Noritaka Ichinohe

Some thoughts on cortical minicolumns

Although a columnar geometry is one of the defining features of cortical organization, major issues regarding its basic nature, key features, and functional significance remain unclear and often controversial. This review is intended to survey some of the basic anatomical features of columnar organization, and in particular the smaller scale dendritic minicolumns. One motive was simply to clarify what seem to be differences in terminology, where “minicolumn” can be used to refer to vertical rows of cells, pyramidal cell modules, or apical dendritic bundles. A second aim was to review anatomical details which over the years have tended increasingly to be overlooked. A third aim was to expand on recent results concerning the border of layers 1 and 2 as a specialized zone with its own micromodular organization. Views on columnar organization have arguably been heavily influenced by a desire for general principles; but re-examination of the complex underlying features may be both timely and worthwhile. We point out that what are defined as dendritic bundles do not extend through the full cortical thickness and are not strictly repetitive, but rather display significant inter- and intra-areal variation.

A di-synaptic projection from the lateral cerebellar nucleus to the laterodorsal part of the striatum via the central lateral nucleus of the thalamus in the rat

We have examined a cerebello-thalamo-striatal pathway from the lateral cerebellar nucleus (LCN) to the laterodorsal part of the striatum (LDS) through the central lateral nucleus (CL) using light and electron microscopy through the employment of a combination of anterograde and retrograde tracing techniques. Biotinylated dextran amine (BDA) was injected into the unilateral LCN, and used as an anterograde tracer. Cholera toxin B subunit (CTb), used for light microscopy, and wheat germ agglutinin-horseradish peroxidase (WGA-HRP), used for electron microscopy, were injected into the contralateral LDS as retrograde tracers. Light microscopic analysis showed a good overlap of the distribution of BDA-labeled axon terminals and CTb-labeled neurons in the middle third of the CL in both dorsoventral and rostrocaudal axes on the LDS injection side. Electron microscopy confirmed the presence of direct synaptic contacts between BDA-labeled terminals and WGA-HRP-labeled dendrites in the CL.

Cortical Connections to Area TE in Monkey: Hybrid Modular and Distributed Organization

To investigate the fine anatomical organization of cortical inputs to visual association area TE, 2–3 small injections of retrograde tracers were made in macaque monkeys. Injections were made as a terminal procedure, after optical imaging and electrophysiological recording, and targeted to patches physiologically identified as object-selective. Retrogradely labeled neurons occurred in several unimodal visual areas, the superior temporal sulcus, intraparietal sulcus (IPS), and prefrontal cortex (PFC), consistent with previous studies. Despite the small injection size (<0.5 mm wide), the projection foci in visual areas, but not in IPS or PFC, were spatially widespread (4–6 mm in extent), and predominantly consisted of neurons labeled by only one of the injections. This can be seen as a quasi-modular organization. In addition, within each projection focus, there were scattered neurons projecting to one of the other injections, together with some double-labeled (DL) neurons, in a more distributed pattern. Finally, projection foci included smaller “hotspots,” consisting of intermixed neurons, single-labeled by the different injections, and DL neurons. DL neurons are likely the result of axons having extended, spatially separated terminal arbors, as demonstrated by anterograde experiments. These results suggest a complex, hybrid connectivity architecture, with both modular and distributed components.

Morphologic changes of dendritic spines of striatal neurons in the levodopa-induced dyskinesia model.

Maladaptive plasticity at corticostriatal synapses plays an important role in the development of levodopa‐induced dyskinesia. Recently, it has been shown that synaptic plasticity is closely linked to morphologic changes of dendritic spines. To evaluate morphologic changes of dendritic spines of two types of striatal medium spiny neurons, which project to the internal segment of globus pallidus or the external segment of globus pallidus, in the levodopa‐induced dyskinesia model, we used 6‐hydroxydopamine‐lesioned rats chronically treated with levodopa. Dendritic spines were decreased and became enlarged in the direct pathway neurons of the model of levodopa‐induced dyskinesia. The same levodopa treatment to normal rats, in which no dyskinesia was observed, also induced enlargement of dendritic spines, but not a decrease in density of spines in the direct pathway neurons. These results suggest that a loss and enlargement of dendritic spines in the direct pathway neurons plays important roles in the development of levodopa‐induced dyskinesia.

