The Origin of Deoxynucleosides in Brain: Implications for the Study of Neurogenesis and Stem Cell Therapy

Abstract

Detection of DNA synthesis in brain employing ((3)H)thymidine (((3)H)dT) or bromo deoxyuridine (BrdU) is widely used as a measure of the "birth" of cells in brain development, adult neurogenesis and neuronal stem cell replacement strategies. However, recent studies have raised serious questions about whether this methodology adequately measures the "birth" of cells in brain either quantitatively or in an interpretable way in comparative studies, or in stem cell investigations. To place these questions in perspective, we review deoxynucleoside synthesis and pharmacokinetics focusing on the barriers interfacing the blood-brain (cerebral capillaries) and blood-cerebrospinal fluid (choroid plexus), and the mechanisms, molecular biology and location of the deoxynucleoside transport systems in the central nervous system. Brain interstitial fluid and CSF nucleoside homeostasis depend upon the activity of concentrative nucleoside transporters (CNT) on the 'central side' of the barrier cells and equilibrative nucleoside transporters (ENT) on their 'plasma side.' With this information about nucleoside transporters, blood/CSF concentrations and metabolic pathways, we discuss the assumptions and weaknesses of using ((3)H)dT or BrdU methodologies alone for studying DNA synthesis in brain in the context of neurogenesis and potential stem cell therapy. We conclude that the use of ((3)H)dT and/or BrdU methodologies can be useful if their limitations are recognized and they are used in conjunction with independent methods.

Vectorial Ligand Transport Through Mammalian Choroid Plexus

“…Cnt3, a sodium-dependent energy-requiring (active) transporter (Table IV), and Ent1, a nitrobenzylthioinosine (NBTI)-sensitive facilitated diffusion transporter are located on the apical and basolateral membranes of CP, respectively (1,31,32). Cnt3 and the Ent1 sequentially transfer both ribo-and deoxyribonucleosides across CP from CSF into blood (see below).…”

“…In previous publications, we and others have described these techniques in detail (1,8,9,13). Now we understand many of these transport processes at both the BBB and the CP.…”

“…These transporters were very likely located in the choroid plexus (CP), since, in vitro, the isolated CP, the locus of the blood-CSF barrier (BCSFB), accumulated these substances in a fashion consistent with the in vivo data. Anatomically, the single layer of tight-junctioned choroid plexus epithelial cells has two sides (7): an apical side facing the CSF and a basolateral side facing the choroid plexus capillary endothelium, which is not joined by tight junctions (1,8). Moreover, there was a widespread and correct belief that for vectorial transport there must be separate transporters at the apical (luminal) and basolateral (interstitial) sides of the CP, as was initially shown for glucose in the tightjunctioned cerebral capillaries that make up the bloodbrain barrier (BBB) (9).…”

Choroid Plexus–Cerebrospinal Fluid Circulatory Dynamics: Impact on Brain Growth, Metabolism, and Repair

“…These substances are needed in nano-to micromolar concentrations over extended periods. Micronutrients transported into CSF include vitamin C, folates (4), deoxyribonucleosides (8), vitamin B 6 (9), and certain trace elements. Active transport pump-like carriers in the CP epithelium pull these micronutrients across the blood-facing membrane of the plexus.…”

“…Transport specificity at the BCSFB points to pharmacologic opportunities (6) for regulating the movement of neurotrophic and reparative agents to neuronal networks. Biomedical devices and strategies for stem cell augmentation (8), as well as CP transplants and transgene manipulation (6,36), may also be feasible by intraventricular instillation (6). Boosting trophic factors, antagonists, and chelators in CSF will benefit neural repair in trauma (37), ischemia (38), hyperthermia (39), and infectious disease.…”