Diffusion of Epidermal Growth Factor in Rat Brain Extracellular Space Measured by Integrative Optical Imaging

Abstract

Epidermal growth factor (EGF) stimulates proliferation, process outgrowth, and survival in the CNS. Understanding the actions of EGF necessitates characterizing its distribution in brain tissue following drug delivery or release from cellular sources. We used the integrative optical imaging (IOI) method to measure diffusion of fluorescently labeled EGF (6,600 M r; 4 μg/ml) in the presence of excess unlabeled EGF (90 μg/ml) to compete off specific receptor binding and reveal the “true” EGF diffusion coefficient following injection in rat brain slices (400 μm). The effective diffusion coefficient was 5.18 ± 0.16 × 10−7(SE) cm 2/s ( n = 22) in rat somatosensory cortex and the free diffusion coefficient, determined in dilute agarose gel, was 16.6 ± 0.12 × 10−7 cm 2/s ( n = 27). Tortuosity (λ), a parameter representing the hindrance imposed on EGF by the convoluted brain extracellular space (ECS), was 1.8, the lowest yet measured by IOI for a protein in brain. Control experiments with fluorescent dextran of similar molecular weight and tetramethylammonium confirmed EGF did not affect local ECS structure. We conclude that transport of smaller growth factors such as EGF through brain ECS is less hindered than that of larger proteins (>10,000 M r, e.g., nerve growth factor) where typically λ > 2.1. Modeling was used to predict that low λ will allow EGF sources in the brain to be further from target cells and still elicit a biological response. High λ values for larger growth factors imply more constrained local biological effects than with smaller proteins such as EGF.