Accessory cells in the gill epithelium of the freshwater rainbow trout Salmo gairdneri

Two types of mitochondria-rich cells were identified in the gill epithelium of the freshwater-adapted rainbow trout, Salmo gairdneri, after selective impregnation of their tubular system with reduced osmium. A first type consisted of large cells with a poorly developed and loosely anastomosed tubular system; thus, that resembled the chloride cells commonly encountered in the gill epithelium of freshwater-adapted euryhaline fishes. A second type comprised smaller cells with an extensively developed and tightly anastomosed tubular system. These never reached the basal lamina of the gill epithelium and were adjacent to chloride cells, to which they were linked by shallow apical junctions (100-200 nm); thus, they resembled accessory cells, which are currently found in the gill epithelium of seawater-adapted fishes but are usually lacking in freshwater living fishes. Transfer of the freshwater-adapted trout into seawater induced the proliferation of the tubular system in the chloride cells and the formation of lateral plasma membrane interdigitations between accessory cells and the apical portion of the chloride cells. The length of the apical junction sealing off this extended intercellular space was reduced to 20-50 nm. The tubular system of the accessory cells was not modified. The extension of the tubular system in the chloride cells of the seawater-adapted fishes indicated that, as in most euryhaline fishes, these cells have a role in the adaptation of the rainbow trout to seawater. In contrast, the function of the presumptive accessory cells in freshwater trout remains to be established.     

The fine structure of the mesonephros of the lamprey,Entosphenus japonicus Martens

Summary The fine structure of the mesonephric kidney of the lamprey, Entosphenus japonicus Martens, has been investigated with the electron microscope and discussed from the viewpoint of comparative morphology of the mesonephros.The structure of the capillary wall of the glomerulus essentially coincides with that of higher vertebrates, though its basement membrane is remarkably thick (300–400 m) because of a dense accumulation of fibrillar material between the endothelium and the basal lamina of epithelial cell. No obvious fenestration of the endothelial cell has been observed in the glomerulus or capillaries in any part of this organ.The kidney tubule is divided into three segments: 1. neck segment composed of ciliated cells with numerous mitochondria and glycogen particles, 2. proximal tubule composed of brush bordered cells provided with extensive pinocytotic vesicles and lysosomal granules in the apical cytoplasm and with lamellar membranes in the basal, and 3. distal tubule characterized by cells which, with their abundant mitochondria and branched tubular endoplasmic reticulum (about 500 Å diameter) with a central core, closely resemble the chloride cells in the gill filament of some teleosts. The possibility that the lamellar membranes in the proximal tubule cells correspond to basal infoldings is discussed.The extensive development of the tubular reticulum and of the mitochondria in the distal tubule cells is believed to reflect the active absorption of urine chloride in the urinary tubule of lamprey mesonephric kidney evidenced by physiologists. The proximal tubule is suggested to take a part also in the urinary transport of water and ions, as the lamellar membranes found in the cells of this portion likely correspond to the basal infoldings in more advanced forms of the kidney.The epithelial cells of the ureteric duct are characterized by granules suggesting a mucous secretion. No fine structure implying an absorptive activity in this duct has been observed.     

Potassium pyroantimonate and osmium-zinc iodide reactivity in the tubular epithelium of the opisthonephric kidney of the sea lamprey,Petromyzon marinus L

Pyroantimonate precipitate indicates that the epithelium of the proximal tubule is the only segment of the tubular nephron of the fresh water lamprey where large accumlations of cations are distributed. Unusually large amounts of reaction product are located within the lateral intercellular spaces and within vesicles closely associated with the plasma membrane at the lateral and basal surfaces. This technique suggests the continuity of these vesicles with the plasma membrane and alludes to the possibility of an endomembranous system of vesicles and the intercellular spaces as vehicles for ion transport. Lateral intercellular spaces of proximal tubules of lower vertebrates may play a different role in kidney function that their counterparts in higher vertebrates. Osmium-zinc iodide has a specificity for certain cells within the proximal, intermediate, and distal segments, but no structural differences are noted when these cells are compared to unstained cells. Smooth endoplasmic reticulum remains unstained in the distal segment but the stain has a strong affinity for elements of the Golgi apparatus, lysosomes, and the nuclear envelope of all cell types. This technique does not suggest a structural or functional similarity between cells of the distal segment and the chloride cells of the gills of teleosts.     

