QUICK SEARCH:   [advanced] Author: Keyword(s):
Home     Help     Feedback     Subscriptions     Archive     Search     Table of Contents

doi: 10.1242/10.1242/dev.00213

This Article Summary Full Text Full Text (PDF) Alert me when this article is cited Alert me if a correction is posted Services Email this article to a friend Similar articles in this journal Similar articles in PubMed Alert me to new issues of the journal Download to citation manager Citing Articles Citing Articles via HighWire Citing Articles via Google Scholar Google Scholar Articles by Luo, R. Articles by Henion, P. D. Search for Related Content PubMed PubMed Citation Articles by Luo, R. Articles by Henion, P. D. Social Bookmarking                         What's this?

Molecular identification of distinct neurogenic and melanogenic neural crest sublineages

¹ Neurobiotechnology Center and Department of Neuroscience, Ohio State University, 105 Rightmire Hall, 1060 Carmack Road, Columbus, OH 43210, USA
² Department of Pathology, Centre Medical Universitaire 1, Rue Michel-Servet, 1211 Geneva 4, Switzerland

View larger version (67K): [in a new window]   Fig. 1. Expression of TrkC and C-Kit during neural crest development in vitro. (A,B) A subset of live cells present in a population of cells 4 hours after the initiation of segregation from neural tube explants. Two cells (arrows) were expressing TrkC as revealed by immunolabeling (A) while the rest of the cells (B) are TrkC-. (C,D) A C-Kit-expressing neural crest cell (C, arrowhead) in a field of C-Kit- (D) cells present in a live 13-16 hour population. (E,F) A double immunolabeled 13-16 hour population of neural crest cells shown after fixation, stained with Hoechst nuclear dye (E) and by phase contrast (F). TrkC-expressing cells (arrows, red) and C-Kit-expressing cells (arrowheads, green) are present in distinct subpopulations of neural crest cells. (G,H) A differentiated (108 hour) culture immunolabeled live with TrkC antiserum (red), fixed and labeled with 16A11 to reveal neurons (green). TrkC is expressed by a subpopulation of neurons (G, yellow) and is not expressed by non-neuronal cells (G,H). (I,J) A region of a live differentiated culture immunolabeled with anti-C-Kit (I, green). A phase-contrast image of the same field shows that the immunolabeled cells are melanocytes (J), whereas unpigmented cells are C-Kit-. Scale bar: 8 µm.

View larger version (32K): [in a new window]   Fig. 2. Experimental design for clonal analysis of molecularly identified cells. (Top) Because we were able to molecularly identify distinct neural crest populations in live cultures (see Fig. 1), we were able to perform clonal analysis of these cells by microinjection of lineage dye (photomicrograph). The lineage dye is inherited by all clonal descendants of the labeled precursor, which permits them to be distinguished in differentiated cultures and phenotypically analyzed using cell type-specific markers. (Bottom) The time periods at which we immunolabeled and injected individual receptor-expressing cells are expressed relative to the initiation of segregation of neural crest cells from neural tube explants (see Results, and Materials and Methods). We labeled individual TrkC-expressing cells in 1-6 and 13-16 hour populations. We labeled C-Kit-expressing cells in 13-16 and 30-36 hour populations. The initial overt differentiation of melanocytes (melanization) does not begin until 54 hours and initial overt neuronal differentiation begins at 72 hours. `Complete' differentiation (>95% of all cells) of the cultures occurs by 108 hours.

View larger version (97K): [in a new window]   Fig. 3. Examples of clones generated by fate-restricted neural crest cells. (A,B) Images of the same field of cells in a differentiated (108 hour) culture. This clone was derived from a single TrkC-expressing neural crest cell labeled during the 13-16 hour time period. The four members of the clone were identified by fluorescent lineage dye (arrows, A) and were found to be neurons by 16A11 immunoreactivity (arrows, B). (C,D) A two-cell clone in a differentiated culture derived from a C-Kit-expressing neural crest cell labeled during the 13-16 hour period. The two members of the clone (arrowheads, C) were melanocytes, as they contained melanin granules (arrowheads, D, phase-contrast). Scale bars: 25 µm in B; 12 µm in D.

CiteULike

Complore

Connotea

Del.icio.us

Digg

Reddit

Technorati

Twitter