Principal component and cluster analysis of layer V pyramidal cells in visual and non-visual cortical areas projecting to the primary visual cortex of the mouse.

The long-distance corticocortical connections between visual and nonvisual sensory areas that arise from pyramidal neurons located within layer V can be considered as a subpopulation of feedback connections. The purpose of the present study is to determine if layer V pyramidal neurons from visual and nonvisual sensory cortical areas that project onto the visual cortex (V1) constitute a homogeneous population of cells. Additionally, we ask whether dendritic arborization relates to the target, the sensory modality, the hierarchical level, or laterality of the source cortical area. Complete 3D reconstructions of dendritic arbors of retrogradely labeled layer V pyramidal neurons were performed for neurons of the primary auditory (A1) and somatosensory (S1) cortices and from the lateral (V2L) and medial (V2M) parts of the secondary visual cortices of both hemispheres. The morphological parameters extracted from these reconstructions were subjected to principal component analysis (PCA) and cluster analysis. The PCA showed that neurons are distributed within a continuous range of morphologies and do not form discrete groups. Nevertheless, the cluster analysis defines neuronal groups that share similar features. Each cortical area includes neurons belonging to several clusters. We suggest that layer V feedback connections within a single cortical area comprise several cell types.

Indirect pathway between the primary auditory and visual cortices through layer V pyramidal neurons in V2L in mouse and the effects of bilateral enucleation.

Visual cortical areas are activated by auditory stimuli in blind mice. Direct heteromodal cortical connections have been shown between the primary auditory cortex (A1) and primary visual cortex (V1), and between A1 and secondary visual cortex (V2). Auditory afferents to V2 terminate in close proximity to neurons that project to V1, and potentially constitute an effective indirect pathway between A1 and V1. In this study, we injected a retrograde adenoviral vector that expresses enhanced green fluorescent protein under a synapsin promotor in V1 and biotinylated dextran amine as an anterograde tracer in A1 to determine: (i) whether A1 axon terminals establish synaptic contacts onto the lateral part of V2 (V2L) neurons that project to V1; and (ii) if this indirect cortical pathway is altered by a neonatal enucleation in mice. Complete dendritic arbors of layer V pyramidal neurons were reconstructed in 3D, and putative contacts between pre-synaptic auditory inputs and postsynaptic visual neurons were analysed using a laser-scanning confocal microscope. Putative synaptic contacts were classified as high-confidence and low-confidence contacts, and charted onto dendritic trees. As all reconstructed layer V pyramidal neurons received auditory inputs by these criteria, we conclude that V2L acts as an important relay between A1 and V1. Auditory inputs are preferentially located onto lower branch order dendrites in enucleated mice. Also, V2L neurons are subject to morphological reorganizations in both apical and basal dendrites after the loss of vision. The A1-V2L-V1 pathway could be involved in multisensory processing and contribute to the auditory activation of the occipital cortex in the blind rodent.

Widespread periodic intrinsic connections in the tree shrew visual cortex.

Intrinsic connections within the tree shrew (Tupaia glis) visual cortex (area 17) are organized in periodic stripelike patterns within layers I, II, and III. This anatomical network resembles the regularly organized stripes of 2-deoxyglucose accumulation seen after stimulation of alert animals with uniformly oriented lines. Such connections imply that widespread lateral interactions are superimposed on the retinotopic organization of area 17 and suggest alternative interpretations of cortical columns.

Complex microstructures of sensory cortical connections.

The neocortex has a distinctive laminar and modular organization. Although important questions remain regarding structure and function at this level of organization, recent studies are addressing a finer scale of synaptic and network microstructure. New findings concerning network properties are rapidly emerging from approaches in which dual or triple intracellular recordings in vitro are combined with analyses of cell and synaptic morphology, as well as from experiments designed to label multiple cell populations.

Zinc-rich transient vertical modules in the rat retrosplenial cortex during postnatal development.

