Evidence for the presence of both peroxisome proliferator-activated receptors alpha and beta in the rat spinal cord

Abstract

Peroxisome proliferator-activated receptors (PPARs) are ligand-activated transcription factors belonging to the nuclear receptor superfamily. Different subtypes of PPARs (α, β, and γ) have been described. Their distinct physiological functions depend on their differential ligand activation profiles but also on their specific tissue expression. Previous studies have described their presence in the central nervous system. However, their expression in the adult rat spinal cord in normal physiological conditions has never been investigated. We demonstrated by using reverse-transcription-polymerase chain reaction, and Western blotting, the mRNA and protein expression of PPARα and PPARβ, but not PPARγ in cervical, thoracic, and lumbar segments of the spinal cord. Using immunohistochemistry, we also showed for the first time the specific cellular distribution of these transcription factors in the different segments of the spinal cord. In the gray matter, the distribution of PPARα was homogenous whereas PPARβ was specifically localized in motoneurons and in medial part of laminae IV, V, VI, VII, VIII, and X. These latter areas are known as nociceptive afferent pathways to supra-spinal structures such as the medulla reticular nucleus and the thalamus. In the white matter, PPARα was localized exclusively in astrocytes while PPARβ was present in oligodendrocytes. The possible functions of PPARα and PPARβ expressed in both white and gray matters of the spinal cord will be discussed but need further studies.

Introduction

The peroxisome proliferator-activated receptors (PPARs) are transcription factors belonging to the nuclear receptor superfamily (Issemann and Green, 1990). Three isotypes of PPARs have been principally described, known as α (NR1C1),? β also called δ (NR1C2), and γ (NR1C3) (Dreyer et al., 1992, Sher et al., 1993, Kliewer et al., 1994). The latter exists as two isoforms γ1 and γ2 resulting from alternative promoter usage and differential splicing (Zhu et al., 1995). PPARs exhibit a variable tissue distribution. PPARα is mainly expressed in the liver, kidney and white adipose tissue, which exhibit high fatty acid metabolism and high peroxisome dependent activities (Kliewer et al., 1994, Mukherjee et al., 1994, Auboeuf et al., 1997). In contrast, PPARβ isoform is ubiquitously distributed (Braissant et al., 1996). PPARγ is mainly expressed in the spleen and adipose tissue (Chawla et al., 1994).

In the central nervous system (CNS), previous studies on PPAR distribution mainly concerned the brain (Kainu et al., 1994, Xing et al., 1995, Braissant et al., 1996, Krémarik-Bouillaud et al., 2000), and to a lesser extent the brainstem (Braissant et al., 1996). In the CNS, PPARβ has been principally associated with neurons of anatomically well-defined regions (Kainu et al., 1994, Xing et al., 1995, Braissant et al., 1996, Krémarik-Bouillaud et al., 2000), whereas PPARα was found in neurons and oligodendrocytes of the corpus callosum (Kainu et al., 1994). The precise localization of PPARs in the white matter was also investigated on cell cultures. Earlier studies have shown that PPARβ is strongly expressed in immature cortical oligodendrocytes (Granneman et al., 1998), and in both neonatal cerebellar and neonatal or adult cortical astrocytes (Cullingford et al., 1998). The expression of PPARα and PPARγ is rather low in astrocytes (Cullingford et al., 1998). Previous studies have also shown that PPARγ was constitutively expressed in rat primary microglial cultures (Bernardo et al., 2000).

Even if PPARα and PPARβ have been described in the rat brain spinal cord during embryonic development from E8.8 to E18.5 (Braissant and Wahli, 1998), the distribution of PPARs in the adult rat spinal cord has not yet been explored. In the CNS, the role of PPARs in the lipid metabolism has been suggested, as a concomitant distribution of acyl CoA synthetase 2 and PPARβ in reaggregated rat brain cell culture has been shown (Basu-Modak et al., 1999). On the other hand, the presence of PPARβ in primary and enriched cultures from neonatal mouse brain oligodendrocytes suggests that this nuclear receptor participates in the differentiation of oligodendrocytes, which are major lipid-producing cells implicated in myelinogenesis (Saluja et al., 2001). Accordingly, PPARβ-null mice display altered myelination of the CNS (Peters et al., 2000). Concerning PPARγ its expression in the brain could be related to modulation of inflammation. Actually, expression of PPARγ has been shown to be increased in Alzheimer's disease brains where inflammatory events are associated with senile plaque formation (Kitamura et al., 1999b). It was also shown that sustained administration of PPARγ agonists reduces the expression of the inducible form of nitric oxide synthase and cell death in the cerebellum (Heneka et al., 2000) or induced neuronal apoptosis (Rohn et al., 2001). Moreover, the expression of PPARγ in the microglia is down-regulated during the activation of these cells (Bernardo et al., 2000). Some PPARγ ligands also reduce NO production and the expression of iNOS in microglial cell cultures stimulated by the endotoxin (Bernardo et al., 2000). To date, no precise function has been postulated for PPARα in the CNS.

In this study, we analyzed the expression of PPARα, β, and γ in the CNS, particularly in the spinal cord by RT-PCR and Western blotting. The precise localization of PPARs should help to clarify their physiological roles. This analysis was also undertaken by immunohistochemistry in the gray and white matter, from cervical to lumbar segments of the spinal cord.

Our results demonstrate the presence of PPARα and PPARβ in the adult rat spinal cord in normal physiological conditions, and we show for the first time that the specific distribution of PPARβ is related to the ascending nociceptive pathways. These results raise the possibility that PPARβ should play a modulating role in pain transmission.