Measurement of brain uptake of radiolabeled substances using a tritiated water internal standard

Uptake transporters in brain microvascular endothelial cells (BMECs) are involved in the penetration of basic (cationic) drugs such as diphenhydramine (DPHM) into the brain. Lipopolysaccharide (LPS)-induced inflammation alters the expression levels and activities of uptake transporters, which change the penetration of DPHM into the brain. A brain microdialysis study showed that the unbound brain-to-plasma partition coefficient (K_p,uu,brain) for DPHM in LPS rats was approximately two times higher than that in control rats. The transcellular transport of DPHM to BMECs was increased when BMECs were cultured with serum from LPS rats. Compared with control rats or BMECs, the brain uptake of DPHM in LPS rats was increased and the intracellular accumulation of DPHM was increased under a high intracellular pH in BMECs from LPS rats, respectively. Treatment of BMECs with transporter inhibitors or inflammatory cytokines had little impact on the intracellular accumulation of DPHM in BMECs. This study suggests that LPS-induced inflammation promotes unidentified proton-coupled organic cation (H⁺/OC) antiporters that improve the penetration of DPHM into rat brain via the blood-brain barrier.

Understanding the pharmacokinetics of therapeutics for disorders of the central nervous system (CNS) is critical for the development and administration of these drugs, as inefficiencies in drug effect are often not due to the drug itself, but instead due to the blood–brain barrier (BBB). The restricted permeability of this barrier hinders transcellular and paracellular transport. Various methods of pharmacokinetic analysis may elucidate the delivery of molecules from the systemic circulation to the CNS. Such analysis provides information on BBB permeability and an indication of the therapeutic potential of a drug. Many BBB manipulation methods are currently being developed; these novel strategies aim to enhance permeability and amplify the efficacy of neurotherapeutics. Herein we provide a succinct overview of traditional pharmacokinetic approaches to ascertain systemic drug delivery of compounds for CNS conditions. We also provide details on current approaches aimed at enhancing drug delivery across the BBB.

The blood-brain barrier (BBB) is a complex and highly selective barrier, made of endothelial cells, pericytes, basement membrane, astrocytes, and tight junctions. BBB protects the central nervous system from aberrant exposure to toxins and even drugs. It is important to determine the extent of drugs that can be delivered to CNS through BBB and for which different in vitro and in vivo models have been utilized. The in vitro models utilized for assessment of drug delivery through BBB are based on synthetic materials or cell cultures originated from animal brains or human stem cells. Also, different animals have been utilized to create in vivo models to assess drug delivery through BBB; commonest are rodents. Various researchers have utilized many analytical techniques in these in vitro and in vivo models, to assess the possible delivery of different drugs that may be delivered through BBB. This chapter covers such in vitro models, in vivo models, and analytical techniques, which can be used in tandem with each other to assess the delivery of variety of agents through BBB.

Increase in the incidence of central nervous system (CNS) diseases in recent years, along with the complexity of the structure of the brain tissue, has led to an important need to develop more effective and safe treatments with CNS drugs used in this field. The CNS drugs need to pass through the blood–brain barrier (BBB) and reach the target area with an effective concentration to induce a pharmacological effect. Due to its position, surface area, and function, BBB plays a protective role in the transport of compounds, which generally cross the barrier via passive diffusion and active transport mechanisms. Other important parameters contributing to effective concentrations of the drug in the brain tissue are the drug penetration rate and the drug penetration degree. In addition, the physicochemical characteristics of drugs and the cerebral blood flow rate (F) are important parameters for affecting the brain penetration as well. The drug penetration rate [unidirectional influx clearance from blood to the brain (CL_in or K_in)], drug penetration degree [the ratio of unbound brain/unbound plasma concentration (K_p,uu,brain)], and drug distribution in the brain [distribution volume of unbound drug in brain (V_u,brain)] are the main parameters that should be determined to verify whether the drug has reached effective concentrations in the brain or not. In vivo, in vitro, in silico, and in situ methods are being used for the determination of these parameters.

Recently tremendous progress has been made in studying choroid plexus (CP) physiology and pathophysiology; and correcting several misconceptions about the CP. Specifically, the details of how CP, a locus of the blood–CSF barrier (BCSFB), secretes and purifies CSF, generates intracranial pressure (ICP), maintains CSF ion homeostasis, and provides micronutrients, proteins and hormones for neuronal and glial development, maintenance and function, are being understood on a molecular level. Unequivocal evidence that the CP secretory epithelium is the predominant supplier of CSF for the ventricles comes from multiple lines: uptake kinetics of tracer ²²Na and ³⁶Cl penetration from blood to CSF, autoradiographic mapping of rapid ²²Na and ³⁶Cl permeation (high permeability coefficients) into the cerebroventricles, CSF sampling from several different in vivo and in vitro CP preparations, CP hyperplasia that increases CSF formation and ICP; and in vitro analysis of CP ability to transport molecules (with expected directionality) and actively secrete fluid against an hydrostatic fluid column. Furthermore, clinical support for this CP–CSF model comes from neurosurgical procedures to remove lateral ventricle CPs in hydrocephalic children to reduce CSF formation, thereby relieving elevated ICP. In terms of micronutrient transport, ascorbic acid, folate and other essential factors are transported by specific (cloned) carriers across CP into ventricular CSF, from which they penetrate across the ependyma and pia mater deeply into the brain to support its viability and function. Without these choroidal functions, severe neurological disease and even death can occur. In terms of efflux or clearance transport, the active carriers (many of which have been cloned and expressed) in the CP basolateral and apical membranes perform regulatory removal of some metabolites (e.g. choline) and certain drugs (e.g. antibiotics like penicillin) from CSF, thus reducing agents such as penicillin to sub-therapeutic levels. Altogether, these multiple transport and secretory functions in CP support CSF homeostasis and fluid dynamics essential for brain function.

During the research and development of new drugs directed at the central nervous system, there is a considerable attrition rate caused by their hampered access to the brain by the blood–brain barrier. Throughout the years, several in vitro models have been developed in an attempt to mimic critical functionalities of the blood–brain barrier and reliably predict the permeability of drug candidates. However, the current challenge lies in developing a model that retains fundamental blood–brain barrier characteristics and simultaneously remains compatible with the high throughput demands of pharmaceutical industries. This review firstly describes the roles of all elements of the neurovascular unit and their influence on drug brain penetration. In vitro models, including non-cell based and cell-based models, and in vivo models are herein presented, with a particular emphasis on their methodological aspects. Lastly, their contribution to the improvement of brain drug delivery strategies and drug transport across the blood–brain barrier is also discussed.