Clearly, the possibility cannot be excluded that a few caveolae (or clusters of cav-eolae) pinch off occasionally and therefore participate in the delivery of CT and other molecules to intracellular compartments, although this must - in terms of endocytic activity - be relatively insignificant. Recent studies conducted by Kirk-ham et al.  showed that a very small fraction (or a subpopulation) of caveolae is actually internalized constitutively. These authors developed an ultrastructural assay which made it possible to quantitate free, budded caveolae versus surface-connected caveolae. For this, they used CT-B-HRP as an endocytic marker, and quenched the HRP reaction of labeled structures connected to the cell surface with ascorbic acid, which is membrane-impermeable. This was combined with an im-munolabeling protocol that allowed them to show that truly free caveolae-like vesicles containing the HRP reaction product were in fact caveolae. In this way, it was shown that about 2% of the total population of caveolae bud off per minute.
Interestingly, the internalization of prion proteins that are GPI-anchored also appears to be a rather slow or inefficient process that can be mediated by cav-eolae . Thus, in an immunogold labeling pulse-chase experiment it was found that approximately 90% of the caveolae at the plasma membrane that were labeled after a 10-min period were still labeled after a 50-min chase; only about 10% of all caveolae were internalized per hour. This slow internalization may be constitutive, although it cannot be excluded that it is somehow stimulated by the prion proteins because the caveolae-mediated pathway was not observed for several other GPI-anchored proteins .
In our FRAP experiments in which the interior of the cells were bleached, only leaving a narrow, fluorescent zone of GFP-tagged caveolin structures, the fluorescence disappeared only very slowly from the peripheral zone (see Fig. 5t in Ref. ). Based on extrapolations of such data, the fluorescence would disappear completely from the periphery within about 3 h. Taken together, the described EM and FRAP data are consistent with a very slow, constitutive internalization of caveolae.
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This ebook provides an introductory explanation of the workings of the human body, with an effort to draw connections between the body systems and explain their interdependencies. A framework for the book is homeostasis and how the body maintains balance within each system. This is intended as a first introduction to physiology for a college-level course.