Elsevier

Neuroscience Research

Volume 105, April 2016, Pages 49-64
Neuroscience Research

Classic cadherin expressions balance postnatal neuronal positioning and dendrite dynamics to elaborate the specific cytoarchitecture of the mouse cortical area

https://doi.org/10.1016/j.neures.2015.09.006Get rights and content

Highlights

  • We confirm expression profiles of classic cadherins in the mouse barrel cortex.

  • We generate a resourceful transgenic mouse illuminating nuclei of barrel neurons.

  • A conditional electroporation method allows us to test postnatal roles of cadherins.

  • Cadherins are involved in elaboration of the barrel area specific cytoarchitecture.

  • Activity dependent machinery might modulate the cadherin expression profiles.

Abstract

A unique feature of the mammalian cerebral cortex is in its tangential parcellation via anatomical and functional differences. However, the cellular and/or molecular machinery involved in cortical arealization remain largely unknown. Here we map expression profiles of classic cadherins in the postnatal mouse barrel field of the primary somatosensory area (S1BF) and generate a novel bacterial artificial chromosome transgenic (BAC-Tg) mouse line selectively illuminating nuclei of cadherin-6 (Cdh6)-expressing layer IV barrel neurons to confirm that tangential cellular assemblage of S1BF is established by postnatal day 5 (P5). When we electroporate the cadherins expressed in both barrel neurons and thalamo-cortical axon (TCA) terminals limited to the postnatal layer IV neurons, S1BF cytoarchitecture is disorganized with excess elongation of dendrites at P7. Upon delivery of dominant negative molecules for all classic cadherins, tangential cellular positioning and biased dendritic arborization of barrel neurons are significantly altered. These results underscore the value of classic cadherin-mediated sorting among neuronal cell bodies, dendrites and TCA terminals in postnatally elaborating the S1BF-specific tangential cytoarchitecture. Additionally, how the “protocortex” machinery affects classic cadherin expression profiles in the process of cortical arealization is examined and discussed.

Introduction

The cerebral cortex (neocortex, isocortex) is a telencephalic region that exhibits a mammal specific laminated cellular arrangement along its radial axis. Recent studies have revealed numerous cellular and molecular machineries that are essential for elaborating the six-layered organization in the developing cerebral cortex. For instance, mouse cortical neurons born in the telencephalic ventricular zone migrate radially toward the marginal cortical plate, where late born neurons go across the layer of early born neurons to form the so-called “inside-out” laminated structure (Molyneaux et al., 2007, Kwan et al., 2012). The secreted molecules and their receptor components that eventually regulate cytoskeletal assemblage have been suggested to play crucial roles in directing the mode of radial cell migration (Frotscher, 2010). In humans, it has been demonstrated that dysfunction of such molecular machinery causes lissencephaly or various types of cortical heterotopia (Liu, 2011). Another remarkable feature of the cerebral cortex is in its architectonic differences along the tangential dimension. It has been proposed that the areal identity of the developing cortex could be determined by an intrinsic “protomap” and/or activity-dependent “protocortex” mechanisms (Rakic, 1988, O’Leary, 1989, Rubenstein and Rakic, 1999, Sur and Leamey, 2001, Mallamaci and Stoykova, 2006, O’Leary et al., 2007, Rakic et al., 2009). For example, a secreted molecule, FGF8, which is highly expressed in the anterior primordium of the mouse cerebral cortex can act as a morphogen to determine areal identities (Fukuchi-Shimogori and Grove, 2001, Grove and Fukuchi-Shimogori, 2003, Shimogori and Grove, 2005). Many transcription factors are currently known to be expressed under the control of a morphogen gradient to sequentially establish the cortical protomap from the ventricular zone to the cortical plate (Greig et al., 2013). Inputs from thalamo-cortical axons (TCAs) also play significant roles in locally driving area-specific gene expressions in the mouse cortical plate at the later developmental stages (Senft and Woolsey, 1991, Vue et al., 2013, Lokmane et al., 2013). Human congenital diseases such as polymicrogyria affect cortical cell arrangement only within restricted sets of functional areas, implicating the dominant roles of cortical area-specific genetic traits among mammalian species (Monuki and Walsh, 2001, Piao et al., 2004, Chen et al., 2011). However, detailed cellular and/or molecular machinery required to strictly define the specific features of each cortical area remains largely unknown.

