Case report: ranitidine-induced bradycardia in a patient with dextrocardia.

Although rare, bradycardia and other cardiac arrhythmias have been associated with the use of H2-receptor antagonists. Ranitidine is among the most frequently prescribed drugs. In this article, the authors observed a ranitidine-mediated sinus bradycardia in a man with dextrocardia (situs inversus) who had acute bleeding from a duodenal ulcer. The bradycardia was resolved after ranitidine was discontinued.

RT-PCR microlocalization of mRNAs for calbindin D28k and vitamin D receptor in the murine nephron.

The spatial relationship between vitamin D receptor (VDR) and calbindin D28k [calcium binding protein D28k (CaBP-D28k)] gene expression within the murine kidney was studied by localizing their mRNAs in discrete nephron structures using reverse transcription-polymerase chain reaction (RT-PCR). Primers for beta-actin mRNA were used as a control for the presence of tissue during RT-PCR for CaBP-D28k mRNA. mRNA for CaBP-D28k was found only in distal convoluted tubules (DCTs), connecting tubules (CNTs), and cortical collecting ducts (CCDs). In contrast, VDR mRNA was detected in glomeruli, S2 proximal convoluted tubules, cortical thick ascending limbs of Henle's loop, DCTs, CNTs, and initial CCDs. The presence of both VDR and CaBP-D28k mRNA in DCTs, CNTs, and CCDs is consistent with the hypothesis that cacitriol acts via the VDR to stimulate CaBP-D28k synthesis. Conversely, the presence of VDR mRNA in other parts of the nephron suggests that calcitriol has genomically mediated actions within the kidney in addition to stimulation of CaBP-D28k synthesis.

Bone histomorphometry of ovariectomized or orchiectomized rats fed a moderately magnesium-deficient fructose diet and treated with exogenous oestrogen or testosterone.

To investigate the effects of sex hormones on bone histomorphometry, and bone density (BMD), 10 week old ovariectomized (OVX), orchiectomized (ORX), and sham-operated (SHAM) rats fed a moderately magnesium-deficient fructose diet were studied. One third of the OVX and ORX rats were injected with beta-oestradiol-3-benzoate; another third, testosterone cypionate; and the remaining SHAM rats, vehicle only. After 14 weeks, a 24 h urine sample was collected for measurements of calcium, phosphorus and cyclic AMP (cAMP). Blood was collected for determination of calcium, phosphorus, and parathyroid hormone (PTH). Femurs and tibias were removed and weighed. Femurs were used to measure bone areas, mineral contents (BMC), and BMD. Tibias were used for bone histomorphometry (that is, trabecular numbers, thicknesses, % areas and separations). Oestrogen treatment increased serum and urine calcium significantly in both OVX and ORX rats, whereas testosterone decreased serum and urine calcium significantly. Oestrogen decreased urinary cAMP and PTH in both OVX and ORX rats, whereas testosterone treatment increased them significantly. Oestrogen treatment increased BMD, trabecular numbers, thicknesses, and % areas, and decreased bone separations in both OVX and ORX rats. In contrast, testosterone did not increase these bone indices in either OVX or ORX rats; rather, it increased bone separations by decreasing bone strength. Testosterone treatment improved trabecular histomorphometry slightly in OVX rats. The results of the present study are concordant with our previous findings that exogenous oestrogen treatment can prevent osteoporosis in either OVX or ORX rats, whereas exogenous testosterone cannot.

Cloning, genetic mapping, and expression analysis of a mouse renal sodium-dependent phosphate cotransporter.

Renal tubular reabsorption of phosphate is critical to the maintenance of phosphate homeostasis in mammals, and the brush-border membrane Na-P(i) cotransport systems in proximal tubules play a major role in this process. We have isolated a cDNA encoding a mouse sodium-dependent phosphate transport protein (Npt1), which is expressed primarily in the kidney. This protein is highly similar to its human and rabbit homologues, based on nucleotide and amino acid comparisons. The presence of potential Asn-linked glycosylation and protein kinase C phosphorylation sites that are conserved among all three homologues suggests that these sites may be important in the function and regulation of this protein. The Npt1 gene was mapped to mouse chromosome 13, close to the Tcrg locus. By both in situ hybridization and reverse transcription-polymerase chain reaction, Npt1 mRNA was localized predominantly to the proximal tubule.