Multicellular complex of chloride cells in the gills of freshwater teleosts

Morphology of branchial chloride cells in the freshwater teleosts Plecoglossus altivelis, Cyprinus carpio, and Oreochromis mossambicus was studied by light and transmission electron microscopy. The chloride cell has an apical membrane directly in contact with the outer medium. Generally, two or more neighboring chloride cells share an apical pit, forming a multicellular complex. The chloride cells form a multicellular complex in which cells differ in cytoplasmic electron density, development of tubular system, and in cell size. Chloride cells are linked by junctions which are shallower than the tight junctions that occur between neighboring pavement cells or between pavement and chloride cells. Multicellular complexes of chloride cells create additional paracellular pathways marked apically by the shallower junctions. Since junctional structure affects transepithelial permeability, development of multicellular complexes of chloride cells in freshwater fishes may be related to the transport of some substances as in the gills of marine fishes.     

In vitro growth and differentiation of human kidney tubular cells on a basement membrane substrate

Summary Kidney cortical tubular cells, mainly proximal tubular cells, isolated from human kidney and grown either on a basement membrane substrate in chemically defined medium or on plastic in serum-supplemented medium, had substantial proliferative potential and could be propagated for more than 10 generations or 8 passages before senescence. Basement membrane produced on a plastic substrate by the HR-9 endodermal cell line could replace serum supplementation in promoting tubular cell growth. Tubular cells grown on an HR-9 basement membrane substrate exhibited stable epithelial morphology over an extended period of time; in the presence of 5% serum they differentiated into organized structures such as hemicysts and cell cords. Cells grown on plastic failed to differentiate and gradually degenerated. Tubular cells on HR-9 basement membrane were characterized by densely packed microvilli, abundant rough endoplasmic reticulum and free polysomes, basal cell membrane interdigitations, a well-developed endocytotic apparatus, and conspicuous junctional complexes—all features of the proximal tubular cell. Compared with cells on plastic substrate, there were higher levels of the brush border enzymes γ-glutamyl transpeptidase,l-leucine aminopeptidase, and alkaline phosphatase in cells maintained on an HR-9 basement membrane substrate, further supporting the conclusion that a basement membrane substrate promoted differentiation of tubular cells. These data and morphological observations indicate that a basement membrane substrate can promote growth and both functional and morphologic differentiation of human kidney tubular cells. This work was supported by the Veterans Administration.     

Functional Expression of p64, an Intracellular Chloride Channel Protein

p64 is a protein identified as a chloride channel by biochemical purification from kidney microsomes. We expressed p64 in HeLa cells using a recombinant vaccinia virus/T7 RNA polymerase driven system. Total cell membranes were prepared from infected/transfected cells and fused to a planar lipid bilayer. A novel chloride channel activity was found in cells expressing p64 and not in control cells. The p64-associated activity shows strong anion over cation selectivity. Single channels show prominent outward rectification with single channel conductance at positive potentials of 42 pS. The chloride channel activity is activated by treatment of the membranes with alkaline phosphatase and inhibited by DNDS and by TS-TM calix(4)arene. Whole membrane anion permeability was determined by a chloride efflux assay, revealing that membranes from cells expressing p64 showed a small but highly significant increase in chloride permeability, consistent with expression of a novel chloride channel activity. Received: 17 November 1997/Revised: 9 February 1998     

Two types of chloride cells in the gill epithelium of a freshwater-adapted euryhaline fish: Lebistes reticulatus; their modifications during adaptation to saltwater

