Article Figures & Data
Figures
- Figure 1.
A–K, Injection sites in the amygdala (A–D) and mediodorsal thalamus (E–K). Scale bars, 1 mm. A, Schematic shows relative position of small injection sites in the amygdala in cases BL-R, BM-R, and BN-R. B–D, Fresh tissue sections show tracer in the amygdala (arrows). E, H, J, Schematics depict location of tracer injections in thalamic mediodorsal nucleus (cases BB-L, BO-R, and BN-R). F, I, K, Fluorescent micrographs show tracer in the thalamus. G, Bright-field photomicrograph shows section with injection in MD magnocellular. BL, Basolateral nucleus; BM, basomedial nucleus; CBL, cascade blue; Ce, central nucleus; Cim, central intermediate nucleus; FE, fluoroemerald; FR, fluororuby; L, lateral nucleus; LY, Lucifer yellow; mc, magnocellular sector; mf, multiform sector; pc, parvicellular sector; Sm, stria medularis; VCo, ventral cortical nucleus.
- Figure 2.
Amygdalar boutons were densest and largest in pOFC. A–C, Areas of study: lateral (A), orbital (B), and medial (C) views of a rhesus monkey brain show areas of interest in areas 9/46, OPro (pOFC), and A32 (ACC). Scale bar, 1 cm. D–F, Nissl-stained columns of cortex in areas 9/46 (D, A9/46); A32 (E); and pOFC (F, area OPro) labeled for amygdalar fibers (G–I, insets). Scale bar: (in D) D–F, 100 μm. G–I, Amygdalar fibers (arrow) at layer 1–2 border in A9/46 (G), A32 (H), and pOFC (I). Scale bar: (in G) G–I, 10 μm. J, Relative density of amygdalar boutons in A9/46, A32, and pOFC, normalized to the highest density in each case (pOFC). K, Relative density of large amygdalar boutons ≥1 μm in diameter in A9/46, A32, and pOFC, normalized to the highest density in each case (pOFC). Vertical lines indicate SE. *p ≤ 0.05.
- Figure 3.
Amygdalar axon boutons in pOFC were larger than in A32 or the thalamic. A, Histogram shows size distribution of amygdalar boutons in A32 (red) and pOFC (blue) across all layers. B, Histogram shows size distribution of amygdalar boutons (blue) and thalamic boutons (yellow) in pOFC across all layers. Vertical lines indicate SE. *p ≤ 0.05.
- Figure 4.
Amygdalar terminals in the upper layers of pOFC were larger than thalamic terminals in the middle layers at the synaptic level. Scale bar: (in A) A–G, 1 μm. A–E, Amygdalar boutons labeled with DAB in the upper layers of pOFC, forming (A) a macular synapse (silhouette arrowheads) with a PV-positive soma labeled with TMB (asterisk); (B) a macular synapse (black arrowheads) with a spine (sp) containing a spine apparatus (arrow); (C) A macular synapse (black arrowheads) with a spine (sp), which also receives an unlabeled symmetric synapse (white arrowhead) from a presumed inhibitory bouton (At); (D) a perforated synapse (black arrowheads) with a spine (sp), which also receives an unlabeled symmetric synapse (white arrowhead) from a presumed inhibitory bouton (At); (E) Two synapses (black arrowheads) with a spine (sp) and a CB-positive dendrite (CB den) labeled with gold (black clumps). F, G, Thalamic boutons labeled with DAB forming (F) a macular synapse (black arrowheads) with a spine (sp) and (G) a macular synapse (black arrowheads) with a CR-positive dendrite (CR den) labeled with TMB. H, Diameters of amygdalar boutons in the upper layers of pOFC (blue) and thalamic boutons in the middle layers of pOFC (yellow), measured in the electron microscope. I, Amygdalar axon boutons in the upper layers of pOFC were larger in volume than thalamic terminals in the middle layers of pOFC. Vertical lines indicate SE. *p ≤ 0.05. amy At, Amygdalar axon terminal; At, axon terminal; den, dendrite; thal At, thalamic axon terminal.
- Figure 5.