Parathyroid hormone action in calcium transport in the distal nephron.

Urinary calcium excretion is regulated homeostatically. Regulation is achieved, in part, by the action of parathyroid hormone on Ca2+ absorption in the distal nephron. Parathyroid hormone increases Ca2+ absorption in the cortical portion of the thick ascending limb of Henle's loop in all species studied, in the murine distal convoluted tubule, and in the rabbit connecting tubule. All of these sites contain parathyroid hormone-stimulated adenylate cyclase. Both cellular and paracellular pathways of Ca2+ absorption are regulated by parathyroid hormone in the cortical portion of the thick ascending limb of Henle's loop. In both distal convoluted and connecting tubule cells, parathyroid hormone regulates transcellular Ca2+ absorption by controlling the insertion and open probability of luminal plasmalemmal Ca2+ ion channels. These channels are stimulated and inhibited by L-type calcium channel agonists and antagonists, respectively, but differ from similar channels in excitable cells in that membrane depolarization does not activate them. Parathyroid hormone also increases the driving force for diffusional Ca2+ ion entry from the luminal fluid into the cytosol by increasing the intracellular negative electrical potential (at least in murine distal convoluted tubule cells) by increasing the chloride ion conductance of the basolateral cell membrane. The effects of parathyroid hormone on the other components of cellular Ca2+ transport, via both protein kinases A and C and their interactions, remain to be examined.

Oestrogen but not testosterone increases bone density in orchiectomized rats more when fed moderately magnesium-deficient fructose than moderately magnesium-deficient cornstarch.

To investigate interactions between circulating sex hormones, dietary fructose and magnesium on bone mineral density and numbers of trabeculae, 10 weeks old orchiectomized and sham-orchiectomized rats were studied. One-third of the orchiectomized animals were injected with beta-oestradiol-3-benzoate twice per week in sesame oil; another one-third, testosterone cypionate; the remaining one-third as well as the sham-orchiectomized animals, sesame oil only. All animals were fed either fructose or cornstarch without added magnesium. After 14 weeks, a 24 h urine sample was collected for measurements of calcium, magnesium, phosphorus, and cAMP. Blood was collected for determinations of calcium, magnesium, phosphorus, 25-monohydroxy and 1,25-dihydroxycholecalciferols, oestrogen, testosterone, and parathyroid hormone. Femurs were used for measurements of bone mineral density, and tibiae, for numbers of trabeculae. Exogenous testosterone interacted with starch and magnesium deficiency to decrease serum calcium concentration significantly, which increased circulating parathyroid hormone. High circulating parathyroid hormone raised urinary cAMP and serum 1,25-dihydroxycholecalciferol. Increased parathyroid hormone, cAMP and 1,25-dihydroxycholecalciferol may be responsible for bone resorption which was noted in reductions of bone mineral density and the numbers of trabeculae in the group. In contrast, exogenous oestrogen interacted with fructose and magnesium deficiency to increase serum calcium concentration which caused a reduction of circulating parathyroid. Low parathyroid hormone, reduced 1,25-dihydroxycholecalciferol and cAMP may explain the increased bone mineral density and the numbers of trabeculae in this group.

Vitamin D receptor gene expression in mammalian kidney.

The vitamin D-receptor protein and its mRNA were localized in microscope sections of paraffin-embedded mammalian kidneys by means of immunocytochemistry and in situ hybridization, respectively. A monoclonal antibody against chicken intestinal vitamin D receptor immunostained the nucleus and cytoplasm of cells within the distal convoluted tubule, connecting segment, and initial cortical collecting duct of both rats and pigs. Although fainter, immunostaining also was present over proximal tubular cells. (35S)UTP-labeled cRNA probes were detected over both the proximal and distal portions of the mouse nephron, but silver grain densities were 5.8-fold greater over the latter. In conclusion, localization of both the vitamin D-receptor protein and its mRNA in both the proximal and distal nephron of adult mammals suggests that the gene for this protein is expressed in cells at both of these sites. The intensity of immunostaining and the density of cRNA-associated silver grains suggest that vitamin D-receptor gene expression is greatest in the distal nephron.

A magnesium-deficient high fructose diet augments bone-sparing action of exogenous oestrogen in ovariectomized rats.