Postnatal development of layer III pyramidal cells in the primary visual, inferior temporal, and prefrontal cortices of the marmoset

Abnormalities in the processes of the generation and/or pruning of dendritic spines have been implicated in several mental disorders including autism and schizophrenia. We have chosen to examine the common marmoset (Callithrix jacchus) as a primate model to explore the processes. As a first step, we studied the postnatal development of basal dendritic trees and spines of layer-III pyramidal cells in the primary visual sensory cortex (V1), a visual association cortex (inferior temporal area, TE), and a prefrontal cortex (area 12, PFC). Basal dendrites in all three areas were longer in adulthood compared with those in the newborn. In particular, rapid dendritic growth occurred in both TE and PFC around the second postnatal month. This early growth spurt resulted in much larger dendritic arbors in TE and PFC than in V1. The density of the spines along the dendrites peaked at 3 months of age and declined afterwards in all three areas: the degree of spine pruning being greater in V1 than in TE and PFC. The estimates of the total numbers of spines in the basal dendrites of a single pyramidal cell were larger in TE and PFC than in V1 throughout development and peaked around 3 months after birth in all three areas. These developmental profiles of spines and dendrites will help in determining assay points for the screening of molecules involved in spinogenesis and pruning in the marmoset cortex.

Parvalbumin positive dendrites co-localize with apical dendritic bundles in rat retrosplenial cortex.

The granular retrosplenial cortex in rats, involved in learning and memory, has a highly modular organization in layer 1. Apical dendrites of layer 2 pyramidal neurons form bundles, which correspond to patchy thalamic input. Here, we demonstrate that dendrites of parvalbumin-immunoreactive (PV-ir) GABAergic neurons in layers 2/3 also form bundles, and that these co-localize in layer 1 with the apical dendritic bundles, as verified by double immunofluorescence for PV and microtuble-associated protein. Deeper, at the border of layers 1/2, the PV-ir bundles merge into a honeycomb-like structure, with walls consisting of PV-ir neuropil. Compartmentalization in layers 1/2 is characteristic of other periallocortical structures. Further work is necessary to determine whether these specializations may be specifically related to learning and memory.

Differential modes of termination of amygdalothalamic and amygdalocortical projections in the monkey

The amygdala complex participates in multiple systems having to do with affective processes. It has been implicated in human disorders of social and emotional behavior, such as autism. Of the interconnected functional networks, considerable research in rodents and primates has focused on connections between the amygdala and orbitofrontal cortex (OFC). The amygdala projects to OFC by both a direct amygdalocortical (AC) pathway and an indirect pathway through mediodorsal thalamus. In the rat, retrograde tracer experiments indicate that the AC and amygdalothalamic (AT) pathways originate from separate populations, and may therefore convey distinctive information, although the characteristics of these pathways remain unclear. To investigate this issue in monkeys we made anterograde tracer injections in the basolateral amygdala complex (BLC; n = 3). Three distinctive features were found preferentially associated with the AT or AC pathways. First, AT terminations are large (average diameter = 3.5 μm; range = 1.2–7.0 μm) and cluster around proximal dendrites, in contrast with small‐bouton AC terminations. Second, AT terminations form small arbors (diameter ≈0.1 mm), while AC are widely divergent (often >1.0 mm long). The AT terminations features are reminiscent of large bouton, “driver” corticothalamic terminations. Finally, AC but not AT terminations are positive for zinc (Zn), a neuromodulator associated with synaptic plasticity. From these results we suggest that AC and AT terminations originate from distinct populations in monkey as well as in rodent. Further work is necessary to determine the degree and manner of their segregation and how these subsystems interact within a broader connectivity network. J. Comp. Neurol. 502:309–324, 2007.

Area-specific substratification of deep layer neurons in the rat cortex

Gene markers are useful tools to identify cell types for fine mapping of neuronal circuits. Here we report area‐specific sublamina structure of the rat cerebral cortex using cholecystokinin (cck) and purkinje cell protein4 (pcp4) mRNAs as the markers for excitatory neuron subtypes in layers 5 and 6. We found a segregated expression, especially pronounced in layer 6, where corticothalamic and corticocortical projecting neurons reside. To examine the relationship between gene expression and projection target, we injected retrograde tracers into several thalamic subnuclei, ventral posterior (VP), posterior (PO), mediodorsal (MD), medial and lateral geniculate nuclei (MGN and LGN); as well as into two cortical areas (M1 and V1). This combination of tracer‐in situ hybridization (ISH) experiments revealed that corticocortical neurons predominantly express cck and corticothalamic neurons predominantly express pcp4 mRNAs in all areas tested. In general, cck(+) and pcp4(+) cells occupied the upper and lower compartment of layer 6a, respectively. However, the sublaminar distribution and the relative abundance of cck(+) and pcp4(+) cells were quite distinctive across areas. For example, layer 6 of the prelimbic cortex was almost devoid of cck(+) neurons, and was occupied instead by corticothalamic pcp4(+) neurons. In the lateral areas, such as S2, there was an additional layer of cck(+) cells positioned below the pcp4(+) compartment. The claustrum, which has a tight relationship with the cortex, mostly consisted of cck(+)/pcp4(−) cells. In summary, the combination of gene markers and retrograde tracers revealed a distinct sublaminar organization, with conspicuous cross‐area variation in the arrangement and relative density of corticothalamic connections. J. Comp. Neurol. 520:3553–3573, 2012.

