The effects of aedeskinin III and analogues on transepithelial electrolyte and fluid secretion in Malpighian tubules of the yellow fever mosquito
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The effects of synthetic aedeskinin III on transepithelial electrolyte and fluid secretion were studied in isolated Malpighian tubules of the yellow fever mosquito, Aedes aegypti. Isolated Malpighian tubules were prepared for measurements of fluid secretion rates by the method of Ramsay. After measuring a control rate of fluid secretion, aedeskinin III was added to the peritubular Ringer solution. Rates of transepithelial fluid secretion significantly increased in a dose-dependent manner at aedeskinin concentrations ranging from 10-5 to 10-9M. Maximal stimulatory effects were observed at 1 ?M aedeskinin III, which increased the fluid secretion rate from 0.667 to 1.451 nl/min. EC50 for aedeskinin III was calculated as 1.028x10-7M. In the presence of 1 ?M aedeskinin III, the Na+ concentration in secreted fluid significantly decreased from 130.5 to 88.2 mM and the Cl- concentration in secreted fluid significantly decreased from 208.6 to 173.5 mM. The increase in the K+ concentration from 82.1 mM to 90.3 mM was not significant. The product of the fluid secretion rate and electrolyte secretion yields the electrolyte secretion rate. Accordingly, the transepithelial secretion rate for Na+ increased from 96.3 pmols/min to 127.6 pmols/min, the K+ secretion rate increased from 61.6 pmols/min to 141.2 pmols/min, and the Cl- secretion rate increased from 153.9 to 257.5 pmols/min. However, only the effects on K+ and Cl- secretion were statistically significant. Synthetic aedeskinin-receptor agonists 1577 and 1460II also stimulated fluid secretion at 1 ?M concentrations, increasing fluid secretion rate from 0.716 to 1.224 nl/min and from 0.889 to 1.615 nl/min, respectively. The in vitro longevity of isolated Malpighian tubules was also assessed. Isolated tubules cultured in Ringer solution and Grace?s insect medium (pH 6.1) did not survive the process and did not secrete fluid one day after isolation. However, when kept in Grace?s insect medium at pH 7.1, the tubules still secreted one day later. These studies confirm that aedeskinin III has diuretic effects in Aedes Malpighian tubules. Rates of transepithelial secretion of NaCl and KCl increased, suggesting that the mechanism of action of aedeskinin III is similar to that of leucokinin. My study documents further and for the first time that synthetic analogs of aedeskinin containing ?-amino acids retain diuretic potency.
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