Perceptual Correlates of Nociceptive Long-Term Potentiation and Long-Term Depression in Humans

Perceptual correlates of the conditioning process

A single train of HFS evoked mild to moderate pain (mean pain rating, 32 of 100 at 10 × T and 44 of 100 at 20 × T), and repetition of such trains (five times at 10 sec intervals) resulted in a gradual increase in the perceived pain (Fig. 2A). Pain ratings for LFS were much lower than for HFS, because LFS consisted of single electrical pulses (first pulse: mean rating, 5 of 100 at 10 × T and 8 of 100 at 20 × T) (Fig. 2B). Pain to LFS initially increased over a period of ∼2.5 min (150 pulses) but then gradually declined until perceived pain was less than the initial pain ratings.

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Figure 2.

Pain sensations evoked by conditioning electrical stimuli. A, HFS (5× 1 sec at 100 Hz; 10 sec intervals between bursts) elicited pain sensations that increased over the five bursts. B, During LFS (1000 pulses at 1 Hz), pain ratings increased over the first 150 pulses and then slowly declined until the end of the conditioning stimulation, when pain ratings were lower than the initial values. Mean ± SEM values across eight subjects are shown. Each filled triangle or filled square represents average pain ratings on a 100 point verbal rating scale. ▴, 10 × T; ▪, 20 × T.

Long-term changes in pain sensitivity at the site of conditioning stimulation

Conditioning HFS led to a significant increase in pain evoked by single electrical test stimuli of 10 × T delivered through the conditioning electrode (Fig. 3A). No differences were found between conditioning stimulus intensities of 10 × T and 20 × T (Table 1); thus data were pooled across intensities. The facilitation took ∼20 min to develop and stabilized at 27% above baseline (p < 0.01) (Table 1). The increase persisted until the end of the observation period (60 min). In two subjects, the observation period was extended to 200 min after conditioning stimulation (20 × T). Hyperalgesia to electrical stimuli stabilized at 36% above baseline after 60 min and remained undiminished until 200 min after conditioning stimulation (39% above baseline).

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Figure 3.

Homotopic effects of conditioning electrical stimuli. A, After LFS, pain evoked by single electrical test pulses (intensity, 10 × T) through the HFS electrode increased to ∼30% above baseline (•). This facilitation lasted until the end of the observation period. Pain evoked by identical test pulses through a remote electrode exhibited a small decrease over time (○). B, After LFS, pain evoked by single electrical test pulses through the LFS electrode decreased to ∼30% below baseline (•). This inhibition was not reversible within the observation period. Mean ± SEM values across seven subjects are shown. Dotted lines indicate mean level of baseline period. Each circle represents normalized pain ratings averaged over a 5 min time window across stimulation intensities (10 × T; 20 × T). Asterisks indicate post hoc paired t tests; conditioned versus control site; p < 0.05.

Conditioning LFS led to a significant decrease in pain (hypoalgesia) evoked by electrical test stimuli that developed continuously during LFS (see above) and persisted at 27% below baseline (p < 0.01) (Table 1) (Fig. 3B). Neither HFS nor LFS changed the pain ratings for stimulation of the remote control site. These findings indicate that HFS and LFS protocols induced LTP- or LTD-like long-term changes in human pain perception mediated by the conditioned pathway (homotopic hyperalgesia and hypoalgesia).

Long-term changes in pain sensitivity adjacent to the site of conditioning stimulation

To test for perceptual correlates of heterotopic LTP and LTD outside of the conditioned pathway, mechanical test stimuli were applied around the conditioned site and around the unconditioned control site. As shown in Figure 4, A and B, HFS induced a significant enhancement of pin prick-evoked pain around the conditioning electrode (secondary hyperalgesia; p < 0.01) (Table 1), which persisted until the end of the observation period (60 min). Pain perception stabilized at a plateau phase of 90% above baseline after 10 × T and of 210% after 20 × T. One hour after conditioning stimulation, the mean area of hyperalgesia was 38 cm² (10 × T) and 56 cm² (20 × T). In three subjects, secondary hyperalgesia was assessed for 200 min after conditioning stimulation (20 × T) when pain perception was still enhanced to ∼220% above baseline (compared with 296% at 60 min). There was no mechanical hyperalgesia around the unconditioned control site. Because the neural pathway for these mechanical test stimuli was spatially segregated from that of the conditioning stimuli, the findings suggest a spread of facilitation to unstimulated inputs of the spinal cord (i.e., heterosynaptic facilitation). No such spread was seen for the inhibitory effect of LFS (Fig. 4C,D). In contrast, LFS caused a small but significant increase in mechanically evoked pain (p < 0.05) (Table 1). The significant effect of the stimulus intensity and significant interaction term reflect the finding that this secondary hyperalgesia occurred only at 20 × T (Fig. 4D) (to 30% above baseline) but not at 10 × T (Fig. 4C) (to 4% below baseline).

