Ribosomal Distributions in Axons of Mammalian Myelinated Fibers Alejandra Kun, 1,2 Leonardo Otero, 1 Jose ´ R. Sotelo-Silveira, 1,2 and Jose ´ R. Sotelo 2 * 1 Departamento de Biologı ´a Celular y Molecular, Facultad de Ciencias, Universidad de la Repu ´blica, Montevideo, Uruguay 2 Departamento de Proteı ´nas y A ´ cidos Nucleicos, Instituto de Investigaciones Biolo ´gicas Clemente Estable, Ministerio de Educacio ´n y Cultura, Montevideo, Uruguay The distribution of ribosomes and polysomes in unin- jured myelinated axons of rat sciatic nerve was ana- lyzed. Ribosomes were identified by immunocytochem- istry at the light and electron microscopic levels. A polyclonal antibody developed against ribosomes rec- ognized both rRNA and ribosomal proteins. The distri- bution of the immunoreaction product was similar to that obtained with human anti-ribosomal P protein. The immunoreaction product distributions were of two types in axons: 1) periodic localization in the cortical region of axoplasm that appeared as a compact structural ag- gregate, consistent with that described as a periaxo- plasmic ribosomal plaques (PARP) domain (Koenig et al. [2000] J. Neurosci. 20:8390–8400), and 2) scattered small immuno-reactive clusters of varying sizes (RNP) within the central core of the axon. The latter observa- tion suggested the possibility that RNP-like particles could be associated with the axonal transport system and in transit. Immunoreaction product was also asso- ciated with a novel structural inclusion, possibly multi- vesicular in makeup that was located in the axon and at the myelin-axon interface, and visible at the light and EM levels. The potential significance of this structural peculiarity is considered. V V C 2007 Wiley-Liss, Inc. Key words: myelinated axons; RNPs; anti-ribosome antibody Although there were early reports of ribosomes in immature axons (Bunge, 1973; Skoff and Hamberger, 1974; Tennyson, 1965, 1970), the earliest reports of ribosomes in adult myelinated fibers were described as ‘‘ribosome-like bodies’’ in proximal axons near the axon hillock region of dorsal root ganglion cells (Ze ´le ´na, 1970, 1972). Pannese and Ledda (1991) showed signifi- cant longitudinal cortical distributions of ribosomes in several myelinated sensory nerve root fibers by means of systematic serial sectioning. Studies of invertebrate axons, using electron spectroscopic imaging (ESI) to image and map ribosomal and RNP phosphorus signals, provided direct supporting evidence of ribosomal distributions in unmyelinated axons (Martin et al., 1989; Giuditta et al., 1991; Crispino et al., 1997). Similar ultrastructural ESI analysis was carried out on axoplasmic whole mounts isolated from the large goldfish myelinated Mauthner fiber (Koenig and Martin, 1996) and from ordinary mammalian myelinated spinal root fibers (Koenig et al., 2000). The latter studies showed that ribosomes and poly- somes were associated with a structural matrix localized to discrete domains that were distributed at intermittent intervals along the periphery of axoplasm. The periodic novel structural formations, rich in actin, were called periaxoplasmic ribosomal plaques (PARP). Many studies have described various components of translational machinery and activity in axons (Alvarez et al., 2000; Giuditta, et al., 2002; Piper and Holt, 2004). Although PARP are apparently absent in non- myelinated axons, including growing axons, RNP par- ticles are distributed in random fashion (Knowles et al., 1996; Bleher and Martin, 2001; Lee and Hollenbeck, 2003). In cultured neurons obtained from human medulloblastoma (P19 cells), a subset of mRNAs was shown to be associated with microtubules. These RNP particles (ELAV complex) form a translational supra complex associated with polysomes and an actin network (Antic and Keene, 1998). RNP particles in axons and growth cones of cultured P19 neurons were shown to contain TAU mRNA (Aronov et al., 2001), HuD (an RNA binding protein), and KIF3A (a kinesin isoform) (Aronov et al., 2002). The presence of HuD associated with these axonal granules suggested that such granules may have a translational role whereas KIF3A could be involved in their microtubule-based transport. Indeed, TAU mRNA was not only present but was shown to be Contract grant sponsor: CSIC-Universidad de la Repu ´blica; Contract grant sponsor: OAS; Contract grant sponsor: PEDECIBA; Contract grant sponsor: MEyC; Contract grant sponsor: JICA; Contract grant sponsor: DAAD (Germany); Contract grant sponsor: CONICyT. *Correspondence to: Dr. Jose ´ Sotelo, Departamento de Proteı ´nas y A ´ cidos Nucleicos, Instituto de Investigaciones Biolo ´ gicas Clemente Esta- ble, Ministerio de Educacio ´n y Cultura, Av. Italia 3318, CP 11600. Montevideo, Uruguay. E-mail: sotelo@iibce.edu.uy Received 19 December 2006; Revised 23 January 2007, 18 February 2007; Accepted 26 February 2007 Published online 22 May 2007 in Wiley InterScience (www. interscience.wiley.com). DOI: 10.1002/jnr.21340 Journal of Neuroscience Research 85:2087–2098 (2007) ' 2007 Wiley-Liss, Inc.