Two types of chloride cells were identified in the gill epithelium of freshwater-adapted guppies. One type, referred to as an "alpha-chloride cell," was a pale, elongated cell located at the base of the secondary lamella in close contact with the arterioarterial pillar capillaries. In its cytoplasm, membranous tubules in continuity with its basolateral plasma membrane formed an extended tridimensional network. The vesiculotubular system (Pisam: Anat. Rec. 200:401-414, 1981) consisted of a few tubules and vesicles located next to the apical plasma membrane. A second type, referred to as a "beta-chloride cell," was a darker, ovoid cell located in the interlamellar region of the primary epithelium facing the central venous sinus. Membranous tubules in continuity with the basolateral plasma membrane were unevenly distributed in the cytoplasm. A prominent vesiculotubular system composed of numerous vesicles and tubules was found between the Golgi apparatus and the apical surface. During seawater adaptation, the alpha-chloride cells increased in size and progressively transformed into characteristic "seawater alpha-chloride cells" with a well-developed, regular, tight tubular network and numerous vesicles and tubules of the vesiculotubular system accumulated below the apical pit. The beta-chloride cells underwent a progressive degeneration and disappeared. Thus, in freshwater-adapted guppies, there are two types of chloride cells, alpha and beta, respectively, related to the arterial and the venous vessels, whereas in seawater-adapted fishes, a single type of cell, the alpha-chloride cell, was related to both the arterial and venous channels.     

Morphology of the kidney of adult bowfin, Amia calva, with emphasis on "renal chloride cells" in the tubule

The nephron of adult bowfin, Amia calva, was described using light and electron microscopic techniques. The kidney of the bowfin possesses an abundant supply of renal corpuscles with each consisting of a glomerulus and a Bowman's capsule of visceral (podocyte) and parietal layers. No juxtaglomerular apparatus is present. The epithelium of the tubule is continuous with the parietal epithelium and is divisible in descending order into neck, first proximal, second proximal, first distal, second distal, and collecting segments. The tubules drain into a complex system of collecting ducts that ultimately unite with the main excretory duct, the archinephric duct. Mucous cells are the dominant cell throughout the entire ductular system. Nephrostomes are dispersed along the kidney capsule. The neck segment has a ciliated epithelium, and while both proximal segments possess a prominent brush border, the fine structure of the first implies involvement in protein absorption and the second in the transport and reabsorption of solutes. The cells of the first distal segment are characterized by deep infolding of the plasma membrane and a rich supply of mitochondria suggesting the presence of a mechanism for ion transport. The second distal segment is composed of cells resembling the chloride cells of fishes and these cells are present in progressively decreasing numbers in the collecting segment and duct system so that only a few are present in the epithelium of the archinephric duct. The "renal chloride cells" possess an abundant network of smooth tubules and numerous mitochondria with a rich supply of cristae. Glycogen is also a conspicuous component of these cells. The presence of "renal chloride cells" in this freshwater holostean, in other relatively primitive freshwater teleosts, and in larval and adult lampreys is discussed with reference to both phylogeny and the need for a special mechanism for renal ion conservation through absorption.     

Kidney atrophy vs hypertrophy in diabetes: which cells are involved?

One of the first structural changes in diabetic nephropathy (DN) is the renal enlargement. These changes resulted in renal hypertrophy in both glomerular and tubular cells. Shrink in the kidney size, which described as kidney atrophy resulted from the loss of nephrons or abnormal nephron function and lead to loss of the kidney function. On the other hand, increase in kidney size, which described as hypertrophy resulted from increase in proximal tubular epithelial and glomerular cells size. However overtime, tubular atrophy and tubulointerstitial fibrosis occurs as subsequent changes in tubular cell hypertrophy, which is associated with the infiltration of fibroblast cells into the tubulointerstitial space. The rate of deterioration of kidney function shows a strong correlation with the degree of tubulointerstitial fibrosis. A consequence of long-standing diabetes/hyperglycemia may lead to major changes in renal structure that occur but not specific only to nephropathy. Identifying type of cells that involves in renal atrophy and hypertrophy may help to find a therapeutic target to treat diabetic nephropathy. In summary, the early changes in diabetic kidney are mainly includes the increase in tubular basement membrane thickening which lead to renal hypertrophy. On the other hand, only renal tubule is subjected to apoptosis, which is one of the characteristic morphologic changes in diabetic kidney to form tubular atrophy at the late stage of diabetes.     

