scholarly journals Histamine treatment induces rearrangements of orthogonal arrays of particles (OAPs) in human AQP4-expressing gastric cells

2001 ◽  
Vol 154 (6) ◽  
pp. 1235-1244 ◽  
Author(s):  
Monica Carmosino ◽  
Giuseppe Procino ◽  
Grazia Paola Nicchia ◽  
Roberta Mannucci ◽  
Jean-Marc Verbavatz ◽  
...  
Keyword(s):  
Cell Line ◽  
Basolateral Membrane ◽  
Water Channel ◽  
Freeze Fracture ◽  
Orthogonal Arrays ◽  
Short Term ◽  
Water Permeability Coefficient ◽  
Cell Surface Biotinylation ◽  
Osmotic Water ◽  
Orthogonal Arrays Of Particles

To test the involvement of the water channel aquaporin (AQP)-4 in gastric acid physiology, the human gastric cell line (HGT)-1 was stably transfected with rat AQP4. AQP4 was immunolocalized to the basolateral membrane of transfected HGT-1 cells, like in native parietal cells. Expression of AQP4 in transfected cells increased the osmotic water permeability coefficient (Pf) from 2.02 ± 0.3 × 10−4 to 16.37 ± 0.5 × 10−4 cm/s at 20°C. Freeze-fracture EM showed distinct orthogonal arrays of particles (OAPs), the morphological signature of AQP4, on the plasma membrane of AQP4-expressing cells. Quantitative morphometry showed that the density of OAPs was 2.5 ± 0.3% under basal condition and decreased by 50% to 1.2 ± 0.3% after 20 min of histamine stimulation, mainly due to a significant decrease of the OAPs number. Concomitantly, Pf decreased by ∼35% in 20-min histamine-stimulated cells. Both Pf and OAPs density were not modified after 10 min of histamine exposure, time at which the maximal hormonal response is observed. Cell surface biotinylation experiments confirmed that AQP4 is internalized after 20 min of histamine exposure, which may account for the downregulation of water transport. This is the first evidence for short term rearrangement of OAPs in an established AQP4-expressing cell line.

Absence of orthogonal arrays in kidney, brain and muscle from transgenic knockout mice lacking water channel aquaporin-4

1997 ◽  
Vol 110 (22) ◽  
pp. 2855-2860 ◽  
Author(s):  
J.M. Verbavatz ◽  
T. Ma ◽  
R. Gobin ◽  
A.S. Verkman
Keyword(s):  
Cho Cells ◽  
Collecting Duct ◽  
Water Channel ◽  
Freeze Fracture ◽  
Orthogonal Arrays ◽  
Knock Out ◽  
Orthogonal Arrays Of Particles ◽  
Freeze Fracture Electron Microscopy ◽  
Knock Out Mice ◽  
And Function

Freeze-fracture electron microscopy (FFEM) of kidney collecting duct, muscle, astrocytes in brain, and other mammalian tissues has revealed regular square arrays of intramembrane particles called orthogonal arrays of particles (OAPs). Their possible role in membrane structure and transport have been proposed, and their absence or decrease has been noted in a variety of hereditary and acquired diseases. A transgenic mouse lacking water channel AQP4 was used to show that AQP4 is the OAP protein. FFEM was done on kidney, skeletal muscle, and brain from AQP4 wild-type [+/+], heterozygous [+/−] and knock-out [-/-] mice. The [-/-] mice did not express detectable AQP4 protein, but were grossly indistinguishable from [+/+] mice. FFEM was done on blinded samples of kidney, brain and muscle from 9 mice. In all 6 kidney samples from [+/+] and [+/−] mice, OAPs similar to those in AQP4-transfected CHO cells were found in basolateral membranes of collecting duct principal cells. In all muscle and brain samples from [+/+] and [+/−] mice, OAPs of identical ultrastructure to those in kidney were seen, but in smaller patch sizes. OAPs were not seen in any sample from [-/-] mice. Label-fracture analysis using a peptide-derived AQP4 polyclonal antibody showed immunogold labeling of OAPs in AQP4-expressing CHO cells. These studies provide direct evidence that AQP4 is required for formation of OAPs and is a component of OAPs, thus establishing the identity and function of OAPs.