The rat retrosplenial cortex is part of a heavily interconnected limbic circuit, considered to have an important role in spatial memory. Interestingly, the granular retrosplenial cortex has an exceptionally distinct system of dendritic bundles, originating from callosally projecting pyramidal neurons in layer II. These can be detected as early as postnatal day 5; and, although their functional significance remains to be elucidated, the existence of these bundles makes the granular retrosplenial cortex an attractive model system for a wide range of development and functional investigations. Here, we report four results concerning the development of modularity in the granular retrosplenial cortex in rats as investigated by neurochemical markers associated to cortico-cortical and thalamo-cortical connections. Emphasis is placed on zinc, an activity-related substance associated with glutamatergic, non-thalamic terminations. 1) Zinc shows a transient strong expression during early postnatal development, but later than the appearance of the upper layer bundles (at postnatal day 5). By postnatal day 11 to postnatal day 15 staining for zinc achieved its most complex pattern; such that layer I had an elaborate organization both in the tangential and radial dimensions. Three sublaminae were distinguished (layers Ia-c): a superficial, thin tier (Ia) with patchy, moderate staining which periodically intruded into the underlying layer Ib ("funnel" modules), a middle band of variable width and light staining (Ib), and a deep, thin band with heavy and patchy staining (Ic) which, at rostral levels, spread upward into layer Ib (as "dome-like" modules). 2) At postnatal day 15, immunohistochemical methods showed that layers Ia, b zinc-funnels were co-localized with glutamate receptor subunits 2/3, GABA receptor type A alpha1 subunit and the thalamo-cortical marker, vesicular glutamate transporter 2. Layer Ic and the zinc dome-like modules were co-labeled for the cortico-cortical marker, vesicular glutamate transporter 1 and calretinin. 3) The spatial coincidence between zinc funnels in layers Ia, b and vesicular glutamate transporter 2 was further investigated by electron microscopy, which demonstrated co-localization of zinc and vesicular glutamate transporter 2 in synaptic boutons. The unusual co-localization of zinc and thalamo-cortical terminations was confirmed by retrograde transport of zinc to neurones in the anterodorsal thalamic nucleus at postnatal day 9 and postnatal day 13, and can thus be considered a transient zinc expression in thalamo-cortical boutons. This was not observed at postnatal day 28 or later. 4) After postnatal day 18, zinc staining started to fade in all layers. Before postnatal day 21, the heavy staining for zinc in the domes had completely disappeared. Zinc staining in layer Ia and the funnels virtually disappeared after postnatal day 28. A transient expression of zinc is reported in at least one other cortical area (layer IV of barrel cortex from postnatal day 5 to postnatal day 14, maximal at postnatal days 9-11). We conclude that the transient expression of zinc can occur in both limbic and sensory areas, and that down-regulation of zinc in cortical modules might be related to synaptic plasticity and remodeling during development.

Strong expression of NETRIN-G2 in the monkey claustrum.

The claustrum is a phylogenetically conserved structure, with extensive reciprocal connections with cortical regions, and has thus been considered important for sensory, motor, emotional, and mnemonic coordination or integration. Here, we show by in situ hybridization that the adult monkey claustrum is strongly positive for NETRIN-G2, a gene encoding a glycosyl phosphatidyl-inositol-linked membrane protein, which constitutes a subfamily with NETRIN-G1 within the netrin/UNC6 family. There is a conspicuous dorsal/ventral differentiation, where the label is stronger in the ventral claustrum. NETRIN-G2 positive neurons are not GABAergic, but rather correspond to claustrocortical projection neurons, as demonstrated by retrograde transport of Fast Blue from cortical injections and by double in situ hybridization for NETRIN-G2 and GAD67. Since NETRIN-G2 is known to be preferentially expressed in cortex, in contrast with the thalamically expressed NETRIN-G1, these results raise the possibility of some functional similarity in regulation of excitatory neural transmission in the claustrum and cortex.

Anatomical organization of primary visual cortex (area 17) in the ferret.

The present report describes the intrinsic and extrinsic cortical connectivity of striate cortex (area 17) in the ferret. Injections of horseradish peroxidase demonstrate periodic intrinsic connections over an extent of 2.5-3.0 mm, mainly in the supragranular layers but also occurring secondarily in layer 5. These connections have a stripelike configuration, with a center-to-center spacing of 0.5-0.7 mm. Their laminar distribution and stripelike configuration resemble the pattern in the cat (Gilbert and Wiesel, '83), another member of the carnivore family, but not that in monkeys. In both macaque and squirrel monkeys, these connections have a bilaminar distribution in layers 2-3 and 4B, and a more complicated latticelike geometry (Rockland and Lund, '83). Their interperiod spacing, of about 0.5 mm, however, is relatively constant across species. Extrinsic connections in the ferret link striate cortex with territories probably homologous to feline areas 18 and 19, and to the suprasylvian region. Callosal connections extend on the lateral surface about 1.5 mm into area 17 and 4.0 mm into area 18 beyond their common border. There are homotopical connections between striate cortices and heterotopical connections from at least areas 18 and 19 to contralateral area 17. In addition to gray matter connections, intracortical injections also result in labeled interstitial neurons in the subgriseal white matter. These occur both subjacent to an injection site in area 17, and below labeled foci in area 18 projecting back to area 17, as if interstitial neurons shared the connectivity of overlying layer 6.

