Induction of endogenous CLN2 mRNA transport by β-adrenergic activation in hippocampal neurons. In situ hybridization was performed with a probe specific for rat CLN2 mRNA. (A and B) Hybridization signal (white silver grains) indicates somatic localization of endogenous CLN2 mRNA under basal conditions. (C and D) After activation of β-ARs with agonist isoproterenol, CLN2 mRNA was found localized along the entire dendritic extent. (E–H) Preincubation of cells with intracellular Ca2+ chelator BAPTA-AM (E and F) or with L-type VDCC blocker nifedipine (G and H) prevented isoproterenol-induced dendritic delivery of CLN2 mRNA. (I and J) Little or no signal was detectable when in situ hybridization was performed with a CLN2 mRNA “sense strand” control probe. Number of cells analyzed: (A and B) 14 neurons, 71 dendrites; (C and D) 14 neurons, 72 dendrites; (E and F) 11 neurons, 53 dendrites; (G and H) 12 neurons, 60 dendrites. Bar, 50 µm. (K) Quantitative analysis. One-way ANOVA, Dunnett’s post hoc analysis (comparison of RNA levels in the basal state with RNA levels after β-adrenergic activation and after β-adrenergic activation in the presence of BAPTA-AM or nifedipine): comparison with isoproterenol (C and D), P < 0.001 for all interval points; comparison with isoproterenol/BAPTA-AM (E and F), P > 0.6 for interval points 50 µm; comparison with isoproterenol/nifedipine (G and H), P > 0.7 for interval points 50 µm. Error bars indicate SEM.
