Jun-Ling Xing

An experimental model for chronic compression of dorsal root ganglion produced by intervertebral foramen stenosis in the rat.

&NA; Under anesthesia and sterile surgery, a small stainless steel rod (4 mm in length and 0.5–0.8 mm in diameter) was inserted into the L5 intervertebral foramen in the rat, developing intervertebral foramen stenosis and hence producing a chronic steady compression of the dorsal root ganglion (DRG). The hind paw on the injured side exhibited a significant reduction in the latency of foot withdrawal to noxious heat and manifested a persistent heat hyperalgesia 5–35 days after surgery. Injection of 1% carrageenan into the intervertebral foramen, presumably causing inflammation of the DRG, also produced hyperalgesia to heat on the hind paw of the injured side 5–21 days after surgery. Extracellular electrophysiological recordings from myelinated dorsal root fibers were performed in vivo. Spontaneous activity was present in 21.5% of the fibers recorded from DRG neurons injured with chronic compression in contrast to 1.98% from uninjured DRG neurons. The pattern of spontaneous activity was periodic and bursting in 75.3% of the spontaneously active fibers. These neurons had a greatly enhanced sensitivity to mechanical stimulation of the injured DRG and a prolonged after discharge. In response to TEA, topically applied to the DRG, excitatory responses were evoked in the injured, but not the uninjured, DRG neurons. Application of this experimental model may further our understanding of the neural mechanisms by which chronic compression of DRG induces low back pain and sciatica.

Blockade of persistent sodium currents contributes to the riluzole-induced inhibition of spontaneous activity and oscillations in injured DRG neurons.

In addition to a fast activating and immediately inactivating inward sodium current, many types of excitable cells possess a noninactivating or slowly inactivating component: the persistent sodium current (INaP). The INaP is found in normal primary sensory neurons where it is mediated by tetrodotoxin-sensitive sodium channels. The dorsal root ganglion (DRG) is the gateway for ectopic impulses that originate in pathological pain signals from the periphery. However, the role of INaP in DRG neurons remains unclear, particularly in neuropathic pain states. Using in vivo recordings from single medium- and large-diameter fibers isolated from the compressed DRG in Sprague-Dawley rats, we show that local application of riluzole, which blocks the INaP, also inhibits the spontaneous activity of A-type DRG neurons in a dose-dependent manner. Significantly, riluzole also abolished subthreshold membrane potential oscillations (SMPOs), although DRG neurons still responded to intracellular current injection with a single full-sized spike. In addition, the INaP was enhanced in medium- and large-sized neurons of the compressed DRG, while bath-applied riluzole significantly inhibited the INaP without affecting the transient sodium current (INaT). Taken together, these results demonstrate for the first time that the INaP blocker riluzole selectively inhibits INaP and thereby blocks SMPOs and the ectopic spontaneous activity of injured A-type DRG neurons. This suggests that the INaP of DRG neurons is a potential target for treating neuropathic pain at the peripheral level.

Evoked bursting in injured Aβ dorsal root ganglion neurons: A mechanism underlying tactile allodynia

Summary The evoked bursting discharges of the Aβ dorsal root ganglion neurons may contribute to the development of allodynia after nerve injury. Abstract Chronic compression of rat dorsal root ganglion (CCD) produced tactile allodynia accompanied with hyperexcitability of the myelinated Aβ dorsal root ganglion (DRG) neurons. The Aβ DRG neuron hyperexcitability exhibits as bursting discharges in response to peripherally evoked action potentials (evoked bursting [EB]). The incidence of EB was significantly increased after chronic compression of DRG (CCD) (43.3%) vs control (13.3%). EB was maintained by oscillation of the membrane potential, and its duration was increased when the membrane potential was depolarized. EB was found to coexist in some neurons with spontaneous bursting (SB), but EB always occurred at a more negative membrane potential than SB. Afterdischarges of the wide dynamic range neurons of the dorsal horn in the spinal cord in response to electrical stimulation of Aβ afferent nerve fibers were suppressed by blocking EB of the DRG neurons. CCD neurons with EB exhibited increased current density of persistent sodium current (INap) and hyperpolarization‐activated cation current (Ih) and decreased α‐dendrotoxin (α‐DTX) sensitive current (IDTX). The increased Ih activated by afterhyperpolarization of peripheral afferent action potential was necessary for EB generation and a balance between IDTX and INap might be necessary for EB maintenance. This study may suggest a role of EB of myelinated DRG neurons in development of allodynia after nerve injury and a potential pharmaceutical therapy in treating neuropathic allodynia.