Morphology of individual axons projecting from area V2 to MT in the macaque.

Efferent axons from area V2 to the middle temporal area (MT) were anterogradely labeled by Phaseolus vulgaris-leucoagglutinin (PHA-L) or biocytin and analyzed in serial reconstructions. Five of seven reconstructed axons had three arbors (each < or = 200 microns in diameter) in layers 3-4, separated by 200-600 microns. Two axons terminated in what was apparently a single focus in layers 3-4. Of 15 additional single arbors analyzed, 12 were concentrated in layers 3-4, and measured 200-250 microns across at their widest point. Three of these arbors were more columnar in shape (about 400 microns in diameter), and extended from layer 4 toward layer 1. This system differs in several features from MT-projecting axons originating from V1. Namely, V2 axons terminating in MT are thinner (approximately 1.0 microns vs. 3.0 microns), their terminal specializations are more delicate, and their arbors are concentrated in layer 4 and overlying layer 3, with no collaterals to layer 6. These differences may reflect the distinctive neuronal populations giving rise to these two connectional systems (different sizes of pyramidal neurons in layer 3 of V2, and a mix of pyramidal and spiny stellate cells in area V1). Differences may have implications for timing factors; that is, impulses from V1, subserved by large-caliber axons, may arrive in MT coincidentally with indirect connections via V2 to MT. Another consideration may be the functional architecture of MT. Regularly spaced clusters of neurons have been described in MT which have similar directionality preferences. The interarbor spacing of cortical efferents is consistent with a columnar organization, but the laminar specificity may indicate recruitment of different combinations of postsynaptic populations by V1 or V2 terminations.

Two types of corticopulvinar terminations: round (type 2) and elongate (type 1).

Corticopulvinar axons were anterogradely labeled by Phaseolus vulgaris-leucoagglutinin injections in the occipitotemporal cortex of the macaque to determine quantitative parameters of divergence and convergence, arbor size and shape, and distribution of terminal specializations. Forty individual axons were analyzed by serial section reconstruction and divided into two major groups. The majority of axons have numerous (typically 500-1,000) small, spinous endings (boutons terminaux). These axons have terminal fields that are beam-like or elongated (E, corresponding to classical type 1) and highly divergent (1.0-3.0 mm). These frequently innervate several of the traditionally designated pulvinar subdivisions; namely inferior pulvinar (PI) and the ventral part of interal pulvinar (PL); medial pulvinar (PM) and dorsal PL, and (one axon) PM, dorsal PL, and PI. Some axons, however (R or round, corresponding to classical type 2), have a small number (typically 70-160) of primarily large, beaded endings (boutons en passant), which concentrate in sharply delimited, round arbors (diameters 100-125 microns). R axons appear to be larger caliber than E axons (1.0-1.5 microns vs. 0.5-1.0 micron, respectively). These differences in phenotype are probably associated with distinct types of projection neurons. In visual areas, corticopulvinar terminations are reported to originate from pyramidal cell subpopulations in layer 5. Indirect evidence, presented here, suggests that the more numerous medium-sized neurons give rise to E axons, and the sparser giant pyramids give rise to R corticopulvinar axons. If this is correct, corticopulvinar connectivity may be involved in multiple transformations. Spatially, axons of giant neurons (with basal dendrites that collect intracortically from a disc-like area, about 1.0 mm in diameter) converge onto a small number of pulvinar neurons. Axons of medium neurons (with basal dendrites that occupy a small intracortical disc, about 0.3 mm in diameter) diverge over 1.0-3.0 mm in the pulvinar and may form many contacts. Giant neurons, although numerically few in relation to medium pyramids (1 or 2: 50?), are likely to have distinctive membrane properties (functionally equivalent to bursting neurons?). Their larger boutons and axon caliber may be associated with a faster transmission that compensates for their small numbers. In primates, the E and R duality does not characterize cortical projections to the caudate, lateral geniculate nucleus, pons, or superior colliculus and thus may be essentially linked to pulvinar-specific processes.

