I examined transport characteristics of isolated Malpighian (excretory) tubules from the tobacco hornworm (Manduca sexta), a Lepidopteran which tolerates high levels of dietary nicotine, to test the hypothesis that alkaloid (e.g. nicotine) transport in the tubules is the consequence of a multidrug transport system similar to that mediated by P-glycoprotein in multidrug resistant cancer cells. P-glycoprotein substrates were applied to the basal (blood) side of the tubule and their subsequent distribution monitored. First, using cannulated tubules in a modification of the Ramsay assay, [3H]-vinblastine levels were measured in samples of bathing and luminal solutions. After 1 h, the tubules had concentrated vinblastine in the lumen 3-fold (from 1 muM). This accumulation was independent of a transepithelial potential, and was inhibited by verapamil, a P-glycoprotein inhibitor, and by nicotine. Second, fluorescent drugs, including daunomycin, rhodamine 123 and acridine orange, were applied to living tubules in a well of saline on a coverslip and examined by confocal microscopy. All of these drugs stained the cells of the tubule within 1 min, but, contrary to expectation, none of the drugs appeared in the lumen even after 1--2 h of incubation. Neither verapamil nor nicotine altered the pattern of daunomycin staining. The fast and intense Malpighian tubule staining was in sharp contrast to that of other tissues, which (for most drugs) stained lightly and only after prolonged exposures. Fluid-phase markers appeared to penetrate the tissue, but more slowly, and remained largely extracellular. The results suggest that the Malpighian tubule clears hemolymph of xenobiotics by passive filtration, active transepithelial transport involving a P-glycoprotein-like pump which handles alkaloids, and a scavenger-like absorption of compounds.
| Identifer | oai:union.ndltd.org:uottawa.ca/oai:ruor.uottawa.ca:10393/4060 |
| Date | January 1998 |
| Creators | Gaertner, Lorin S. |
| Contributors | Morris, Catherine, |
| Publisher | University of Ottawa (Canada) |
| Source Sets | Université d’Ottawa |
| Detected Language | English |
| Type | Thesis |
| Format | 108 p. |
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