The interactive effects of hypoxia and nitric oxide on catecholamine secretion in rainbow trout (Oncorhynchus mykiss)

Experiments were performed to test the hypothesis that exposure of rainbow trout to repetitive hypoxia would result in a decreased capacity of chromaffin cells to secrete catecholamines owing to increased production of nitric oxide (NO), a potent inhibitor of catecholamine secretion. A partial sequence of trout neuronal nitric oxide synthase (nNOS) was cloned and its mRNA was found to be present in the posterior cardinal vein (PCV), the predominant site of chromaffin cells in trout. Using heterologous antibodies, nNOS and endothelial NOS (eNOS) were localized in close proximity to the chromaffin cells of the PCV. Exposure of trout to acute hypoxia (5.33 kPa for 30 min) in vivo resulted in significant increases in plasma catecholamine and NO levels. However, after 4 days of twice-daily exposures to hypoxia, the elevation of plasma catecholamine levels during hypoxia was markedly reduced. Associated with the reduction in plasma catecholamine levels during acute hypoxia was a marked increase in basal and hypoxia-evoked circulating levels of NO that became apparent after 2-4 days of repetitive hypoxia. The capacity of the chromaffin cells of the hypoxia-exposed fish to secrete catecholamine was assessed by electrical stimulation of an in situ saline-perfused PCV preparation. Compared with control (normoxic) fish, the PCV preparations derived from fish exposed to repeated hypoxia displayed a significant reduction in electrically evoked catecholamine secretion that was concomitant with a marked increased in NO production. This additional rise in NO secretion in preparations derived from hypoxic fish was prevented after adding NOS inhibitors to the perfusate; concomitantly, the reduction in catecholamine secretion was prevented. The increased production of NO during hypoxia in vivo and during electrical stimulation in situ was consistent with significant elevations of nNOS mRNA and protein; eNOS protein was unaffected. These results suggest that the reduced capacity of trout chromaffin cells to secrete catecholamines after repeated hypoxia reflects an increase in the expression of nNOS and a subsequent increase in NO production during chromaffin-cell activation.