To investigate interactions between circulating oestrogen, high dietary fructose, and low dietary magnesium on bone mineral density and numbers of trabeculae, 10 week old ovariectomized (OVX) and sham-ovariectomized (SOVX) rats were studied. The OVX animals were divided into three groups: one-third of the animals were injected with beta-oestradiol-3-benzoate dissolved in sesame oil twice a week; another one-third were injected with testosterone cypionate; and the remaining OVX and all of the SOVX animals were injected with sesame oil only. One-half of the animals in each group were fed cornstarch without magnesium and the other half, fructose without magnesium. After a 14 week experimental period, a 24 h urine sample was collected for measurements of Ca, Mg, P and cAMP. Blood was collected for determination of Ca, Mg, P, 25-hydroxy- and 1,25-dihydroxy-cholecalciferol. Femurs were used for determination of bone density, and tibiae for numbers of trabeculae. Testosterone-treated and OVX control animals fed cornstarch diets had the lowest bone density, whereas oestrogen-treated and SOVX control rats fed fructose had the greatest bone density. Oestrogen-treated animals fed fructose without magnesium had the highest serum and urinary Ca, whereas testosterone-treated animals fed cornstarch without magnesium had the lowest serum and urinary Ca. Serum alkaline phosphatase was higher in OVX- and testosterone-treated and starch-fed animals as compared to their respective counterparts. High urinary cAMP in OVX- and testosterone-treated animals may reflect the action of increased circulating concentrations of PTH, which could be responsible for bone resorption. The results show that high dietary fructose without magnesium interacts with endogenous or exogenous oestrogen to decrease bone mineral loss significantly in ovariectomized rats.

Calbindin-D28k gene expression in the developing mouse kidney.

Calbindin-D28k appears in the metanephric kidney during embryogenesis. We studied the temporal appearance and spatial distribution of calbindin-D28k mRNA in the developing kidneys of 12-day fetal through 21-day postnatal mice by in situ hybridization. 35S-UTP-labeled antisense (cRNA) probe to calbindin-D28k mRNA hybridized to the ureteric buds of 12-day embryos, whereas adjacent metanephrogenic tissue was unlabeled. By embryonic day 13, Y-shaped bodies of "advancing" ureteric buds were labeled intensely. In 16-day embryos, ampullae of ureteric buds were located immediately beneath the renal capsule and labeled strongly, in contrast to metanephric tubules and S-shaped bodies. The former were unlabeled and the latter were labeled only at points of contact with the ampullae. Subsequently, the ampullae of the metanephric ureteric buds hybridized with the cRNA probe, and from the 18th embryonic to the 21st postnatal day, this labeling was intense. The cRNA probe did not hybridize with the renal vesicles, proximal tubules, or tubular segments of Henle's loop derived from nephrogenic blastema, but it did label distal nephron segments. By the 21st postnatal day, collecting ducts and ureter no longer were labeled. In conclusion, calbindin-D28k mRNA is present in the developing mouse kidney, and its distribution during nephrogenesis is identical to that of calbindin-D28k per se. Collectively, these findings show that the calbindin-D28k gene is transcribed and its message is translated by the cells of the ureteric bud during the initial stage of renal morphogenesis.

Immunocytochemical localization of sodium-calcium exchanger in canine nephron.

The sodium-calcium exchanger was localized in tissue sections of canine kidneys with two different polyclonal antisera raised against purified sodium-calcium exchanger protein derived from dog cardiac sarcolemma. The sodium-calcium exchanger was prominent in the basolateral plasmalemma of the majority of cells in all connecting tubules. In contrast, it was observed rarely and inconspicuously over the basolateral aspects of proximal tubular cells. The sodium-calcium exchanger was undetectable in all other portions of the nephron.

Calcium and phosphorus fluxes during hemodialysis with low calcium dialysate.

