Microinjection studies on the effect of furosemide on the rat nephron

The tubular site of furosemide (F) action was studied by the technique of diuretic microinjection (MIJ) into proximal tubules of the rat nephron. F was injected into the last proximal superficial loop of 51 proximal tubules, at the concentration of 3 · 10^-4 mol/l in an infusate that contained ¹⁴C-inulin. Collections were performed at the early distal tubule before and after MIJ. Single nephron filtration rate (SNGFR) remained unchanged, while the percent of filtered volume reabsorbed up to the site of collection was 85 ± 2 before, 79 ± 2% after MIJ, p < 0.005. The calculated concentration of F in the collected distal tubular fluid during the post-MIJ measurements averaged 3 · 10^-5 mol/l. This experiment was repeated by injecting F into the early proximal convolutions of 43 nephrons, while the collections were performed at the last proximal segments. In these studies of proximal volume absorption, SNGFR was 34 ± 3 before, 35 ± 4 nl/min after F (p > 0.6). The respective percent reabsorptions were 70 ± 3 and 73 ± 3% (p ± 0.3). In order to determine whether the technique per se was suitable to detect changes in reabsorption, the proximal MIJ study was repeated by using the carbonic anhydrase inhibitor dichlorphenamide 3 · 10^-5 mol/l in the microinjectate: while SNGFR remained unchanged, percent reabsorption fell from 63+5 to 45 ± 7% during injection of the diuretic, p < 0.03. We conclude that the technique is adequate to examine the effects of drugs, and that F does not reduce proximal volume absorption at concentrations of 3 · 10^-5 mol/l. The loop diuretic decreases distal volume absorption by abolishing the osmotic gradient between blood and tubular fluid along the early distal convoluted tubule.