A reticular pattern of intrinsic connections in primate area V2 (area 18).

A system of periodic intrinsic connections is demonstrated in area V2 (area 18) of squirrel and macaque monkeys by large injections of tritiated amino acids, horseradish peroxidase (HRP), and fluorescent latex beads. These connections originate from pyramidal neurons concentrated in layers 3 and 5. Terminations occur in all cortical layers, largely coextensive with labeled neurons but more restricted in layer 4. This multilaminar distribution contrasts with the mainly supragranular localization of periodic intrinsic connections in V1 (area 17), and may imply a close interaction, in V2, of periodic intrinsic connections with pulvinocortical, as well as with corticocortical terminations (concentrated, respectively, in layers 3 and 5, and in lower 3 and 4). As in V1, the tangential configuration of these connections in V2 is reticular or latticelike, and is detectable for 2.5-3.0 mm from an injection site of HRP, 3H amino acids, or latex beads. Cross-sectional widths of labeled regions vary from 250 to 800 micron in squirrel monkey and from 400 to 1,000 micron in macaque, depending on which portion of the lattice is measured. When periodic intrinsic connections are compared with stripes labeled histochemically by cytochrome oxidase (CO), no clear relationship is obvious between the two systems. This result contrasts with the orderly tangential alignment reported between CO-reactive zones in V2 and certain extrinsic connections; namely, pulvinocortical terminations (Livingstone and Hubel, '82) and clusters of neurons projecting to area V4 (DeYoe and Van Essen, '84). Other extrinsic connections, however, such as backgoing connections from V2 to V1, do not seem to have a periodic distribution. Thus, although some discontinuous cortical connections relate to each other in a precise mosaic fashion, intrinsic and some extrinsic connections may observe different modes of organization.

Convergence and branching patterns of round, type 2 corticopulvinar axons.

Corticopulvinar connections consist of at least two morphologically distinct subpopulations. In one subgroup (E, type 1), axons have an "elongated" terminal field and thin, spinous terminations; in the other (R, type 2), axons have a small, round arbor and large, beaded terminations. Previous work (Rockland, 1996) indicates that E-type axons from several occipitotemporal areas branch extensively within and sometimes between pulvinar subdivisions, but that R-type axons tend to have spatially delimited arbors. The present report is a further investigation of R-type axons from areas V1 and MT and was initiated to test the generality of the previous findings. There are four main results: 1) By serial section reconstruction of anterogradely labeled axons, 10 of 25 axons originating in area V1 had two or three spatially separate arbors (8 and 2 axons, respectively). Sixteen axons analyzed from area MT, however, all had single arbors, although the arbors were often formed by the convergence of widely separate branches. 2) Multiple (at least 2-5) R-type corticopulvinar axons, from V1 or from MT, can converge in a single focus. 3) R-type axons originating from both areas V1 and MT can branch to other structures; namely, the superior colliculus, the pretectal area, and/or the reticular nucleus of the thalamus. 4) Finally, corticopulvinar terminations from area V1 are predominantly R-type, whereas those from MT are more predominantly E-type. These results thus provide additional evidence of the special relationship of area V1 to the pulvinar. They also emphasize that the idea of corticopulvinocortical "feedback loops," although convenient as a shorthand nomenclature, does not adequately convey the full complexity of the system.

Terminal arbors of individual "feedback" axons projecting from area V2 to V1 in the macaque monkey: a study using immunohistochemistry of anterogradely transported Phaseolus vulgaris-leucoagglutinin.

In the present study, the anterograde tracer Phaseolus vulgaris-leucoagglutinin (PHA-L) was injected into area V2 in order to demonstrate the precise morphology of individual axons from area V2 to V1. On the basis of 28 complete axon reconstructions, several characteristic features have been identified. 1) Individual axons arborize in multiple layers: 1, 2, 5, and (inconstantly) 3. A single axon may have numerous terminal clusters in layers 1 and 2, but at most one in layer 3. 2) Axons typically ascend to layer 1, turn asymmetrically in one direction, and travel for long distances in this layer (1.10-4.30 mm; dimensions uncorrected for shrinkage). A few axons (three of 28 reconstructed) were found to have a single terminal cluster (0.3-0.5 mm wide) in layers 1 and 2. 3) Collaterals in layer 5 seem to extend over shorter distances (0.60 mm or less). 4) Delicate sprays of boutons (both beads and spines) are clustered along the main trunk. Spacing is variable but usually ranges from 0.35 mm to 0.65 mm. 5) In addition to clustered boutons, there can be linear collaterals, continuously studded with boutons, parallel to the main axon in layer 1. These results indicate that axons from V2 have complex radial and tangential distributions in V1. Terminations in different layers may be directed to different sets of neurons or to different portions of the dendritic tree (for example, distal portions of pyramidal neuron apical dendrites in layers 1 and 2, but more proximal portions in layer 3). Clustered terminations over wide tangential areas may imply a divergent innervation by a single axon of multiple compartmental structures, such as ocular dominance columns or cytochrome oxidase patches.

