The synaptic connections between cortical areas V1 and V2 in macaque monkey

Abstract

The primary visual cortex (V1) and V2 together form approximately 24% of the total neocortex of the macaque monkey and have each other as their major partners. The major target of the V1 projection to V2 is layer 4, where it forms clusters of boutons, which form asymmetric (excitatory) synapses mainly with dendritic spines (75%). The remainder form synapses with dendritic shafts. The synapses found on spines were often more complex, perforated postsynaptic densities than those found on dendritic shafts. The reciprocal projection from V2 to V1 targeted layers 1, 2/3, and 5 and was formed of axons of different morphologies. One axon type, originating from superficial layer pyramidal cells, had a morphology resembling those of local pyramidal cell collaterals. These axons arborized in layers 1, 2/3, and 5 of V1. Another type of axon, arborizing in layer 1, was slender (0.3 microm), unbranched, unmyelinated, and uniformly covered with boutons terminaux and formed asymmetric synapses mainly with slender spines. Yet a third type of axon also confined to layer 1, was thick (>1 microm), branched, heavily myelinated, and formed separate small clusters of large ( approximately 1 microm) en passant multisynaptic boutons that formed asymmetric synapses mainly with large flat spines. These data show the existence of a reciprocal excitatory loop between V1 and V2 that is formed by different axonal types, each with preferred layers of termination.

Figure 1.

Location of injection site in V1 and labeled boutons in V2. A, Low-power sketch of a parasagittal section through the lunate gyrus showing the approximate location of a BDA injection site (solid circle) in V1, the location of BDA-labeled boutons (asterisk), and the V1–V2 border (filled arrowhead). B, Light photomontage of cortical area V2 showing BDA-labeled axons and their terminals in layer 4 that project from an injection of BDA made in area V1. Laminae and their boundaries are indicated to the bottom left and top right. Scale bar: B (bottom right), 15 μm.

Figure 2.

Location of injection site in V2. A, Light photomontage of a parasagittal section through the cortex at the border between V1 and V2 (arrow) showing the injection site in V2 (white arrowhead) and the location of a weak retrogradely labeled pyramidal cell (arrowhead). V1 laminae are indicated to the right. B, Light microscopic drawing of the section shown in A. Anterogradely labeled axons projecting from the injection site (open arrowhead) cross the border into V2. Axons in the immediate vicinity of the injection site were not drawn because the density of label was too intense. The position of the weakly labeled pyramidal cell shown in A is indicated (arrowhead). Laminae and their borders are indicated to the left and right. Scale bars, 300 μm.

Figure 3.

Two “feedback” axons labeled with PHA-L, confined to layer 1 and showing terminations in area V1. A, B, Low-power sketches of lunate gyrus showing approximate path of two axons through layer 1 and earliest appearance of axon (asterisk). The white matter (wm) and V1–V2 border are also indicated (arrow). Axon shown in A and C–E is referred to as the “thin” axon. C, Light microscopic reconstruction of the thin axon showing a long, unbranched, bouton encrusted (small filled arrowheads) collateral. D, High-magnification (100× oil objective) light microscopic drawing showing detail of thin axon and frequently located boutons (filled arrowheads). E, Light microscopic photomontage of thin axon in layer 1. Axon shown in F–H is referred to as the “thick” axon. F, Light microscopic reconstruction of thick, branched axon showing clusters of boutons (solid arrowheads). The computer assisted 3-D reconstruction of the axon is rotated to provide the view from the surface of the brain. G, High-magnification light microscopic drawing showing detail from one bouton cluster. H, Light microscopic photomontage of thick axon and bouton cluster. Scale bars: C, F, 0.5 mm; D, G, 25 μm; E, H, 10 μm.

Figure 4.

Electron photomicrographs taken from cortical area V2 showing BDA-labeled boutons in layer 4. A, A labeled bouton en passant forms an asymmetric synapse (solid arrowhead) with a spine (sp). B, A spine (sp) forms an asymmetric synapse (solid arrowhead) with a labeled bouton and a symmetric synapse (open arrowhead) with an unidentified bouton. The asymmetric synapse of the labeled bouton has been cut rather obliquely obscuring the synaptic cleft and smearing the postsynaptic density. C, A labeled bouton forms two asymmetric synapses (solid arrowheads) with a spine (sp) and a small caliber dendritic shaft (d). D, A large-caliber dendritic shaft (d) forms an asymmetric synapse (solid arrowhead) with a labeled bouton. The dendrite (d) forms a further two asymmetric synapses (small solid arrowheads) with two unidentified boutons. Serial section reconstruction of the dendrites shown in C and D revealed that both formed more synapses and contain numerous mitochondria. These features are characteristic of GABA-containing smooth cells. E, A lower-power electron micrograph showing a labeled bouton forming an asymmetric synapse (solid arrowhead) with a medium-caliber dendritic shaft (d). The dendrite produces a spine (sp), opposite the labeled bouton, that forms an asymmetric synapse (small solid arrowhead). An unidentified bouton forms a symmetric synapse (open arrowhead) with the dendritic shaft. Dendrites showing few shaft synapses, few mitochondria, and forming spines are features of spiny cells, probably pyramidal cells. Scale bars: A–E, 0.5 μm.