Membrane organization and function of M1 and M23 isoforms of aquaporin-4 in epithelial cells

2004 ◽  
Vol 287 (3) ◽  
pp. F501-F511 ◽  
Author(s):  
Claudia Silberstein ◽  
Richard Bouley ◽  
Yan Huang ◽  
Pingke Fang ◽  
Nuria Pastor-Soler ◽  
...  
Keyword(s):  
Water Permeability ◽  
Single Channel ◽  
Phosphorylation Site ◽  
Freeze Fracture ◽  
Parietal Cells ◽  
Aquaporin 4 ◽  
Osmotic Water Permeability ◽  
Cellular Expression ◽  
Water Permeability Coefficient ◽  
Osmotic Water

Aquaporin-4 (AQP4) water channels exist as heterotetramers of M1 and M23 splice variants and appear to be present in orthogonal arrays of intramembraneous particles (OAPs) visualized by freeze-fracture microscopy. We report that AQP4 forms OAPs in rat gastric parietal cells but not in parietal cells from the mouse or kangaroo rat. Furthermore, the organization of principal cell OAPs in Brattleboro rat kidney is perturbed by vasopressin (arginine vasopressin). Membranes of LLC-PK1 cells expressing M23-AQP4 showed large, abundant OAPs, but none were detectable in cells expressing M1-AQP4. Measurements of osmotic swelling of transfected LLC-PK1 cells using videomicroscopy, gave osmotic water permeability coefficient ( Pf) values (in cm/s) of 0.018 (M1-AQP4), 0.019 (M23-AQP4), and 0.003 (control). Quantitative immunoblot and immunofluorescence showed an eightfold greater expression of M1- over M23-AQP4 in the cell lines, suggesting that single-channel pf (cm3/s) is much greater for the M23 variant. Somatic fusion of M1- and M23-AQP4 cells ( Pf = 0.028 cm/s) yielded OAPs that were fewer and smaller than in M23 cells alone, and M1-to-M23 expression ratios (∼1:4) normalized to AQP4 in M1 or M23 cells indicated a reduced single-channel pf for the M23 variant. Expression of an M23-AQP4-Ser111E mutant produced ∼1.5-fold greater single-channel pf and OAPs that were up to 2.5-fold larger than wild-type M23-AQP4 OAPs, suggesting that a putative PKA phosphorylation site Ser111 is involved in OAP formation. We conclude that the higher-order organization of AQP4 in OAPs increases single-channel osmotic water permeability by one order of magnitude and that differential cellular expression levels of the two isoforms could regulate this organization.

Molecular characterization of water-selective AQP (EbAQP4) in hagfish: insight into ancestral origin of AQP4

2007 ◽  
Vol 292 (1) ◽  
pp. R644-R651 ◽  
Author(s):  
Goro Nishimoto ◽  
Go Sasaki ◽  
Eishin Yaoita ◽  
Masaaki Nameta ◽  
Huiping Li ◽  
...  
Keyword(s):  
Rnase Protection Assay ◽  
Water Channel ◽  
Molecular Characteristics ◽  
Rnase Protection ◽  
Pavement Cells ◽  
Molecular Phylogenetic Tree ◽  
Osmotic Water ◽  
Basolateral Side ◽  
Orthogonal Arrays Of Particles ◽  
Molecular Phylogenetic