Feedback connections from area MT of the squirrel monkey to areas V1 and V2.

Area MT/V5 is reciprocally connected with both V1 and V2; but, despite extensive anatomical and physiological investigations, detailed information on the feedback component of these connections is still not available. The present report uses serial section reconstruction of single axons, labeled by anterograde tracers injected in area MT of squirrel monkeys, to characterize these connections further. As with other feedback systems, MT axons terminating in both areas V1 (n = 9) and V2 (n = 6) are widely divergent. In area V1, MT fields are larger than those from V2 and are about comparable to those from V4 or TEO. Terminations in V1, unlike other feedback connections described so far, terminate in several laminar combinations: only layer 1 (n = 2); only layer 4B (n = 3); layers 1 and 4B (n = 1); and layers 1, 4B, and 6 (n = 3). In V2, they occur mainly in layers 1 and 5 or 6. Terminations have two patterns even within a single axon: strung along collateral segments and grouped within small clusters. There are no apparent differences in the size, shape, or density of terminal specializations in V1 or V2, and, consistently with previous double-labeling experiments (Kennedy and Bullier [1985] J Neurosci 5:2815-2830), some axons can branch to both areas. This result, along with the laminar evidence for subtypes of feedback connections, argues against an exclusively hierarchical organization based on "pairwise" connectivity. For V1 and MT, there may be directly reciprocal loops between feedforward and feedback projecting neurons, but this is less likely to be so for V2 and MT.

Axon collaterals of Meynert cells diverge over large portions of area V1 in the macaque monkey.

Patchy intrinsic connections, originating mainly from horizontal collaterals of pyramidal neurons, have been demonstrated in area V1 and many other cortical areas. In this article, we identify a network of intrinsic connections concentrated in layer 6 of area V1. These are visualized by extracellular injections of anterograde tracers in V1, which label small clusters of large terminal boutons in layer 6, in conjunction with thick axon segments. These segments can be traced back to infragranular Meynert cells (n = 10), which are retrogradely labeled from the injections. By using serial section analysis, we identified the following features of this distinctive system of Meynert cell collaterals: (1) terminal clusters are relatively small (<100 microm); (2) each cluster has a small number of rather large boutons (up to 3.0 microm); (3) there is typically a termination-free zone in the immediate vicinity (0.5-2.0 mm) of the cell body; (4) a single neuron has multiple branches that can extend up to 8.0 mm from the soma; and (5) the collaterals are concentrated in layer 6. These features are different from those of horizontal intrinsic connections in the supragranular layers of area V1. They are consistent with fast dynamics and a possible role in wide-field motion processing, such as has been associated with Meynert cells from other studies.

Collateralized divergent feedback connections that target multiple cortical areas.

Nonreciprocal feedback connections from ventromedial areas TE and TF have previously been reported to visual areas V1 and V2 (Kennedy and Bullier, 1985; Rockland and Van Hoesen, 1994). The present report confirms these earlier observations by utilizing anterograde label in conjunction with serial section analysis. Furthermore, it directly demonstrates the divergent configuration and range of these terminal fields. Thirteen axons were analyzed from ventromedial TE (4) or area TF (9) to occipitotemporal areas, and two from area TF to the upper bank of the intraparietal sulcus (IPS). All these axons have narrow, elongated fields that range from 4.0-21.0 mm. Terminations are distributed linearly along the axon or, in some cases, concentrated in irregularly spaced clusters. Most of these axons have terminations concentrated in layer 1. The two axons in the IPS have a bistratified terminal distribution (in layers 1-3 and 6) in their anterior field, but a distinctly different laminar pattern (with terminations concentrated in layer 1) in their distal 2.0 mm. These fields probably correspond to different areas, most likely MIP and PO. Axons projecting from higher order to early visual areas may contribute to extraperceptual, complex processes within area V1, such as activation in response to visual imagery, and are a possible substrate for synchronous linkage of spatially discrete assemblies of neurons. In summary, these results demonstrate 1) that some neurons in ventromedial TE and TF are in direct communication with early visual areas, including V1 and V2, and 2) that some feedback axons target several areas, sometimes with different laminar termination patterns. These results emphasize that cortical areas are interrelated by multiple direct and indirect pathways, not all of which are strictly hierarchical.

