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We have previously shown that continuous intravenous infusion of NaHCO3 for 24 h (∼1000 μmol kg-1 h-1) results in the relocation of V-H+-ATPase from the cytoplasm to the basolateral membrane in the gills of the Pacific dogfish. To further investigate this putative base-secretive process we performed similar experiments with the addition of colchicine, an inhibitor of cytoskeleton-dependent cellular trafficking processes. Blood pH and plasma total CO2 were significantly higher in the colchicines-treated, HCO3--infused fish compared with fish infused with HCO3- alone. The effect of colchicine was highest after 24 h of infusion (8.33±0.06 vs 8.02±0.03 pH units, 15.72±3.29 vs 6.74±1.34 mmol CO2 l-1, N=5). Immunohistochemistry and western blotting confirmed that colchicine blocked the transit of V-H+-ATPase to the basolateral membrane. Furthermore, western blotting analyses from whole gill and cell membrane samples suggest that the short-term (6 h) response to alkaline stress consists of relocation of V-H+-ATPases already present in the cell to the basolateral membrane, while in the longer term (24 h) there is both relocation of pre-existent enzyme and upregulation in the synthesis of new units. Our results strongly suggest that cellular relocation of V-H+-ATPase is necessary for enhanced HCO3- secretion across the gills of the Pacific dogfish. [ABSTRACT FROM AUTHOR]