Appendix B. Blood–brain barrier permeabilities of mannitol, sucrose and inulin and the identification of markers for perivascular elimination
Evidences that various substances are markers for perivascular elimination and that elimination to CSF or lymph from the parenchyma is primarily perivascular are intertwined. One of the principal arguments that many substances leave the parenchyma by a convective process is that the clearances for these are all similar despite their being a large range of sizes, e.g. from mannitol and sucrose on one hand, to serum ablumin and many of the polyethyleneglycols (PEGs) and dextrans on the other. These arguments were first advanced by Cserr and associates [126, 127, 129]. However, the data on which they based their argument was obtained under barbiturate anaesthesia, which is now known to greatly suppress the perivascular efflux process.
It is thus important that Groothuis et al. [131] have reexamined the rates of elimination for a range of polar substances including sucrose (MW 342), inulin (MW 5500) and dextran-70 K (MW 70,000) (see Table 1) and have found again that there is no variation in the rate constant for elimination. Others have also measured efflux rate constants, for mannitol and inulin and have found similar values (see Table 1). For many of these substances there is no evidence for transport across the blood–brain barrier. However, for mannitol, sucrose and inulin there appear to be measurable rates of influx, so it must be asked whether they should be included in the list of markers.
The measured rate constant for elimination keff − total, of the putative markers from the parenchyma is the sum of the efflux rate constants for the perivascular and blood–brain barrier routes. Groothuis et al. [131] found values close to 0.24 h−1 = 0.003 min−1. The rate constant for the passive efflux of a neutral solute across the blood–brain barrier can be calculated from the permeability-surface area product, PS, and the volume of distribution, VD, as keff,BBB= PS/VD (see [127, 131] and Appendix A). Because these substances are restricted to ISF within the parenchyma, VD can be taken to be 0.2 mL g−1. PS can be measured from the initial rate of accumulation when the substance is added to the blood. Values are tabulated in Table 7. There is obviously considerable variation in the values found in different studies, which probably reflects the difficulties inherent in measuring small permeabilities. However, several features are apparent. Firstly in each of the three studies that compared mannitol, sucrose and inulin, mannitol had the highest permeability, inulin the least. Secondly the same order is apparent in the averages, 0.0040 ± 0.0013 min−1, 0.0011 ± 0.0002 min−1 and 0.00027 ± 0.0001 min−1. Thirdly for sucrose and inulin the rate constants for elimination by transport across the blood–brain barrier are substantially less than the total rate constant for elimination, 0.003 min−1, implying that some other mechanism accounts for most of the elimination. It is at present unclear whether or not mannitol is suitable as a marker for perivascular elimination.
Preston et al. [531] pointed out a major difficulty that occurs in the measurement of very small permeabilities using radiotracers. If the sample of labelled substance contains small quantities of labelled impurities that are more permeable than the principal substance, the impurities will make a disproportionately large contribution to permeability measured by accumulation of the radiolabel. They showed that further purification by thin layer chromatography decreased the measured permeability for mannitol and sucrose (see Table 7). Miah et al. [532] have taken this one step further and have compared the uptake of radiolabel from a sample of 14C-sucrose with the uptake of 13C-sucrose measured by mass spectrometry. Their measurements using a sample of radiolabelled sucrose yielded one of the higher measured permeabilities, while that obtained using assay of sucrose yielded the lowest. It is thus plausible that perivascular elimination accounts for an even larger fraction of the elimination of sucrose than indicated by comparison of the average of the rate constants in the table with the total rate constant of elimination.
The available evidence strongly supports the commonly held view that sucrose, inulin, albumin, and a number of dextrans are suitable markers for elimination from the brain parenchyma by perivascular convection.