REGULATION OF THE ELECTROGENIC H-MEMBRANE OF NEUTROPHILS - POSSIBLE ROLE OF PHOSPHOLIPASE-A2, INTERNAL AND EXTERNAL PROTONS( CHANNEL IN THEPLASMA)

Possible factors regulating the opening of and the rate of H+ flux thr ough a recently described, Cd2+-sensitive, phorbol ester- and arachido nic acid (AA)-activatable H+-conducting pathway in the plasma membrane of neutrophil granulocytes were investigated. (1) The phospholipase A 2 blocker p-bromophenacyl bromide (BPB) inhibited the phorbol 12-myris tate 13-acetate (PMA)-induced activation of this channel in a concentr ation-dependent manner (IC50, 4 muM). (2) Neither BPB nor the protein kinase C (PKC) inhibitor staurosporine influenced the AA-elicited stim ulation of this route. (3) Intracellular acidification (cytoplasmic pH below 6.9) itself is capable of activating an electrogenic, Cd2+-sens itive H+ efflux indicating that protons can open up this route in the absence of any other stimulator. (4) PMA significantly decreases the i ntracelullar H+ concentration ([H+]i) threshold for the opening of the channel, thus providing a conductive state at resting pH values, and elevates the rate of H efflux at any [H+]i. (5) Changes in external pH also modify the operation of the channel: above an extracellular pH ( pH(o)) value of 7.4. the H+-flux/driving force relationship is approx. 5-fold greater than below this value. Our results suggest a multifact orial regulation of the electrogenic H+ channel: most probably PKC act ivates the channel indirectly, via stimulation of phospholipase A, tha t subsequently liberates AA. In addition to this, the channel conducta nce seems to be promoted by internal H+ and inhibited by external H+.