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. 2015 Feb 18;6:6319. doi: 10.1038/ncomms7319

Figure 6. Effects of dendritic cAMP or dopamine release on nuclear PKA.

Figure 6

Live imaging of nuclear PKA activity shows a nonlinear, non-monotonic relationship with the distance of dendritic cAMP or dopamine release from the cell body. (a) Striatal neurons in culture were infected with Sindbis-AKAR3 and 6 h later loaded with DMNB-caged-cAMP. cAMP was released with a brief (1 s) ultraviolet pulse (d=1 μm) at the indicated locations. Two stimulations at different locations (randomized order between proximal, medial and distal locations) were carried out per neuron, with an interstimulus interval of at least 30 min. Scale bar, 10 μm. (b) Time course of normalized 480 nm/535 nm fluorescence ratio of nuclear AKAR3, following the local release of cAMP at various locations in dendrites. Data are means±s.e.m., n=9–12 in four different experiments. (c) The experimental points (area under the curve (AUC) in b, colour) are plotted with the model prediction (Fig. 3f, grey dashed line; the correspondence of ordinate values is arbitrary). Comparison of the experimental points grouped as indicated by the colour code (horizontal bars correspond to means: proximal, 12±2 μm from the nucleus, mean±s.e.m., n=12, medial, 53±3 μm, n=9, and distal 108±7 μm, n=10), one-way analysis of variance (ANOVA), F(2,28)=5.89, P=0.007; Tukey’s test, *P<0.05, **P<0.01. (d) Striatal neurons transfected with D1R receptor, expressing AKAR3 as in a, were incubated with 100 μM ((N)-1-(2-nitrophenyl)ethylcarboxy-3,4-dihydroxyphenethylamine-caged-dopamine, which was photo-released as in a at the indicated distances from the nucleus. Comparison of the experimental points grouped as proximal, medial and distal, as indicated by the colour code (horizontal bars correspond to means: proximal, 10.3±0.7 μm, n=6, medial, 48.9±4.9 μm, n=9, distal, 101.6±4.9 μm, n=5, from 3 different experiments). FRET responses: 1-way ANOVA, F(2,17)=11.40, P=0.0007; Tukey’s test, **P<0.01.