Mami Ishida

Organization and development of corticocortical associative neurons expressing the orphan nuclear receptor Nurr1

The developmental mechanism that contributes to the highly organized axonal connections within the cerebral cortex is not well understood. This is partly due to the lack of molecular markers specifically expressed in corticocortical associative neurons during the period of circuit formation. We have shown previously that latexin, a carboxypeptidase A inhibitor, is expressed in intrahemispheric corticocortical neurons from the second postnatal week in the rat (Arimatsu et al. [1999] Cereb. Cortex 9:569–576). In the present study, we first demonstrate in the adult rat that the orphan nuclear receptor Nurr1 is coexpressed in latexin‐expressing neurons located in layer V, sublayer VIa, and the white matter of the lateral sector of the neocortex, and also in latexin‐negative early born neurons in sublayer VIb of the entire neocortex. Virtually all Nurr1‐expressing neurons exhibit immunoreactivity for phosphate‐activated glutaminase but not for γ‐aminobutyric acid, suggesting that they are glutamatergic‐excitatory neurons. By combining Nurr1 immunohistochemistry and 5‐bromo‐2′‐deoxyuridine‐birthdating, we then show that Nurr1 is expressed in (early born) subplate neurons and (later born) presumptive latexin‐expressing neurons from embryonic day 18 onward. Finally, by combination of Nurr1 immunohistochemistry and retrograde tracing, we show that Nurr1‐expressing neurons, including those in sublayer VIb, contribute predominantly to long‐range intrahemispheric corticocortical projections. These results raise the possibility that Nurr1 plays a role in the establishment and maintenance of normal corticocortical circuitry and function. J. Comp. Neurol. 466:180–196, 2003.

Area- and lamina-specific organization of a neuronal subpopulation defined by expression of latexin in the rat cerebral cortex.

The aim of the present study was to investigate the density, laminar distribution, size, morphology, and neurotransmitter phenotype of rat cortical neurons expressing latexin, an inhibitor of carboxypeptidase A. Immunohistochemical analyses established that latexin-immunoreactive neurons are restricted essentially to the infragranular layers of lateral cortical areas in the rat. The overall density, laminar or sublaminar localization, and cell size distribution of latexin-positive neurons differed substantially across cytoarchitectonic areas within lateral cortex. Numerous latexin-positive neurons had the morphology of modified pyramidal cells especially of layer VI. The vast majority of latexin-positive neurons were glutamate-immunoreactive in the six lateral neocortical areas examined, while neurons immunoreactive for both latexin and GABA were virtually absent. Thus the majority of latexin-positive neurons are likely to be excitatory projection neurons. The area- and lamina-specific distribution of the latexin-expressing subpopulation of glutamate-immunoreactive neurons is a distinctive feature that may contribute to the functional specialization of the lateral cortical areas.

SUMO-1 immunoreactivity co-localizes with phospho-Tau in APP transgenic mice but not in mutant Tau transgenic mice

Sumoylation is a post-translational modification process that is supposed to be implicated in the pathogenesis of several neurodegenerative diseases. Recently, the microtubule-associated protein Tau was identified as a target for sumoylation in the analysis of the transfected cells. We investigated the localization of SUMO-1 protein in APP transgenic mice and mutant Tau transgenic mice, and found that SUMO-1 immunoreactivity was co-localized with phosphorylated Tau aggregates in amyloid plaques of APP transgenic mice. By contrast, no SUMO-1 immunoreactivity was observed in phosphorylated Tau aggregates of mutant Tau transgenic mice. The contribution of sumoylation to the neurodegeneration in Alzheimers disease will be further elucidated via the analysis of APP transgenics.