Subthreshold membrane potential oscillation mediates the excitatory effect of norepinephrine in chronically compressed dorsal root ganglion neurons in the rat

Injured dorsal root ganglion (DRG) neurons often develop adrenergic sensitivity. To investigate the mechanisms of this phenomenon, the effects of norepinephrine (NE) on membrane potential of large‐ and medium‐sized A‐type neurons from chronically compressed DRG were recorded electrophysiologically in vitro. NE induced a depolarization in both control (26/36) and injured (56/62) neurons, whereas the incidence and amplitude of NE‐induced depolarization in the injured neurons were significantly higher than that in controls. Following NE‐induced depolarization, a subthreshold membrane potential oscillation (SMPO) was triggered or enhanced that initiated or increased repetitive firing in a fraction of injured neurons (15/56). After the SMPO was selectively abolished by application of tetrodotoxin (TTX), NE‐induced depolarization failed to produce repetitive firing, even with a greater depolarization. Application of Rp‐cAMPS (500 &mgr;M), a selective inhibitor of protein kinase A (PKA), decreased both SMPO and repetitive firing evoked by NE application or by intracellular current injection. Conversely, Sp‐cAMPS (500 &mgr;M), a PKA activator, had a facilitating effect on both the SMPO and the repetitive firing. These results strongly suggest that a PKA mediated triggering and enhancement of SMPO may be responsible for the excitatory effects of NE on sensory neurons in neuropathic rats.

Effects of gabapentin on spontaneous discharges and subthreshold membrane potential oscillation of type A neurons in injured DRG

&NA; Ectopic spontaneous discharges play a critical role for both initiation and maintenance of the neuropathic pain state. Gabapentin (GBP) has been shown to be effective in animal models of neuropathic pain as well as in chronic pain patients. To investigate the peripheral mechanisms of GBP, the effects of GBP on spontaneous discharges and subthreshold membrane potential oscillation (SMPO) of chronically compressed dorsal root ganglion (DRG) were examined electrophysiolocally in vitro. The rate of spontaneous discharges was transitorily enhanced when GBP was applied to the DRG. When the concentration was under 5 μM, only enhanced effect was observed, while spontaneous discharges were completely suppressed when the concentration of GBP was beyond 5 μM. The similar doses of GBP blocking the spontaneous discharges failed to block the propagation of impulses by electrical nerve stimulation. Furthermore, we found that the SMPO of injured DRG cells can be selectively abolished by GBP without interrupting spike propagation. The results suggest that the inhibitory effect of GBP on SMPO might be one of the membrane mechanisms of action of GBP. This may partially explain the antinociceptive action of GBP by directly suppression nociceptive afferent input to the spinal cord.

Adrenergic sensitivity of neurons with non-periodic firing activity in rat injured dorsal root ganglion.

In this study, we compared the sensitivity of non-periodically and periodically active neurons in chronically compressed dorsal root ganglion in rats to norepinephrine and sympathetic stimulation. Forty-nine of 58 (84.5%) neurons with non-periodic activity showed responses to norepinephrine, whereas only five of 48 (10.4%) neurons with periodic activity displayed any response. The dose-response relationship of norepinephrine to the irregular burst pattern neurons shifted towards the left significantly compared to that of the periodic activity neurons. Responses to norepinephrine became apparent in eight neurons after their periodic firing activity was transformed into the non-periodic firing activity through the increase in Ca(2+). Changes in the time-response curves indicate a higher sensitivity of irregular burst pattern neurons to sympathetic stimulation than the periodic activity neurons. Finally, deterministic dynamics contained within the interburst interval series for non-periodic activity were identified. From these results, we suggest that the non-periodic activity neurons have a higher adrenergic sensitivity than those displaying periodic activity, and that this sensitivity may depend on the deterministic chaos within its firing dynamic system.

Saikosaponin a Mediates the Anticonvulsant Properties in the HNC Models of AE and SE by Inhibiting NMDA Receptor Current and Persistent Sodium Current

