The successful development of drugs to treat diseases of the central nervous system will require adequate transport of these agents across the blood–brain and/or blood–cerebrospinal fluid barriers (i.e., BBB and BCSFB) (Taylor, 2002
; Begley, 2004
; Ohtsuki and Terasaki, 2007
). These barrier systems probably arose from the necessity to maintain strict homeostasis of the extracellular fluid and CSF for normal brain function. Although originally conceived as a physical barrier formed by the linking tight junctions of brain capillary endothelial cells and epithelial cells of the choroid plexus and arachnoid membrane, the BBB and BCSFB also include degradative enzymes as well as efflux transporters that can affect the delivery of these agents to specific regions in brain (Smith et al, 2004
). This is important not only for drug delivery but also for endogenous neuropeptides that are released within the brain. Beyond the BBB and BCSFB, the pharmacology of drugs or the physiology of neuromodulating peptides depends on their movement through the interstitial fluid into adjacent brain tissue, their uptake and binding by brain cells, their exchange between brain and CSF at the ependyma and glia limitans, and their distribution through the ventricles and subarachnoid space.
PEPT2 (SLC15A2), a high-affinity, low-capacity carrier of the proton-coupled oligopeptide transporter family, facilitates the movement of substrates across biologic membranes through an electrochemical membrane gradient (Brandsch et al, 2008
; Rubio-Aliaga and Daniel, 2008
). PEPT2 is the most studied protein of the proton-coupled oligopeptide transporter family found in brain where it is expressed at the apical surface of choroid plexus epithelia (i.e., CSF-facing), as well as in astrocytes (newborn rats) and neuronal cells (newborn and adult rats) (Shen et al, 2004
). Residing at the blood–CSF interface, PEPT2 has a significant role in limiting the exposure of peptides/mimetics and peptide-like drugs in CSF by transporting these substrates into choroid plexus tissue (Shu et al, 2002
; Teuscher et al, 2004
; Shen et al, 2005
). In accordance with this, the synthetic dipeptide glycylsarcosine (GlySar) (Ocheltree et al, 2005
), the aminocephalosporin antibiotic cefadroxil (Shen et al, 2007
), and the naturally occurring dipeptide carnosine (Kamal et al, 2009
) were found to have reduced CSF levels in vivo
after intravenous bolus administration, but increased choroid plexus levels in wild-type mice as compared with Pept2
null animals. Moreover, pharmacologic differences linked to PEPT2 expression in brain have been observed. For instance, this protein protects against 5-aminolevulinic acid (5-ALA) neurotoxicity after subcutaneous dosing in wild-type mice (Hu et al, 2007
) and produces dissimilar antinociceptive responses between wild-type and Pept2
null mice after intracerebroventricular (icv) administration of kyotorphin (Jiang et al, 2009
). Such findings show that PEPT2 expression in brain can have profound effects on both the disposition and dynamics of peptide/mimetic substrates and peptide-like drugs.
In a study of convectional and diffusional movement in the CSF–brain system, the intracranial distribution of [14
C]sucrose after icv injection was examined in rats by quantitative autoradiographic analysis (Ghersi-Egea et al, 1996
). The resulting data indicated that the circulatory and delivery functions of CSF were complex and varied among CSF compartments. Rapid convective (bulk) flow of CSF and CSF-entrained [14
C]sucrose were observed throughout the ventricular system and through the velum interpositum and superior medullary velum into subarachnoid cisterns, but such flow was rather slow and marginal to and over the cerebral and cerebellar cortices. Results also indicated a CSF–brain barrier to sucrose diffusion in some areas of the glia limitans but not ependyma. Sucrose was also found to accumulate in and/or around the blood vessels within the subarachnoid cisterns, suggesting that the pia-arachnoid investing such vessels is different than that at cerebral and cerebellar cortical surfaces.
The preceding pharmacologic and physiologic findings are limited and indicate how little is known about the effects of specific transporters on the circulation and distribution of drugs or endogenous peptide/mimetic substrates in CSF and brain parenchyma. This is not surprising because the first mammalian ion-coupled drug transporters were cloned around the early mid-1990s (Fei et al, 1994
; Gründemann et al, 1994
) and genetically modified mice were developed even more recently (Schinkel et al, 1994
; Jonker et al, 2002
; Shen et al, 2003
). In view of this, the objectives of this study were to (1) define the CSF clearance kinetics of PEPT2 substrates (i.e., GlySar and cefadroxil) after icv administration, (2) determine if regional differences exist in the choroid plexus uptake of GlySar across the lateral, third, and fourth ventricles, and (3) evaluate how PEPT2 might alter the penetration of GlySar from CSF into parenchymal tissue in different regions of the brain. To address these objectives, we performed experiments in wild-type and Pept2
null mice, a unique resource to evaluate the role of a single gene on drug disposition and dynamics (Kamal et al, 2008
); in addition, quantitative autoradiography (QAR) was applied for the accurate localization of radioactive drug within parts of the CSF system and specific regions of brain tissue.