Fig 1: Multiphoton images illustrating the immunocytochemical location of SyGCaMP2-mCherry in the CA1 region of the hippocampus. An antibody raised against mCherry was used to establish the location of SyGCaMP2-mCherry with respect to the presynaptic marker bassoon (A,B) and the postsynaptic marker PSD-95 (C,D). Images in green show the expression patterns of bassoon and PSD-95 and images in red illustrate the location of mCherry. The third column shows a merged image of green and red fluorescence in each case. Scale bars represent 50 µm in the upper and 10 µm in the lower rows. The positions of the s. radiatum (sr), s. pyramidale (sp) and s. oriens (so) are indicated. Panels (B,D) illustrate fluorescence values extracted from lines drawn through punctate regions of the higher magnification images illustrated in panels (A,B) respectively. Absolute fluorescence values are not shown since they were dependent on both the microscope settings for each fluorophore and the relative proportion of the merge value selected. To the right are graphs showing the relationship between each synaptic marker and mCherry along with the Pearson correlation and P values in each case (****P < 0.0001). Panels (E–H) illustrate images obtained using antibodies against vGluT1 and vGaT which identify glutamatergic and GABA-ergic terminals respectively. Data are presented in the same way as panels (A–D).