Figure 5.

Electron photomicrographs taken from cortical area V1 showing PHA-L-labeled boutons in layer 1. All examples are taken from the “thin” axon (Fig 1; A–C). A–C, Examples of small boutons forming asymmetric synapses (solid arrowheads) with spines (sp). The majority of labeled boutons and their targets were of similar dimensions. D, A labeled bouton forms a synapse (solid arrowhead) with a small caliber dendrite (d). The dendrite formed a second synapse with another labeled bouton from the same axon. The ultrastructural characteristics of the dendrite (variable diameter, numerous mitochondria, and forming many synapses) indicate that it is from a GABAergic neuron with smooth dendrites. Scale bars, 0.5 μm.

Figure 6.

Electron micrographs of PHA-L-labeled boutons forming synapses in layer 1 of area V1. All examples are taken from the “thick” axon (Fig. 1; D–F). A, A large labeled bouton forms an asymmetric synapse (solid arrowhead) with a small spine (sp). The spine and parent dendrite (d) are weakly labeled with retrogradely transported PHA-L. B, Two large spines (sp) each form a perforated asymmetric synapse (solid arrowheads) with a large labeled bouton. C, A medium sized bouton forms a perforated asymmetric synapse (solid arrowheads) with a spine (sp) that can be followed back to the small caliber parent dendrite (d). D, A small bouton terminaux forms an asymmetric synapse (solid arrowhead) with a medium-caliber dendrite (d). The dendrite contained few mitochondria, formed no other synapses and showed little variation in diameter. These features are characteristics of neurons with spiny dendrites, probably pyramidal cells. E, A large bouton en passant forms an asymmetric synapse (solid arrowhead) with a large-caliber dendrite (d). The dendrite was unusual in that it formed synapses with the shaft and with small and slender spinous processes that each formed at least one asymmetric synapse (small solid arrowheads). The shaft also contained numerous mitochondria. Scale bars, 0.5 μm.

Figure 7.

Three-dimensional reconstruction from serial ultrathin sections of PHA-L-labeled boutons in layer 1 of V1 showing synaptic targets. A, Thin axon (blue) showing numerous projecting boutons terminaux each forming a single synapse (yellow) in most cases with a small spine (transparent brown). Near the top of the reconstruction a bouton terminaux branches to form two boutons, one of which forms a synapse with a dendritic shaft (transparent mauve). A spine projects from the uppermost surface of the dendrite and forms another synapse with the labeled axon. This same spine also forms a symmetric synapse (red) with an unidentified bouton. B, Thick axon (blue) showing a string of three boutons en passant and their synaptic targets; spines (transparent brown) and dendritic shafts (transparent mauve). Postsynaptic densities, often complex, are shown in yellow. The two uppermost boutons each form two synapses, one each with a spine and the second with a dendritic shaft that passes between the two boutons. The dendrite also receives an asymmetric synapse from an unidentified bouton. The lower bouton forms four synapses; two with spines and two with a dendritic shaft. One of the spines can be traced back to the parent dendrite showing more spines and forms an asymmetric synapse (red) on the shaft. Scale bars, 2 μm.

Figure 8.

Reconstructed spines found postsynaptic to labeled boutons of thin axon (A) and thick axon (B). The spines have been ordered roughly by size. Each spine has been rotated to present its broadest face. Scale bars, 0.5 μm.

Figure 9.

Two-dimensional projection of the reconstructed postsynaptic densities found on spines, soma, and dendrites postsynaptic to V1-labeled boutons in layer 4 of area V2. The densities are ordered by increasing surface area. Scale bar, 1 μm.

Figure 10.

Two-dimensional projection of the reconstructed postsynaptic densities found on spines and dendrites postsynaptic to V2-labeled boutons in layers 1 and 2/3 of area V1. The densities are ordered by increasing surface area. Scale bar, 1 μm.

Figure 11.

Two-dimensional projection of the reconstructed postsynaptic densities found on the spines and dendrites postsynaptic to labeled boutons in layer 1 of area V1 from thin axon (A) and thick axon (B). The densities are ordered by increasing surface area. Scale bar, 1 μm.

Figure 12.

Histogram showing the distributions of postsynaptic areas (in square micrometers) formed with spines and dendrites by labeled V1 boutons in layer 4 of V2.

Figure 13.

Histograms of the distributions of postsynaptic density areas (in square micrometers) formed by labeled V2 boutons in superficial layers of V1. Shown are thin axon (black, n = 44), thick axon (white, n = 61), and layer 3 pyramidal cells (gray, n = 27).

Figure 14.

A, Histogram of the synaptic targets of labeled V1 boutons in area V2. Spine n = 91, dendrite n = 34, soma n = 1. B, Histogram of the synaptic targets of the boutons of two labeled V2 axons in layer 1 of area V1 and a group of labeled synapses in layers 1 and 2/3 of V1 from superficial layer 3 pyramidal cells in V2. For the thin axon, n = 44, and for the thick axon, n = 61. For the group of V2-labeled layer 3 cell boutons in V1, n = 60.

Figure 15.

A, Histogram of the number of synapses formed per labeled V1 bouton in layer 4 of area V2. B, Histogram of the number of synapses formed per labeled V2 bouton in superficial layers of area V1.