Figure 4. Telencephalic outputs are widely copied onto the hippocampus.

(A) Experimental strategy. A MAPseq virus library was injected into deep medial entorhinal cortex (MEC; n = 3 mice). Forty-four hours later, mice were sacrificed and brains were serially sectioned in the sagittal plane at 400 μm thickness. Tissue from five major divisions (isocortex, CNU, Olf/Ctxsp, dHip, and vHip), as well as dorsal and ventral MEC were then dissected and collected in tubes. Brainstem and spinal cord tissue was collected as a negative control. Tissue was further processed for RNA extraction and next generation sequencing. (B) Nearly all barcodes detected in CNU, Olf/Ctxsp, and isocortex were also detected in the hippocampus (isocortex: 98.6% ± 0.06%; CNU: 99.2% ± 0.08%; Olf/Ctxsp: 99.0% ± 0.4%; n = 3 mice, 1603, 3493, and 1677 barcodes from brains 1, 2, and 3, respectively). (C) Dorso-ventral topography of projections revealed by relative barcode counts. For each barcode, counts were normalized to show the relative projection strength of each neuron to dorsal and ventral hippocampus (n = 3 mice).

Figure 4—figure supplement 1. Verification of viral expression and further MAPseq data visualization.

(A) Brightfield and epifluorescence microscope images of the injection site on a sagittal brain section showing the spread of neurons infected by the MAPseq virus library in wild-type mice. (B) A neuron’s position in the dorso-ventral axis does not affect the likelihood of collateralization of its axons to the hippocampus (paired two-tailed Wilcoxon signed rank test, p = 1, median proportion of collateralizing barcodes in dEC = 99.3%, vEC = 99.8%, n = 3 mice).