Divergent cortical connections to entorhinal cortex from area TF in the macaque.

The entorhinal cortex (EC) is an important component of the medial temporal lobe memory system in the primate and is often viewed as a "gatekeeper" area that passes on highly convergent cortical inputs toward the hippocampus. Further analysis of these connections at a microcircuitry level regarding the actual size and shape of arbors and terminations is not yet available, but may contribute to understanding the role of the EC in memory or other functions. The main emphasis of this report was on serial section analysis of anterogradely labeled axons that project from area TF (lateral parahippocampal cortex; Bonin and Bailey, 1947) to the EC (n = 12). By way of evaluating network organization, other projections from area TF--to TH (in the medial parahippocampal gyrus; n = 5) and to posterior visual areas (n = 3)--were also investigated. All three systems were found to terminate heavily in layer 1, as expected from previous investigations, but some terminations were verified in layer 6 of the EC as well. This technique further demonstrated that terminal fields are widely divergent and elongated. In the EC, terminal fields extended over 6-11 mm and spanned multiple cell islands and interislands. These axons resemble "feedback" cortical connections by virtue of their layer 1 terminations and their markedly divergent geometry, but not by their origin from layer 3. Spatially extended terminal fields recall the nontopographic, distributed character of olfactory connections and raise questions of how these features might be related to the memory functions attributed to medial temporal regions.

Intrinsic laminar lattice connections in primate visual cortex.

Intracortical injections of horseradish peroxidase (HRP) reveal a system of periodically organized intrinsic connections in primate striate cortex. In layers 2 and 3 these connections form a reticular or latticelike pattern, extending for about 1.5-2.0 mm around an injection. This connectional lattice is composed of HRP-labeled walls (350-450 microns apart Saimiri and about 500-600 microns in macaque) surrounding unlabeled central lacunae. Within the lattice walls there are regularly arranged punctate loci of particularly dense HRP label, appearing as isolated patches as the lattice wall labeling thins further from the injection site. A periodic organization has also been demonstrated for the intrinsic connections in layer 4B, which are apparently in register with the supragranular periodicities, although separated from these by a thin unlabeled region. The 4B lattice is particularly prominent in squirrel monkey, extending for 2-3 mm from an injection. In both layers, these intrinsic connections are demonstrated by orthogradely and retrogradely transported HRP and seem to reflect a system of neurons with long horizontal axon collaterals, presumably with arborizations at regularly spaced intervals. The intrinsic connectional lattice in layers 2 and 3 resembles the repetitive array of cytochrome oxidase activity in these layers; but despite similarities of dimension and pattern, the two systems do not appear identical. In primate, as previously described in tree shrews (Rockland et al., '82), the HRP-labeled anatomical connections resemble the pattern of 2-deoxyglucose accumulation resulting from stimulation with oriented lines, although the functional importance of these connections remains obscure.

Inferior parietal lobule projections to the presubiculum and neighboring ventromedial temporal cortical areas.

