The Drosophila melanogaster homologue of an insect calcitonin-like diuretic peptide stimulates V-ATPase activity in fruit fly Malpighian tubules

The Drosophila melanogaster homologue of an insect calcitonin-like diuretic hormone was identified in a BLAST search of the Drosaphila genome database , The predicted 31-residue amidated peptide (D. melanogaster DH31; Drome-DH 31) was synthesised and tested for activity on fruit Ay Malpighian tubules, It increases tubule secretion by approximately 35 % of the response obtain ed with a myokinin from the housefly Musca domestica (muscakinin; Musdo-K) and has an EC50 of 4.3 nmol l(-1), The diuretic activities of Drome-DH31 and Musdo-K were additive when tested at threshold and supra-maximal concentrations, which suggests that they ta rget different transport processes, In support of this, Drome-DH31 increase d the rate of secretion by tubules held in bathing fluid with a reduced Cl- concentration, whereas Musdo-K did so only in the presence of Drome-DH31. Stimulation with Drome-DH31 increased the lumen-positive transepithelial po tential in the main secretory segment of the tubule, This was attributed to activation of an apical electrogenic proton-translocating V-ATPase in prin cipal cells, since it was associated with hyperpolarisation of the apical m embrane potential and acidification of secreted urine by 0.25 pH units, Exogenous 8-bromo-cyclic AMP and cyclic GMP increased tubule secretion to t he same extent as Drome-DH31 and, when tested together with the diuretic pe ptide, their activities were not additive, Stimulation with Drome-DH31 resu lted in a dose-dependent increase in cyclic AMP production by tubules incub ated in saline containing 0.5 mmol l(-1) 3-isobutyl-1-methylxanthine, where as cyclic GMP production was unchanged, Taken together, the data are consis tent with Drome-DH31 activating an apical membrane V-ATPase via cyclic AMP, Since the K+ concentration of the secreted urine was unchanged, it is like ly that Drome-DH31 has an equal effect on K+ and Na+ entry across the basol ateral membrane.