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

First published online 16 November 2005
doi: 10.1242/dev.02167

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 Related articles in Development 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 Glasgow, S. M. Articles by Johnson, J. E. Search for Related Content PubMed PubMed Citation Articles by Glasgow, S. M. Articles by Johnson, J. E. Social Bookmarking                         What's this?

Ptf1a determines GABAergic over glutamatergic neuronal cell fate in the spinal cord dorsal horn

¹ Center for Basic Neuroscience, University of Texas Southwestern Medical Center, Dallas, TX 75390, USA
² Molecular Biology Department, University of Texas Southwestern Medical Center, Dallas, TX 75390, USA
³ Department of Cell and Developmental Biology, and Program in Developmental Biology, Vanderbilt University School of Medicine, Nashville, TN 37232, USA

View larger version (108K): [in a new window]   Fig. 1. Ptf1a is in precursors to multiple neural tissues and overlaps with Mash1 in the dorsal neural tube. The Ptf1a lineage was visualized by ß-gal staining of Ptf1aCre/+;R26R-stop-lacZ+/- mice. (A) E11.5 whole mount, showing ß-gal staining in the dorsal neural tube (inset) from the mid-hindbrain to the tail (arrowheads). Staining in the pancreas is indicated by the arrow. (B) brain and spinal cord dissected from an E16.5 embryo, stained for ß-gal, showing expression in cerebellum (arrow) and dorsal spinal cord (arrowhead). Ventral diencephalon also reveals cells from the Ptf1a lineage (inset). (C,D) Adult cervical spinal cord, showing ß-gal stained cells primarily in laminae I-IV. The boxed area in C is shown at higher magnification in D. Arrows (C) highlight labeled cells found ventral to boxed area. (E) Adult cerebellum with ß-gal-stained Purkinje, stellate, basket and Golgi cells. (F) Coronal section of the adult brain reveals ß-gal-stained cells in the ventral hypothalamus. Tissue is counterstained with Nuclear Fast Red (C,D,F) and Nissl (E). Immunofluorescence on transverse neural tube sections of E10.5 (G,H) and E11.5 (I,J) wild-type mice. (G,I) Ptf1a (red), Mash1 (green) and their co-labeling (yellow) in a subset of cells. (H,J) Ptf1a (red) and BrdU incorporation (green) are largely non-overlapping. The asterisk in H indicates a rare BrdU-incorporating Ptf1a cell. Scale bar: 1 mm in A; 50 µm in D-J; 12.5 µm in the insets. ba, basket cells; dI3-dI5, dorsal interneurons 3-5; Go, Golgi cells; MZ, marginal zone; Pu, Purkinje cells; st, stellate cells; VZ, ventricular zone.

View larger version (75K): [in a new window]   Fig. 2. dI4 are trans-fated to dI5 interneurons in Ptf1a-deficient embryos. Immunofluorescence on neural tube transverse sections of Ptf1aCre/+ (A,C) and Ptf1acre/cre (B,D) mouse E10.5 embryos. (A,B) dI4 neurons marked in yellow by co-labeling of Lhx1/5 (red) and Pax2 (green) are lost in the Ptf1a null (arrows). (C,D) Lmx1b (red) labeling dI5 is expanded in Ptf1a null embryos, while Isl1 (green) labeling dI3 are not affected. (E-H) Anti-GFP antibody was used to detect YFP in E10.5 Ptf1aCre;R26R-stop-YFP embryos. YFP acts as a lineage marker for cells that have expressed the Ptf1a locus. YFP (green) co-localizes with the Lhx1/5 (dI4;red), but fails to do so in Ptf1 null embryos (compare E and F). (G) Lmx1b (dI5;red) does not co-localize with YFP in the presence of Ptf1a, but they do co-localize in the null (H). The ventricle is to the left in all panels, as shown in a schematic representation of the markers used to distinguish individual neuronal populations (I). (J) Cell counts for each neuronal population. (K) Summary diagram showing that Ptf1a is required for dI4 and suppression of dI5 interneurons (see Results). Scale bar: 50 µm in A-D; 25 µm in E-H.

