摘要:

A simple device for fluorescence measurements on kidney surface in situ (FKS) is described. The device consists of (1) an illuminating unit, (2) a detector, (3) an evaluation unit. The device was developed for measurement of tissue content of anionic sulfofluorescein and cationic 4(4-dimethylaminostyryl)-N-methylpyridinium (ASP) during its secretion by proximal renal tubular cells.

ISSN:1420-4096    

DOI:10.1159/000173788

出版商:S. Karger AG    

年代:1994

数据来源: Karger

摘要:

The influence of passage number and different culture conditions on the effect of ATP and angiotensin II (A II) on membrane voltage (Vm) of rat mesangial cells (MC) was examined with the patch clamp technique in slow and fast whole cell recordings. MC were characterized immunologically and grown in standard medium in primary culture (PC) and long-term culture up to passage 21 in the presence of 90 g/l fetal calf serum (LTC/ +FCS) or without or with 5 g/l FCS for 1-3 days (LTC/-FCS). In all three series the studies were performed in a FCS-free Ringer-like solution. Vm of MC did not differ in the series (PC: -49 ± lmV, n = 151; LTC/+FCS: -52 ± 1 mV, n = 49; LTC/-FCS: -51 ± 1 mV, n = 44). In primary culture and long-term cultured MC up to passage 8, FCS (ED50≈5 g/l), ATP (ED50 ≈ 2 × 10-6 mol/l) and AII (ED50 ≈ 5 × 10-10 mol/l) induced a depolarization of Vm. Reduction of extracellular Cl- concentration (from 145 to 32 mmol/l) had no effect on Vm but led to an increased depolarization of Vm by FCS, ATP and A II. In long-term cultured MC above passage 8 grown with 90 g/l FCS both ATP and AII induced a concentration-dependent hyperpolarization of Vm, which was attenuated in increased extracellular K+ concentration (from 3.6 to 33.6 mmol/l). In long-term cultured MC beyond passage 8, grown without or with a reduced FCS concentration of 5 g/l, ATP and A II led to a transient depolarization of Vm, which was increased in the presence of 32 mmol/l extracellular Cl-. The depolarization was followed by a hyperpolarization, which was attenuated in the presence of increased extracellular K+. The data indicate that vasoactive agents depolarize Vm of MC in primary culture by activating a Cl- conductance whereas they hyperpolarize Vm by activation of a K+ conductance in long-term cultured MC grown with FCS. The latter effect was partially reversed when FCS w

ISSN:1420-4096    

DOI:10.1159/000173789

出版商:S. Karger AG    

年代:1994

数据来源: Karger

摘要:

Nord et al. [Am J Physiol 1986;250:F539-F550] proposed a method to give a high yield of proximal tubule cells by exposing a suspension of rabbit cortical tubules to a hypotonic shock in calcium-free media. The present study desribes the effects of both amplitude and duration of the hypotonic treatment on some transport-related characteristics of individual cells as compared to the starting tubule suspension. The averaged cell yield increased by an order of magnitude when the osmolality of the hypotonic solution was varied in four steps from 200 (C200 cells) to 70 mosm/kg H2O (C70 cells) while the proportion of trypan blue-positive cells progressively decreased from 33% for C200 cells to 9.5% for C70 cells. An increase in duration of the hypotonic shock from 0.5 to 6 min did not change the cell yield of C200 cells while it significantly increased that of C70 cells by 61%. Basal and ouabain-sensitive oxygen consumption (QO2) increased by 57 and 155%, respectively, from C70 to C200 cells but was approximately one order of magnitude smaller than the QO2 measured for tubule suspension. Intracellular ATP content averaged 5.5 ± 0.8 nmol/mg for the starting tubule suspension, 4.6 ± 0.8 nmol/mg for C70 cells but only 1.3 ± 0.1 nmol/mg for C200 cells. The maximal velocity for phloridzin-sensitive α-methyl glucose transport averaged 13.7 ± 1.7 nmol min-1 mg-1 for C70 cells and only 6.3 ± 1.3 nmol min-1 mg-1 for C200 cells which is approximately one order of magnitude smaller than what can be expected from a tubule presenting a good access to luminal membrane. We conclude from these results that, in the process of isolating individual cells from a polarized epithelium, membrane transport rates have decreased by one order of magnitude and this reduction is intensified by a large hypotonic shock. In comparison with C200 cells, the cells obtained with a large hypotonic shock give a high yield, a larger proportion of trypan blue-negative cells and their lower overall transport rate allows the cells to maintain a better electrochemical gradient for Na and a higher intracellular ATP

ISSN:1420-4096    

DOI:10.1159/000173790

出版商:S. Karger AG    

年代:1994

数据来源: Karger

摘要:

The ability of a renal epithelial cell line of porcine origin (LLC-PK1) to synthesize dopamine (DA) from L-dopa and release DA into the culture media was studied. L-Dopa was rapidly taken up by LLC-PK1 cells and converted to DA which was then released into the media. L-Dopa uptake and conversion to DA was competitively inhibited by other aromatic amino acids (Tyr and Phe). Studies were conducted to assess L-dopa uptake, DA synthesis and DA release in LLC-PK1 cells in response to changes in the ionic composition of the Krebs-Ringer bicarbonate (KRB) buffer media and to various drugs known to alter renal transport function. Sodium chloride addition to KRB media resulted in enhanced DA release into the media that was significantly attenuated by both quinine (1 mM) and benzamil (0.1 mM). Other sodium salts (sodium acetate and sodium phosphate) also enhanced DA release from LLC-PK1 cells. Salts containing chloride (LiCl, NH4Cl, choline chloride) stimulated a greater efflux of DA from LLC-PK1 cells than sodium salts at equimolar concentrations. Osmotic stimuli (sucrose, mannitol and dextran, 50-200 mM) also elicited increased DA release from LLC-PK1cells into the media but not to the same extent as inorganic salts. DA release (secretion) from LLC-PK1 cells was strongly inhibited (25-50%) by drugs known to be substrates for the renal cation transport system. These studies have characterized extensively the factors that govern and regulate the synthesis of DA from L-dopa and the transport system responsible for the secretion (release) of intracellular DA into the media.

ISSN:1420-4096    

DOI:10.1159/000173791

出版商:S. Karger AG    

年代:1994

数据来源: Karger

摘要:

Serial serum atrial natriuretic peptide (ANP) determinations were performed in 15 uninephrectomized Charles River rats and in 15 sham-operated control animals during the 60 days following surgery. In a second group of 7 control and 7 uninephrectomized animals housed in metabolism cages, serum ANP, body weight, 24-hour urine volume, osmolality and sodium excretion were serially measured. In a third group of uninephrectomized and control rats the effect of acute salt loading 24 h, 6 and 60 days after surgery on serum ANP was studied. No significant changes in ANP levels were observed during the 60 days following surgery in control animals. In the uninephrectomized animals a sharp drop in basal ANP levels was evident 24 and 48 h after surgery, but increased levels of serum ANP were seen from day 6 to 28. Thereafter ANP returned to baseline levels for the rest of the study period. Urinary sodium excretion decreased in the nephrectomized animals on days 1 and 2 following surgery. No such change was seen in the control animals during the same period. Body weight, 24-hour urine volume and urine osmolality were not statistically different in the nephrectomized vs. control rats at any time and remained constant in each group throughout the experimental period. Central venous pressure (CVP) did not change significantly in both groups 24 h and 6 days following surgery. CVP rose similarly in both groups immediately following saline loading and returned to preload levels 1 h later. Nephrectomized animals showed no change in serum ANP 1 h after saline loading performed 24 h after surgery, while in control animals acute saline load resulted, as expected, in a significant rise in serum ANP. 6 and 60 days following surgery, augmented ANP release 1 h after acute salt loading was evident in both the nephrectomized and control groups. We suggest that some as yet unknown mechanism exists through which the renal mass contributes to the control of ANP secretion by the cardiac atria.

ISSN:1420-4096    

DOI:10.1159/000173792

出版商:S. Karger AG    

年代:1994

数据来源: Karger

摘要:

We studied basal serum atrial natriuretic peptide (ANP) levels and the response to acute salt loading in rats with different grades of functional renal mass reduction. The six groups of rats studied included 2 controls, i.e. unoperated (n = 12) and sham-operated (n = 24) groups. Each of the 4 experimental groups (n = 24 in each) underwent one of the following procedures: bilateral nephrectomy; unilateral nephrectomy; bilateral ureteral ligation; unilateral ureteral ligation. Basal ANP was assessed in the intact controls and operated groups 4, 8,16 and 24 h after surgery. In addition, ANP was determined in the sham-operated controls and in the experimental groups 1 h following acute intravenous saline loading performed 4 h after surgery with central venous pressure monitoring. Basal ANP dropped significantly following bilateral nephrectomy but was not significantly altered after unilateral nephrectomy or the two modalities of ureteral ligation . In all 4 experimental groups ANP failed to rise after saline loading. We conclude that substantial renal damage results in early impairment in ANP secretion suggesting the existence of a renal physiological stimulus controlling ANP release by cardiac atria.

ISSN:1420-4096    

DOI:10.1159/000173793

出版商:S. Karger AG    

年代:1994

数据来源: Karger