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Figure 4.

Heterotopic effects of conditioning electrical stimuli on pin prick-evoked pain. A, B, Conditioning HFS induced a significant enhancement of pin prick-evoked pain adjacent to the conditioning electrode (•) but not adjacent to the control electrode (○). This secondary hyperalgesia occurred after conditioning stimulus intensities of 10 × T and 20 × T. Pain perception was significantly increased already at 5 min and remained potentiated throughout the 60 min observation period (10 × T) or even increased further (20 × T). C, Conditioning LFS at 10 × T induced no changes in pin prick-evoked pain at any test site. D, For conditioning LFS at 20 × T, a small but significant increase in pin prick-evoked pain was found near the conditioning electrode but not the control electrode. Mean ± SEM values across eight subjects are shown. Dotted lines indicate mean level of baseline period. Each circle represents the normalized average of pain ratings across all seven stimulus intensities over a 5 min time window. Asterisks indicate post hoc paired t tests; conditioned versus control site; p < 0.05.

HFS also induced touch-evoked pain (allodynia) around the HFS electrode but not around the remote unconditioned control site (Fig. 5A,B). Light tactile stimuli elicited mildly painful sensations in 9 of 16 experiments after HFS (p < 0.05) (Table 1). LFS at 10 × T did not elicit allodynia (Fig. 5C), but one subject reported touch-evoked pain after 20 × T (Fig. 5D).

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Figure 5.

Heterotopic effects of conditioning electrical stimuli on touch-evoked pain. A, B, Conditioning HFS induced a state in which these tactile, normally non-noxious stimuli became painful adjacent to the conditioning electrode (•) (allodynia) but not adjacent to the control electrode (○). Allodynia gradually developed after HFS and persisted throughout the 60 min observation period. C, Conditioning LFS at 10 × T elicited no allodynia. D, For conditioning LFS at 20 × T, one subject developed allodynia near the conditioning electrode but not the control electrode. Mean ± SEM values across eight subjects are shown. Each circle represents the normalized average of pain ratings across all three stimulus intensities over a 5 min time window. Asterisks indicate post hoc paired t tests; conditioned versus control site; p < 0.05.

The stimulus–response functions of the two different mechanosensitive inputs are shown in detail in Figure 6. The change in perceived quality of light tactile stimuli toward a burning and painful sensation is represented by a significant upward shift (F_(1,7) = 10.97; p < 0.05) (Fig. 6A) that was homogenous across all three test stimuli (F_(2,14) = 1.34; NS). This effect was absent at the remote control site (Fig. 6C). Punctate probes of graded intensities induced graded pain sensations both before and after conditioning stimulation (F_(6,42) = 79.1; p < 0.001) (Fig. 6B). The effect of conditioning HFS was a parallel leftward shift of the stimulus response function (F_(1,7) = 33.21; p < 0.001) (Fig. 6B), indicating both a decrease in pain thresholds and an increase in pain ratings elicited by suprathreshold stimuli. This effect was confined to skin areas adjacent to the conditioning electrode, as shown by the complete absence of a shift in the stimulus–response function at the control site (Fig. 6D).

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Figure 6.

Stimulus–response functions for punctate probes and light tactile stimuli after HFS at 20 × T. A, Allodynia. Light tactile stimuli were not painful when moved across normal skin before conditioning stimulation (○). Adjacent to the conditioning electrode, the same stimuli were mildly painful after HFS at 20 × T (•). CW, 3 mN; QT, 100 mN; BR, 400 mN. B, Secondary hyperalgesia. The stimulus–response function for punctate probes was approximately linear in log–log coordinates in normal skin (○). After HFS, there was a parallel leftward shift of this function adjacent to the conditioning electrode (•). C, D, There was neither allodynia nor hyperalgesia adjacent to the unstimulated control site. Mean ± SEM values across eight subjects are shown. Each circle represents average pain ratings on a 100 point verbal rating scale. Asterisks indicate post hoc paired t tests; preconditioning versus postconditioning stimulation; p < 0.05.

