Altered density of glomerular binding sites for atrial natriuretic factor in bile duct-ligated rats with ascites.

The renal response to atrial natriuretic factor is blunted in cirrhosis with ascites. This might be due to alterations of renal receptors for atrial natriuretic factor. Therefore density and affinity of glomerular atrial natriuretic factor binding sites of bile duct-ligated rats with ascites (n = 10) and of sham-operated controls (n = 10) were determined. Glomerular atrial natriuretic factor binding sites were identified to be of the B-("biologically active") and C-("clearance") receptor type. Discrimination and quantitative determination of B and C receptors for atrial natriuretic factor were achieved by displacement experiments with atrial natriuretic factor(99-126) or des(18-22)atrial natriuretic factor(4-23), an analogue binding to C receptors only. Density of total glomerular atrial natriuretic factor binding sites was significantly increased in bile duct-ligated rats (3,518 +/- 864 vs. 1,648 +/- 358 fmol/mg protein; p less than 0.05). This was due to a significant increase of C-receptor density (3,460 +/- 866 vs. 1,486 +/- 363 fmol/mg protein; p less than 0.05), whereas density of B receptors was not significantly different in bile duct-ligated rats (58 +/- 11 vs. 162 +/- 63 fmol/mg protein). Affinity of atrial natriuretic factor to its glomerular binding sites did not differ significantly between both groups. These data suggest that an altered glomerular atrial natriuretic factor receptor density could be involved in the renal resistance to atrial natriuretic factor in cirrhosis with ascites.

Discrimination and quantification of glomerular receptor subtypes for atrial natriuretic factor (ANF).

Binding sites for atrial natriuretic factor (ANF) were determined on isolated rat glomeruli as well as on glomerular membranes. To define optimal conditions, binding of ANF was investigated varying incubation time, temperature and protein concentration. Binding conditions were found to be best at 4 degrees C for 5 hours with 15 micrograms of glomerular protein. Saturation and affinity cross-linking experiments confirmed the presence of two distinct receptor subtypes-the B-receptor (130 kDa) and the C-receptor (65 kDa). Quantitative differentiation of both ANF binding sites was achieved by competitive displacement with two different unlabeled ANF ligands: a) rANF(99-126) (homologous displacement), b) des(18-22)rANF(4-23)NH2 (heterologous displacement). Intact glomeruli and glomerular membranes did not differ significantly in receptor density for the B-receptor (71 +/- 37 vs. 94 +/- 53 fmol/mg protein) or the C-receptor (976 +/- 282 vs. 966 +/- 167 fmol/mg protein) or in affinity constants for the B-receptor (43 +/- 36 vs. 52 +/- 44 pM) or the C-receptor (876 +/- 377 vs. 307 +/- 36 pM). Glomerular membranes compared to glomeruli showed less nonspecific binding and less intra-assay variation of measuring points done in triplicates. This method of selective displacement should allow to study the influence of various physiological and pathophysiological conditions on the binding properties of B- and C-receptors for ANF.

Dehydration increases the density of C receptors for ANF on rat glomerular membranes.

The present study determined the presence of two types of binding sites for atrial natriuretic factor (ANF), the B and C receptor, on rat glomerular membranes. The effect of short-term salt loading and dehydration on these two receptor populations was investigated consecutively. Salt-loaded rats did not show significant changes in plasma ANF concentrations or in the number of ANF binding sites. Water-deprived rats presented significantly lower plasma ANF concentrations (22.0 +/- 1.9 vs. 34.4 +/- 3.8 fmol/ml, P less than 0.01) and an increase in total receptor density (1,860 +/- 398 vs. 987 +/- 143 fmol/mg protein) as compared with the control group. Differentiation of both receptor populations showed that it was the C receptors that accounted for this increase (1,772 +/- 369 vs. 901 +/- 151 fmol/mg protein, P less than 0.05), whereas B-receptor density was unchanged (89 +/- 31 vs. 87 +/- 44 fmol/mg protein). These data suggest that C receptors for ANF are affected by changes of body fluid volume.

Mu-induced rifamycin-resistant mutations not located in the rpoB gene of Escherichia coli.

A new class of rifamycin-resistant mutants of Escherichia coli was obtained by lysogenic insertions of bacteriophage Mu Amp DNA. Rifamycin resistance is closely linked to the ampicillin resistance conferred by the prophage. Mapping by conjugation with auxotrophic markers revealed that the rifamycin-resistant mutations are located between 28 and 37 min on the E. coli chromosome standard map, some distance from the rpoB gene at 89.5 min. The DNA-dependent RNA polymerase of these mutants is highly sensitive to rifampicin.