Hagfish ( Eptatretus burgeri) are agnathous and are the earliest vertebrates still in existence. Pavement cells adjacent to the mitochondria-rich cells show orthogonal arrays of particles (OAPs) in the gill of hagfish, a known ultrastructural morphology of aquaporin (AQP) in mammalian freeze-replica studies, suggesting that an AQP homolog exists in pavement cells. We therefore cloned water channels from hagfish gill and examined their molecular characteristics. The cloned AQP [ E. burgeri AQP4 (EbAQP4)] encodes 288 amino acids, including two NPA motifs and six transmembrane regions. The deduced amino acid sequence of EbAQP4 showed high homology to mammalian and avian AQP4 (rat, 44%; quail, 43%) and clustered with AQP4 subsets by the molecular phylogenetic tree. The osmotic water permeability of Xenopus oocytes injected with EbAQP4 cRNA increased eightfold compared with water-injected controls and was not reversibly inhibited by 0.3 mM HgCl2. EbAQP4 mRNA expression in the gill was demonstrated by the RNase protection assay; antibody raised against the COOH terminus of EbAQP4 also detected (by Western blot analysis) a major ∼31-kDa band in the gill. Immunohistochemistry and immunoelectron microscopy showed EbAQP4 localized along the basolateral membranes of gill pavement cells. In freeze-replica studies, OAPs were detected on the protoplasmic face of the split membrane comprising particles 5–6 nm long on the basolateral side of the pavement cells. These observations suggest that EbAQP4 is an ancestral water channel of mammalian AQP4 and plays a role in basolateral water transport in the gill pavement cells.

scholarly journals Indirect Role of AQP4b and AQP4d Isoforms in Dynamics of Astrocyte Volume and Orthogonal Arrays of Particles

Cells ◽  
2020 ◽  
Vol 9 (3) ◽  
pp. 735 ◽  
Author(s):  
Marjeta Lisjak ◽  
Maja Potokar ◽  
Robert Zorec ◽  
Jernej Jorgačevski
Keyword(s):  
Plasma Membrane ◽  
Cell Volume ◽  
Volume Regulation ◽  
Water Channel ◽  
Regulatory Volume Decrease ◽  
Cell Volume Regulation ◽  
Orthogonal Arrays ◽  
Cell Swelling ◽  
Early Endosomes ◽  
Orthogonal Arrays Of Particles

Water channel aquaporin 4 (AQP4) plays a key role in the regulation of water homeostasis in the central nervous system (CNS). It is predominantly expressed in astrocytes lining blood–brain and blood–liquor boundaries. AQP4a (M1), AQP4c (M23), and AQP4e, present in the plasma membrane, participate in the cell volume regulation of astrocytes. The function of their splicing variants, AQP4b and AQP4d, predicted to be present in the cytoplasm, is unknown. We examined the cellular distribution of AQP4b and AQP4d in primary rat astrocytes and their role in cell volume regulation. The AQP4b and AQP4d isoforms exhibited extensive cytoplasmic localization in early and late endosomes/lysosomes and in the Golgi apparatus. Neither isoform localized to orthogonal arrays of particles (OAPs) in the plasma membrane. The overexpression of AQP4b and AQP4d isoforms in isoosmotic conditions reduced the density of OAPs; in hypoosmotic conditions, they remained absent from OAPs. In hypoosmotic conditions, the AQP4d isoform was significantly redistributed to early endosomes, which correlated with the increased trafficking of AQP4-laden vesicles. The overexpression of AQP4d facilitated the kinetics of cell swelling, without affecting the regulatory volume decrease. Therefore, although they reside in the cytoplasm, AQP4b and AQP4d isoforms may play an indirect role in astrocyte volume changes.

FITC-dextran as a probe for endosome function and localization in kidney

1990 ◽  
Vol 258 (2) ◽  
pp. C309-C317 ◽  
Author(s):  
W. I. Lencer ◽  
P. Weyer ◽  
A. S. Verkman ◽  
D. A. Ausiello ◽  
D. Brown
Keyword(s):  
Epithelial Cells ◽  
Water Channel ◽  
Fluorescein Isothiocyanate ◽  
Endocytic Pathway ◽  
Water Permeability Coefficient ◽  
Osmotic Water ◽  
Endocytic Vesicles ◽  
Excised Tissue ◽  
Specific Membrane ◽  
Endocytic Pathways