The association between familial distal renal tubular acidosis and mutations in the red cell anion exchanger (band 3, AE1) gene.

In distal renal tubular acidosis (dRTA) the tubular secretion of hydrogen ion in the distal nephron is impaired, leading to the development of metabolic acidosis, frequently accompanied by hypokalemia, nephrocalcinosis, and metabolic bone disease. The condition can be familial, when it is usually inherited as an autosomal dominant, though there is a rarer autosomal recessive form associated with nerve deafness. It has been shown that the autosomal dominant form of dRTA is associated with a defect in the anion exchanger (AE1) of the renal collecting duct intercalated cell. This transporter is a product of the same gene (AE1) as the erythrocyte anion exchanger, band 3. In this review we will look at the evidence for this association. Studies of genomic DNA from families with this disorder have shown, both by genetic linkage studies and by DNA sequencing, that affected individuals are heterozygous for mutations in the AE1 gene whilst unaffected family members have a normal band 3 sequence. Mutations have been found in the region of proposed helices 6 and 7 of the membrane domain of band 3 and involve amino acids Arg-589 and Ser-613, and in the COOH-terminal domain of band 3. Studies of red cell band 3 from these families have provided information on the effect these mutations have on the structure and function of erythrocyte band 3. Expression studies of the erythroid and kidney isoforms of the mutant AE1 proteins, in Xenopus laevis oocytes, have shown that they retained chloride transport activity, suggesting that the disease in the dRTA families is not related simply to the anion transport activity of the mutated proteins. A possible explanation for the dominant effect of these mutant AE1 proteins in the kidney cell is that these mutations affect the targeting of AE1 from the basolateral to the apical membrane of the alpha-intercalated cell.     

Structural adaptation to altered electrolyte metabolism by cortical distal segments

The four different renal cell types in the cortical segments beyond the macula densa--distal convoluted tubule (DCT) cell, the connecting tubule (CNT) cell, the principal (P) cell, and the intercalated (I) cell--each respond to prolonged specific stimuli with changes in cell membrane area. Increases in basolateral cell membrane area, reflecting the transport capacity of the cell, are associated in DCT cells with luminal, chronically high sodium load, in CNT cells with mineralocorticoids and tubular solute load, and in P cells with mineralocorticoids. In I cells the luminal cell membrane area seems to be influenced by the local luminal environment of the cells as well as by the peritubular environment. These structural findings indicate that tubular fluid composition as well as peritubular factors (e.g., mineralocorticoid levels) may have a role in regulating the transport capacity of distal cell types in the kidney.     

A dense cell in the epithelium of the gill lamellae of the brook trout, Salvelinus fontinalis (Mitchill)

A cell was found in freshwater brook trout which was similar to a chloride cell as it was mitochondria-rich with an extensive tubular network, but differed in having a dark cytoplasm, large round vesicles in the apical cytoplasm, and large membrane-bound bodies near the nucleus. The base of the cell was separated from the basement membrane by cytoplasm from another epithelial cell. The cell was more rarely found in trout adapted to brackish water and salt water.     

Relationship of the tubular system with other organelles in chlorride cells of the channel catfish,Ictalurus punctatus

The morphology of chloride cells in the channel catfish, Ictalurus punctatus, has been studied by transmission electron microscopy. The chloride cell possesses abundant tubules, mitochondria, and granules. The employment of a special membrane stain in conjunction with a two- or tridimensional analysis reveals a complex interjoining and interlocking ring system of tubules. Tubular sides constituting the complex rings frequently lack granules. The tubular rings join with tubulous mitochondrial profiles and other cytoplasmic components. © 1995 Wiley-Liss, Inc.     

