Role of ion conductance changes and of the sodium‐pump in adrenaline‐induced hyperpolarization of rat diaphragm muscle fibres

Abstract

The ionic mechanism of membrane hyperpolarization induced by adrenaline in rat diaphragm muscle fibres was studied. Removal of the extracellular K+ ([K+]o) from Krebs-Ringer solution initially increased the resting membrane potential and then caused an increase in the intracellular Na+ activity ([Na+]i) and a decrease in the intracellular K+ activity ([K+]i). All the changes were maintained for more than 3 h. Application of ouabain (0.1 mM) or lowering the temperature rapidly reduced the resting potential by about 10 mV in the K+-free solution. It then produced further progressive decreases in resting potential and in [K+]i and a progressive increase in [Na+]i. These observations indicate that an electrogenic Na-pump operates in the K+-free solution. Removal of most of the Cl- in the K+-free solution did not affect the resting potential or the magnitude of the initial decrease produced by ouabain, despite an increased input resistance; this result implies a passive distribution of Cl-. Adrenaline (30-60 microM) either added to the bathing solution or applied to the membrane by ionophoresis produced a hyperpolarization (3-10 mV: adrenaline hyperpolarization), the amplitude of which was decreased with a rise in [K+]o and increased with a reduction in [K+]o, but unaffected by the removal of Cl-. Adrenaline produced an increase in input resistance, the relative magnitude (17-18%) of which was constant whether external K+ or Cl- was removed. In contrast, a conditioning membrane hyperpolarization hardly affected the resistance. Ouabain (0.1 mM) or low temperature (8-10 degrees C) abolished both the hyperpolarization and the increased input resistance induced by adrenaline. The [K+]i, [Na+]i and the peak of the action potential remained unchanged after a 20 min exposure to adrenaline (30 microM). The hyperpolarization induced by the replacement of all Na+ with Tris (Tris-hyperpolarization) in the K+-free solution was depressed by 39% during the early period (4-31 min) of exposure to adrenaline (30 microM), while it was enhanced by 26% during the later period (80-130 min). The initial depression suggested a decrease in the ratio of the membrane permeability for Na+ (PNa) to that for K+ (PK). These results suggest that the adrenaline hyperpolarization is generated largely by a decrease in PNa/PK, which is associated with the activity of the Na-pump.