Specificity in the Projections of Prefrontal and Insular Cortex to Ventral Striatopallidum and the Extended Amygdala

A1–C3, Photomicrographs showing single- and double-labeled cells in orbital cortex of a CPu–AcbC case visualized by immunofluorescence with single-band filters (A1, A2), immunofluorescence with a dual-band filter (B), and confocal microscopy (C1–C3). Double-labeled cells (asterisks) are yellow. A double-labeled neuron in which the two fluorophores were unevenly exhibited appear slightly yellow (caret in C3). Note that B is not a merged image but rather shows, with the dual-band filter, the same field illustrated in A1 and A2 with single band filters. Alternatively, C3 is the merged image of C1 and C2.

Plots of retrograde labeling in the prefrontal cortex produced in representative cases for each of the following injection pairs: A, CPu–AcbC; B, CPu–AcbS; C, BST–AcbS; D, CeA–BST. DBL, Double-labeled neurons; AC, anterior cingulate; PrC, precentral cortex; PL, prelimbic; IL, infralimbic; DP, dorsal peduncular; AI, agranular insular; VO, ventral orbital; LO, lateral orbital; DLO, dorsolateral orbital.

Plots of retrograde labeling in the insular cortex at anterior, middle, and posterior levels in representative cases for the following injection pairs: A, CPu–AcbC; B, CPu–AcbS; C, BST–AcbS; D, CeA-BST. DBL, Double-labeled neurons; GI, granular insular; DI, dysgranular insular; AI, agranular insular.

Mean (±SEM) retrograde single labeling in prefrontal and insular cortices produced by homotypic and heterotypic pairs of tracer injections in ventral striatal and extended amygdala sites. Within the medial prefrontal cortex, ventral striatal and extended amygdala displayed distinguishing characteristics in the amount of retrograde labeling (A) (F_(4,35) = 5.53; p < 0.005), dorsoventral distribution of labeling (B)(F_(12,143) = 12.10; p < 0.001), and anteroposterior labeling distribution within prelimbic (C)(F_(4,71) = 6.37; p < 0.001) and infralimbic (D)(F_(4,71) = 7.79; p < 0.001) cortices. E, AcbC-induced labeling dominated the lateral prefrontal cortex compared with all other sites examined (F_(4,35) = 5.92; p = 0.001). F, CPu and AcbC injections produced the most labeling in orbital cortex (F_(4,35) = 10.16; p < 0.001), which followed a medial-to-lateral gradient (G) (F_(2,35) = 43.67; p < 0.001). Retrograde labeling in the insular cortex was strongest after AcbC and CeA injections (H) (F_(4,35) = 6.42; p < 0.001), localized mainly within the agranular insular cortex (I) (F_(2,107) = 38.87; p < 0.001), and displayed different anteroposterior labeling distributions for striatal versus extended amygdala injection sites (J)(F_(8,107) = 4.05; p < 0.001). AC, Anterior cingulate; PrC, precentral cortex; PL, prelimbic; IL, infralimbic; DP, dorsal peduncular; GI, granular insular; DI, dysgranular insular; AI, agranular insular; VO, ventral orbital; LO, lateral orbital; DLO, dorsolateral orbital. ^#p < 0.05 compared with CeA; *p < 0.05 compared with CPu; **p < 0.05 compared with the highest bar for each site; ^†p < 0.05 compared with posterior; ^‡p < 0.05 compared with AcbC; *** p < 0.05 compared with VO.

Mean (±SEM) double labeling produced by injection pairs throughout prefrontal and insular cortices. CPu–AcbC pairs produced significantly more double labels than any other injection pair in the medial prefrontal cortex (A)(F_(5,17) = 11.60; p < 0.001), lateral prefrontal cortex (B) (F_(5,17) = 2.61; p = 0.08), and orbital cortex (C) (F_(5,17) = 9.31; p < 0.001), where CPu–AcbC double labels were found along a strong medial-to-lateral gradient (D) (F_(2,8) = 39.97). In the agranular insular cortex, both CPu-AcbC and CeA-BST pairs produced strong double labeling (E) (F_(5,17) = 7.26; p < 0.005) that differed strikingly in distribution along the anterior–posterior axis (F)(F_(2,17) = 10.70; p < 0.005). PrC, Precentral cortex; AC, anterior cingulate; PL, prelimbic; IL, infralimbic; DP, dorsal peduncular; AI, agranular insular; VO, ventral orbital; LO, lateral orbital; DLO, dorsolateral orbital. *p < 0.05 compared with CPu-AcbC group, **p < 0.05 compared with VO.

Photomicrographs of retrograde labeling produced in the insular cortex by a CeA–AcbC injection pair. A, Distribution of single labeling in the rostral insular cortex produced by CeA (black) and AcbC (brown) injections. B, Enlargement of A. C, Posterior insular level. D, Enlargement of C. Note that the distributions of retrogradely labeled neurons produced by the two injections interdigitate and that double-labeled neurons are negligible.