Fluorescein isothiocyanate (FITC)-labeled endosomes were localized in kidney epithelial cells after tissue fixation and sectioning, and specific membrane transport properties of isolated endocytic vesicles were measured using the same probe. Rats were infused intravenously with 10 kDa FITC-dextran, and kidneys were fixed with paraformaldehyde lysine periodate. FITC-labeled vesicles were visualized in semithin (1 micron) frozen sections of excised tissue by epifluorescent microscopy and by electron microscopy after a photoconversion reaction. Most FITC-labeled endosomes were apically located in epithelial cells lining the urinary tubules. By immunocytochemistry the anti-lysosomal glycoprotein LGP 120 was absent from most of the FITC-labeled vesicles, although some colocalization was noted. The limiting membrane of FITC-labeled endosomes contained a vacuolar proton pump (pHmin = 6.23 +/- 0.033) and a water channel (osmotic water permeability coefficient, Pf = 0.052 +/- 0.005 cm/s) and was highly permeable to ethylene glycol and urea but relatively impermeable to glucose. Methods allowing the attribution of specific membrane functions to vesicles that can be visualized in the apical endocytic pathway of epithelial cells should be of general use for the study of endocytic pathways in a variety of systems.

scholarly journals Aquaporin-4 Dynamics in Orthogonal Arrays in Live Cells Visualized by Quantum Dot Single Particle Tracking

2008 ◽  
Vol 19 (8) ◽  
pp. 3369-3378 ◽  
Author(s):  
Jonathan M. Crane ◽  
Alfred N. Van Hoek ◽  
William R. Skach ◽  
A. S. Verkman
Keyword(s):  
Plasma Membranes ◽  
Freeze Fracture ◽  
Orthogonal Arrays ◽  
Cell Types ◽  
Single Particle Tracking ◽  
Aquaporin 4 ◽  
Live Cells ◽  
Restoring Force ◽  
Orthogonal Arrays Of Particles ◽  
Freeze Fracture Electron Microscopy

Freeze-fracture electron microscopy (FFEM) indicates that aquaporin-4 (AQP4) water channels can assemble in cell plasma membranes in orthogonal arrays of particles (OAPs). We investigated the determinants and dynamics of AQP4 assembly in OAPs by tracking single AQP4 molecules labeled with quantum dots at an engineered external epitope. In several transfected cell types, including primary astrocyte cultures, the long N-terminal “M1” form of AQP4 diffused freely, with diffusion coefficient ∼5 × 10−10 cm2/s, covering ∼5 μm in 5 min. The short N-terminal “M23” form of AQP4, which by FFEM was found to form OAPs, was relatively immobile, moving only ∼0.4 μm in 5 min. Actin modulation by latrunculin or jasplakinolide did not affect AQP4-M23 diffusion, but deletion of its C-terminal postsynaptic density 95/disc-large/zona occludens (PDZ) binding domain increased its range by approximately twofold over minutes. Biophysical analysis of short-range AQP4-M23 diffusion within OAPs indicated a spring-like potential, with a restoring force of ∼6.5 pN/μm. These and additional experiments indicated that 1) AQP4-M1 and AQP4-M23 isoforms do not coassociate in OAPs; 2) OAPs can be imaged directly by total internal reflection fluorescence microscopy; and 3) OAPs are relatively fixed, noninterconvertible assemblies that do not require cytoskeletal or PDZ-mediated interactions for formation. Our measurements are the first to visualize OAPs in live cells.

Particle arrays on insect nerve membranes

1983 ◽  
Vol 60 (1) ◽  
pp. 289-301
Author(s):  
N.E. Flower ◽  
J.D. Briers
Keyword(s):  
Freeze Fracture ◽  
Orthogonal Arrays ◽  
Basic Structure ◽  
Optical Diffraction ◽  
Electron Micrograph ◽  
Clear Cut ◽  
Large Numbers ◽  
Orthogonal Arrays Of Particles ◽  
Initial Impressions ◽  
Antennal Nerve

Freeze-fracture studies of Lepidopteran antennae have revealed the presence of orthogonal arrays of particles on antennal nerve membranes. Initial impressions were that several different arrays were present. Optical diffraction was used to examine the arrays on the electron micrograph negatives. This technique showed that all the arrays were derived from the same basic structure, suggesting that the superficial differences in appearance were due to shadowing effects. The particles are arranged in a lattice with spacings of 10.7 nm X 9.1 nm. The arrays are not clear-cut but tend to break up, producing a disorganized region around their edges. The particles are shown to have depressions in them. However, the evidence available suggests that the arrays do not have the other characteristics of gap junctions. The arrays appear not to be present on most of the nerve membrane faces, occurring only in localized regions of the nerve membranes where they are present in large numbers. This suggests that the arrays may have a specialized local function.

