Further investigation of ionic diffusive properties and of NH ⁺₄ pathways inXenopus laevis oocyte cell membrane

Abstract

To study the ionic diffusive properties and the NH ⁺₄ pathways in theXenopus laevis oocyte cell membrane, we recorded the effects of various inhibitors on membrane potential (V _m) and membrane resistance (R _m); intracellular acidification was taken as an index of NH ⁺₄ influx from the bath to the cytoplasm. The following results were obtained: in the control state, barium and quinine (Q) ions depolarizedV _m and raisedR _m, consistent with inhibition of K⁺ conductance(s). Diphenylamine-2-carboxylic acid (DPC), 3′, 5′-dichlorodiphenylamine-2-carboxylic acid (DCDPC) and gadolinium ions hyperpolarizedV _m and increasedR _m, suggesting the inhibition of nonselective cationic conductance(s). In the presence of 20 mmol/l NH₄Cl,V _m depolarized,R _m fell, and intracellular pH (pH_i) decreased, consistent with an NH ⁺₄ influx. In the presence of DPC, the same manoeuvre induced a biphasicV _m change (i.e. a spike depolarization followed by a membrane hyperpolarization) and a fall ofR _m; in most oocytes, intracellular acidification persisted and was reversible upon adding ouabain (Oua). These results indicate that a DPC-sensitive conductance is not the unique NH ⁺₄ pathway and that Na, K-ATPase may also mediate NH ⁺₄ influx. However, Oua did not prevent theR _m decrease, suggesting that ouabain-insensitive rheogenic pathway(s) are activated. Thus, we investigated theV _m change induced by NH₄Cl addition in the presence of DPC: the spike depolarization followed by secondary hyperpolarization became a plateau depolarization when Q was added, suggesting involvement of Q-sensitive pathway(s) in the above described biphasicV _m change. In the presence of DPC, Q, and Oua, intracellular acidification upon adding NH₄C1 persisted consistent with further NH ⁺₄ influx through quinine-, DPC- and Oua-insensitive pathway (s).