Morphological study of the tegmental pedunculopontine nucleus, substantia nigra and subthalamic nucleus, and their interconnections in rat organotypic culture

The morphological organization of the tegmental pedunculopontine nucleus, midbrain extrapyramidal area, substantia nigra and subthalamic nucleus and their interrelationships were studied in rat organotypic culture using immunohistochemistry and NADPH-diaphorase histochemistry. Three coronal sections, one containing the tegmental pedunculopontine nucleus/midbrain extrapyramidal area, another with the substantia nigra and the third with the subthalamic nucleus, were obtained from postnatal 1–2-day-old rats. These sections were co-cultured for 3–4 weeks using the roller-tube technique. In the tegmental pedunculopontine nucleus/midbrain extrapyramidal area, the distribution pattern of cholinergic neurons was similar to that found in the in vivostudy. We could, therefore, identify the subdivisions of the tegmental pedunculopontine nucleus (i.e., pars compacta and pars dissipata) and the midbrain extrapyramidal area. As in the in vivosituation, glutamate immunoreactive neurons were also located in these areas. Approximately 10% of NADPH-diaphorase positive neurons in the tegmental pedunculopontine nucleus, were glutamate immunoreactive. In the substantia nigra, as in the in vivo, tyrosine hydroxylase immunoreactive (putative dopaminergic) neurons were identified predominantly in the substantia nigra pars compacta, and parvalbumin immunoreactive neurons (putative GABAergic) mainly in the substantia nigra pars reticulata. The subthalamic nucleus was ladened with glutamate immunoreactive neurons. NADPH-diaphorase stained axons originating from the tegmental pedunculopontine nucleus were traced into the substantia nigra and subthalamic nucleus. They were often in close apposition to tyrosine hydroxylase immunoreactive neurons in the substantia nigra. Parvalbumin immunoreactive fibers from the substantia nigra projected heavily to the midbrain extrapyramidal area, but only sparsely to the tegmental pedunculopontine nucleus and the subthalamic nucleus. These findings indicate that the tegmental pedunculopontine nucleus/midbrain extrapyramidal area, substantia nigra and subthalamic nucleus in the organotypic culture have retained a basic morphological organization and connectivity similar to those seen in the in vivosituation. Therefore, this preparation could be a useful model to conduct further studies to investigate functional circuits among the structures represented.

Zinc-enriched amygdalo- and hippocampo-cortical connections to the inferotemporal cortices in macaque monkey

Synaptic zinc (Zn), a co-factor in some glutamatergic synapses, has been implicated in plasticity effects, as well as in several excitotoxic and other pathophysiological conditions. In this study, we provide information about the distribution of Zn in inferotemporal cortex, a region at the interface of the visual and hippocampal networks. In brief, we found a lateral to medial increase in Zn, where TEad, a unimodal visual area, showed low levels of Zn; TEav, intermediate levels; and perirhinal cortex, a multimodal limbic area, high levels. The distribution of parvalbumin, a calcium binding protein, showed a reverse gradient to that of Zn. The neurons of origin of the Zn+ termination were identified by making intracortical injections sodium selenite (Na2SeO3). This substance interacts with Zn to form precipitates of ZnSe and in this form is transported retrogradely to the soma. A mixed population of labeled neurons was visualized, which included Zn+ neurons in CA1 of the hippocampus and in several amygdala subnuclei. In CA1, Zn+ neurons were restricted to the upper part of stratum pyramidale. Zn is thought to contribute to activity-dependent synaptic plasticity. The specifically high level in perirhinal cortex, and its origin from neurons in CA1 and the amygdala, may relate to cellular events involved in visual long-term memory formation.