Autonomic and motor neuron death is progressive and parallel in a lumbosacral ventral root avulsion model of cauda equina injury

Abstract

Injuries to the cauda equina of the spinal cord result in autonomic and motor neuron dysfunction. We developed a rodent lumbosacral ventral root avulsion injury model of cauda equina injury to investigate the lesion effect in the spinal cord. We studied the retrograde effects of a unilateral L5-S2 ventral root avulsion on efferent preganglionic parasympathetic neurons (PPNs) and pelvic motoneurons in the L6 and S1 segments at 1, 2, 4, and 6 weeks postoperatively in the adult male rat. We used Fluoro-Gold-prelabeling techniques, immunohistochemistry, and quantitative stereologic analysis to show an injury-induced progressive and parallel death of PPNs and motoneurons. At 6 weeks after injury, only 22% of PPNs and 16% of motoneurons remained. Furthermore, of the neurons that survived at 6 weeks, the soma volume was reduced by 25% in PPNs and 50% in motoneurons. Choline acetyltransferase (ChAT) protein was expressed in only 30% of PPNs, but 80% of motoneurons remaining at 1 week postoperatively, suggesting early differential effects between these two neuronal types. However, all remaining PPNs and motoneurons were ChAT positive at 4 weeks postoperatively. Nuclear condensation and cleaved caspase-3 were detected in axotomized PPNs and motoneurons, suggesting apoptosis as a contributing mechanism of the neural death. We conclude that lumbosacral ventral root avulsions progressively deplete autonomic and motor neurons. The findings suggest that early neuroprotection will be an important consideration in future attempts of treating acute cauda equina injuries.

Cell Proliferation is Influenced by Bulbectomy and Normalized by Imipramine Treatment in a Region-Specific Manner

“…Delayed apoptosis is an accepted phenomenon in the nervous system. From stroke patients, it is known that the structural lesion solidifies over weeks recruiting additional parts of the brain into infarction (Dirnagl et al, 1999); and for peripheral nerve injury or ventral root avulsion, an injury-induced progressive death of motor neurons was shown (Hoang et al, 2003). To clarify the nature of apoptotic cell death in our paradigm, further experiments with different time windows will be essential.…”

At-level neuropathic pain is induced by lumbosacral ventral root avulsion injury and ameliorated by root reimplantation into the spinal cord

“…The VRA surgical procedures have been previously described at length (Hoang et al, 2003, Hoang et al, 2006a. Briefly, a midline incision was made along the back under general anesthesia (2% isoflurane).…”

“…Post-operative tests were performed initially at 5 days after surgery, and weekly thereafter from 2-7 wks. Seven wks was chosen as the endpoint for these studies as we have extensive time course data for motoneuron and parasympathetic preganglionic neuron (PPN) cell death up to 6 wks post-avulsion (Hoang et al, 2003). The experimenter was blinded as to group identification during behavioral testing for allodynia and hyperalgesia.…”

“…Adjacent sections (every 5 th section, 4-8 sections/rat) in the L5 spinal segment were processed for glial fibrillary acidic protein (GFAP; Chemicon, Temecula, CA), allograft inflammatory factor-1 (AIF-1; Abcam, Inc., Cambridge, MA), or ED-1 (Chemicon), indicating the presence of astrocytes, microglia, and macrophages, respectively. Additionally, as motoneurons are nearly absent at the level of the lesion by 6 wks post-VRA (Hoang et al, 2003), immunohistochemistry for choline acetyltransferase (ChAT) was used to verify the normal and symmetric presence of motoneurons as further verification that the spinal level being examined was rostral to the lesion. Free-floating sections were rinsed in 0.1 M PBS, blocked for 1 hr in 5% normal donkey serum (Jackson Immuno Research Labs, Inc. Westgrove, PA), and incubated in primary antibodies (anti-rabbit GFAP, 1:1000; anti-rabbit AIF-1, 1:000; anti-mouse ED-1 1:500) with 0.3% triton overnight at room temperature.…”

“…Although we have extensively examined autonomic and motoneuron cell death (Hoang et al, 2003), as well as inflammation (Ohlsson et al, 2006) in the ventral horn at the level of injury in this model, it is unknown whether VRA alters the processing of sensory information in the dorsal horn at or beyond the injured segment. As neuropathic pain is a consequence of this type of spinal cord injury in the clinical setting, we propose that the VRA injury may contribute to the development of neuropathic pain.…”