The urothelium of a hibernator: the American black bear.

The American black bear undergoes a 3-5 month winter hibernation during which time bears do not eat, drink, defecate, or urinate. During hibernation renal function (GFR) is 16-50% of normal but urine is reabsorbed across the urinary bladder (UB) urothelium thus enabling metabolic recycling of all urinary constituents. To elucidate the mechanism(s) whereby urine is reabsorbed, we examined the UBs of five nonhibernating wild bears using light, electron (EM), and confocal immunofluorescent (IF) microscopy-concentrating on two components of the urothelial permeability barrier - the umbrella cell apical membranes and tight junctions (TJ). Bear UB has the same tissue layers (serosa, muscularis, lamina propria, urothelia) and its urothelia has the same cell layers (basal, intermediate, umbrella cells) as other mammalians. By EM, the bear apical membrane demonstrated a typical mammalian scalloped appearance with hinge and plaque regions - the latter containing an asymmetric trilaminar membrane and, on IF, uroplakins Ia, IIIa, and IIIb. The umbrella cell TJs appeared similar to those in other mammals and also contained TJ proteins occludin and claudin - 4, and not claudin -2. Thus, we were unable to demonstrate urothelial apical membrane or TJ differences between active black bears and other mammals. Expression and localization of UT-B, AQP-1 and -3, and Na(+), K(+)-ATPase on bear urothelial membranes was similar to that of other mammals. Similar studies of urothelia of hibernating bears, including evaluation of the apical membrane lipid bilayer and GAGs layer are warranted to elucidate the mechanism(s) whereby hibernating bears reabsorb their daily urine output and thus ensure successful hibernation.

Hydration status affects sodium, potassium, and chloride transport across rat urothelia.

Recent data suggest possible net transport of urinary constituents across mammalian urinary tract epithelia (urothelia). To evaluate the effect of animal hydration status on such transport, we instilled urine collected during 2-day water deprivation, water loading, or ad libitum water intake into isolated in situ bladder(s) of groups of rats undergoing one of the same three hydration states. After 1-h bladder dwell, we retrieved the urine and measured differences in volume and solute concentrations between instilled and retrieved urine. We previously reported results regarding changes in urine volume and net urea and creatinine transport and herein report the results of net urinary sodium, potassium, and chloride transport in the same animals. During water-loading conditions, urinary concentrations of Na, K, and Cl rose 4.9 (30.7%), 2.6 (16.5%), and 6.0 meq/l (26.8%), respectively, indicating urothelial secretion into urine. During ad libitum water intake, urinary K and Cl concentrations fell 33.6 (14.8%) and 28.4 meq/l (12%), respectively (Na did not change), and during water deprivation urine Na, K, and Cl concentrations fell dramatically by 53.2 (18.6%), 159.4 (34.6%) and 133.7 meq/l (33.8%), respectively, reflecting urothelial reabsorption of each ion. For each ionic species, two factors independently influenced transport: instilled urinary ion concentration and animal hydration state. These results demonstrate significant regulated ion transport across mammalian urothelia, support the notion that lower urinary tract modifies final urine, and suggest that the lower urinary tract may play a role in local and whole animal solute homeostasis.

Dietary protein affects urea transport across rat urothelia.

Recent evidence suggests that regulated solute transport occurs across mammalian lower urinary tract epithelia (urothelia). To study the effects of dietary protein on net urothelial transport of urea, creatinine, and water, we used an in vivo rat bladder model designed to mimic physiological conditions. We placed groups of rats on 3-wk diets differing only by protein content (40, 18, 6, and 2%) and instilled 0.3 ml of collected urine in the isolated bladder of anesthetized rats. After 1 h dwell, retrieved urine volumes were unchanged, but mean urea nitrogen (UN) and creatinine concentrations fell 17 and 4%, respectively, indicating transurothelial urea and creatinine reabsorption. The fall in UN (but not creatinine) concentration was greatest in high protein (40%) rats, 584 mg/dl, and progressively less in rats receiving lower protein content: 18% diet, 224 mg/dl; 6% diet, 135 mg/dl; and 2% diet, 87 mg/dl. The quantity of urea reabsorbed was directly related to a urine factor, likely the concentration of urea in the instilled urine. In contrast, the percentage of instilled urea reabsorbed was greater in the two dietary groups receiving the lowest protein (26 and 23%) than in those receiving higher protein (11 and 9%), suggesting the possibility that a bladder/urothelial factor, also affected by dietary protein, may have altered bladder permeability. These findings demonstrate significant regulated urea transport across the urothelium, resulting in alteration of urine excreted by the kidneys, and add to the growing evidence that the lower urinary tract may play an unappreciated role in mammalian solute homeostasis.