Calmodulin-mediated gating of lens gap junction channels in vesicles

1984 ◽  
Vol 42 ◽  
pp. 134-137
Author(s):  
Camillo Peracchia ◽  
Stephen J. Girsch
Keyword(s):  
Gap Junctions ◽  
Freeze Fracture ◽  
Orthogonal Arrays ◽  
Eye Lens ◽  
Lens Fiber ◽  
Cell Coupling ◽  
Particle Spacing ◽  
Fiber Cells ◽  
Gap Junction Channels ◽  
Communicating Junctions

The fiber cells of eye lens communicate directly with each other by exchanging ions, dyes and metabolites. In most tissues this type of communication (cell coupling) is mediated by gap junctions. In the lens, the fiber cells are extensively interconnected by junctions. However, lens junctions, although morphologically similar to gap junctions, differ from them in a number of structural, biochemical and immunological features. Like gap junctions, lens junctions are regions of close cell-to-cell apposition. Unlike gap junctions, however, the extracellular gap is apparently absent in lens junctions, such that their thickness is approximately 2 nm smaller than that of typical gap junctions (Fig. 1,c). In freeze-fracture replicas, the particles of control lens junctions are more loosely packed than those of typical gap junctions (Fig. 1,a) and crystallize, when exposed to uncoupling agents such as Ca++, or H+, into pseudo-hexagonal, rhombic (Fig. 1,b) and orthogonal arrays with a particle-to-particle spacing of 6.5 nm. Because of these differences, questions have been raised about the interpretation of the lens junctions as communicating junctions, in spite of the fact that they are the only junctions interlinking lens fiber cells.

scholarly journals Transepithelial water and urea permeabilities of isolated perfused Munich-Wistar rat inner medullary thin limbs of Henle's loop

2014 ◽  
Vol 306 (1) ◽  
pp. F123-F129 ◽  
Author(s):  
C. Michele Nawata ◽  
Kristen K. Evans ◽  
William H. Dantzler ◽  
Thomas L. Pannabecker
Keyword(s):  
Water Channel ◽  
Wistar Rat ◽  
Structural Features ◽  
Urea Transporter ◽  
Outer Medulla ◽  
Aquaporin 1 ◽  
Passive Mechanism ◽  
Short Loop ◽  
Osmotic Water ◽  
Gene Expression Levels

To better understand the role that water and urea fluxes play in the urine concentrating mechanism, we determined transepithelial osmotic water permeability ( Pf) and urea permeability ( Purea) in isolated perfused Munich-Wistar rat long-loop descending thin limbs (DTLs) and ascending thin limbs (ATLs). Thin limbs were isolated either from 0.5 to 2.5 mm below the outer medulla (upper inner medulla) or from the terminal 2.5 mm of the inner medulla. Segment types were characterized on the basis of structural features and gene expression levels of the water channel aquaporin 1, which was high in the upper DTL (DTLupper), absent in the lower DTL (DTLlower), and absent in ATLs, and the Cl-1 channel ClCK1, which was absent in DTLs and high in ATLs. DTLupper Pf was high (3,204.5 ± 450.3 μm/s), whereas DTLlower showed very little or no osmotic Pf (207.8 ± 241.3 μm/s). Munich-Wistar rat ATLs have previously been shown to exhibit no Pf. DTLupper Purea was 40.0 ± 7.3 × 10−5 cm/s and much higher in DTLlower (203.8 ± 30.3 × 10−5 cm/s), upper ATL (203.8 ± 35.7 × 10−5 cm/s), and lower ATL (265.1 ± 49.8 × 10−5 cm/s). Phloretin (0.25 mM) did not reduce DTLupper Purea, suggesting that Purea is not due to urea transporter UT-A2, which is expressed in short-loop DTLs and short portions of some inner medullary DTLs close to the outer medulla. In summary, Purea is similar in all segments having no osmotic Pf but is significantly lower in DTLupper, a segment having high osmotic Pf. These data are inconsistent with the passive mechanism as originally proposed.

Sign in / Sign up

Export Citation Format

Share Document