LXR and TSPO as new therapeutic targets to increase the levels of neuroactive steroids in the central nervous system of diabetic animals Nico Mitro a , Gaia Cermenati a , Silvia Giatti b , Federico Abbiati a , Marzia Pesaresi b , Donato Calabrese b , Luis Miguel Garcia-Segura c , Donatella Caruso a , Roberto C. Melcangi b,⇑ a Dept. of Pharmacological Sciences, Università degli Studi di Milano, Milano, Italy b Dept. of Endocrinology, Pathophysiology and Applied Biology – Center of Excellence on Neurodegenerative Diseases, Università degli Studi di Milano, Milano, Italy c Instituto Cajal, C.S.I.C., Madrid, Spain article info Article history: Available online 3 March 2012 Keywords: Streptozotocin Liquid chromatography-tandem mass spectrometry Central nervous system Steroid level Cholesterol Rat abstract Neuroactive steroid levels are decreased in the central nervous system (CNS) of streptozotocin (STZ) dia- betic rats. In agreement, they exert protective effects in this experimental model, counteracting degener- ative events occurring in the CNS. Therefore, an interesting therapeutic strategy could be to increase their levels directly in the CNS. In this study we have evaluated whether activation of translocator protein- 18 kDa (TSPO) or liver X receptors (LXRs) may affect the levels of neuroactive steroids present in the CNS of diabetic and non-diabetic animals. We observed that the treatment with either Ro5-4864 (i.e., a ligand of TSPO) or with GW3965 (i.e., a ligand of LXRs) induced an increase of neuroactive steroids in the spinal cord, the cerebellum and the cerebral cortex of STZ-rats, but not in the CNS of non-patho- logical animals. Interestingly, the pattern of induction was different among the three CNS areas analyzed and between the two pharmacological tools. In particular, the activation of LXRs might represent a prom- ising neuroprotective strategy, because the treatment with GW3965, at variance to Ro5-4864 treatment, did not induce significant changes in the plasma levels of neuroactive steroids. This suggests that activa- tion of LXRs may selectively increase the CNS levels of neuroactive steroids avoiding possible endocrine side effects exerted by the systemic treatment with these molecules. Interestingly GW3965 treatment induced an increase of dihydroprogesterone in the spinal cord of diabetic animals in association with an increase of myelin basic protein expression. Thus we demonstrated that LXR activation was able to rescue CNS symptoms of diabetes. Ó 2012 Elsevier Ltd. All rights reserved. 1. Introduction The finding that neuroactive steroids, such as progesterone (PROG) and its derivatives, dihydroprogesterone (DHP) and tetrahy- droprogesterone (THP), testosterone and its derivatives, dihydrotes- tosterone (DHT) and 5a-androstane-3a,17b-diol (3a-diol), dehydroepiandrosterone (DHEA) and estrogens, are neuroprotec- tive agents in central and peripheral nervous system is now a consol- idated concept. Indeed, in experimental models of Alzheimer’s disease, Parkinson’s disease, multiple sclerosis, traumatic brain in- jury, stroke, autism, schizophrenia, mood disorders and peripheral neuropathy, these molecules have been observed to exert neuropro- tective effects (Melcangi and Garcia-Segura, 2010; Melcangi and Panzica, 2009; Schumacher et al., 2007). Nervous system is not only a target for the action of neuroactive steroids, but it is also able to synthesize and to metabolize them (Melcangi et al., 2008). There- fore, an interesting therapeutic strategy could be to increase the lev- els of neuroactive steroids directly in the nervous system. This therapeutic approach could be extremely interesting because it may avoid possible endocrine side effects exerted by the systemic treatment with neuroactive steroids. With this perspective, ligands of translocator protein-18 kDa (TSPO) may represent a promising option. TSPO is mainly present in the mitochondrial outer mem- brane, where it promotes the translocation of cholesterol to the in- ner mitochondrial membrane, allowing the transformation of cholesterol into pregnenolone (PREG) and the increase of steroid levels (Papadopoulos et al., 1997, 2006a,b). We recently analyzed whether treatment with a TSPO ligand, such as Ro5-4864, increases neuroactive steroid levels in sciatic nerve of diabetic animals and ex- erts neuroprotective effects. Data obtained by liquid chromatogra- phy–tandem mass spectrometry (LC–MS/MS) show that the treatment with Ro5-4864 is able to significantly stimulate the low levels of pregnenolone (PREG), PROG and DHT observed in the sciatic nerves of diabetic rats and to counteract neurophysiological, 0197-0186/$ - see front matter Ó 2012 Elsevier Ltd. All rights reserved. doi:10.1016/j.neuint.2012.02.025 ⇑ Corresponding author. Address: Dept. of Endocrinology, Pathophysiology and Applied Biology – Center of Excellence on Neurodegenerative Diseases, Università degli Studi di Milano, Via Balzaretti 9, 20133 Milano, Italy. Tel.: +39 02 50318238; fax: +39 02 50318204. E-mail address: roberto.melcangi@unimi.it (R.C. Melcangi). Neurochemistry International 60 (2012) 616–621 Contents lists available at SciVerse ScienceDirect Neurochemistry International journal homepage: www.elsevier.com/locate/nci