Perceptual correlate of homosynaptic LTP

Conditioning stimulation of one specific input pathway to the human spinal cord was sufficient to induce a long-term change in human pain perception mediated by this pathway. As demonstrated by spreading vasodilatation (Fig. 1C), the input pathway encompassed peptidergic cutaneous afferents containing substance P and calcitonin gene-related peptide (Brain and Williams, 1988), the central terminals of which preferentially synapse in lamina II (outer part) of the spinal cord dorsal horn (Hunt and Mantyh, 2001). LTP has been described after conditioning HFS of the synaptic connection between peptidergic C-fiber afferents and the nociceptive lamina I spinoparabrachial neurons (Ikeda et al., 2003) that mediate hyperalgesia in behaving animals (Khasabov et al., 2002). HFS also induced LTP in lamina II (Randic et al., 1993; Liu and Sandkühler, 1997) and in deep dorsal horn neurons (Svendsen et al., 1998).

In addition to the spinal cord, synaptic LTP in other parts of the CNS (e.g., the thalamus or neocortex) or altered descending inhibitory or facilitating control may have contributed to the observed long-term changes in pain perception. However, human models of cortical LTP involve the motor rather than the sensory cortex, and they require either simultaneous activation of two input pathways (Bütefisch et al., 2000) or direct depolarization of cortical neurons (Nitsche and Paulus, 2001). Moreover, high intensities of electrical stimulation, such as with the HFS protocol, activate the inhibitory rather than the facilitating descending pathways (Porreca et al., 2002; Suzuki et al., 2002). Therefore, spinal LTP is the most likely mechanism of the long-term enhancement of pain perception observed at the conditioned skin site.

Perceptual correlate of homosynaptic LTD

Conditioning pulse trains at 1 Hz (LFS) resulted in a long-term decrease in pain perception (hypoalgesia) mediated by the conditioned pathway throughout the observation period. This LTD-like reduction of human pain perception was similar to LTD at the synapses between nociceptive afferents and superficial dorsal horn neurons in rats, both in magnitude and in its restriction to the conditioned pathway (Chen and Sandkühler, 2000). Pain perception during conditioning LFS exhibited a biphasic time course. The initial increase in pain is attributable to the action potential “wind-up” in dorsal horn neurons. However, wind-up in the dorsal horn in animals reaches a plateau after 10–100 electrical pulses and outlasts conditioning stimulation only for seconds (Woolf, 1996; Herrero et al., 2000). The subsequent decline in pain ratings in our study is compatible with a delayed onset of LTD (Mizuno et al., 2001).

The LTD-like reduction of human pain perception observed in this study is unlikely to be attributable to descending inhibitory mechanisms such as diffuse noxious inhibitory control, which would have affected all sites outside of the conditioned site, including the unstimulated control site (Gozariu et al., 1997). However, the reduction of pain perception was confined strictly to the conditioned pathway.

Some clinical methods of stimulation-induced analgesia may at least partly be attributable to LTD in the spinal cord, such as spinal cord stimulation (North et al., 1993), cutaneous field stimulation (Nilsson et al., 2003), and the low-frequency, high-intensity mode of transcutaneous electrical nerve stimulation (acupuncture-like TENS) (Sandkühler, 2000b). In contrast, the short-lasting effect of nonpainful, high-frequency, low-intensity TENS (conventional TENS) is commonly attributed to segmental spinal inhibition induced by activation of Aβ fibers (Garrison and Foreman, 1996; Sandkühler, 2000b), as suggested in the gate control theory.

