A vesicle preparation for resolving single-channel currents in tegument of male Schistosoma mansoni.

Abstract

A tegumental vesicle preparation from adult male Schistosoma mansoni was developed that allows the resolution of single ion-channel currents. Adult male schistosomes were exposed to a low pH (3.75) medium for a period of approximately 30 min at 37 degrees C. During this period smooth vesicles formed from the tegument. Fluorescence microscopy following staining of the tegument with the dye, 5-N-[octadecanoyl]aminofluorescein (AF-18), transmission electron microscopy and scanning electron microscopy revealed that the vesicles were produced from the outer tegumental membrane. The fluorescence studies showed the presence of the double bilayer structure of the outer membrane in > 41% of the vesicles. These studies suggested that the preparation is suitable for single-channel recording with the patch-clamp technique. Cell-attached and isolated inside-out patch recordings of ion-channel activity were obtained with giga-ohm resistance seals. Different types of ion-channel were recorded from tegumental vesicles from male schistosomes, illustrating the potential of the technique. The channels observed included: a non-selective cation channel (360 pS); a K+ channel (with a conductance of 115 pS in high bath-K conditions); and a Cl- selective channel (20 pS). The currents of these ion-channels may cross the double bilayer of the outer tegumental membrane.