Transient weakening of the blood-brain barrier in neonatal mice

Tight junctions (TJs) are a major structural component of the blood-brain barrier (BBB), contributing to brain homeostasis by restricting paracellular diffusion. Although BBB maturation begins during embryogenesis, the timing and dynamics of functional barrier maturation perinatally remain unclear. This question is particularly relevant for the striatum, a metabolically demanding brain region that undergoes rapid postnatal maturation and is vulnerable to neonatal injury.

We investigated developmental dynamics of BBB tracer permeability in mouse striatum using in situ microperfusion of capillaries (ISMICAP) combined with two-photon microscopy. Small-molecule tracers were applied from late embryonic (E18) to adolescent (P25) stages. Bovine serum albumin (BSA) was applied as a macromolecular tracer at P2 and P25.

Small-molecule tracers, namely 7-hydroxycoumarin-3-carboxylic acid (7HCC), sulforhodamine 101 (SR101), and biocytin-tetramethylrhodamine, revealed a pronounced but transient increase in extravascular fluorescence during the neonatal phase (P0-P2), followed by progressive restriction by P12 and P25. During the neonatal permeability window, 7HCC labeled perivascular cells located ~ 2.8 μm from the endothelium and distinct from NT500/525-labeled pericytes, whereas SR101 accumulated within endothelial cytoplasm, indicating tracer-specific vascular and perivascular accumulation patterns. The membrane probe FM1-43 showed a similar temporal pattern, with enhanced diffusion to abluminal membranes and labeling of pericyte-like mural cells. In contrast, BSA showed low extravascular fluorescence at P2 comparable to P25, indicating that increased neonatal small-molecule permeability did not extend to BSA-sized macromolecules. At P25, small-molecule tracer permeability was higher in the striatum than in the cortex, whereas FM1-43 showed the opposite regional pattern.

BBB maturation in the striatum is not a linear tightening process but includes a transient neonatal phase of increased TJ-associated permeability to small-molecule tracers, while remaining restrictive to BSA-sized macromolecules. Although tracer-specific vascular and perivascular accumulation patterns were observed, the overall temporal profile supports a discrete perinatal window of decreased barrier restriction. This dynamic permeability window may reflect physiological remodeling of barrier function during the perinatal transition. Defining this window mechanistically may improve understanding of neonatal brain vulnerability and may inform strategies for temporally targeted CNS drug delivery.