Epilepsy is one of the most common neurological disorders, yet its treatment remains unsatisfactory. Saikosaponin a (SSa), a triterpene saponin derived from Bupleurum chinensis DC., has been demonstrated to have significant antiepileptic activity in a variety of epilepsy models in vivo. However, the electrophysiological activities and mechanisms of the antiepileptic properties of SSa remain unclear. In this study, whole-cell current-clamp recordings were used to evaluate the anticonvulsant activities of SSa in the hippocampal neuronal culture (HNC) models of acquired epilepsy (AE) and status epilepticus (SE). Whole-cell voltage-clamp recordings were used to evaluate the modulation effects of SSa on NMDA-evoked current and sodium currents in cultured hippocampal neurons. We found that SSa effectively terminated spontaneous recurrent epileptiform discharges (SREDs) in the HNC model of AE and continuous epileptiform high-frequency bursts (SE) in the HNC model of SE, in a concentration-dependent manner with an IC50 of 0.42 µM and 0.62 µM, respectively. Furthermore, SSa significantly reduced the peak amplitude of NMDA-evoked current and the peak current amplitude of INaP. These results suggest for the first time that the inhibitions of NMDA receptor current and INaP may be the underlying mechanisms of SSa’s anticonvulsant properties, including the suppression of SREDs and SE in the HNC models of AE and SE. In addition, effectively abolishing the refractory SE implies that SSa may be a potential anticonvulsant candidate for the clinical treatment of epilepsy.

Subthreshold membrane oscillations underlying integer multiples firing from injured sensory neurons.

Integer multiples firing (IMF), a special temporal pattern of firing, was recently observed in spontaneous discharge from injured dorsal root ganglion (DRG) neurons. To investigate the mechanism underlying IMF, the injured DRG neurons of rat were recorded intracellularly. Of 64 recorded A-neurons discharging spontaneously, eight fired spikes in the IMF pattern. Interspike interval (ISI) time series of IMF showed a structure of distinct bands on scatter map. Regular subthreshold membrane oscillations (SMOs) with relatively stable amplitude and frequency were observed on all eight IMF neurons. IMF could be induced from the neurons in periodic firing by local application of tetrodotoxin (TTX), a Na+ channel antagonist. During this process, the amplitude of SMOs varied markedly. Some SMOs were below action potential threshold so that they did not trigger spikes. Nor did some SMOs, though their amplitude were obviously beyond the threshold measured from nearby spikes. The results indicate that regular SMOs existent in injured DRG A-neurons underlie IMF, and that TTX-induced transformation of firing pattern from periodic to integer multiples may occur in two ways: decreasing the amplitude of SMOs and/or elevating action potential threshold.

Conduction Failures in Rabbit Saphenous Nerve Unmyelinated Fibers

Recent experimental and theoretical data indicate that the functional capabilities of axons with specialized structures are much more diverse than traditionally thought. However, few observations were concerned with the main axons without arborization. In the present study, electrical stimulation of the saphenous nerve at different frequencies (2, 5, 10, 20 Hz) was used to test the role of activity-dependent effects on the pattern of action potentials that propagate along individual unmyelinated fibers (C fibers) within the trunk of the saphenous nerve in rabbits. Three basic types of C fiber responses to repetitive stimulation were observed: type-1 fibers showed an entrained response without conduction failure; type-2 fibers discharged with intermittent conduction failures; while only sporadic conduction failures happened in type 3. The failure modality in type-2 and type-3 fibers is closely related to the conductive distance as well as the frequency and duration of stimuli which lead to a critical level of conduction velocity slowing. A novel fluctuation in interspike intervals was always observed immediately before the occurrence of the failures, implying that the fluctuation of conduction velocity is correlated with imminent failures. Both the 4-aminopyridine-sensitive potassium current and hyperpolarization-activated cation current were recognized to be involved in the regulation of conduction failure patterns. The results confirmed, at least in part, the existence of conduction failures in the main axon of C fibers, suggesting that axonal operations may also be determinants for adaptation phenomenon and information processing in peripheral nervous system.

MEMBRANE CURRENT-BASED MECHANISMS FOR EXCITABILITY TRANSITIONS IN NEURONS OF THE RAT MESENCEPHALIC TRIGEMINAL NUCLEI

Since Hodgkins first description of three classes of excitability in crustacean nerve axons (1948), theoretical studies have used mathematical models to demonstrate that small changes in the parameters describing ionic currents could result in transitions between classes of membrane excitability. However, these transitions have rarely been investigated experimentally. Here, we show that states of excitability in rat mesencephalic V (Mes V) neurons can be classified into three groups, with manipulations of the 4-aminopyridine sensitive K(+) current (I(4-AP)) or persistent Na(+) current (I(NaP)) leading to the corresponding transitions. However, alterations in the hyperpolarization-activated cation current (I(h)), tetraethylammonium (TEA)-sensitive K(+) current, or Cd(2+)-sensitive Ca(2+) current were ineffective in causing these transitions. These results provide experimental evidence for the excitability transitions predicted by Hodgkin and characterize their ionic mechanisms in Mes V neurons.