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. 2010 Jan;24(1):102-9.
doi: 10.1016/j.bbi.2009.09.001. Epub 2009 Sep 6.

Minimal penetration of lipopolysaccharide across the murine blood-brain barrier

William A Banks  1 , Sandra M Robinson
Affiliations

Minimal penetration of lipopolysaccharide across the murine blood-brain barrier

William A Banks et al. Brain Behav Immun. 2010 Jan.

Abstract

LPS given peripherally or into the brain induces a neuroinflammatory response. How peripheral LPS induces its effects on brain is not clear, but one mechanism is that LPS crosses the blood-brain barrier (BBB). Alternatively, LPS acts outside the BBB by stimulating afferent nerves, acting at circumventricular organs, and altering BBB permeabilities and functions. Here, we labeled LPS with radioactive iodine (I-LPS) and coinjected it with radioactively labeled albumin (I-Alb) which acted as a vascular space marker. Measurable amounts of I-LPS associated with the BBB, most reversibly bound to brain endothelia. Brain endothelia also sequestered small amounts of I-LPS and about 0.025% of an intravenously injected dose of I-LPS crossed the BBB to enter the CNS. Disruption of the BBB with repeated injections of LPS did not enhance I-LPS uptake. Based on dose-response curves in the literature of the amounts of LPS needed to stimulate brain neuroimmune events, it is unlikely that enough peripherally administered LPS enters the CNS to invoke those events except possibly at the highest doses used and for the most sensitive brain functions. I-LPS injected into the lateral ventricle of the brain entered the circulation with the reabsorption of cerebrospinal fluid (bulk flow) as previously described. In conclusion, brain uptake of circulating I-LPS is so low that most effects of peripherally administered LPS are likely mediated through LPS receptors located outside the BBB.

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Figures

Figure 1
Figure 1
Upper Panel: Clearance of I-LPS administered by intravenous injection. Results are expressed as the log of the level of radioactivity per ml of serum vs time. The relation was highly significant between log(serum cpm) and time (p<0.001) demonstrating clearance of I-LPS from blood; the half-time clearance was calculated to be 259 min with a volume of distribution of 1.5 ml. Lower Panel: Uptake by brain of I-LPS after its intravenous administration. There was no statistically significant correlation between brain/serum ratios and exposure time, indicting that there was no measurable BBB penetration of I-LPS by this method. The mean brain/serum ratio for I-LPS of 11.3 was in the upper range of the vascular space for brain.
Figure 2
Figure 2
Effect of inclusion of unlabeled LPS (100 µg/mouse) or washout of brain vascular space on brain/serum ratios of I-LPS 1, 2, and 5 min after the iv injection of I-LPS. Values are corrected for vascular space by including I-Alb in the iv injection and subtracting the brain/serum ratios for I-Alb from those for I-LPS. The lack of an effect for unlabeled LPS indicates no saturable component to I-LPS uptake. A decrease in brain/serum ratios for I-LPS with washout further to that seen with correction for the I-Alb space indicates a reversible binding of I-LPS to brain endothelial cells.
Figure 3
Figure 3
Capillary depletion demonstrates the relation of capillary sequestration and BBB penetration of I-LPS. The results show that about half of the I-LPS associated with brain had permeated the BBB and about half was sequestered by the capillary bed.
Figure 4
Figure 4
Effect of 3 injections of LPS over 24 h on the uptake of I-Alb and I-LPS by brain. Upper panel shows that LPS treatments increased I-Alb uptake (p<0.05), demonstrating BBB disruption. Lower panel shows I-LPS results corrected for the I-Alb space and shows that LPS treatment had no statistical effect.
Figure 5
Figure 5
Brain perfusion followed with or without washout. Without washout, uptake of I-LPS was measured to be 8.65 ± 1.96 µl/g-min (n = 9); this compares to an unmeasurable uptake by brain after iv injection as shown in Figure 1, lower panel. This indicates that circulating factors retarded uptake. With vascular washout, this uptake was greatly reduced, demonstrating that most of the uptake is likely reversible binding to the luminal surface of brain endothelial cells.
Figure 6
Figure 6
Brain-to-blood efflux of I-LPS after intracerebroventricular injection. Main panel shows efflux rate of 33.7 min. Each point represents the mean of 3 mice. Inset shows that inclusion of unlabeled LPS did not alter efflux of I-LPS from brain (n = 6–8/group).
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References

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