Electrophysiological properties of three types of granulocytes in circulating blood of the newt

Abstract

1. The electrophysiological properties of three subtypes of granulocytes obtained from the circulating blood of the newt, Triturus pyrrhogaster, were studied using the whole-cell variation of the patch-electrode voltage-clamp technique. 2. Neutrophils were identified by their multilobulated nucleus in the cytoplasm. Basophils and eosinophils, having characteristic granular structures in their cells, could definitely be distinguished from other leucocytes. The reliability of cellular identification using phase-contrast microscopy was confirmed by fixing and staining the granulocytes with Wright's solution. 3. In neutrophils under current-clamp conditions, a hyperpolarization-induced conductance increase was observed. With depolarization, however, no changes in regenerative potential were detected. When voltage clamped in standard saline (containing 96 mM-NaCl), hyperpolarizing voltage pulses to a potential more negative than -90 mV evoked slowly decaying inward currents. 4. The hyperpolarization-evoked membrane currents in neutrophils were identified as anomalous rectifying K+ currents, since (1) externally applied Cs+ (0.1 or 1 mM) or Ba2+ (1 mM) produced suppressive effects on the currents, (2) replacement of external Na+ with choline ions eliminated the decay of macroscopically observed currents, and (3) both the amplitude and kinetic properties of the currents were strongly dependent on membrane potential as well as on external K+ concentration; the activation of the conductance depended on the electrochemical force for K+ rather than on membrane potential alone. The magnitude of steady-state conductance was roughly proportional to the square root of the external K+ concentration. 5. In basophils and eosinophils, no major time- or voltage-dependent increase in conductance was detected at voltages between +20 and -130 mV. However, under current-clamp conditions, spontaneous fluctuation of zero-current potentials was clearly apparent, presumably due to the activities of some ion channels generating a small amount of current flux through the membranes of these cells. 6. It was concluded that the three subtypes of granulocytes in the newt differ considerably not only in appearance and structure but also in the electrical properties of their membranes. The anomalous rectifying K+ channels in neutrophils may serve to determine the resting potential of the cell at K+ equilibrium potential. The closure of the channels at depolarization might facilitate the maintenance of depolarization triggered by stimuli accompanying current influx.