Candidaalbicans serotype a strains grow in yeast extract-added sabouraud liquid medium at pH 2.0, elaborating mannans without β-1,2 linkage and phosphate group

Cultivation of three Candida albicans strains, NIH A-207, J-1012, and NIH B-792, abbreviated as A-, J-, and B-strains, respectively, in yeast extract-enrich Sabouraud liquid medium at pH 2.0 provided the following findings, i.e., the two former strains belonging to serotype A were able to grow in this medium in almost the same rates as those in the same medium of pH 5.9, while B-strain cells did not proliferate under the former condition. The cells of A- and J-strains cultivated at pH 2.0 did not undergo agglutination with the factor serum 6 in a commercially available factor serum kit, Candida Check, corresponding to C. albicans serotype A-specific epitope. It was also revealed by 1H-13C correlation spectra of the mannans isolated from the cells of A- and J-strains contained neither phosphate group nor beta-1,2-linked mannopyranose unit, although these mannans retained non-reducing terminal alpha-1,3 linked mannopyranose units, providing a substantiating evidence that the serotype A-specific epitope contains a non-reducing terminal beta-1,2-linked mannopyranose unit.

Chemically defined feedback connections from infragranular layers of sensory association cortices in the rat

The primary visual (V1), auditory (AI), and somatosensory (SI) cortices are reciprocally connected with their respective sensory association cortices. In the rat, we have previously demonstrated that some of the connections arising from the secondary somatosensory (SII) and parietal insular (PA) cortices and terminating in the SI, are characterized by the expression of latexin, a candidate protein of carboxypeptidase A inhibitor. Here, by using retrograde tracing and latexin-immunohistochemistry, we show that latexin-expressing neurons in other association cortices of different sensory modalities also contribute to the feedback projections to the corresponding primary sensory cortices. These are the lateral part of the secondary visual cortex (V2L), temporal association cortex, and the dorsal and ventral (AIIv) parts of the secondary auditory belt cortex. Within sublayer VIa of the V2L, AIIv and SII, the majority of the V1-, AI- and SI-projecting neurons respectively, are latexin-immunopositive. In contrast to feedback connections, far fewer latexin-expressing neurons participate in callosal or intrahemispheric feedforward connections. The latexin-expressing neurons constitute a virtually completely different population from corticothalamic neurons within the infragranular layers. Given that latexin might participate in the modulation of neuronal activity by controlling the protease activity, latexin-expressing feedback pathways would play a unique role in the modulation of sensory perception.

Distinct neuronal populations specified to form corticocortical and corticothalamic projections from layer VI of developing cerebral cortex

Layer VI of the cerebral cortex contains heterogeneous populations of pyramidal neurons whose axons project either cortically or subcortically. It has been shown that a subset of layer VI neurons expressing latexin projects ipsilaterally to other cortical areas but does not contribute to the corticothalamic projections. Taking advantage of the connectional specificity of latexin-expressing neurons, we here determine whether corticocortical and corticothalamic neurons are generated at different times, and at which stage the connectional distinction develops in corticogenesis. Our experimental findings indicate that: (1) thalamic-projecting neurons in layer VI of the rat secondary somatosensory cortex (SII) are born at embryonic day 14 or before while latexin-expressing neurons in the same layer are generated at embryonic day 15 or later; (2) axonal invasion by SII neurons into ipsilateral cortical areas and into the posterior dorsal thalamus mainly takes place early in the postnatal period; (3) latexin-expressing neurons never project toward the dorsal thalamus in normal development; (4) presumptive latexin-expressing neurons in the neonatal SII are able to grow into a cortical slice in vitro, but do not invade a thalamic slice even transiently; (5) thalamic-projecting neurons, on the other hand, fail to simultaneously establish connections with a cortical slice. Taken together, our findings suggest that the time frame in which presumptive corticocortical and corticothalamic neurons are generated differs, and that the two populations are restricted in connectional fate potential by the perinatal period prior to target innervation.

Early patterning of the rat cerebral wall for regional organization of a neuronal population expressing latexin