We evaluated the acute effects of varying dialysate calcium concentration on plasma concentrations and dialyzer fluxes of calcium and phosphorus in adult hemodialysis patients. Seven individuals with stable end-stage renal failure were dialyzed 4 hours, three times weekly. The effects of dialysates containing 1.75, 1.25, or 0.75 mmol/L (70.1, 50.1, or 30.1 mg/L) of calcium were compared. Each patient was studied once at each bath calcium concentration. Compared with the predialysis mean value of 2.27 mmol/L (9.1 mg/dL), plasma total calcium concentration increased, remained constant, or decreased with the 1.75-, 1.25-, or 0.75-mmol/L calcium dialysates, respectively. The 0.75-mmol/L calcium dialysate did not cause signs or symptoms of hypocalcemia (and the plasma calcium concentration did not fall below 1.80 mmol/L [7.2 mg/dL]). Plasma phosphorus concentrations decreased equally from a predialysis mean value of 2.16 mmol/L (6.7 mg/dL), regardless of the dialysate calcium concentration. After 4 hours of treatment with the three different dialysates, the cumulative calcium fluxes were significantly different. With 1.75 mmol/L calcium, mean bodily calcium accumulation was 21.9 mmol (879 mg). With 1.25 mmol/L, there was no net calcium flux. With 0.75 mmol/L, mean patient calcium loss was 5.8 mmol (231 mg). Mean phosphorus removal after 4 hours was 32.5 mmol (1,006 mg) and was unaffected by dialysate calcium concentration.(ABSTRACT TRUNCATED AT 250 WORDS)

Characterization of the renal and intestinal adaptations to dietary calcium deprivation in growing female rabbits.

To characterize the time courses and efficiencies of the renal and intestinal adaptations to dietary Ca deprivation, growing female albino rabbits were fed a low-Ca diet for up to 44 days while they were housed in metabolism cages. Urinary Ca excretion decreased markedly within 8 h of Ca restriction and became essentially undetectable after 14 days. This persisted as long as dietary Ca was low. The intestinal Ca absorption observed during the predeprivation control period was initially changed to net secretion with dietary Ca restriction. However, after 21 days of the low-Ca diet, intestinal Ca absorption was observed. Fecal Ca contents decreased appropriately with Ca deprivation, but never became undetectable. Hyperphosphaturia, which occurred within 8 h of Ca deprivation and persisted as long as dietary Ca was restricted, diminished positive P balance, despite increased intestinal P absorption. Intestinal hyperabsorption and renal conservation of Ca were observed after dietary Ca repletion, suggesting the presence of driving forces for the restitution of Ca deficits incurred by Ca deprivation during growth. We conclude that there are appropriate intestinal and renal homeostatic adaptations to dietary Ca deprivation in the growing female rabbit and that the latter are more rapidly induced and ultimately more efficient than the former.

Basolateral cell membrane Ca-Na exchange in single rabbit connecting tubules.

Studies of cortical proximal nephrons and plasma membrane vesicles suggest that a Ca-Na exchanger regulates intracellular Ca2+ concentration ([Ca2+]i) in renal tubular cells. We tested this hypothesis in isolated perfused rabbit connecting segments by measuring [Ca2+]i with fura-2. Within 2 min of replacing bath NaCl with mannitol, [Ca2+]i rose from a base line of approximately 100 nM to a peak of approximately 650 nM, then declined to a plateau of approximately 500 nM for approximately 5 min before rising to a second peak of approximately 600 nM. [Ca2+]i returned toward base line after restoring bath NaCl. Substitution of choline Cl or tetraethylammonium chloride for bath NaCl reproduced the rise in [Ca2+]i, implicating the Na+ as the mediator. Selective bath (but not lumen) Ca removal or lumen Na deletion virtually abolished these effects, suggesting that bath Na deletion causes peritubular Ca influx by a process that depends on lumen Na. Lumen Na removal lowered, whereas its repletion increased, [Ca2+]i. Smaller increments in [Ca2+]i were produced by raising lumen [Na] from 0 to 35-55 mM or from 20 to 120 mM, but not from 55 to 150 mM. Clamping bath [Ca] at approximately 100 nM abolished the rise in [Ca2+]i produced by lumen Na, corroborating the role of peritubular Ca. These results suggest a Ca influx across the basolateral membrane that is driven by a cell-to-bath [Na] gradient and that can be activated by changes in lumen [Na]. We propose that this process, in part, regulates [Ca2+]i in the rabbit connecting tubule.

cAMP-stimulated rise of [Ca2+]i in rabbit connecting tubules: role of peritubular Ca.

