Occludin and Claudin-5 are Comparably Abundant and Co-localized in the Rat’s Blood Brain Barrier from Late Gestation to Adulthood

Background: The role of tight junction proteins (TJPs) in maintaining blood brain barrier (BBB) permeability after hypoxic-ischemic (H-I) cerebral injury has recently gained significant interest. In recent animal studies of such injury, TJP degradation by matrix metalloproteinase (MMP) has been postulated as a mechanism of BBB disruption. Although variation in abundance of occludin and claudin-5 is frequently used as a marker of BBB disruption, in the rat brain, the abundance of these proteins throughout development has not been well characterized. Objective: To characterize the abundance and localization of cerebral occludin and claudin-5 in the rat brain across multiple stages of development, as a first step in understanding the ontogeny of the BBB. Methods: Sprague-Dawley rats of different developmental ages: embryonic day 17 (E17), postnatal day 7 (P7), and postnatal day 70 (P70) were used as subjects for the study. Cerebral occludin and claudin-5 expression and tissue localization was assessed with immunoblotting, ELISA, and immunofluorescence. One-way ANOVA was used to compare the level of protein expression between the groups. Descriptive data was expressed as mean and SEM. Results: In our study, western blotting, ELISA and immunofluorescence showed abundance of occludin and claudin-5 in the rat brain across a variety of developmental ages. No differences between age groups were observed with ELISA. Immunofluorescence revealed that occludin and claudin-5 are co-localized in cortical vessels comprising the rat BBB at every study age. Conclusions: Our findings of occludin and claudin-5 co-expression at every study age suggest that the rat BBB is relatively intact early in development. The confirmation of occludin and claudin-5 co-localization in the rat brain cortical vessels further strengthens this hypothesis.