The entorhinal and perirhinal cortices have long been accorded a special role in the communications between neocortical areas and the hippocampal formation. Less attention has been paid to the presubiculum, which, however, is also a component of the parahippocampal gyrus, receives dense inputs from several cortical areas, and itself is a major source of connections to the entorhinal cortex (EC). In part of a closer investigation of corticohippocampal systems, the authors applied single-axon analysis to the connections from the inferior parietal lobule (IPL) to the presubiculum. One major result from this approach was the finding that many of these axons (at least 10 of 14) branch beyond the presubiculum. For 4 axons, branches were followed to area TF and to the border between the perirhinal and entorhinal cortices, raising the suggestion that these areas, which sometimes are viewed as serial stages, are tightly interconnected. In addition, the current data identify several features of presubicular organization that may be relevant to its functional role in visuospatial or memory processes: 1) Terminations from the IPL, as previously reported for prefrontal connections (Goldman-Rakic et al. [1984] Neuroscience 12:719-743), form two to four patches in the superficial layers. These align in stripes, but only for short distances ( approximately 1.5 mm). This pattern suggests a strong compartmentalization in layers I and II that is also indicated by cytochrome oxidase and other markers. 2) Connections tend to be bistratified, terminating in layers I-II and deeper in layer III. 3) Single axons terminate in layer I alone or in different combinations of layers. This may imply some heterogeneity of subtypes. 4) Individual axons, both ipsilateral projecting (n = 14 axons) and contralateral projecting (n = 6 axons), tend to have large arbors (0.3-0.8 mm across). Finally, the authors observe that projections from the IPL, except for its anteriormost portion, converge at the perirhinal-entorhinal border around the posterior tip of the rhinal sulcus. These projections partially overlap with projections from ventromedial areas TE and TF, and this convergence may contribute to the severe deficits in visual recognition memory resulting from ablations of rhinal cortex.

Single axon analysis of pulvinocortical connections to several visual areas in the macaque.

The pulvinar nucleus is a major source of input to visual cortical areas, but many important facts are still unknown concerning the organization of pulvinocortical (PC) connections and their possible interactions with other connectional systems. In order to address some of these questions, we labeled PC connections by extracellular injections of biotinylated dextran amine into the lateral pulvinar of two monkeys, and analyzed 25 individual axons in several extrastriate areas by serial section reconstruction. This approach yielded four results: (1) in all extrastriate areas examined (V2, V3, V4, and middle temporal area [MT]/V5), PC axons consistently have 2-6 multiple, spatially distributed arbors; (2) in each area, there is a small number of larger caliber axons, possibly originating from a subpopulation of calbindin-positive giant projection neurons in the pulvinar; (3) as previously reported by others, most terminations in extrastriate areas are concentrated in layer 3, but they can occur in other layers (layers 4,5,6, and, occasionally, layer 1) as collaterals of a single axon; in addition, (4) the size of individual arbors and of the terminal field as a whole varies with cortical area. In areas V2 and V3, there is typically a single principal arbor (0.25-0.50 mm in diameter) and several smaller arbors. In area V4, the principal arbor is larger (2.0- to 2.5-mm-wide), but in area MT/V5, the arbors tend to be smaller (0.15 mm in diameter). Size differences might result from specializations of the target areas, or may be more related to the particular injection site and how this projects to individual cortical areas. Feedforward cortical axons, except in area V2, have multiple arbors, but these do not show any obvious size progression. Thus, in areas V2, V3, and especially V4, PC fields are larger than those of cortical axons, but in MT/V5 they are smaller. Terminal specializations of PC connections tend to be larger than those of corticocortical, but the projection foci are less dense. Further work is necessary to determine the differential interactions within and between systems, and how these might result in the complex patterns of suppression and enhancement, postulated as gating mechanisms in cortical attentional effects, or in different states of arousal.

Anatomical binding of intrinsic connections in striate cortex of tree shrews (Tupaia glis).

The intrinsic connectivity of striate cortex was investigated by injecting horseradish peroxidase (HRP) into this area in tree shrews. Such HRP injections demonstrated periodically organized, stripelike connections within area 17. These stripes occur in layers I-IIIA and consist of a small number or retrogradely filled neurons, some clearly pyramidal, together with HRP-labeled axon terminals. HRP-filled axons trunks run between labeled stripes, interconnecting adjacent and distant regions of the stripe pattern. Correlation with Golgi-stained tissue suggests that these stripes are horizontally interconnected by pyramidal neurons with long intracortical axon collaterals (followed for distances over 1 mm from the soma). The HRP-labeled strips measure about 230 micrometers in width, with a center-to-center repeat distance of 450--500 micrometers. They have been mapped over an 8 mm2 area of striate cortex and would thus seem capable of effecting lateral interactions over considerable portions of the retinotopic map. In their dimensions and overall pattern, these anatomical stripes resemble the 2-deoxyglucose (2-DG) bands resulting from visual stimulation of trees shrews with stripes of a single orientation. While the functional role of the HRP-labeled stripes is unclear, their similarities with the 2-DG pattern raise the intriguing possibility that they may be related to orientation selectivity. The striking regularity of these extensive lateral interconnections emphasizes the importance of horizontal intralaminar connections within the cortex.