Neuroscience Letters 630 (2016) 45–52 Contents lists available at ScienceDirect Neuroscience Letters journal homepage: www.elsevier.com/locate/neulet Research article Survival of corticostriatal neurons by Rho/Rho-kinase signaling pathway Kenta Kobayashi a,b,∗ , Hiromi Sano b,c , Shigeki Kato d , Keisuke Kuroda e , Shinichi Nakamuta e , Tadashi Isa a,b,f , Atsushi Nambu a,b,c , Kozo Kaibuchi e , Kazuto Kobayashi d a Section of Viral Vector Development, National Institute for Physiological Sciences, Okazaki 444-8585, Japan b SOKENDAI (The Graduate University for Advanced Studies), Hayama 240-0193, Japan c Division of System Neurophysiology, National Institute for Physiological Sciences, Okazaki 444-8585, Japan d Department of Molecular Genetics, Institute of Biomedical Sciences, Fukushima Medical University School of Medicine, Fukushima 960-1295, Japan e Department of Cell Pharmacology, Nagoya University Graduate School of Medicine, Nagoya 466-8550, Japan f Department of Developmental Physiology, National Institute for Physiological Sciences, Okazaki 444-8585, Japan h i g h l i g h t s • A novel conditional genetic approach using a dual viral vector system was applied. • C3 transferase or Rho-K DN was conditionally expressed in corticostriatal neurons. • Inhibition of Rho/Rho-kinase signaling caused corticostriatal neuron apoptosis. a r t i c l e i n f o Article history: Received 4 June 2016 Received in revised form 7 July 2016 Accepted 13 July 2016 Available online 15 July 2016 Keywords: Rho GTPase Rho-kinase Corticostriatal neuron Neuronal survival Viral vector a b s t r a c t Developing cortical neurons undergo a number of sequential developmental events including neu- ronal survival/apoptosis, and the molecular mechanism underlying each characteristic process has been studied in detail. However, the survival pathway of cortical neurons at mature stages remains largely uninvestigated. We herein focused on mature corticostriatal neurons because of their important roles in various higher brain functions and the spectrum of neurological and neuropsychiatric disorders. The small GTPase Rho is known to control diverse and essential cellular functions through some effector molecules, including Rho-kinase, during neural development. In the present study, we investigated the role of Rho signaling through Rho-kinase in the survival of corticostriatal neurons. We performed the conditional expression of Clostridium botulinum C3 ADP-ribosyltransferase (C3 transferase) or dominant-negative form for Rho-kinase (Rho-K DN), a well-known inhibitor of Rho or Rho-kinase, respectively, in corticos- triatal neurons using a dual viral vector approach combining a neuron-specific retrograde gene transfer lentiviral vector and an adeno-associated virus vector. C3 transferase markedly decreased the number of corticostriatal neurons, which was attributed to caspase-3-dependent enhanced apoptosis. In addition, Rho-K DN produced phenotypic defects similar to those caused by C3 transferase. These results indicate that the Rho/Rho-kinase signaling pathway plays a crucial role in the survival of corticostriatal neurons. © 2016 Elsevier Ireland Ltd. All rights reserved. Abbreviation: NeuRet vector, neuron-specific retrograde gene transfer lentivi- ral vector; AAV vector, adeno-associated virus vector; C3 transferase, Clostridium botulinum C3 ADP-ribosyltransferase; Rho-K DN, dominant-negative form for Rho- kinase. ∗ Corresponding author at: Section of Viral Vector Development, National Institute for Physiological Sciences, 38 Nishigonaka Myodaiji, Okazaki, Aichi, 444-8585, Japan. E-mail address: kobaya@nips.ac.jp (K. Kobayashi). 1. Introduction Developing cortical neurons undergo a number of sequential developmental events [1–4]. Neuronal survival/apoptosis is one of the characteristic developmental processes, and its molecu- lar machinery has been studied in detail. Neurotrophin signaling through TrkB receptors [5] and serotoninergic neurotransmission [6] are known to be essential for the survival of developing corti- cal neurons. The blockade of NMDA receptors triggers widespread http://dx.doi.org/10.1016/j.neulet.2016.07.020 0304-3940/© 2016 Elsevier Ireland Ltd. All rights reserved.