Degeneration and death,by apoptosis and necrosis,of the pavement and chloride cells in the gills of the teleost Oreochromis mossambicus

Summary Degeneration and death of branchial epithelial cells were studied in an African cichlid fish. In both freshwater and seawater fish the superficially located pavement cells are sloughed off at the end of their lifecycle. This process is preceded by degeneration via a process of cytoplasmic shrinkage and condensation related to apoptotic (physiologically controlled) cell death. The chloride cells are pleomorphic, i.e., accessory, mature, and degenerating cells. Degeneration of chloride cells mainly occurs by apoptosis. Degenerating cells show shrinkage and densification of cytoplasm and nuclei, and swelling of the tubular system; these cells are then separated from the ambient water by pavement cells. They are finally phagocytosed and digested by macrophages. Apoptosis of chloride cells, but not of pavement cells, is greatly stimulated when the fish are in seawater; this reflects an increase in cellular turnover of the chloride cells. Accidental cell death (necrosis) of pavement cells or chloride cells is rarely observed in fully adapted freshwater and seawater fish. Its incidence increases in the first few days following transfer of fish from fresh water to seawater.     

The osmoregulatory tissue around the afferent blood vessels of the coxal gills in the estuarine amphipods, Grandidierella japonica and Melita setiflagella

By electron microscopy of the coxal gills in two species of estuarine amphipod crustaceans, Grandidierella japonica and Melita satifragella, we found a patch-like, specialized tissue area which consisted of unique cells closely resembling the salt-excreting cells in the gill of the brine shrimp and so-called chloride cells in teleost gills. These cells were characterized by an abundance of mitochondria, two kinds of extensive networks of cytoplasmic tubules, well-developed lamellar infoldings of the basal cell membrane, sparse microvillous projections of the apical border, and numerous large vacuoles with several incomplete partitions. The large (60 nm in diameter) and the small (30 nm) cytoplasmic tubular networks were found in the basal and the apical portions of the cell, respectively. The large networks, which were both directly and indirectly (through the lamellar system) continuous with the basal cell membrane, were regarded as extensions of the cell membrane. Both the outer walls and the partition walls of the vacuoles were reinforced with a parallel array of microtubules. The results suggest that this unique tissue plays an important role in the active transport of electrolytes to maintain a constant osmotic pressure of the hemolymph under widely fluctuating salinities of the estuarine environments.     

The cellular origin and proliferative status of regenerating renal parenchyma after mercuric chloride damage and erythropoietin treatment

OBJECTIVES: In this study, we have sought to establish the cellular origin and proliferative status of the renal parenchyma as it regenerates after damage induced by mercuric chloride, with or without erythropoietin treatments, that might alter the response. MATERIALS AND METHODS: Female mice were irradiated and male whole bone marrow was transplanted into them. Six weeks later recipient mice were assigned to one of four groups: control, mercuric chloride treated, erythropoietin treated and treated with mercuric chloride plus erythropoietin. RESULTS: Tubular injury scores were high 3 days after mercuric chloride and had recovered partially after 14 days, in line with serum urea nitrogen levels. Confocal microscopy confirmed the tubular location of bone marrow-derived cells. A 'four-in-one' analytical technique (identifying cell origin, tubular phenotype, tubular basement membranes and S-phase status) revealed that tubular necrosis increased bone marrow derivation of renal tubular epithelium from a baseline of approximately 1.3% to approximately 4.0%. Erythropoietin increased the haematocrit, but no other effects were detected. CONCLUSION: As 1 in 12 proximal tubular cells in S-phase was derived from bone marrow, we conclude that in the kidney, the presence of bone marrow-derived cells makes a minor but important regenerative contribution after tubular necrosis.     

