Fig 1.
A: Chemical structure of tribendimidine.
B: Diagram of the two-micropipette current-clamp technique used to record the membrane potential (v) and to inject (i) 40 nA hyperpolarizing 500 ms current pulses at 0.3 Hz. P is the microperfusion pipette used to apply and wash off the drugs. C: Application of 3 μM acetylcholine and then 0.1, 1 and 10 μM tribendimidine in the same preparation. Notice that 1 μM tribendimidine produces a bigger depolarization response (upward movement) and conductance increase (reduction in the voltage responses to current injection, producing a narrowing of the width of the trace) than 3 μM acetylcholine.
Fig 2.
A: Top, representative recording of the application, for 10 seconds, of different concentrations of tribendimidine which produces a depolarization (upward movement of the thickest line) and increase in membrane conductance (decrease in the width of the trace).
Bottom, representative recording of the application for 10 seconds, of tribendimidine in the presence of 3 μM mecamylamine, a nicotinic antagonist, with inhibition of the tribendimidine responses. B: Plots of the mean ± s.e. (n = 4) of the tribendimidine-concentration-depolarization-responses in the absence and presence of 3 μM mecamylamine. The control plot was fitted to the Hill equation allowing estimation of the control EC50 as 0.83 μM.
Fig 3.
Contraction of Ascaris suum muscle strips produced by tribendimidine.
A: Diagram of the isometric contraction technique used to measure the force of contraction and representative trace of a cumulative-tribendimidine-contraction-response. B: Plot of the mean ± s.e. (n = 6 or more) cumulative-tribendimidine-contraction-response and Hill equation fit with and EC50 of 0.2 μM. The EC50 for contraction is, as expected, is lower than for the membrane potential response because the amplification in the depolarization-contraction signaling cascade of muscle.
Fig 4.
Effect of antagonists of the tribendimidine-concentration-contraction plots allowing the estimation of pA2 values.
A: Effect of derquantel pA2 = 6.42 ± 0.12 mean ± s.e. (n = 108). B: Effect of paraherquamide pA2 = 7.21 ± 0.13 mean ± s.e. (n = 90). C: Effect of methyllycaconitine (MLA) pA2 = 6.61 ±0.09 mean ± s.e. (n = 90).
Fig 5.
A: Similarity Cluster analysis of pA2s of the antagonists, derquantel, paraherquamide and methyllycaconitine against the agonists beph (bephenium), trib (tribendimidine), then (thenium), lev (levamisole), pyr (pyrantel), oxa (oxantel), meth (methyridine), nic (nicotine) (ordinate: similarity, averaged, squared Euclidian, standardized variable, estimated using Minitab 13.2 (State College, PA) showing that tribendimidine does not group with levamisole and is closer to bephenium.
B: Schematic illustration of the selectivity of tribendimidine and levamisole for B-subtype (green) and L-subtype (red) nAChRs.
Fig 6.
Larval migration inhibition assays on O. dentatum L3 levamisole-sensitive (SENS) and levamisole-resistant (LEVR) isolates.
A: Levamisole is less effective on levamisole-resistant isolates than on the levamisole-sensitive isolates and the difference is significant (p<0.001, F-test). B: tribendimidine is more potent on levamisole-resistant isolates than on levamisole-sensitive isolates and the difference is significant (p<0.001, F-test).