Disrupted axon-glia interactions at the paranode in myelinated nerves cause axonal degeneration and neuronal cell death in the aged Caspr mutant mouse shambling

Abstract

Emerging evidence suggests that axonal degeneration is a disease mechanism in various neurodegenerative diseases and that the paranodes at the nodes of Ranvier may be the initial site of pathogenesis. We investigated the pathophysiology of the disease process in the central and peripheral nervous systems of a Caspr mutant mouse, shambling (shm), which is affected by disrupted paranodal structures and impaired nerve conduction of myelinated nerves. The shm mice manifest a progressive neurological phenotype as mice age. We found extensive axonal degeneration and a loss of neurons in the central nervous system and peripheral nervous system in aged shm mice. Axonal alteration of myelinated nerves was defined by abnormal distribution and expression of neurofilaments and derangements in the status of phosphorylated and non/de-phosphorylated neurofilaments. Autophagy-related structures were also accumulated in degenerated axons and neurons. In conclusion, our results suggest that disrupted axon-glia interactions at the paranode cause the cytoskeletal alteration in myelinated axons leading to neuronal cell death, and the process involves detrimental autophagy and aging as factors that promote the pathogenesis.

Introduction

In human neurodegenerative diseases, pathophysiological processes and disease mechanisms remain elusive and are often controversial, despite great efforts over the past a few decades (Garden and La Spada, 2012, Liu et al., 2011, Morfini et al., 2009, Yuan et al., 2012). Central efforts have been undertaken during the investigation of these diseases, focusing on studies of protein misfolding, mitochondrial dysfunction, and oxidative stress. Recent progress in neurobiology, however, suggests that axonal degeneration is caused by impaired axonal transport and is a disease substrate in various neurodegenerative diseases such as Alzheimer's disease (AD), Huntington's disease (HD), Parkinson's disease (PD), and amyotrophic lateral sclerosis (ALS) (Lamberts et al., 2015, Millecamps and Julien, 2013, Morfini et al., 2009). Axonal transport is an essential function of the neuron to maintain neuronal integrity and survival of the cell and is achieved through the functions of cytoskeletal and motor proteins (Maday et al., 2014, Millecamps and Julien, 2013). Impaired axonal transport implicates an early manifestation and progression of pathologies in axonal degeneration of the adult-onset neurodegenerative disease (Dadon-Nachum et al., 2011).

Nodes of Ranvier, which are specialized axonal domains in myelinated nerves (Arancibia-Carcamo and Attwell, 2014, Buttermore et al., 2013, Eshed-Eisenbach and Peles, 2013), have been shown to be critical sites for the pathogenesis of various neurodegenerative diseases (Arancibia-Carcamo and Attwell, 2014, Buttermore et al., 2013). Dysfunction and/or disruption of the nodes of Ranvier (node/paranode/juxtaparanode) have been demonstrated extensively (Cifuentes-Diaz et al., 2011, Devaux and Scherer, 2005, Howell et al., 2006, Wolswijk and Balesar, 2003). The paranodes flank both sides of the node and form septate-like adhesive junctions (paranodal junctions) between myelin loops and the axonal membrane. The paranodal junction is composed of contactin and Caspr (contactin-associated protein) on the axonal membrane (axolemma) and the 155-kDa isoform of neurofascin (Nfasc 155) on the glial side.

Previously, we reported that a mutated gene in the neurological mouse mutant shambling (shm) encodes Caspr, a trans-membrane protein (Sun et al., 2009). We found that, in shm mice, paranodal junctions are completely or partially absent from peripheral nervous system (PNS) and central nervous system (CNS) myelinated nerves and that nerve impulse conduction is impaired in both the PNS and CNS (Sun et al., 2009). Other paranodal mutants that are deficient in paranodal and myelin molecules [Caspr KO (Bhat et al., 2001, Einheber et al., 2006, Pillai et al., 2007), contactin KO (Boyle et al., 2001, Davisson et al., 2011), Nfasc 155 KO (Sherman et al., 2005), ceramide galactosyl transferase (CGT) KO (Dupree et al., 1998, Garcia-Fresco et al., 2006), and cerebroside sulfotransferase (CST) KO (Ishibashi et al., 2015)] also lack a normal paranodal junction and display ataxia, motor deficits, and dramatically reduced nerve conduction velocities.

In the course of our last study, we noticed that shm mice exhibited intriguing neurological phenotypes, including symptomatic worsening as the mice age (Sun et al., 2009). Here, we focus on the progressive motor disabilities and the pathological processes involved during the aging of shm mice. Further, we address whether disrupted paranodal junctions cause alterations in axons and neurons themselves. For this purpose, we investigated the pathophysiology of the disease in myelinated nerves and neurons by following the ages of shm mice from infancy to advanced age. We found that disrupted paranodal junctions in shm mice caused cytoskeletal alterations in the axonal cytoplasm of myelinated nerves and led to a loss of neurons. By verifying this pathophysiological process, we propose a causal mechanism underlying neuronal death accompanied by progressive neurological defects in aging shm mice and, consequently, attempt to provide a novel prospective in the study of human neurodegenerative diseases.