Background
The second messenger signaling pathways in primary afferent nociceptors that mediate hypersensitivity to mechanical stimuli differ between models of painful peripheral neuropathies [
1]. Two extreme examples of this are the neuropathies induced by chronic ethanol consumption, and by acquired immunodeficiency disease syndrome (AIDS) therapy (nucleoside reverse transcriptase inhibitors). In alcohol-induced neuropathy, protein kinase Cε(PKCε) has a major contribution to mechanical hyperalgesia [
2], whereas in AIDS therapy neuropathy, Ca
++, caspase signaling and mitochondrial electron transport [
3‐
5] but not PKCε or a number of other second messenger signaling pathways (i.e., protein kinase A, protein kinase G, extracellular signal-regulated kinases 1/2 or nitric oxide) contribute [
3].
Enhanced activity in sensory neurons is thought to contribute to pain reported by patients with small-fiber peripheral neuropathies. Microneurography techniques have demonstrated pathological responses such as sensitization to mechanical stimuli, in patients with trigeminal neuralgia [
6], traumatic nerve injury [
7], entrapment neuropathy [
8], phantom limb [
9] and erythromelalgia [
10]. However, there are practical limitations in performing microneurography in patients, including inability to classify fiber functions fully, small numbers of fibers that can be evaluated in an individual patient and the potential for inducing further injury by introducing a microelectrode into an already damaged nerve. Furthermore, in spite of the fact that in most patients, metabolic abnormalities, toxins, drugs or infectious organisms are producing the neuropathic conditions, most microneurography studies have been done in patients with a traumatic nerve injury [
7‐
9].
Single-fiber electrophysiology has been performed in animal models of metabolic and toxic, as well as traumatic nerve injury-associated painful peripheral neuropathy. Following traumatic nerve injury it has been reported that there is increased spontaneous activity occurring in irregular bursts [
11‐
13]; in diabetic neuropathy, in addition to increased spontaneous activity, a decrease in threshold and increase in response to supra-threshold stimulation has been reported [
14‐
19]; in models of cancer chemotherapy neuropathy, C-fibers have been reported to be hyperresponsive and to fire irregularly [
1,
20]; in alcohol neuropathy, C-fibers also demonstrate a decrease in threshold and increased response to stimulation [
2]; and, in nucleoside reverse transcriptase inhibitor-induced AIDS-therapy neuropathy, a change in post-stimulus interspike interval (ISI) histogram, without change in threshold or number of action potentials in response to threshold or suprathreshold mechanical stimulus has been reported [
3]. In this study, we have performed a side-by-side comparison of evoked C-fiber activity in models of two frequently co-morbid forms of peripheral neuropathy, alcohol and AIDS therapy-induced painful peripheral neuropathy, which differ markedly in the nociceptor second messenger signaling pathways involved [
2,
3].
Discussion
While it is generally accepted that enhanced activity in primary afferent nociceptors plays an important role in the pain experienced by patients with peripheral neuropathy [
6‐
10], changes in activity in primary afferent nociceptors have received little attention, including direct comparisons between changes in primary afferent nociceptor function in different forms of painful peripheral neuropathy. In this study, we have compared mechanically evoked C-fiber activity in rat models of alcohol and AIDS therapy-induced peripheral neuropathy, for which enhanced nociception has been shown to be dependent on different second messenger signaling pathways.
While the mechanical hyperalgesia observed in these two models of painful peripheral neuropathy are of similar magnitude [
2,
3], the changes in C-fiber function differ markedly, being fairly well restricted to a decrease in conduction velocity for AIDS therapy, while many aspects of mechanically-evoked activity were effected by alcohol. Although clinical studies show slowed conduction velocity in many types of peripheral neuropathy [
21,
23‐
25], the mechanism underlying slowing of conduction velocity remains to be established. Available data suggest that changes in ionic currents, most especially for voltage-gated ion channels, contribute to conduction velocity abnormalities [
21,
26,
27]. The most well studied model with respect to mechanisms involved in changes in conduction velocity is diabetic neuropathy [
21], in which I
Na+, I
K+ and I
h have been shown to decrease [
26,
27] and I
Ca2+ to increase [
28‐
30]. However, depending on the composition of other ion channels in the membrane of the sensory neuron, one may observe either enhanced or attenuated sensation [
21]. Since slowing of nerve conduction velocity is the major change in C-fiber function in the rat model of ddC-induced painful peripheral neuropathy, direct analysis of ionic currents in dorsal root ganglion neurons treated with AIDS therapy could provide important insights into the mechanisms involved in the pain associated with this class of neuropathies.
