Age-dependence of the lateral diffusion coefficient of Concanavalin-A receptors in the plasma membrane of ex vivo prepared brain cortical nerve cells of BN/BiRijHsd rats

Abstract 

A new method has been developed for ex vivo preparation of brain cortical cells of BN/BiRijHsd rats to make them suitable for the measurement of the lateral diffusion coefficient of the membrane components by means of fluorescence recovery after photobleaching (FRAP). The method involves chopping the brain cortex into pieces of less than 1 mm. These parts are stained with a fluorescent label (e.g., concanavalin-A-fluorescein, Con-A-FL conjugate) and then gently pressed onto a microscope slide using the coverslip. In the resulting specimen, the largest cells of the cortex can be recognized in phase-contrast image, sufficiently stained by the label and ready for the FRAP measurement. The lateral diffusion coefficient of Con-A-receptor proteins (D _p) was measured in such brain cell preparations of 15 female rats in four age groups (5.6–31.8 months) and 11 males in three age groups (13.8–31.8 months). Highly significant negative, linear age correlation of D _p (R=−0.9958 in females, and −0.9956 in males) were found, the regression equations being D _p,=(8.8311–0.1425 X)⁻¹⁰ and D _p█=(9.3240−0.1630 X)⁻¹⁰ cm²/s, respectively, where X is age in months. The data confirm that the lateral mobility of plasma membrane proteins represents an important biomarker of cellular aging in the brain cortical cells of BN/BiRijHsd rats.