Peritubular membrane potential in kidney proximal tubular cells of spontaneously hypertensive rats

Peritubular membrane potential in kidney proximal tubular cells of spontaneously hypertensive rats (SHR-Okamoto strain adult rats) was measured with conventional 3 mol KCl microelectrodes, in vivo. Peritubular cell membrane potential was not different in SHR (-66.5 ± 0.7 mV) as compared with normotensive control Wistar rats (-67.5 ± 1.2 mV). To test the effects of possible altered sodium membrane transport in SHR on proximal tubule peritubular membrane potential, we allowed SHR and control rats to drink 1% NaCl for two weeks. Again, proximal tubule peritubular membrane potential was not different in SHR on 1% NaCl (-67.0 ± 1.0 mV) as compared with control rats on 1% NaCl (-64.7 ± 1.3 mV). From these results we concluded that peritubular membrane potential in kidney proximal tubular cells of SHR was not different from normotensive Wistar control rats, and if some alteration of sodium transport in kidney proximal tubular cells of SHR could exist, that was not possible to evaluate from the measurements of peritubular membrane potential in kidney proximal tubular cells.     

Ion channels in Madin-Darby canine kidney cells.

Ion channels in Madin-Darby canine kidney cells serve transepithelial chloride transport and probably cell volume regulation. Three distinct potassium channels and one anion channel have been revealed by patch clamp studies in Madin-Darby canine kidney cells. The potassium channels are activated by an increase in intracellular calcium activity. A number of hormones activate the potassium channels by an increase in intracellular calcium activity. However, under certain conditions the hormones hyperpolarize the cell membrane without increasing intracellular calcium activity sufficiently to activate the calcium-sensitive potassium channels. Thus, the hormones may activate potassium channels via another, as yet undefined, intracellular mechanism. The anion channel is stimulated by cAMP. Another factor modifying channel activity is cell volume: cell swelling leads probably to subsequent activation of potassium and anion channels. The net result is a variable transient hyperpolarization followed by a sustained depolarization of the cell membrane.     

The surface epithelium of teleostean fish gills. Cellular and junctional adaptations of the chloride cell in relation to salt adaptation

Various species of teleostean fishes were adapted to fresh or salt water and their gill surface epithelium was examined using several techniques of electron microscopy. In both fresh and salt water the branchial epithelium is mostly covered by flat respiratory cells. They are characterized by unusual outer membrane fracture faces containing intramembranous particles and pits in various stages of ordered aggregation. Freeze fracture studies showed that the tight junctions between respiratory cells are made of several interconnecting strands, probably representing high resistance junctions. The organization of intramembranous elements and the morphological characteristics of the junctions do not vary in relation to the external salinity. Towards the base of the secondary gill lamellae, the layer of respiratory cells is interrupted by mitochondria-rich cells ("chloride cells"), also linked to respiratory cells by multistranded junctions. There is a fundamental reorganization of the chloride cells associated with salt water adaptation. In salt water young adjacent chloride cells send interdigitations into preexisting chloride cells. The apex of the seawater chloride cell is therefore part of a mosaic of sister cells linked to surrounding respiratory cells by multistranded junctions. The chloride cells are linked to each other by shallow junctions made of only one strand and permeable to lanthanum. It is therefore suggested that salt water adaptation triggers a cellular reorganization of the epithelium in such a way that leaky junctions (a low resistance pathway) appear at the apex of the chloride cells. Chloride cells are characterized by an extensive tubular reticulum which is an extension of the basolateral plasma membrane. It is made of repeating units and is the site of numerous ion pumps. The presence of shallow junctions in sea water-adapted fish makes it possible for the reticulum to contact the external milieu. In contrast in the freshwater-adapted fish the chloride cell's tubular reticulum is separated by deep apical junctions from the external environment. Based on these observations we discuss how solutes could transfer across the epithelium.