Viral Delivery of NR2D Subunits Reduces Mg²⁺Block of NMDA Receptor and Restores NT-3-Induced Potentiation of AMPA-Kainate Responses in Maturing Rat Motoneurons

Abstract

. Viral delivery of NR2D subunits reduces Mg 2ϩ block of NMDA receptor and restores NT-3-induced potentiation of AMPA/kainate responses in maturing rat motoneurons. J Neurophysiol 92: 2394 -2404, 2004. First published May 19, 2004 10.1152 10. /jn.00278.2004 responsiveness of motoneurons declines during the initial 2 postnatal weeks due to increasing Mg 2ϩ block of NMDA receptors. Using gene chip analyses, RT-PCR, and immunochemistry, we have shown that the NR2D subunit of the NMDA receptor (NMDAR), known to confer resistance to Mg 2ϩ block, also declines in motoneurons during this period. We injected a viral construct (HSVnr2d) into the lumbar spinal cord on postnatal day 2 in an attempt to restore NMDAR function in motoneurons during the second postnatal week. Following HSVnr2d injection, we detected elevated levels of NR2D mRNA in spinal cord samples and NR2D protein specifically in motoneurons. These molecular changes were associated with marked functional alterations whereby NMDAR-mediated responses in motoneurons associated with both dorsal root (DR) and ventrolateral funiculus (VLF) inputs returned to values observed at E18 due to decreased Mg 2ϩ blockade. Viruses carrying the ␤-galactosidase gene did not induce these effects. NT-3 is known to potentiate AMPA-kainate responses in motoneurons if the response has an NMDAR-mediated component and thus is normally ineffective during the second postnatal week. Restoration of NMDAR-mediated responsiveness in the second postnatal week was accompanied by a return of the ability of neurotrophin-3 (NT-3) to potentiate the AMPA-kainate responses produced by both DR and VLF synaptic inputs. We conclude that delivery of the gene for a specific NMDA subunit can restore properties characteristic of younger animals to spinal cord motoneurons. This approach might be useful for enhancing the function of fibers surviving in the damaged spinal cord.

Changes in Motoneuron Properties and Synaptic Inputs Related to Step Training after Spinal Cord Transection in Rats

“…Considering first the strengthening of the sEPSP, it is known that muscle spindle afferents are very active during stepping movements (Loeb and Duysens, 1979;Prochazka et al, 1979) and that strengthening of synaptic connections can occur via synchronized firing of the presynaptic and postsynaptic neurons (Gustafsson et al, 1987). This strengthening involves the action of NMDA receptors known to be functional on motoneurons, particularly in neonates (Ziskind-Conhaim, 1990;Arvanian et al, 2004). Furthermore, motoneuron NMDA receptor activation is enhanced by administration of the neurotrophins NT-3 and BDNF (Arvanov et al, 2000;Arvanian and Mendell, 2001), both of which are elevated in hindlimb muscle and/or spinal cord by step training (Gomez-Pinilla et al, 2001; Hutchinson et al, 2004).…”

Chondroitinase ABC Combined with Neurotrophin NT-3 Secretion and NR2D Expression Promotes Axonal Plasticity and Functional Recovery in Rats with Lateral Hemisection of the Spinal Cord

“…HSV-1-gfp amplicons encoding NR2D or control ␤-galactosidase, were injected intraspinally (2 injections of 1 l each; ϳ10 4 viral particles/l) into the left and right ventral horn just caudal to the injury region. HSV-1 amplicons have a transgene capacity sufficient to carry the NR2D cDNA and the coexpressed GFP reporter gene (Arvanian et al, 2004). HSV-1-gfpmediated gene expression peaks during the first 24 -48 h (Bowers et al, 2000(Bowers et al, , 2001.…”

“…The motoneuron infection was determined by using double immunolabeling with peripherin (motoneurons marker, green) and with GFP (red). The immunolabeling procedure and analyses have previously been described (Arvanian et al, 2004). We found a substantial level of infection of motoneurons in the thoracic cord and even at a remote lumbar level, measured 7 d after injection of HSV-NR2D-gfp.…”

“…Initiation of this neurotrophininduced plasticity may require functional NMDA receptors in motoneurons and ceases shortly after birth, at which time NR2D expression is downregulated at these synapses (Arvanian et al, 2004). In later postnatal animals the initial plasticity in response to NT3 can be restored if Mg 2ϩ is depleted or by injecting viruses expressing the NR2D gene (Arvanian et al, 2004).…”

“…Previous studies in newborn animals have shown that the strength of the excitatory synaptic connection from VLF axons to lumbar motoneurons can be enhanced by NT3 (Arvanian et al, 2003), but novel polysynaptic connections required to provide detours around lesions are not . Enhancing expression of the NR2D subunit of NMDA receptor, normally downregulated in neonatal rats (Arvanian et al, 2004), promotes the appearance of novel polysynaptic connections in neonates .…”