A SAGE-based screen for genes expressed in sub-populations of neurons in the mouse dorsal root ganglion

<p>Abstract</p> <p>Background</p> <p>The different sensory modalities temperature, pain, touch and muscle proprioception are carried by somatosensory neurons of the dorsal root ganglia. Study of this system is hampered by the lack of molecular markers for many of these neuronal sub-types. In order to detect genes expressed in sub-populations of somatosensory neurons, gene profiling was carried out on wild-type and TrkA mutant neonatal dorsal root ganglia (DRG) using SAGE (serial analysis of gene expression) methodology. Thermo-nociceptors constitute up to 80 % of the neurons in the DRG. In TrkA mutant DRGs, the nociceptor sub-class of sensory neurons is lost due to absence of nerve growth factor survival signaling through its receptor TrkA. Thus, comparison of wild-type and TrkA mutants allows the identification of transcripts preferentially expressed in the nociceptor or mechano-proprioceptor subclasses, respectively.</p> <p>Results</p> <p>Our comparison revealed 240 genes differentially expressed between the two tissues (P < 0.01). Some of these genes, CGRP, Scn10a are known markers of sensory neuron sub-types. Several potential markers of sub-populations, Dok4, Crip2 and Grik1/GluR5 were further analyzed by quantitative RT-PCR and double labeling with TrkA,-B,-C, c-ret, parvalbumin and isolectin B4, known markers of DRG neuron sub-types. Expression of Grik1/GluR5 was restricted to the isolectin B4+ nociceptive population, while Dok4 and Crip2 had broader expression profiles. Crip2 expression was however excluded from the proprioceptor sub-population.</p> <p>Conclusion</p> <p>We have identified and characterized the detailed expression patterns of three genes in the developing DRG, placing them in the context of the known major neuronal sub-types defined by molecular markers. Further analysis of differentially expressed genes in this tissue promises to extend our knowledge of the molecular diversity of different cell types and forms the basis for understanding their particular functional specificities.</p

Collagen XXVIII Is a Distinctive Component of the Peripheral Nervous System Nodes of Ranvier and Surrounds Nonmyelinating Glial Cells

Growing evidence indicates that collagens perform crucial functions during the development and organization of the nervous system. Collagen XXVIII is a recently discovered collagen almost exclusively expressed in the peripheral nervous system (PNS). In this study, we show that this collagen is associated with nonmyelinated regions of the PNS. With the notable exception of type II terminal Schwann cell in the hairy skin, collagen XXVIII surrounds all nonmyelinating glial cells studied. This includes satellite glial cells of the dorsal root ganglia, terminal Schwann cells type I around mechanoceptors in the skin, terminal Schwann cells around proprioceptors in the muscle spindle or at the neuromuscular junction and olfactory ensheathing cells. Collagen XXVIII is also detected at nodes of Ranvier where the myelin sheath of myelinated fibers is interrupted and is thus a distinctive component of the PNS nodal gap. The correlation between the absence of myelin and the presence of collagen XXVIII is confirmed in a mouse model of Charcot-Marie-Tooth characterized by dysmyelinated nerve fibers, in which enhancement of collagen XXVIII labeling is observed. (C) 2010 Wiley-Liss, Inc

Collagen XVI is a neural component of the developing and regenerating dorsal root ganglia extracellular matrix.

International audienceCollagen XVI is a homotrimeric molecule harbouring similarities to the FACIT (fibril-associated collagens with interrupted triple helices) family of collagens (Grassel et al., 1996). Collagen XVI is expressed in skin and cartilage where it is integrated into tissue specific aggregates (Grassel et al., 1999; Kassner et al., 2003). In the nervous system, collagen XVI has been detected at lowlevel in the brain and a strong expression was also reported in spinal root fibers during development (Lai and Chu, 1996). In dorsal root ganglia (DRG), analysis of SAGE banks performed by our group during development and after nerve injury (Mechaly et al., 2006) shows a fluctuation of collagen XVI expression between the different conditions and prompted us to study it further. DRGs contain the cell bodies of neurons, the axons of which transmit sensory information from the periphery to the central nervous system. While it is well known that during development and regeneration, neurites require extracellular matrix molecules for growth and guidance (Hari et al., 2004), the composition and the role of the matrix surrounding neurons in the ganglia itself have solicited little interest. Here, we show that collagen XVI is a component of the developingDRG extracellular matrix, that following nerve injury, its expression is increased around neuronal cell bodies and that neurons express collagen XVI in the peripheral nervous system

Double in situ hybridization was carried out by using fluorescein/fast red detection for TrkA (A, E), TrkB (B, F), TrkC (C, G), c-ret (D, H) and DIG/NBT-BCIP for Dok4 (A, B, C, D) and Crip2 (E, F, G, H)

<p><b>Copyright information:</b></p><p>Taken from "A SAGE-based screen for genes expressed in sub-populations of neurons in the mouse dorsal root ganglion"</p><p>http://www.biomedcentral.com/1471-2202/8/97</p><p>BMC Neuroscience 2007;8():97-97.</p><p>Published online 19 Nov 2007</p><p>PMCID:PMC2241628.</p><p></p> In situ signals were converted into pseudo colors and images were superimposed to show co-labelling of cells. Dok4 co-localised with all major subtypes of DRG neurons, Crip2 was specifically excluded from TrkC population. Scale bar 50 μm

Gene expression determined by real-time PCR on P0 and P0 TrkA mutant mouse lumbar DRG

<p><b>Copyright information:</b></p><p>Taken from "A SAGE-based screen for genes expressed in sub-populations of neurons in the mouse dorsal root ganglion"</p><p>http://www.biomedcentral.com/1471-2202/8/97</p><p>BMC Neuroscience 2007;8():97-97.</p><p>Published online 19 Nov 2007</p><p>PMCID:PMC2241628.</p><p></p> TrkA and Ube2e3 (ubiquitin-conjugating enzyme E2E 3) were used as controls. Data (means ± SEM) were calculated by the delta-CT method [37] on three independent experimental replicates. The arithmetic means of the expression levels of two genes (Polr2j, Ddx48) whose expression do not change in the course of development and in TrkA-/- DRG were used to normalize the expression levels. Data were analyzed using the Mann Whitney U-test (*P < 0.05). ND: Not detected