For quantification of GIRK2 fluorescence in dendritic processes, only infected neurons that were clearly separated from other infected cells were evaluated. MAP2 labeling was used to identify clearly the dendrites; 2–3 relatively straight segments (35–50 microns) of primary, secondary and tertiary dendrites exhibiting even MAP2 labeling from each neuron were selected for analysis. The primary dendrite fragments were located at least 20 microns from the soma. GIRK2 fluorescence intensity measurements within these processes were acquired following background subtraction, and data are presented as arbitrary units (AU). For GIRK2 isoform and PSD-95 co-localization analysis, the soma and surrounding area (20 microns from the edge of the soma) was removed from the image prior to analysis. Quantification was performed using the NIH ImageJ 1.29 plug-in, Puncta Analyzer. MAP2 labeling was used to delineate dendritic processes, and red (GIRK2) and green (PSD-95) channels were thresholded to highlight visible puncta without introduction of background noise. The percentage of co-localized GIRK puncta overlapping with total PSD-95 puncta was calculated for each analyzed neuron.