Hydration status affects urea transport across rat urothelia.

Although mammalian urinary tract epithelium (urothelium) is generally considered impermeable to water and solutes, recent data suggest that urine constituents may be reabsorbed during urinary tract transit and storage. To study water and solute transport across the urothelium in an in vivo rat model, we instilled urine (obtained during various rat hydration conditions) into isolated in situ rat bladders and, after a 1-h dwell, retrieved the urine and measured the differences in urine volume and concentration and total quantity of urine urea nitrogen and creatinine between instilled and retrieved urine in rat groups differing by hydration status. Although urine volume did not change >1.9% in any group, concentration (and quantity) of urine urea nitrogen in retrieved urine fell significantly (indicating reabsorption of urea across bladder urothelia), by a mean of 18% (489 mg/dl, from an instilled 2,658 mg/dl) in rats receiving ad libitum water and by a mean of 39% (2,544 mg/dl, from an instilled 6,204 mg/dl) in water-deprived rats, but did not change (an increase of 15 mg/dl, P = not significant, from an instilled 300 mg/dl) in a water-loaded rat group. Two separate factors affected urea nitrogen reabsorption rates, a urinary factor related to hydration status, likely the concentration of urea nitrogen in the instilled urine, and a bladder factor(s), also dependent on the animal's state of hydration. Urine creatinine was also absorbed during the bladder dwell, and hydration group effects on the concentration and quantity of creatinine reabsorbed were qualitatively similar to the hydration group effect on urea transport. These findings support the notion(s) that urinary constituents may undergo transport across urinary tract epithelia, that such transport may be physiologically regulated, and that urine is modified during transit and storage through the urinary tract.

The ROMK potassium channel is present in mammalian urinary tract epithelia and muscle.

There is increasing evidence that mammalian urinary tract epithelial cells utilize membrane channels and transporters to transport solutes across their apical (luminal) and basalateral membranes to modify solute concentrations in both cell and urine. This study investigates the expression, localization, and regulation of the ROMK (K(ir) 1.1) potassium channels in rat and dog ureter and bladder tissues. Immunoblots of homogenates of whole ureter, whole bladder, bladder epithelial cells, and bladder smooth muscle tissues in both rat and dog identified approximately 45- to 50-kDa bands characteristic of ROMK in all tissues. RT-PCR identified ROMK mRNA in these same tissues in both animal species. ROMK protein localized by immunocytochemistry was strongly expressed in the apical membranes of the large umbrella cells lining the bladder lumen and to a lesser extent in the cytoplasm of epithelial cells and smooth muscle cells in the rat bladder. ROMK protein and mRNA were also discovered in cardiac, striated, and smooth muscle in diverse organs. There was no difference in immunoblot expression of ROMK abundance in bladder homogenates (whole bladder, epithelial cell, or muscle cell) or ureteral homogenates between groups of rats fed high- or low-potassium diets. Although the functional role of ROMK in urinary tract epithelia and smooth muscle is unknown, ROMK may participate in the regulation of epithelial and smooth muscle cell volume and osmolality, in the dissipation of potassium leaked or diffused from urine across the epithelial cell apical membranes or tight junctions, and in net or bidirectional potassium transport across urinary tract epithelia.

High urea and creatinine concentrations and urea transporter B in mammalian urinary tract tissues.

