Volume regulatory mechanisms of human granulocytes in hypoosmotic media

Human neutrophil granulocytes, when placed into hypoosmotic media, first swell, than show a regulatory volume decrease (RVD) reaction. This RVD is the most effective at pH 7.5 when inosine is used as a metabolic substrate. The RVD reaction is blocked by the inhibitors of the calcium-induced K+ transport, such as quinine, quinidine, trifluoperazine (TFP), as well as by those of the conductive Cl-transport (dipyridamole, SITS, oligomycin C). Removal of external calcium by EGTA, or addition of the calcium channel inhibitor, verapamil, reduce the RVD. Ouabain affects RVD only after long preincubation and furosemide or phloretin have hardly any effect on this process. In a hypoosmotic KCl medium neutrophils show a TFP-, and SITS-sensitive secondary volume increase. All these data indicate the opening of Ca2+-induced K+ channels and of conductive Cl- channels in hypoosmotically shocked neutrophils. Under these conditions, however, direct measurement of cytoplasmic calcium by Indo-1 does not show any major change in the overall cytoplasmic free Ca2+ levels. In neutrophils, the reduced calcium signal evoked by formyl-methionyl-leucyl-phenylalanine (fMLP) after a hypoosmotic shock, suggests that cellular calcium metabolism is altered under these conditions.