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First published online June 6, 2005
Journal of Experimental Biology 208, 2227-2236 (2005)
Published by The Company of Biologists 2005
doi: 10.1242/jeb.01617

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Specificity of the fluorescein transport process in Malpighian tubules of the cricket Acheta domesticus

Douglas S. G. Neufeld^1,*, Ross Kauffman² and Zachary Kurtz¹

¹ Department of Biology, Eastern Mennonite University, Harrisonburg, VA 22802, USA
² Ohio State University, School of Public Health, Columbus, OH 43210, USA

View larger version (14K): [in a new window]   Fig. 1. Example fluorescence microscopy traces of fluorescein uptake in the presence and absence of a representative OAT inhibitor, methotrexate. During an uptake trial, the signal trace shows an initial rapid increase in fluorescence due to the change-out of chamber with fluid containing FL (a), followed by a slower increase representing accumulation in the tubule (b).

View larger version (75K): [in a new window]   Fig. 2. Brightfield (left) and fluorescence (right) micrographs showing FL accumulation in Acheta Malpighian tubules. In some cases, fluorescence was stronger in the lumen (A), while in others it was more generally dispersed through lumen and cell (B). In (C) FL is present in both the Malpighian tubule (right) and the larger ureter (left) that connects Malpighian tubules to intestine.

View larger version (31K): [in a new window]   Fig. 3. Inhibition of FL accumulation by mono- and dicarboxylic acids representing a range of sizes (as indicated by the number of carbons in the backbone). Sample sizes range from 3 to 7 tubules, except for `no inhibitor', which represents all control trials (N=27). *Change in uptake rate (second measurement period relative to first measurement period) in the presence of inhibitor significantly different from uptake rate in absence of inhibitor; P<0.05, paired t-test.

View larger version (20K): [in a new window]   Fig. 4. Inhibition of FL accumulation by glutathione and a size range of its conjugates. All inhibitor concentrations are 0.2 mmol l–1; sample sizes are 6 tubules, except for `no inhibitor, which represents all control trials (N=23). *Change in uptake rate (second measurement period relative to first measurement period) in presence of inhibitor significantly different from uptake rate in absence of inhibitor; P<0.05, paired t-test.

View larger version (12K): [in a new window]   Fig. 5. FL accumulation in Malpighian tubules of Drosophila, and its inhibition by several potential substrates. N=3 tubules for each. PBD, 3-phenobenzoic acid (1 mmol l–1); PAH, p-aminohippuyric acid (3 mmol l–1); PBA, probenecid (0.5 mmol l–1). *Change in uptake rate (second measurement period relative to first measurement period) in presence of inhibitor significantly different from uptake rate in absence of inhibitor; P<0.05, paired t-test.

View larger version (20K): [in a new window]   Fig. 6. Uptake of insecticide metabolites 3-phenoxybenzoic acid (PBA) and malathion monocarboxylic acid (MMA) in Malpighian tubule bundles, as detected by HPLC. Uptake in presence of inhibitor conditions (FL, 2 mmol l–1; PBD, 10 mmol l–1; or water pre-exposure) is plotted relative to control uptake (uptake with metabolite alone in Ringer's). N=4 samples for each. *Uptake of metabolite in presence of inhibitor condition significantly different from uptake in absence of inhibitor condition.

View larger version (17K): [in a new window]   Fig. 7. Primary metabolic fates known for malathion in animal tissues (including insects).