Membrane recycling and endocytosis in Paramecium confirmed by horseradish peroxidase pulse-chase studies

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Abstract
Paramecium caudatum cells were pulse-chased in horseradish peroxidase (HRP) using pulses of 30 s to follow endosome (endocytotic vesicle) formation from defaecating digestive vacuoles and 3-min pulses to follow the movement and fate of these vesicles in the cell. Endosomes formed during the 30 s in HRP are heavily labelled with HRP reaction product and are mostly flattened. Some align along microtubules that point toward the cytopharynx. Diskoidal vesicles at the cytopharynx are unlabelled in unchased cells. Cells exposed to a 3-min HRP pulse contain varied amounts of HRP-labelled diskoidal vesicles at the cytopharynx but no labelled vesicles near their closed cytoprocts. Labelled vesicles are also found aligned along the cyto-pharyngeal microtubular ribbons. Diskoidal vesicles are no longer labelled after a 47-min chase. Use of pulse-chase HRP cytochemistry supports the hypothesis that the membrane of digestive vacuoles retrieved at the cytoproct is moved along microtubular ribbons directly to the cytopharynx where the membrane enters the diskoidal vesicle pool. This membrane appears to enter neither the Golgi nor lysosomal systems in its passage. HRP reaction product is also found in vesicles near the parasomal sacs, in some secondary lysosomes and in small spherical vesicles that are probably trichocyst membrane fragments. Possibly some membrane from parasomal sacs or condensing digestive vacuoles may also enter the diskoidal vesicle pool but apparently not membrane from trichocysts.