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Free keywords:
Tubular Cell; Fluid Absorption; Proximal Tubular Cell; Polyethyleneimine; Protamine Sulfate
Abstract:
The present study was initiated with the hope of clarifying the role of negative charges in the luminal brush border membrane in the overall process of transepithelial isotonic sodium and water absorption. Using micropuncture techniques, cationic polyamino acids such as polylysine (mol wt 100,000, 17,000 and 1,500–5,000, 1 mg/ml), tetralysine, polyornithine (mol wt 100,000, 1 mg/ml), polyethyleneimine (2 mg/ml), polymyxin B (2 mg/ml), protamine sulfate (25 mg/ml) and histone (0.5 mg/ml) were perfused through the segments of rat kidney proximal tubule for 30 sec to 2 min. The rate of isotonic fluid absorption was measured before and after each perfusion with the Gertz's split drop method using Ringer's solution as a shrinking drop. Polylysine 100,000 and 17,000 and polyornithine were the most potent, inhibiting isotonic reabsorption by 93%. The sequence of inhibitory effect was: polylysine 100,000≃polyornithine 100,000≃polylysine 17,000>polyethyleneimine>polylysine 1,500–5,000≃polymyxin B> protamine sulfate ≃ histone. In contrast, tetralysine (2 mg/ml) showed no inhibitory effect. Electrical potential difference (p.d.) of the proximal tubular cells was destroyed within 10 sec of luminal perfusion with polylysine 100,000 (1 mg/ml). Simultaneously with the drop in p.d., electrical resistance of the luminal brush border membrane was nearly totally eliminated, whereas transepithelial input resistance remained unaltered. Furthermore, trypan blue dye was taken up by polylysine 100,000-perfused tubular cells but not by normal cells. Expanding drop analysis (mannitol solution as a split drop) was performed as a screening test to examine if the permeability for water and sodium in the lateral paracellular pathway is altered by polylysine 100,000. No significant difference was observed in the velocity of split drop expansion between untreated and polylysine-perfused tubules. A lower concentration of polylysine 100,000 (0.1 mg/ml) showed a much less inhibitory effect on fluid absorption and on cell p.d. These observations indicate that the strong inhibition on proximal tubular fluid absorption exerted by polylysine and perhaps also by other cationic polyamino acids is due not to modification of membrane negative charges but to the lysis of tubular cells by these polycations.