View larger version (97K): [in a new window]   Fig. 3. dILA trans-fate to dILB interneurons in Ptf1a null embryos. Immunofluorescence on neural tube transverse sections of Ptf1aCre/+ (A,C) and Ptf1aCre/Cre (B,D) mouse E12.5 embryos. (A,B) dILA neurons, marked by Lhx1/5 (red) and Pax2 (green), are lost specifically in the dorsal neural tube, but not the ventral neural tube in the Ptf1a null (dashed line indicates the position of the ventricle). (C,D) dILB neurons, marked by Lmx1b (red) increase in the Ptf1a null, but there is no change in the number of Isl1 (green) cells. (E-J) Anti-GFP antibody was used to detect YFP at E12.5 in Ptf1aCre;R26R-stop-YFP embryos. YFP acts as a lineage marker for cells that have expressed the Ptf1a locus. (E-F') In the presence of Ptf1a, YFP (green) co-localizes (yellow) with the dILA marker, Lhx1/5 (red), but it does not do so in Ptf1a null embryos. (G-H') Lmx1b (dILB; red) and (I-J) Tlx3 (dILB; red) rarely co-localizes with YFP in embryos with Ptf1a, while substantially more Lmx1b/YFP and Tlx3/YFP double positive cells appear in Ptf1a-deficient embryos (yellow). The boxed regions in E-H are shown at higher magnification in E'-H'. (K) Cell counts of specific neuronal populations. (L) Summary diagram showing that Ptf1a is required for the dILA interneuron cell fate, while suppressing dILB. Scale bar: 50 µm in A-H; 25 µm in E'-H',I,J.

View larger version (113K): [in a new window]   Fig. 4. PTFla is required for generation of dorsal horn GABAergic neurons. Transverse sections through spinal cord cervical regions of Ptf1aCre/+ and Ptf1aCre/Cre mouse E16.5 embryos were processed for mRNA in-situ hybridization with the GABAergic marker gene Gad1 (A,B).Note the complete absence of Gad1 in the dorsal regions in the absence of Ptf1a. Loss of ventral Gad1 was not consistently observed. (C-H) Anti-GFP antibody was used to detect YFP in Ptf1aCre;R26R-stop-YFP E16.5 embryos. YFP acts as a lineage marker for cells that have expressed the Ptf1a locus. (C) The lineage reporter YFP is detected largely in the dorsal regions in wild type. (D) By contrast, in the mutant, YFP is detected at higher levels in more cells and with a different organization (arrowheads). The boxed area in C is the approximate area shown in E-H. Immunofluorescence for GAD67 (E) and the neurotransmitter GABA (G) co-localize with YFP in embryos heterozygous for Ptf1a (arrows) but not in embryos lacking Ptf1a (F,H). Panels E-H are all from dorsal horn regions. Scale bar: 110 µm in A,B; 13 µm in E-H.

View larger version (51K): [in a new window]   Fig. 5. PTFla is required for suppression of dorsal horn glutamatergic neurons. Transverse sections through spinal cord cervical regions of Ptf1aCre/+ and Ptf1aCre/Cre mouse E16.5 embryos were processed for mRNA in-situ hybridization for the glutamatergic marker Vglut2 gene (A,B) or for immunofluorescence for the protein VGLUT2 (C,D). Note the increase in Vglut2 specifically in the dorsal regions in the absence of Ptf1a. The arrows in C,D indicate superficial laminae that have substantial increase in VGLUT2 in the mutant. The dashed line in C,D indicates the midline. (E,F) Anti-GFP antibody was used to detect YFP in Ptf1aCre;R26R-stop-YFP E16.5 embryos. VGLUT2 and YFP seldom co-localize in embryos containing Ptf1a (E); however, co-localization of VGLUT2 and YFP is detected in distal processes in the Ptf1a null (F). The density of Tlx3+ cells is increased in dorsal regions in the mutant (G,H). (E-H) are all from dorsal horn regions indicated by the box in C. (I) A model for the role of Ptf1a in GABAergic and glutamatergic neurons in the dorsal horn. Ptf1a acts in two ways: determination of GABAergic neurons by inducing Pax2, which is required for the expression of the GABAergic phenotype in these cells, and suppressing Tlx3, which is required for specifying glutamatergic neurons by inducing glutamatergic specific genes and suppressing Pax2 (Cheng et al., 2004). Scale bar: 110 µm in A,B; 50 µm in C,D; 13 µm in E-H.

CiteULike

Complore

Connotea

Del.icio.us

Digg

Reddit

Technorati

Twitter