Classic cadherins are transmembrane proteins that confer subclass-specific adhesiveness to cells (Nose et al., 1988, Takeichi, 1988, Steinberg and Takeichi, 1994, Takeichi, 2007, Hirano and Takeichi, 2012). As many as 20 classic cadherin subclasses encoded by different genes have thus far been identified, and each subclass shows spatio-temporally regulated expression patterns during development (Takeichi, 1988, Hirano and Takeichi, 2012). A previous series of studies has confirmed the roles of this molecular family in assembling the elaborate structure of germ layers, tissue and/or organs via selective cell sorting machinery (Detrick et al., 1990, Fujimori et al., 1990, Nakagawa and Takeichi, 1998, Gumbiner, 2005). In the developing vertebrate central nervous system, cadherin expression occurs in a manner specific to neuromeres, layers and/or nuclei (Matsunami and Takeichi, 1995, Redies, 1995, Redies and Takeichi, 1996, Inoue et al., 1997). Accumulated evidence has shown that such restricted expression profiles are indeed required to form and/or maintain the cellular arrangement or neuronal connectivity (Manabe et al., 2000, Inoue et al., 2001, Suzuki et al., 2007, Osterhout et al., 2011, Williams et al., 2011). In the postnatal mouse brain, for instance, three type-II classic cadherins, cadherin-6 (Cdh6), Cdh8 and Cdh11, differentially demarcate functional circuitries such as somatosensory, visual and auditory systems (Suzuki et al., 1997, Inoue et al., 1998). In the perinatal mouse cerebral cortex, those cadherin gene expression profiles seem to subdivide the cortical plate into tens of tangential domains (Suzuki et al., 1997). In our CreERT2-based genetic fate tracing experiment, it has been demonstrated that a sharp Cdh6 gene expression boundary established at around postnatal day 5 (P5) in the mouse cortical plate precisely corresponds to the future medial boundary along the barrel field of the primary somatosensory area (S1BF) (Terakawa et al., 2013). We have also revealed that ectopic expression of Cdh6 in the postnatal cortical plate perturbs the characteristic TCA terminal structures formed in the S1BF layer IV (Terakawa et al., 2013). This suggests that the Cdh6 expression profile per se might have functional significance with regard to determining the areal boundary and/or area-specific cytoarchitecture. By using a dominant negative molecule for all subclasses of classic cadherins, it has been reported that some types of classic cadherins could do mediate selective local intracortical circuit formation among layer II/III and layer IV neurons in the mouse S1BF (Wakimoto et al., 2015). Although expression and/or functional redundancy has minimized gene knock out phenotypes of classic cadherins in the developing or postnatal mouse brain, those neural circuits dominantly expressing small sets of cadherin subclasses are definitely affected in mouse mutants, highlighting the crucial roles of this molecular family in selective synapse formation (Manabe et al., 2000, Osterhout et al., 2011, Williams et al., 2011, Suzuki et al., 1997). Incidentally, several mutations in human classic cadherin loci have been demonstrated to harbor linkages with neurodevelopmental psychiatric disorders (Wang et al., 2009, Pagnamenta et al., 2011, Redies et al., 2012, Crepel et al., 2014).

In the present study, we first confirmed differential expression profiles of classic cadherins in the tangential plane of postnatal mouse S1BF and revealed the developmental dynamics of the barrel area-specific cytoarchitecture by generating a new Cdh6::nlsEGFP-BAC Tg mouse line that selectively illuminates the cell nuclei of Cdh6-expressing cortical layer IV spiny stellate neurons. By means of the ingenious conditional electroporation methodology, we next found that both gain- and loss-of-function analyses for various classic cadherin subclasses results in perturbation of the characteristic barrel cell positioning and/or dendrite arborization patterns along the cortical tangential dimension. These findings suggest the value of classic cadherin-mediated sorting machinery among neuronal cell bodies, dendrites and TCA terminals in elaborating the areal identity. Further we provided evidence that such area-specific classic cadherin expression profiles might depend on neuronal activity. From these results, we propose that cell–cell/axon–dendrite interactions balanced by classic cadherin expressions could do organize the cortical area-specific cytoarchitecture, possibly under the control of neuronal activity.

Section snippets

Animals

All animal experiments in this study have been approved by the Animal Care and Use Committee of the National Institute of Neuroscience, NCNP, Japan. Generation of Cdh6::nlsLacZ-BAC-Tg and Cdh6::GAP-EGFP-BAC-Tg mouse lines were detailed previously (Terakawa et al., 2011, Terakawa et al., 2013). To establish Cdh6::nlsEGFP-BAC-Tg mouse lines, RP23-78N21, the same BAC clone utilized in Cdh6::nlsLacZ-BAC-Tg mouse line, was modified by means of homologous recombination so that enhanced green

Expression mapping of classic cadherins in the mouse S1BF along the tangential dimension of cortical layer IV at P7

Our group and others have already reported that many of classic cadherin subclasses are expressed by distinct sets of cortical layers/cells in the mouse S1BF during the postnatal stages (Suzuki et al., 1997, Terakawa et al., 2013, Inoue et al., 2008b, Krishna et al., 2011, Lefkovics et al., 2012). In the present study, we first examined expression profiles of classic cadherins in the mouse S1BF at P7 especially along the tangential dimension of cortical layer IV by utilizing available in situ

Discussion

In order to elaborate area-specific cytoarchitecture after radial migration, neurons must control their own spatial occupancy and axon/dendrite extensions based on intrinsic and/or extrinsic areal pattering information in each cortical layer. It is hence likely that the cellular and/or molecular machinery required for precise cell–cell recognitions plays pivotal role in the developmental processes. In the present study, we detailed classic cadherin expression profiles in the mouse S1BF layer IV

Competing interest

The authors declare no competing financial interests.

Authors’ contribution

S.F.E. and T.I. designed the project. S.F.E., Y.U.I., J.A., Y.W.T. and T.I. performed the experiments with help from M.H. and S.F.E. and T.I. analyzed the data. T.I. wrote the manuscript with help from S.F.E.

Acknowledgements

We thank Dr. Neal Copeland for the recombineering related materials including EL250 bacterial stain, and Drs. Koichi Kawakami and Yoshiko Takahashi for Tol2 related materials. We also acknowledge Drs. Takayuki Sota, Shinichi Kohsaka, Keiji Wada and M.H.’s lab members for discussions and encouragements. This work was supported by grants from Takeda Science Foundation, Intramural Research Grant (24-12, 26-9, 27-7) for Neurological and Psychiatric Disorders of NCNP and Grant-in-Aid for Scientific

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