Parathyroid hormone (PTH) increases cytosolic free Ca concentration ([ Ca2+]i) by mechanisms that depend on extracellular Ca in both cultured renal proximal tubules and isolated rabbit connecting tubules (CNTs). In CNTs 8-bromoadenosine 3',5'-cyclic monophosphate (8-BrcAMP) mimics this action, implicating cAMP as a second messenger, and part of the rise, due to increased luminal membrane Ca entry, is likely related to Ca absorption. In cultured proximal tubules the rise in [Ca2+]i, presumably mediated by increased Ca entry across the basolateral plasmalemma, activates gluconeogenesis and shortens microvilli. In the present study we examined cAMP-mediated Ca entry across the basolateral membranes of CNT cells, an effect potentially related to cell activation. Single CNTs were dissected from rabbit kidneys and loaded with fura-2. [Ca2+]i was measured by dual-wavelength excitation during perfusion of isolated segments in vitro. With 1.8 or 2.0 mM Ca in the lumen and the bath, suffusate 8-BrcAMP increased [Ca2+]i within minutes in a dose-dependent fashion. The increase persisted as long as 8-BrcAMP was present and reversed on its withdrawal. With 0.1 microM Ca in the lumen and the bath, 8-BrcAMP, but not ionomycin, failed to increase [Ca2+]i, implying that extracellular Ca is the major source. In tubules perfused with 2 mM ethylene glycol-bis(beta-aminoethyl ether)-N,N,N',N'-tetraacetic acid to eliminate luminal Ca, but suffused with 1.8 or 2.0 mM Ca, 8-BrcAMP increased [Ca2+]i (though less so than with Ca in the lumen), implying Ca entry across basolateral cell membranes. This rise in [Ca2+]i was attenuated markedly by the presence of 50 microM LaCl3 in the bath.(ABSTRACT TRUNCATED AT 250 WORDS)

Effects of moderate dietary phosphorus restriction on intestinal absorption and external balances of phosphorus and calcium in growing female rabbits.

The effects of moderate dietary P restriction on intestinal net fluxes and external balances of P and Ca were studied in growing female albino rabbits that were fed a P-deficient diet for 10 consecutive days while they were housed in metabolism cages. Intestinal P secretion occurred during the first 24 h of P restriction and thereafter changed to absorption. During recovery, when the rabbits were consuming a normal diet, P absorption was significantly greater than either prerestriction control values or than in a separate group of time control rabbits. Phosphorus balance was negative during the 1st day of P restriction but thereafter became positive. This pattern occurred because intestinal P secretion changed to absorption and because urinary losses of P were negligible. Intestinal Ca absorption increased within 24 h of P restriction, reached maximal values by 4 days, and remained elevated for each of the remaining 6 days that dietary P was low. It also was elevated compared to P-sufficient time controls for 8 days after replenishment of dietary P. Despite increased intestinal absorption, Ca balance was significantly reduced during P restriction because of substantial hypercalciuria. Thus, selective dietary P restriction reduced the positive balances of both P and Ca that are characteristic of growth. We conclude that in growing rabbits moderate dietary P restriction induces both intestinal and renal adaptations that conserve this mineral; concomitantly, positive Ca balance is reduced. With dietary P replenishment, adaptations persist to restore the positive mineral balances that were lost because of dietary P restriction during growth.

Ca2+ absorption in the pars recta of cortical S2 rabbit proximal tubules: role of diffusion.

Individual partes rectae of cortical S2 proximal tubules were dissected from rabbit kidneys and perfused in vitro. The bath was a protein-free, plasmalike solution, whereas the perfusate simulated "late" proximal tubular fluid (high [Cl-], low [HCO3-], absence of Na+-cotransported solutes). Transepithelial voltages at the perfusion and collection ends were measured simultaneously in doubly cannulated tubules. Ionized calcium concentrations in perfusates, collectates, and the bath were calculated from measurements of calcium ion activities made in situ with Ca2+-selective microelectrodes. Epithelial calcium permeability was estimated from the bath-to-lumen movement of 45Ca. Total calcium concentrations of perfused and collected fluids were measured by continuous-flow microcolorimetry. Under these conditions there was a lumen-positive transepithelial voltage that was equal at both ends of the tubules. Luminal [Ca2+] [corrected], which decreased along the length of the tubule, was greater than that of the bath. Using the measured calcium permeability, we calculated the diffusional component of the net calcium absorptive flux from the mean transepithelial delta [Ca2+] and the measured transepithelial voltage. We found that ionic diffusion accounted for the net flux, suggesting that diffusion is the primary mechanism of calcium absorption in this segment of the rabbit nephron.

