The impact of target mutations on compound binding enzyme efficiency

The primary chemical agents currently in use for controlling mosquitoes are insecticides that target the nervous system. Although the development of insecticides such as DDT and pyrethroids, which modulate the CYM 9484 activity of ion channels in the central nervous system of insects, offered promise for the eradication of mosquitoes in the 20th century, the emergence of resistance in mosquito populations has reduced their efficacy. Currently, there are not many alternatives, because no new insecticides for public-health use have been developed in over 30 years. Thus, new chemicals and new approaches to control mosquitoes are urgently needed. A physiological process in the mosquito that has not yet been targeted by insecticides is the excretion of urine. The renal tubules generate urine via the transepithelial secretion of NaCl, KCl, other solutes, and water from the extracellular fluid to the tubule lumens. The tubules empty their secretions into the hindgut where solute and/ or water is removed or added to the final urine before it is ejected via muscular contractions of the hindgut. Thus, inhibiting the function of Malpighian tubules causing renal failure is expected to disrupt extracellular fluid homeostasis with detrimental consequences to normal functions in the mosquito. Female mosquitoes would be particularly vulnerable to renal failure, because they would not be able to excrete the unwanted salt and water ingested during a blood meal. The aim of the present study is to elicit renal failure in adult female mosquitoes using a small-molecule inhibitor of inward-rectifying potassium channels. Kir channels are a phylogenetically ancient family of barium-sensitive channels that play fundamental roles in nerve, muscle, endocrine, and epithelial function in diverse organisms. In mosquito Malpighian tubules, Kir channels of the basolateral membrane are considered one of the two major routes for the uptake of the epithelium. The Malpighian tubules of Ae. CPCCOEt aegypti express three cDNAs encoding Kir channel subunits, which we have cloned and functionally characterized. AeKir1 mediates robust K + currents when expressed in Xenopus oocytes, whereas AeKir2B and AeKir3 produce relatively small and nominal K + currents, respectively. Thus, we focused on inhibiting AeKir1 in the present study. With few exceptions, the small-molecule pharmacology of the Kir channel family is undeveloped. In an effort to discover new modulators of human Kir1.1, we previously performed a high-throughput screen of approximately 225,000 small molecules from the National Institutes of Health Molecular Libraries Small- Molecule Repository. The screen revealed compound VU573, which inhibits several human Kir channels. Thus, we tested whether VU573 also inhibits the mosquito Kir channel, AeKir1. To assess whether VU573 inhibits the production of urine by Malpighian tubules we used the method of Ramsay. As shown in Figure 2D, isolated Malpighian tubules bathed in a highpotassium Ringer solution spontaneously secrete fluid at a rate of,0.6 nl/min in the first 30 min.

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