Effect of protamine sulfate on the permeability properties of the mammalian urinary bladder

Protamine sulfate (PS, an arginine-rich protein of molecular weight 5,000) has been reported to affect the ionic permeability of gallbladder epithelium, the permeability of cultured epithelial cells to mannitol, and the permeability of endothelial cell layers to albumin. Although the effect of PS has been widely investigated, the mechanism of its action on membrane permeability is presently unknown. The effect of PS on the rabbit urinary bladder epithelium was studied using both transepithelial and intracellular microelectrode techniques in conjunction with equivalent circuit analysis. The addition of 100 micrograms/ml of PS to a NaCl-containing mucosal solution caused (over a 40-min period) a large increase in the transepithelial conductance (Gt) and a transient hyperpolarization of the transepithelial voltage (Vt) followed by a depolarization of Vt. This secondary depolarization of Vt was not present if the mucosal solution was a KCl or a K-gluconate Ringer. The PS-induced increase in Gt was due to an increase in the apical membrane permeability to both cations (Na+ or K+) and anions (Cl- or gluconate). Further studies revealed the following features of the PS-induced conductance. (i) Trypsin inhibits the PS effect; however, this was due to PS hydrolysis by trypsin and not a membrane effect. (ii) Mucosal PS partially inhibited the PS-induced apical membrane conductance. (iii) The ability of PS to increase the membrane conductance was enhanced when the apical membrane potential was cell interior negative. (iv) The rate of conductance change (at any given membrane potential) was a saturating function of the PS concentration. This finding suggests that PS must interact with a membrane binding site before it can induce a change in the membrane conductance. (v) Lanthanum inhibited the PS-dependent conductance by two different mechanisms. One was as a reversible blocker of the PS-induced conductance. The other was by inhibiting the interaction between PS and a membrane binding site. A kinetic model is developed to describe the steps involved in the increase in membrane conductance.