Evidence for cAMP-dependent protein kinase in mediating the parathyroid hormone-stimulated rise in cytosolic free calcium in rabbit connecting tubules.

We investigated cellular mechanisms mediating the parathyroid hormone (PTH)-induced increase in cytosolic free Ca2+ concentration ([Ca2+]i) in isolated perfused rabbit connecting tubules. Prior and/or concomitant exposure to 0.5 mM of N-[2-(methylamino)ethyl]-5-isoquinolinesulfonamide dihydrochloride (H-8), a cyclic nucleotide-dependent protein kinase inhibitor, abolished the rise in [Ca2+]i produced by 0.1 nM PTH in five connecting tubules and suppressed it by approximately 50% in another five. In the latter, there was a delayed onset in the rise of [Ca2+]i. Such responses contrasted to the prompt increase in [Ca2+]i in PTH-stimulated control tubules. However, when H-8 was withdrawn, [Ca2+]i rose within minutes to reach a plateau value similar to the uninhibited response to PTH in controls, indicating rapidly reversible inhibition by H-8. In an otherwise identical protocol, 0.5 mM H-8 also reversibly suppressed the rise in [Ca2+]i induced by 0.175 mM 8-Br-cAMP. In contrast to the stimulatory effect of 8-Br-cAMP on [Ca2+]i, 1 mM 8-Br-cGMP caused no increase. At a concentration of 0.4 mM, the Rp diastereomer of adenosine cyclic 3',5'-phosphorothioate (Rp-cAMPS), a well-characterized cAMP-dependent protein kinase inhibitor, totally abolished the rise in [Ca2+]i caused by 0.1 nM PTH. We conclude that a cAMP-dependent protein kinase plays an important role in the PTH-stimulated rise in [Ca2+]i in the rabbit connecting tubule. Since the increase in [Ca2+]i was shown previously to depend on extracellular Ca2+, we propose that cAMP-dependent protein phosphorylation is important in mediating PTH-stimulated Ca2+ fluxes across plasma membranes of connecting tubule cells.

Effects of parathyroid hormone on cytosolic free calcium concentration in individual rabbit connecting tubules.

PTH stimulates active Ca reabsorption in isolated perfused rabbit kidney connecting tubules (CNTs). The existence of PTH-sensitive adenylate cyclase and the reproduction of increased epithelial Ca transport by dibutyryl-cAMP suggest that cAMP is the mediator. Accordingly, we studied the effects of PTH and 8-bromoadenosine 3',5'-cAMP (8-Br-cAMP) on cytosolic free calcium concentration [( Ca2+]i) in individual rabbit CNTs. [Ca2+]i was estimated by continuous epifluorescence microscopy of single fura-2-loaded tubules during dual wave-length excitation. In nonperfused controls at 37 degrees C, [Ca2+]i decreased with time. In contrast to vehicle controls, synthetic bovine (1-34) PTH (0.1 nM) increased [Ca2+]i within 4 min, produced a maximal effect in 7.2 min, and sustained its effect for at least 2 min after washout. 8-Br-cAMP (1 mM) mimicked the effect of PTH, but with an earlier onset of action. To test the hypothesis that lumen Ca is the predominant source of the rise in [Ca2+]i, we studied singly perfused CNTs. In the absence of bath and lumen Ca, PTH elicited no rise in [Ca2+]i, implying that intracellular Ca stores are not the major source. In contrast, there was a rise when Ca was replenished in both media. In the continuous presence of bath Ca, lumen Ca was estimated to contribute 65% of the total rise in [Ca2+]i in response to PTH when it was first deleted and then replenished. However, when the sequence of lumen Ca manipulation was reversed, the contributions by lumen and bath Ca were found to be essentially equal. We conclude (a) at a physiologic concentration, PTH increases [Ca2+]i in rabbit CNTs, (b) 8-Br-cAMP mimics this action, implicating cAMP as a second messenger, and (c) the PTH-stimulated rise in [Ca2+]i depends importantly on both bath and tubular luminal fluid Ca.