Although the mammalian urinary tract is generally held to be solely a transit and storage vehicle for urine made by the kidney, in vivo data suggest reabsorption of urea and other urine constituents across urinary tract epithelia. To determine whether urinary tract tissue concentrations are increased as a result of such reabsorption, we measured urea nitrogen and creatinine concentrations and determined whether urea transporter B (UT-B) was present in bladder, ureter, and other tissues from dogs and rats. Mean urea nitrogen and creatinine concentrations in dogs and rats were three- to sevenfold higher in urinary tract tissues than in serum and were comparable to those in renal cortex. In water-restricted or water-loaded rats, urea nitrogen concentrations in bladder tissues fell inversely with the state of hydration, were proportional to urine urea nitrogen concentrations, and were greater than the corresponding serum urea nitrogen concentration in every animal. Immunoblots of rat and dog urinary tract tissues demonstrated the presence of UT-B in homogenates of bladder and ureter, and immunocytochemical analysis localized UT-B to epithelial cell membranes. These findings are consistent with the notion that urea and creatinine are continuously reabsorbed from the urine across the urothelium, urea in part via UT-B, and that urine is thus altered in its passage through the urinary tract. Urea reabsorption across urinary tract epithelia may be important during conditions requiring nitrogen conservation and may contribute to pathophysiological states characterized by high blood urea nitrogen, such as prerenal azotemia and obstructive uropathy.

Successful treatment of calciphylaxis with intravenous sodium thiosulfate.

Calciphylaxis is a dreaded complication of renal failure characterized by nodular subcutaneous calcification and painful tissue necrosis often leading to ulceration, secondary infection, and high mortality rates. The case of a woman receiving continuous ambulatory peritoneal dialysis who had a typical clinical presentation of calciphylaxis confirmed by x-ray and technetium scan findings is described. After nonresponse to conventional therapy, treatment with intravenous sodium thiosulfate 3 times weekly was begun, and she had rapid and dramatic relief of signs and symptoms and improvement of technetium scans. Prolonged treatment was well tolerated without serious side effects and accompanied by continued clinical improvement.

Expression, localization, and regulation of urea transporter B in rat urothelia.

Although mammalian urothelia are generally considered impermeable to urinary constituents, in vivo studies in several species suggest urothelial transport of water, urea, and solutes under certain conditions. This study investigates the expression, localization, and regulation of urea transporter-B (UT-B) in rat renal pelvis, ureter, and bladder tissues. Immunoblots of homogenates of tissues identified characteristic approximately 40- to 55- and approximately 32-kDa bands in the ureter, bladder, and renal inner medulla, but not renal cortex. UT-B was localized by immunocytochemistry and was strongly expressed in all cell membranes (and to a limited extent in intracellular vesicles in the cytoplasm) of epithelial cells lining the rat bladder, ureter, and renal pelvis lumens except the apical membrane of the umbrella cells. It was also present in single-layer papillary surface epithelial cells. There was no difference in immunoblot expression of UT-B in the bladder or ureteral homogenates between groups of rats fed high- or low-protein or high- or low-sodium diets. Water restriction resulted in an increase in UT-B expression in ureters (49%, P = 0.001) but not in bladders (14%, P = not significant). The functional role of UT-B in the genitourinary tract epithelia is unknown. UT-B may participate in the regulation of epithelial cell volume and osmolality, in the dissipation of urea gradients, and in possible net urea transport across uroepithelia.

Expression, localization, and regulation of aquaporin-1 to -3 in rat urothelia.

Although mammalian urothelia are generally considered impermeable to constituents of urine, in vivo studies in several species indicate urothelial transport of water and solutes under certain conditions. This study investigates the expression, localization, and regulation of aquaporin (AQP)-1, -2, and -3 in ureteral and bladder tissues in 48-h dehydrated and water-loaded female Wistar rats. Immunoblots of homogenates of whole ureter and bladder identified characteristic approximately 28- and 35- to 44-kDa bands for AQP-1, -2, and -3. AQP-1 was localized to capillary and arteriole endothelial cells, whereas AQP-2 and -3 circumferentially lined the epithelial cell membranes except for the apical membrane of the epithelial cells adjacent to the lumens of both ureter and bladder. AQP-2 was also present in epithelial cell cytoplasm. Dehydration resulted in 160-200% increases of AQP-3 signal and 24-49% increases of AQP-2 signal but no change in AQP-1 signal on immunoblots of homogenates of ureters and bladders. AQPs in genitourinary tract urothelia likely play a role in the regulation of epithelial cell volume and osmolality and may play a role in bulk water movement across urothelia.