EFFECT OF PROTAMINE ON ION CONDUCTANCE OF UPPER PORTION OF DESCENDING-LIMB OF LONG-LOOPED NEPHRON FROM HAMSTERS

To estimate the contribution of paracellular shunt pathway to the cation-selective permeability in the upper portion of the descending limb of long-looped nephron (LDL(u)) of hamsters, we observed the effect of protamine on salt-diffusion voltage (DELTA-V(T)) and transmural resistance (R(T)). DELTA-V(T) generated on reduction of lumen NaCl concentration was decreased from 12.0 +/- 1.4 to 7.3 +/- 1.2 mV when 100-mu-g/ml protamine were added to the lumen. Although the effect of protamine persisted after removal of the agent from the lumen, addition of 30 U/ml heparin reversed the DELTA-V(T) toward the control level. The effect of protamine was dose dependent in the range from 3 to 1,000-mu-g/ml. Protamine was without effect from the bath. Studies on single salt dilution voltage revealed that 100 and 300-mu-g/ml protamine inhibited relative Na+ to Cl- permeability from 4.03 +/- 0.38 to 2.14 +/- 0.21 and from 3.75 +/- 0.37 to 1.36 +/- 0.09, respectively. Protamine markedly decreased the apparent transference number for Na+ but slightly increased the value for Cl-. Protamine also inhibited permeabilities for K+, Rb+, and Li+ relative to Cl-, indicating that the inhibitory effect of protamine was not confined to Na+ but was generalized to cations. Transmural cable analysis showed that 100-mu-g/ml protamine increased R(T) from 14.0 +/- 1.1 to 19.3 +/- 1.2 OMEGA.cm2, with the effect being reversed by 30 U/ml heparin. Because the effect of protamine on R(T) was unaffected by ouabain in the bath, changes in R(T) may mainly represent those of the paracellular shunt resistance. Cable analysis with cell puncture in combination with BaCl2 effect further confirmed this view. Protamine at 100-mu-g/ml increased shunt resistance from 34.0 +/- 8.3 to 44.0 +/- 10.5 OMEGA.cm2 without affecting apical and basolateral membrane resistances. From these observations, we conclude that the use of protamine provides a useful tool to study the contribution of the paracellular shunt pathway and that at least approximately 50% of total conductance of the LDL(u) is accounted for by the cation-selective paracellular permeability.