Analytical and biological methods for probing the blood-brain barrier

Abstract

The blood-brain barrier (BBB) is an important interface between the peripheral and central nervous systems. It protects the brain against the infiltration of harmful substances and regulates the permeation of beneficial endogenous substances from the blood into the extracellular fluid of the brain. It can also present a major obstacle in the development of drugs that are targeted for the central nervous system. Several methods have been developed to investigate the transport and metabolism of drugs, peptides, and endogenous compounds at the BBB. In vivo methods include intravenous injection, brain perfusion, positron emission tomography, and microdialysis sampling. Researchers have also developed in vitro cell-culture models that can be employed to investigate transport and metabolism at the BBB without the complication of systemic involvement. All these methods require sensitive and selective analytical methods to monitor the transport and metabolism of the compounds of interest at the BBB.

Figure 1

The cells that make up the blood-brain barrier and the neurovascular unit. Reprinted with permission from Reference 3.

Figure 2

Mechanisms of transport across the blood-brain barrier.

Figure 3

(a) Whole-body positron emission tomography (PET) acquired over two minutes after an intravenous injection of ¹⁸F-labeled fluoropyridinyl (FPy) derivatives. (b) In vivo cerebral pharmacokinetics determined using the PET images. Reprinted with permission from Reference 22.

Figure 4

Microdialysis sampling system. The probe is implanted in the brain of the rat. Small molecules and peptides of interest from the extracellular fluid of the brain diffuse across the probe membrane and are transported in the perfusate to a fraction collector or online analytical system.

Figure 5

In vivo monitoring of the transport of MPD into the brain correlated with dopamine (DA) release and behavior. Abbreviation: MPD, methylphenidate. Reprinted with permission from Reference 30.

Figure 6

On-rat sampling system for monitoring drug transport and behavior. Abbreviations: DOPAC, 3,4-dihydroxyphenylacetic acid; FEP, fluorinated ethylene propylene; 5-HIAA, 5-hydroxyindoleacetic acid; HVA, homovanillic acid. Reprinted with permission from Reference 64.

Figure 7

Simultaneous monitoring of blood-brain barrier (BBB) integrity and excitatory amino acids in the brain via microdialysis sampling coupled to microchip electrophoresis. Fluorescein (Fl) was used as a marker of BBB integrity. Abbreviations: D, aspartic acid; E, glutamic acid. Reproduced from Reference 68.

Figure 8

Systems used for monitoring in vitro transport across the blood-brain barrier. (a) In the Side-by-Side™ diffusion chamber, cells are grown on polycarbonate membranes (PCMs) that are subsequently mounted between two water-jacketed, thermally controlled chambers. (b) In the Transwell® system, a coculture setup is demonstrated in a six-well format in which brain endothelial cells are grown in the Transwell insert and astrocytes are grown in the bottom of the multiwall plates. Abbreviation: BBMEC, bovine brain microvessel endothelial cell.

Figure 9

Accumulation of Rhodamine 123 in bovine brain microvessel endothelial cells following treatment with (a) 5 μM of Rhodamine 123 alone and (b) 5 μM of Rhodamine 123 with 10 μM of Paclitaxol, an anticancer drug and known P-gp substrate. Decreased Rhodamine 123 efflux is observed in the presence of Paclitaxol (76).

Figure 10

Metabolism of substance P (SP) by bovine brain microvessel endothelial cells. The cells were grown in multiwell culture plates and incubated with SP; then the aliquots were removed over time. The time course and appearance of each SP metabolite are shown. Reprinted with permission from Reference 63.

Figure 11

Transport of dynorphin A 1–11 amide by bovine brain microvessel endothelial cells. The lag time observed is indicative of a receptor-mediated process (90). (a) Blood to brain. (b) Brain to blood. (c) Effect of temperature on transport, indicating an active transport process.

Figure 12

Microfluidic system for the investigation of nitric oxide release by endothelial cells in the presence of erythrocytes. Reproduced with permission from Reference 103.