Figure 1.

Ndnf Is a Selective Marker of Neocortical Layer 1 Interneurons

(A) Circuit diagram of neocortex. The connections of L1 NDNF-INs are not known and are the subject of this study.

(B) RiboTag-seq indicates that Ndnf-expression is highly enriched in GABAergic neurons, but not in IN subtypes that express Pv, Sst, or Vip.

(C and D) Ndnf-expressing GABAergic neurons are concentrated in L1. RNAscope FISH for Ndnf, Gad1, and IN subtype markers was done in the adult auditory cortex.

(C) Ndnf-expressing cells are concentrated in L1 (scale bar, 200 μm).

(D) The distribution of Ndnf-expressing INs in the adult auditory cortex differs from the distribution of other INs. The distance from the pia was determined for each cell expressing a given marker and plotted as a histogram (for Ndnf) or the corresponding probability density function (PDF; for Ndnf, Pv, Sst, and Vip; dashed line indicates the L1 border).

(E–H) Ndnf-expressing neurons constitute the majority of L1 GABAergic neurons and do not overlap with Pv, Sst, or Vip.

(E) Representative image of FISH for Ndnf and Gad1 in the auditory cortex (co-expressing neurons are indicated by arrowheads, and DAPI-labeled nuclei are in blue; scale bar represents 100 μm).

(F) Percentage of L1 Ndnf neurons that co-express the respective marker.

(G) Percentage of L1 neurons expressing the respective subtype marker that co-expresses Ndnf.

(H) Percentage of Gad1-positive L1 neurons expressing each subtype marker.

(I) A newly generated mouse line allows for temporally controlled selective labeling of L1 NDNF-neurons. Auditory cortex section of an Ndnf-Ires-CreERT2 mouse injected with an AAV-construct that drives Cre-dependent tdTomato expression (AAV-hSyn-Flex-tdTomato) upon tamoxifen application (scale bar, 200 μm).

Data are presented as mean ± SEM.