Perceptual correlates of heterosynaptic LTP

LTP in the spinal cord has been proposed as the underlying mechanism of central sensitization (Randic et al., 1993; Miletic and Miletic, 2000; Sandkühler, 2000a; Willis, 2002). Sensitization in the rat or monkey spinal cord was specific for mechanically evoked input, whereas heat-evoked input was not enhanced (Simone et al., 1991; Dougherty et al., 1998; Pertovaara, 1998). Likewise, central sensitization in humans led to an increase in pain sensitivity to mechanical but not to heat stimuli (secondary hyperalgesia and allodynia) in a large area around the injured skin site (LaMotte et al., 1991; Treede et al., 1992; Ali et al., 1996). We conclude that secondary hyperalgesia and allodynia to mechanical stimuli after the HFS protocol also represent nociceptive spinal LTP. However, they differ from homosynaptic LTP on several accounts. First, the facilitated input comes from adjacent skin sites and thus is spatially remote from the conditioning input (heterotopic). Second, both secondary hyperalgesia and allodynia involve the interaction of two input pathways: the conditioning input is carried by C fibers, whereas the facilitated input is carried by Aδ fibers and Aβ fibers, respectively (LaMotte et al., 1991; Kilo et al., 1994; Ziegler et al., 1999; Magerl et al., 2001). Third, whereas the conditioning pathway expresses the capsaicin-sensitive receptor TRPV1, both facilitated pathways do not (Magerl et al., 2001). These lines of evidence suggest that the central sensitization underlying secondary hyperalgesia is not restricted to the activated synapse (homosynaptic LTP) but also spreads to adjacent synapses (heterosynaptic LTP) (cf. Woolf and Salter, 2000). Because hyperalgesia in neuropathic pain involves facilitation of the same input pathways as secondary hyperalgesia (Treede et al., 1992; Fields et al., 1998; Baumgärtner et al., 2002), it may also be related to heterosynaptic LTP in the spinal cord.

Convergence of A- and C-fiber nociceptors on nociceptive spinal neurons could provide the anatomical basis for heterosynaptic LTP. Alternatively, mechanosensitive inputs may be facilitated at adjacent neurons by a diffusable factor that reaches extrasynaptic receptors (Liu et al., 1994). Nociceptive neuropeptides are likely candidates for this type of volume transmission, because substance P may spread over considerable distances (Duggan et al., 1995). The action of substance P on neurokinin 1 (NK1) receptors was essential both for the induction of LTP in spinal cord neurons (Liu and Sandkühler, 1997; Ikeda et al., 2003) and for the induction of secondary hyperalgesia (Laird et al., 2001). Moreover, NK1 receptor-bearing spinal cord neurons were shown to be essential for the development of hyperalgesia (Mantyh et al., 1997; Khasabov et al., 2002) and for the induction of LTP (Ikeda et al., 2003). For behaviorally relevant consequences, homosynaptic LTP at glutamatergic synapses in the rat hippocampus needs to be supplemented by heterosynaptic facilitation (Bailey et al., 2000). Thus the behaviorally relevant consequences of LTP in the spinal cord may also require heterosynaptic facilitation.

Heterosynaptic LTP appears to be induced not only by HFS but also by LFS protocols given at C-fiber intensity, as shown bythe induction of secondary hyperalgesia in our data (Fig. 4D). Secondary hyperalgesia also developed during prolonged LFS (5 Hz for 120 min) by an intracutaneous needle electrode (Koppert et al., 2001). Recent electrophysiological data indicate that some spinal neurons (those projecting to the periaqueductal gray) may develop LTP after LFS at 2 Hz (Ikeda and Sandkühler, 2003). These phenomena, however, deserve further study.

Conclusions

The present findings provide the missing link between clinical observations in humans (neurogenic hyperalgesia after tissue damage or lesions of the nervous system) and a basic neurobiological mechanism (synaptic LTP) that was proposed to turn acute pain into chronic pain. It may not be surprising that long-term memory in the human injury-detection system is found to be related to the highly conserved mechanism of learning (LTP), considering the fact that during evolution, strong injury-related selection pressures have shaped the nervous systems of all species (Walters, 1994). Research in many areas of neuroscience uses experimental paradigms that simulate aspects of actual or threatened injury to study mechanisms of learning and memory (e.g., conditioning of the gill withdrawal reflex in Aplysia) (Illich and Walters, 1997; Kandel, 2001). Our data show that neurogenic hyperalgesia, which is an aspect of injury-related behavior in humans, shares characteristics with both homosynaptic and heterosynaptic types of LTP.