Visualization of OTR-expressing astrocytes in the central amygdala
(A) Schematic depiction of the location of OTR-positive astrocytes in the central lateral amygdala.
(B) For the visualization of OTR-positive astrocytes after RNAScope and IHC, 25 μm thick z-stacks should be obtained at the confocal microscope.
(C) It is important to adjust the laser intensity prior to imaging using the negative control probe. All channels should be adjusted so that virtually no signal is visible for the negative RNA control probe to minimize the chances of false positive detection.
(D) Confocal image shows OTR-positive astrocytes labeled via GFAP, GS, DAPI and OTR mRNA probes. No drastic differences were observed when incubating 48 instead of 24 h with the primary antibodies. This figure was created with BioRender.com.