The exact timing of regional patterning in the developing cerebral cortex and other telencephalic structures remains to be elucidated. In the present study, we addressed this issue by comparing the distribution and density of neuronal population expressing latexin in the adult rat telencephalon, with the regional pattern in the fetal cerebral wall as to the potential to generate latexin-expressing neurons. Immunohistochemical analyses on adult animals have shown that latexin-expressing neurons are restricted to a lateral cortical field, within which they are most abundant at the middle level, decreasing in number rostrally and caudally. Substantial numbers of latexin-immunopositive neurons were recorded in the claustrum and endopiriform nuclei, both of which are located from rostral to middle level in the lateral telencephalon. By examining the number and density of latexin-immunopositive neurons in organotypic slice cultures from various portions of the developing rat cerebral wall, it has been shown that the regional pattern within the early cerebral wall as to the potential to generate latexin-expressing neurons matches well the distribution and density of latexin-expressing neurons in the adult telencephalon. Thus, in cultures derived from either embryonic day 13 or 16 fetuses, latexin-immunopositive neurons appeared most prominently in those from rostral-to-middle portions of the lateral cerebral wall, decreasing in number rostrally and caudally. In cultures from the dorsal cerebral wall, the number was generally very low. In light of our previous finding that most prospective latexin-expressing neurons are still dividing at embryonic day 13, it can be concluded that some kind of pattern formation event occurs within the early cerebral wall even prior to the genesis of the postmitotic neurons that would be later allocated in a region-specific manner.

Latexin expression in smaller diameter primary sensory neurons in the rat.

Most of the smaller diameter neurons of dorsal root and trigeminal ganglia in adult rats expressed latexin, which has the inhibitor activity of carboxypeptidase A. Most of the dorsal root ganglion (DRG) neurons containing either calcitonin gene-related peptide (CGRP), substance P (SP) or somatostatin (SST) coexpressed latexin. Latexin was widely distributed in the cytoplasm of the cell body and in axonal fibers of cultured DRG neurons which were sensitive to capsaicin. In addition, latexin-immunoreactivity was observed throughout lamina II of the spinal cord in normal animals, but was lost following sciatic nerve-axotomy, suggesting the presence of latexin-immunoreactive axonal fibers and/or terminals from DRG neurons. Immunoelectron microscopy indeed revealed latexin-immunoreactive axonal terminals and thinly myelinated and unmyelinated axonal fibers within the dorsal horn. These observations suggest that latexin may be involved in nociceptive information transmission or its modulation.

Reduced pain sensitivity in mice lacking latexin, an inhibitor of metallocarboxypeptidases

Latexin, the endogenous protein inhibitor of the A/B subfamily of metallocarboxypeptidases, is expressed in small nociceptive neurons in sensory ganglia and in a subset of neurons in the telencephalon. In this study, we generated latexin-deficient mice that exhibited increased tail-flick latency compared to wild-type animals upon noxious heat stimulation. The reduced pain sensitivity in the mutants was rescued by the systemic administration of a plant carboxypeptidase inhibitor that inhibits the A/B subfamily of metallocarboxypeptidases. These findings suggest that latexin is involved in the transmission of pain.

Expression of the semaphorins Sema 3D and Sema 3F in the developing parathyroid and thymus

We have identified two Semaphorin genes, Sema 3D and Sema 3F, as being specifically expressed in the developing parathyroid and thymus. The thymus and parathyroid originate from a common primordium that develops from the third pharyngeal pouch in mice. The expression of Sema 3D and Sema 3F was compared with that of gcm2, Pax1, and Neuropilin‐1 and ‐2, and genes encoding a variety of Plexins by in situ hybridization during mouse pharyngeal development. Sema 3D was specifically expressed in the dorsal and cranial portions of the primordium developing from the third pouch at mouse embryonic day (E) 10.5. That the expression pattern of Sema 3D was consistent with that of gcm2 indicates that Sema 3D was expressed in the parathyroid‐specific domains of the developing third pouch. The parathyroid‐specific pattern of Sema 3D expression continued until E17.5. By contrast, Sema 3F was expressed in both the parathyroid and thymic domains of the common primordium at E10.5, and the expression levels in both tissues were decreased during development. The Semaphorin receptors were expressed in the blood vessels, nerves, and mesenchymal cells adjacent to the common primordium or the separated parathyroid and thymus during development. Our results show that Sema 3D and Sema 3F genes were overlappingly and distinctly expressed in the developing parathyroid and thymus, respectively, and that Semaphorin signaling might play roles in the interactions between these organs and surrounding tissues such as nerves and blood vessels, and in the recruitment of lymphoid cells. Developmental Dynamics 237:1699–1708, 2008.