Neutralization of Inhibitory Molecule NG2 Improves Synaptic Transmission, Retrograde Transport, and Locomotor Function after Spinal Cord Injury in Adult Rats

Abstract

NG2 belongs to the family of chondroitin sulfate proteoglycans that are upregulated after spinal cord injury (SCI) and are major inhibitory factors restricting the growth of fibers after SCI. Neutralization of NG2's inhibitory effect on axon growth by anti-NG2 monoclonal antibodies (NG2-Ab) has been reported. In addition, recent studies show that exogenous NG2 induces a block of axonal conduction. In this study, we demonstrate that acute intraspinal injections of NG2-Ab prevented an acute block of conduction by NG2. Chronic intrathecal infusion of NG2-Ab improved the following deficits induced by chronic midthoracic lateral hemisection (HX) injury: (1) synaptic transmission to lumbar motoneurons, (2) retrograde transport of fluororuby anatomical tracer from L5 to L1, and (3) locomotor function assessed by automated CatWalk gait analysis. We collected data in an attempt to understand the cellular and molecular mechanisms underlying the NG2-Ab-induced improvement of synaptic transmission in HX-injured spinal cord. These data showed the following: (1) that chronic NG2-Ab infusion improved conduction and axonal excitability in chronically HX-injured rats, (2) that antibody treatment increased the density of serotonergic axons with ventral regions of spinal segments L1-L5, (3) and that NG2-positive processes contact nodes of Ranvier within the nodal gap at the location of nodal Na ϩ channels, which are known to be critical for propagation of action potentials along axons. Together, these results demonstrate that treatment with NG2-Ab partially improves both synaptic and anatomical plasticity in damaged spinal cord and promotes functional recovery after HX SCI. Neutralizing antibodies against NG2 may be an excellent way to promote axonal conduction after SCI.

Combination of chondroitinase ABC and AAV-NT3 promotes neural plasticity at descending spinal pathways after thoracic contusion in rats

“…The choice of this dual treatment was based on the knowledge that accumulation of chondroitin sulfate proteoglycans (CSPGs;Dou and Levine 1994;Galtrey and Fawcett 2007;Silver and Miller 2004;Snow et al 1990) and insufficient neurotrophin support (Arvanian 2013;Fortun et al 2009;Grill et al 1997;Mendell et al 2001;Schnell et al 1994) are major reported obstacles known to restrict better recovery after SCI. Degradation of CSPGs with the enzyme ChABC was shown to promote sprouting of fibers following cervical crush (Barritt et al 2006;Bradbury et al 2002), hemisection (Houle et al 2006Iseda et al 2008), transection (Fouad et al 2005, and contusion (Caggiano et al 2005;Tom et al 2009, but see Iseda et al 2008 injuries.…”

“…Transverse sections (40 m) of T 6 -T 8 were used for reconstructions of acute lesions and L 1 -L 5 for 5-HT immunostaining. Cresyl violet staining was used to determine spinal cord segmental levels and reconstructions of injury and acute lesions as previously described (Arvanian et al 2006;Petrosyan et al 2013). For immunofluorescence staining, sections were washed with 0.3% Triton X-100 in PBS, blocked with 5% normal goat serum in 0.3% Triton X-100 in PBS for 2 h at room temperature, and then incubated in appropriate primary antibodies: rabbit anti-5-HT [1:1,000, ImmunoStar for serotonin (5-HT) immunoreactivity] or mouse antibody to chondroitinase-generated C-4-S (2B6, 1:200, Mdbioproducts) overnight at 4°C.…”

“…Thresholded pixel mean area was calculated by eliminating the background and quantifying 5-HT-positive fibers (Petrosyan et al 2013;Tom et al 2009) within a fixed area of dorsal and ventral horns, respectively (see Fig. 3A), using ImageJ software (National Institutes of Health).…”

“…Gait parameters, such as regularity index and base of support, were collected and compared between groups. These parameters have been reported as objective measurements of locomotor performance and coordination after SCI (Koopmans et al 2005;Petrosyan et al 2013).…”

“…After completion of electrophysiological recordings, cross sections at lumbar L 1 -L 5 segments were immunostained for serotonergic fibers and immunoreactivity was quantified in the dorsal and ventral horns. We measured 5-HT immunoreactivity as thresholded pixel area, as described previously (Petrosyan et al 2013;Tom et al 2009). As shown in Fig.…”