Spastic Long-Lasting Reflexes of the Chronic Spinal Rat Studied In Vitro

Abstract

Over the months following sacral spinal cord transection in adult rats, a pronounced spasticity syndrome emerges in the affected tail musculature, where long-lasting muscle spasms can be evoked by low-threshold afferent stimulation (termed long-lasting reflex). To develop an in vitro preparation to examine the neuronal mechanisms underlying spasticity, we removed the whole sacrocaudal spinal cord of these spastic chronic spinal rats (>1 mo after S(2) sacral spinal transection) and maintained it in artificial cerebral spinal fluid in a recording chamber. The ventral roots were mounted on monopolar recording electrodes in grease, and the reflex responses to dorsal root stimulation were recorded and compared with the reflexes seen in the awake chronic spinal rat. When the dorsal roots were stimulated with a single pulse, a long-lasting reflex occurred in the ventral roots, with identical characteristics to the long-lasting reflex in the awake spastic rat tail. The reflex response was low threshold (T), short latency, long duration ( approximately 2 s), and enhanced by repeated stimulation. Brief high-frequency stimulation trains (0.5 s, 100 Hz, 1.5 x T) evoked even longer duration responses (5-10 s), with repeated bursts of activity that were similar to the repeated muscle spasms evoked in awake rats with stimulation trains or manual skin stimulation. Stimulation of a given dorsal root evoked long-lasting reflexes in both the ipsilateral and contralateral ventral roots. Long-lasting reflexes did not occur in the sacrocaudal spinal cord of acute spinal rats (S(2) transection), which is similar to the areflexia seen in awake acute spinal rats. However, long-lasting reflexes could be made to occur in the acute spinal rat by altering K(+) (7 mM) or Mg(2+) (0 mM) concentrations, or by application of high doses of the neuromodulators norepinephrine (NE, >20 microM) or serotonin (5-HT, >20 microM). In chronic spinal rats, much lower doses of these neuromodulators (0.1 microM) enhanced the long-lasting reflexes, suggesting a denervation supersensitivity to 5-HT and NE following injury. Higher doses of NE or 5-HT produced a paradoxical inhibition of the long-lasting reflexes. The high dose inhibition by NE was mimicked by the alpha(2)-adrenergic receptor agonist clonidine but not the alpha(1)-adrenergic receptor agonist methoxamine. In summary, the sacral spinal in vitro preparation offers a new approach to the study of spinal cord injury and analysis of antispastic drugs.

Spastic Long-Lasting Reflexes in the Awake Rat After Sacral Spinal Cord Injury

“…Importantly, the above characteristics of the long-lasting tail muscle reflexes are identical to the reflexes measured from ventral root recordings of the same chronic spinal rats, but with the affected sacrocaudal spinal cord maintained in vitro (Li et al 2004b). Thus in as much as the long-lasting reflex represents spastic/spasm behavior, these reflexes and spasticity in general can be studied with the in vitro sacrocaudal spinal cord preparation.…”

“…These low-threshold polysynaptic EPSPs are N-methyl-D-aspartate (NMDA)-dependent (Bennett et al 2001b;Clarke et al 2002) and particularly important in triggering spasms after chronic injury, because a single EPSP is sufficiently long (Ͼ200 ms) to evoke the slowly activating PICs in motoneurons that cause a many second long discharge (Bennett et al 2001c;Li et al 2004a). Likewise, the normal depression of mono-and polysynaptic reflexes with repeated stimulation is lost after injury (Mailis and Ashby 1990;Thompson et al 1992), and the cutaneous polysynaptic reflexes are instead enhanced with repeated stimulation (windup of C-fiber and low threshold reflexes; Clarke et al 2002;Gozariu et al 1997;Li et al 2004b). Thus repeated inputs can summate to produce sufficiently long EPSPs to trigger PICs and spasms in chronic injury (Li et al 2004a).…”

Combining Schwann Cell Bridges and Olfactory-Ensheathing Glia Grafts with Chondroitinase Promotes Locomotor Recovery after Complete Transection of the Spinal Cord

“…However, our results do not necessarily have to be interpreted as increased pain or allodynia. Paralyzed rats showed signs comparable with spasticity in human patients, and thus the increased responses to tactile stimuli could simply be a sign of increased spasticity triggered by disinhibition of spinal pathways (Calancie et al, 1993) or changes in neuronal properties Li et al, 2004). Increased sensory feedback has been suggested to accentuate locomotor recovery after SCI (Pearson, 2001).…”

Differential contributions of somatic and dendritic calcium-dependent potassium currents to the control of motoneuron excitability following spinal cord injury

“…(Li and Bennett 2003)). However, it is unlikely that such activity could induce spasm-like high frequency discharge in motoneurons (e.g., (Li et al 2004b)). Available evidence also suggests that elimination of I h does not alter estimations of 5-HT induced Ca 2?…”

“…Given this state of chronic PIC facilitation, mechanisms that activate or suppress activation of PICs are potentially important factors in causing or reducing motoneuron hyperexcitability. PIC activation may be facilitated by increased excitatory reflexes following spinal cord injury (Baker and Chandler 1987;Li et al 2004b). Synaptic inhibition has been shown to be a key mechanism in the control of motoneuron discharge and dendritic PIC activation (Bennett et al 1998;Hultborn et al 2003), particularly for dendritic sources of inhibition (Bui et al 2008;Venugopal et al 2011).…”