Rapid ADP-evoked currents in human platelets recorded with the nystatin permeabilized patch technique.
"Whole-cell" patch recordings using nystatin permeabilization were made from single human platelets during application of agonists from a "puffer" pipette. In platelets clamped near the resting potential and bathed in Na+ saline, 40 microM ADP activated a transient inward current within tens of milliseconds. At -73 mV the current lasted between 0.1 and 1 s and had a peak of between 13 and 31 pA in different cells. Ion substitution experiments indicated that the channel is permeable to Na+,K+, and Ba2+ and presumably also to Ca2+, but is not permeable to Cl-. The single channel conductance was 15 pS (near the resting potential) in nominally Ca(2+)-free saline and 11 picosiemens in BaCl2 saline. Thrombin, at 1 unit/ml, did not elicit detectable currents during a 3-s application in platelets bathed in 1 mM Ca2+, Na+ saline. Under the same conditions, in fura-2-loaded cells, thrombin-evoked Ca2+ entry (monitored by Mn2+ quench) was detectable after a delay of 1.4 s. This suggests that early thrombin-evoked Ca2+ entry occurs via small conductance channels, below the resolution of the patch clamp technique, or by an electroneutral pathway. The ADP-evoked channel has the requisite speed of activation to account for the rapid Ca2+ influx observed during stopped-flow studies of agonist-evoked changes in [Ca2+]i.
Mahaut-Smith
M P
MP
Physiological Laboratory, Cambridge, United Kingdom.
Sage
S O
SO
Rink
T J
TJ
D000244:Adenosine Diphosphate / Q000494:pharmacology*; D001464:Barium / Q000097:blood; D001792:Blood Platelets / Q000187:drug effects / Q000502:physiology*; D002118:Calcium / Q000097:blood; D002462:Cell Membrane / Q000187:drug effects / Q000502:physiology; D002463:Cell Membrane Permeability; D002712:Chlorides / Q000097:blood; D004594:Electrophysiology / Q000379:methods; D005071:Evoked Potentials / Q000187:drug effects; D016257:Fura-2; D006801:Humans; D008564:Membrane Potentials / Q000187:drug effects; D009761:Nystatin / Q000494:pharmacology*; D011188:Potassium / Q000097:blood; D012964:Sodium / Q000097:blood; D013050:Spectrometry, Fluorescence; D013917:Thrombin / Q000494:pharmacology
D002712:Chlorides; D009761:Nystatin; D001464:Barium; D000244:Adenosine Diphosphate; D012964:Sodium; D013917:Thrombin; D011188:Potassium; D002118:Calcium; D016257:Fura-2