While the relatively small change in single fiber electrophysiological properties of primary afferent nociceptors observed in rats with ddC neuropathy might suggest that changes in the peripheral terminal of sensory neurons make a minor contribution to AIDS therapy-induced pain, we have previously shown that peripheral administration, at the site of nociceptive testing, of antagonists of intracellular calcium [
3], caspase signaling [
4] and the mitochondrial electron transport chain [
5], which in control animals have no effect on mechanical nociceptive threshold, reverses ddC-induced mechanical hyperalgesia. Taken together these findings provide support for the suggestion that changes in primary afferent nociceptor function, not tested for in the present study, may play a role in the decreased behavioral mechanical nociceptive threshold in the ddC-induced painful peripheral neuropathy. Alternatively, since the mechanism of action of nucleoside reverse transcriptase inhibitor-induced neurotoxicity is via their effects on mitochondrial function [
31,
32], it may be that a fraction of mitochondria are affected in most neurons, leading to a smaller change in function in a larger percentage of sensory neurons. In contrast, in alcohol, diabetic [
16,
17] and vincristine [
20] peripheral neuropathy, the toxic insult appears to produce an all-or-none change in activity, in a subset of neurons (i.e., the high-firing fibers) not observed in AIDS therapy neuropathy.
Decrease in mechanical threshold and increase in number of action potentials elicited by the same intensity stimulus contribute to inflammatory pain [
33,
34], which is characterized by mechanical hyperalgesia. In the present study we found a decrease in mechanical threshold and increase in number of action potentials produced by threshold and suprathreshold stimulation in rats consuming alcohol, but not in ddC-treated rats. The increase in number of short ISIs, in response to both threshold and suprathreshold mechanical stimulation, in alcohol fed rats, will increase temporal summation in postsynaptic spinal dorsal horn neurons; increasing the range of ISIs, near 100 ms, as observed in rats consuming alcohol, causes greater temporal summation of C-fiber-evoked excitatory postsynaptic currents in dorsal horn neurons [
35], and in the same range of ISIs, temporal summation of afferent activity appears to be an important factor in human pain perception [
36‐
41].
While pattern of activity in a presynaptic neuron can dramatically affect activity in its postsynaptic neurons [
42‐
46], much less attention has been given to the importance of the pattern of activity elicited by mechanical stimulation of primary afferents in the pain associated with peripheral neuropathy. In previous studies of painful peripheral neuropathy we have observed that changes in primary afferent nociceptor function occur in an all-or-nothing fashion. Thus, in models of diabetic [
16,
17] and vincristine [
20] neuropathy, we found enhanced activity restricted to a subpopulation of C-fibers (i.e., high-firing fibers), the function of the remaining C-fibers being similar to those in control rats. In alcohol-induced neuropathy this dichotomy was also present. Therefore, in our analysis of variability in action potential timing we also separately evaluated the change in activity pattern in high- and low-firing C-fibers. Marked alteration in the distribution of Cv2 values was observed in high-firing C-fibers in alcohol-induced painful peripheral neuropathy; however, this change was different from that in high-firing C-fibers in diabetic and vincristine-treated rats, in that there was a marked decrease in maximum Cv2 in rats with alcohol-induced neuropathy. While the mechanism underlying these changes is unknown, the lower Cv2 value can be generated by a repetitively bursting pattern of activity [
42]. The functional significance of variability in neuronal discharge patterns has been the focus of study in somatosensory cortex and other sensory areas [
42‐
44,
46]. It has been suggested that such "variability may not be so much a flaw as a feature that the brain puts to good use" to "provide the dynamic range for rapid modulation of synaptic efficacy" [
45]. This may be relevant to the function of nociceptors as afferent activity-dependent plasticity in spinal nociceptive pathways is thought to be a crucial feature of pain signaling [
47], and may contribute to the progressive increase in pain during a prolonged stimulus, even while adaptation decreases the mean firing frequency of nociceptive nerve fibers [
48].
In summary, in two models of painful peripheral neuropathies that differ markedly based on the involvement of second messenger signaling mechanisms in primary afferents, we have found marked differences in C-fiber activity. Our findings raise the question; does activity in sensory neurons from different forms of peripheral neuropathy have unique signatures? Since alcohol consumption and AIDS are common co-morbid conditions [
49‐
51], the possibility that they produce painful peripheral neuropathy by different mechanisms raises the question are symptoms more severe in AIDS patients who chronically consume alcohol? One step in developing an understanding of the importance of these mechanisms would be to directly activate individual second messengers in primary afferent nociceptors, to determine their effect on mechanically-evoked nociceptor activity, and then to study specific ion channels in dorsal root ganglion neurons,
in vitro, to determine the ionic basis of these differences.
In vitro studies of the effect of ddC on specific ionic conductance may be especially important in furthering our understanding of the functional alterations in AIDS therapy neuropathy, which does not appear to markedly alter function of individual primary nociceptors.
Authors' contributions
XC participated in the design of the study, carried out all the experiment, performed the statistical analysis and drafted the manuscript. JDL participated in the design of the study and drafted the manuscript. All authors read and approved the final manuscript.