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United States Department of Agriculture

Agricultural Research Service

Title: Voltage Clamping Single Cells in Intact Malpighian Tubules of Mosquitoes

Authors
item Masia, R - CORNELL UNIVERSITY
item Aneshansley, D - CORNELL UNIVERSITY
item Nagel, W - UNIVERSITY OF MUNICH
item Nachman, Ronald
item Beyenbach, K - CORNELL UNIVERSITY

Submitted to: American Journal of Physiology - Renal Fluid and Electrolyte Metabolism
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: May 31, 2000
Publication Date: N/A

Interpretive Summary: Because of problems with the development of resistance to conventional pesticides, there is a critical need for new concepts and alternative approaches in controlling such pests. The basic premise of this research is that peptides (short chains of amino acids) serve as potent internal messengers in insects to regulate vital functions. Peptides themselves are eunsuitable for control measures due to their instability to enzymes in the circulatory and digestive systems of the insect. New, selective control measures may be developed by designing metabolically stable mimics of these neuropeptides that actively inhibit or over-stimulate functions regulated by them, resulting in disruption of the internal environment of the insect. One of the important pieces of information required to develop neuropeptide-based pest control strategies is to determine the effects of the natural neuropeptides on critical bodily functions and/or behavior. In nthis paper, we describe the details of the operation of the Malpighian tubules, analogous to kidneys in mammals. The paper further describes the mechanism of action of the kinin neuropeptides on Malpighian tubules and how this can influence the regulation of water balance in insects. This information will aid in the development of enzyme-resistant mimics of the kinin neuropeptides that can disrupt the regulation of water balance, a process that is critical for insect survival. This work leads us one step closer to the development of practical neuropeptide-like chemicals that will be effective in controlling certain pest insects in an environmentally friendly fashion.

Technical Abstract: Principal cells of the Malpighian tubule of the yellow fever mosquito were studied with the methods of two-electrode voltage clamp (TEVC). Intracellular voltage (Vpc) was -86.7 mV, and input resistance (Rpc) was 388.5 ko (n=49 cells). In six cells, Ba**2+ (15 mM had negligible effects on VPC, but it increased R from 325.3 to 684.5 ko (P<0.001). In the presence of Ba**2+, leucokinin-VIlI (1 uM) increased V to -101.8 mV (P<0.001) and reduced R to 340.2 ko (p<0.002). Circuit analysis yields the following: basolateral membrane resistance, 652.0 ko; apical membrane resistance, 340.2 ko; shunt resistance (Rsh), 344.3 ko; trans-cellular resistance, 992.2 ko. The fractional resistance of the apical membrane (0.35) and the ratio of transcellular resistance and Rsh (3.53) agree closely with values obtained by cable analysis in isolated perfused tubules and confirm the usefulness of TEVC methods in single principal cells of the eintact Malpighian tubule. Dinitrophenol (0.1 mM) reversibly depolarized Vp from -94.3 to -10.7 mV (P<0.001) and reversibly increased Rpc from 412 to 2,879 ko (P<0.001), effects that were duplicated by cyanide (0.3 mM). Significant effects of metabolic inhibition on voltage and resistance suggest a role of ATP in electrogenesis and the maintenance of conductive transport pathways.

Last Modified: 7/22/2014
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