Examples of amygdalar and thalamic axons targeting inhibitory neurons in pOFC. A, Distribution of presumed inhibitory neurons shown in pseudocolor in pOFC, where CR neurons (green) and CB neurons (red) were found mostly in the uppers layers and PV neurons (blue) were found mostly in the middle–deep layers. Scale bar, 100 μm. B, Amygdalar axons (red) in the upper layers of pOFC; one is closely apposed to a dendrite of a CB neuron (green, white arrows). Scale bar: (in B, left) B–E, left, 10 μm. Right, Sequential optical sections through apposition and 3D rotation through apposition. Scale bar: (B, inset) B–E, insets, 1 μm. C, Amygdalar axons (red) in the upper layers of pOFC; one close apposition with a dendrite of a CR neuron (green, white arrows). D, Thalamic axon (green) in the middle layers of pOFC; example of a close apposition with a PV neuron (red, white arrows). E, Thalamic axon (green) in the middle layers of pOFC formed a close apposition with a CR neuron (red, white arrows). F, Amygdalar terminals targeted more CB and CR inhibitory neurons in the upper layers of pOFC (open bars), and more PV neurons in the middle layers (filled bars). G, Thalamic terminals (open bars) targeted more PV and CR neurons in the middle layers compared with amygdalar terminals (filled bars). Vertical lines indicate SD. *p ≤ 0.05.
- Figure 6.
Excitatory and inhibitory postsynaptic targets of amygdalar boutons in pOFC. A–C, Amygdalar boutons (blue) form synapses (yellow) with (A) a smooth dendrite of a presumed inhibitory neuron (red), (B) a smooth dendrite (red) labeled for CB and a spiny dendrite of a presumed excitatory neuron (green), and (C) a spiny dendrite of a presumed excitatory neuron (green). Scale: cubes are 1 μm3. D, Left, Amygdalar boutons most frequently innervated single spines in the upper layers of pOFC, and a significant proportion were multisynaptic. Right, Thalamic boutons also formed synapses mostly with spines, but thalamic pathways contained fewer multisynaptic boutons in the middle layers of pOFC. E, Amygdalar boutons with ≥1 inhibitory target were larger than amygdalar boutons with only excitatory targets. F, Among labeled dendrites, amygdalar boutons mostly formed synapses with CB-positive dendrites of presumed inhibitory neurons in the upper layers of pOFC. G, Amygdalar (blue diamonds, solid line) and thalamic (yellow squares, dashed line) bouton volume was positively correlated with PSD area. H, Amygdalar (blue diamonds, solid line) and thalamic (yellow squares, dashed line) bouton volume was positively correlated with spine volume. *p ≤ 0.05.
- Figure 7.
Summary of features in amygdalar pathway to pOFC, ACC, and areas 9/46. A, Thickness of arrows depicts the relative density and terminal size of amygdalar pathways to pOFC, ACC, and areas 9/46. B, Amygdalar terminals were larger and formed synapses more often with excitatory neurons in the upper layers of pOFC compared with thalamic axons in the middle layers. Among inhibitory neurons, amygdalar axons targeted CB and CR neurons in the upper layers, while thalamic axons targeted more PV and CR neurons in the middle layers. Large amygdalar synapses may drive excitatory neurons in pOFC, while synapses on CB and CR neurons may reduce background excitation, or noise. Excitatory spiny stellate neurons (SS) are a major target of thalamic pathways. BL, Basolateral; BM, basomedial; Co, cortical; mc, magnocellular sector; P, pyramidal neuron; pc, parvicellular sector.
Tables
Case; hemisphere Injection site; division Tracer Sex Age (years) BB; left (BB-L) Amygdala; basomedial, basolateral, anterior BDA Female 2 BB; right (BB-R) Amygdala; basolateral, anterior, basomedial, cortical BDA Female 2 BD; left (BD-L) Amygdala; basolateral, central, lateral, basomedial BDA Male 2 BD; right (BD-R) Amygdala; cortical, medial, basomedial, basolateral BDA Male 2 BL; right (BL-R) Amygdala; cortical, basomedial FR Male 3 BM; right (BM-R) Amygdala; basomedial, cortical FR Female 3.5 BN; right (BN-R) Amygdala; basomedial, basolateral FE Male 2 BB; left (BB-L) Thalamus; mediodorsal magnocellular, mediodorsal parvicellular FE Female 2 BO; right (BO-R) Thalamus; mediodorsal magnocellular, paraventricular, central intermediate LY Male 3 BN; right (BN-R) Thalamus; mediodorsal parvicellular, habenula CBL Male 2
