Figure 4. Biased cortical synaptic input to the direct and indirect pathways.

(A–B) Cortical inputs from somatosensory cortex onto both direct and indirect pathway MSNs arise almost exclusively from superficial layer 5, though overall input density appeared to be higher onto direct pathway MSNs. Scale bar = 100μm and applies to panels A–F.

(C–D) Motor cortex inputs onto both pathways arise primarily from superficial layer 5, but some superficial layer cells also contribute input. Motor cortex preferentially innervates the indirect pathway.

(E–F) Inputs from the insular and orbital cortices (labeled PFC) arise from both superficial and deep layers, and appear to innervate both the direct and indirect pathway similarly.

(G) Cortical inputs were segregated into four major input streams. Direct pathway inputs are colored red, whereas indirect pathway inputs are shaded in blue. Individual p values indicate two-tailed t-test comparison of direct vs. indirect pathway input for each stream. Error bars indicate 1 SEM.

(H) Center of gravity analysis demonstrates that cortical input to the direct pathway is significantly caudal to that for the indirect pathway. Individual anterior-posterior centers of gravity are indicated by colored circles at the top of the graph. The two-dimensional chart shows center of gravity for cortical inputs to the direct and indirect pathways in both the anterior-posterior and dorsal-ventral dimensions. Bar width indicates ±1 SEM. The plane of slice is 2.04 mm lateral to the midline. The border between primary somatosensory and primary motor cortex at this plane of slice is indicated by the dashed line for reference. One mouse with significantly different center of gravity is excluded in the D1R-Cre cohort, and is indicated by the light circle.

(I) Summary diagram showing the relative contribution of different cortical layers to striatal input for four different cortical areas in the plane of slice containing the cortical center of gravity (ordered from rostral to caudal). Within a given cortical region, each cortical layer provided similar proportions of synaptic input to the direct and indirect pathways, even though total proportion of inputs could be dramatically different (see figure 3). More rostral brain structures provided more superficial input than more caudal cortical areas. ND: not determined, representing the portion of labeled neurons whose layer identity could not be accurately determined, largely due to plane of slice providing ambiguous layer information.

(J) Summary image diagrams both the relative strength of layer input from major cortical input structures (shaded in tan) as well as the relative amount of input streaming from each cortical area onto direct (red) and indirect (blue) motor pathway MSNs, as well as the downstream targets of these MSNs.

See also Figures S2 and S3.