citing Articles via Web of Science (1)
Right arrow Citing Articles via Google Scholar
Google Scholar
Right arrow Articles by Voronina, S. G.
Right arrow Articles by Tepikin, A. V.
Right arrow Search for Related Content
PubMed
Right arrow PubMed Citation
Right arrow Articles by Voronina, S. G.
Right arrow Articles by Tepikin, A. V.

HOME HELP FEEDBACK SUBSCRIPTIONS ARCHIVE SEARCH TABLE OF CONTENTS
Visit Other APS Journals Online
Copyright © 2007 by the American Physiological Society.