GUINEA-PIG MEGAKARYOCYTES CAN RESPOND TO EXTERNAL ADP BY ACTIVATING CA-2+-DEPENDENT POTASSIUM CONDUCTANCE

1. The responses of megakaryocytes to adenosine diphosphate (ADP) were studied using whole‐cell patch electrodes and a Ca2(+)‐sensitive fluorescent dye, Fura‐2. Megakaryocytes (diameter, 17‐42 microns) were mechanically dissociated from the bone marrow of adult guinea‐pigs and ADP (1‐10 microM) was pressure‐applied to megakaryocytes under recording. 2. In megakaryocytes immersed in standard saline, ADP evoked an obvious outward current at a membrane potential of ‐63 mV. The current was identified as a K(+)‐carried current, since the reversal potential depended distinctly on the external K+ concentration, but it showed no changes after removal of external Na+. The amplitude of evoked K+ currents showed considerable intercell variation, which is presumably due to differences of current density in the membrane. 3. During application of ADP, the evoked K+ current was not sustained but slowly decayed to become negligible within 10‐20 s, suggesting the appearance of desensitization. The response of the megakaryocyte to ADP recovered slowly and returned to an original level after 4‐5 min of continuous washing. 4. When the intracellular free Ca2+ concentration ([Ca2+]i) was measured using the Ca2(+)‐sensitive fluorescent dye, Fura‐2, application of 10 microM‐ADP induced an increase of [Ca2+]i by about 5‐fold, which was followed by a gradual decay to the original level within 30‐50 s. Roles of internal Ca2+ for activating the K+ current were confirmed by observing (1) enhancement of evoked currents by the use of internal saline containing no Ca2+ chelators and (2) generation of prolonged K+ current by application of a Ca2+ ionophore, A23187, to the megakaryocyte. 5. In a fraction of the megakaryocytes, spontaneous hyperpolarization of the resting membrane potential was observed. The hyperpolarization seemed to result from the activation of K+ channels in the membrane, which was caused by spontaneous release of Ca2+ from the internal storage site. 6. It was concluded that megakaryocytes of the guinea‐pig can respond to external ADP by increasing [Ca2+]i and consequently by activating Ca2(+)‐dependent K+ channels in the membrane. © 1990 The Physiological Society