Wide Dynamic Range Cells Research Articles

Hypothalamic Paraventricular Stimulation Inhibits Nociceptive Wide Dynamic Range Trigeminocervical Complex Cells via Oxytocinergic Transmission.

Oxytocinergic transmission blocks nociception at the peripheral, spinal, and supraspinal levels through the oxytocin receptor (OTR). Indeed, a neuronal pathway from the hypothalamic paraventricular nucleus (PVN) to the spinal cord and trigeminal nucleus caudalis (Sp5c) has been described. Hence, although the trigeminocervical complex (TCC), an anatomical area spanning the Sp5c, C1, and C2 regions, plays a role in some pain disorders associated with craniofacial structures (e.g., migraine), the role of oxytocinergic transmission in modulating nociception at this level has been poorly explored. Hence, in vivo electrophysiological recordings of TCC wide dynamic range (WDR) cells sensitive to stimulation of the periorbital or meningeal region were performed in male Wistar rats. PVN electrical stimulation diminished the neuronal firing evoked by periorbital or meningeal electrical stimulation; this inhibition was reversed by OTR antagonists administered locally. Accordingly, neuronal projections (using Fluoro-Ruby) from the PVN to the WDR cells filled with Neurobiotin were observed. Moreover, colocalization between OTR and calcitonin gene-related peptide (CGRP) or OTR and GABA was found near Neurobiotin-filled WDR cells. Retrograde neuronal tracers deposited at the meningeal (True-Blue, TB) and infraorbital nerves (Fluoro-Gold, FG) showed that at the trigeminal ganglion (TG), some cells were immunopositive to both fluorophores, suggesting that some TG cells send projections via the V1 and V2 trigeminal branches. Together, these data may imply that endogenous oxytocinergic transmission inhibits the nociceptive activity of second-order neurons via OTR activation in CGRPergic (primary afferent fibers) and GABAergic cells.

The role of the endocannabinoid 2-arachidonoylglycerol in the in vivo spinal oxytocin-induced antinociception in male rats

Oxytocin receptor (OTR) activation at the spinal level produces antinociception. Some data suggest that central OTR activation enhances social interaction via an increase of endocannabinoids (eCB), but we do not know if this could occur at the spinal level, modulating pain transmission. Considering that oxytocin via OTR stimulates diacylglycerol formation, a key intermediate in synthesizing 2-arachidonylglycerol (2-AG), an eCB molecule, we sought to test the role of the eCB system on the spinal oxytocin-induced antinociception. Behavioral and electrophysiological experiments were conducted in naïve and formalin-treated (to induce long-term mechanical hypersensitivity) male Wistar rats. Intrathecal RHC 80267 injections, an inhibitor of the enzyme diacylglycerol lipase (thus, decreasing 2-AG formation), produces transient mechanical hypersensitivity, an effect unaltered by oxytocin but reversed by gabapentin. Similarly, in in vivo extracellular recordings of naïve spinal wide dynamic range cells, juxtacellular picoinjection of RHC 80267 increases the firing of nociceptive Aδ-, C-fibers, and post-discharge, an effect unaltered by oxytocin. Interestingly, in sensitized rats, oxytocin picoinjection reverses the RHC 80627-induced hyperactivity of Aδ-fibers (but not C- or post-discharge activity). In contrast, a sub-effective dose of JZL184 (a monoacylglycerol lipase inhibitor, thus favoring 2-AG levels), which does not have per se an antinociceptive effect in the formalin-induced hypernociception, the oxytocin-induced antinociception is boosted. Similarly, electrophysiological experiments suggest that juxtacellular JZL184 diminishes the neuronal firing of nociceptive fibers, and co-injection with oxytocin prolongs and enhances the antinociceptive effect. These data may imply that 2-AG formation may play a role in the spinal antinociception induced by oxytocin.

The Rostral Agranular Insular Cortex, a New Site of Oxytocin to Induce Antinociception.

The rostral agranular insular cortex (RAIC) is a relevant structure in nociception. Indeed, recruitment of GABAergic activity in RAIC promotes the disinhibition of the locus ceruleus, which in turn inhibits (by noradrenergic action) the peripheral nociceptive input at the spinal cord level. In this regard, at the cortical level, oxytocin can modulate the GABAergic transmission; consequently, an interaction modulating nociception could exist between oxytocin and GABA at RAIC. Here, we tested in male Wistar rats the effect of oxytocin microinjection into RAIC during an inflammatory (by subcutaneous peripheral injection of formalin) nociceptive input. Oxytocin microinjection produces a diminution of (1) flinches induced by formalin and (2) spontaneous firing of spinal wide dynamic range cells. The above antinociceptive effect was abolished by microinjection (at RAIC) of the following: (1) L-368899 (an oxytocin receptor [OTR] antagonist) or by (2) bicuculline (a preferent GABAA receptor blocker), suggesting a GABAergic activation induced by OTR. Since intrathecal injection of an α2A-adrenoceptor antagonist (BRL 44408) partially reversed the oxytocin effect, a descending noradrenergic antinociception is suggested. Further, injection of L-368899 per se induces a pronociceptive behavioral effect, suggesting a tonic endogenous oxytocin release during inflammatory nociceptive input. Accordingly, we found bilateral projections from the paraventricular nucleus of the hypothalamus (PVN) to RAIC. Some of the PVN-projecting cells are oxytocinergic and destinate GABAergic and OTR-expressing cells inside RAIC. Aside from the direct anatomic link between PVN and RAIC, our findings provide evidence about the role of oxytocinergic mechanisms modulating the pain process at the RAIC level.SIGNIFICANCE STATEMENT Oxytocin is a neuropeptide involved in several functions ranging from lactation to social attachment. Over the years, the role of this molecule in pain processing has emerged, showing that, at the spinal level, oxytocin blocks pain transmission. The present work suggests that oxytocin also modulates pain at the cortical insular level by favoring cortical GABAergic transmission and activating descending spinal noradrenergic mechanisms. Indeed, we show that the paraventricular hypothalamicnucleus sends direct oxytocinergic projections to the rostral agranular insular cortex on GABAergic and oxytocin receptor-expressing neurons. Together, our data support the notion that the oxytocinergic system could act as an orchestrator of pain modulation.

Recurrent antinociception induced by intrathecal or peripheral oxytocin in a neuropathic pain rat model.

The search for new ligands to treat neuropathic pain remains a challenge. Recently, oxytocin has emerged as an interesting molecule modulating nociception at central and peripheral levels, but no attempt has been made to evaluate the effect of recurrent oxytocin administration in neuropathic pain. Using male Wistar rats with spinal nerve ligation, we evaluated the effects of recurrent spinal (1nmol; given by lumbar puncture) or peripheral (31nmol; given by intraplantar injection in the ipsilateral paw to spinal nerve ligation) oxytocin administration on pain-like behavior in several nociceptive tests (tactile allodynia and thermal and mechanical hyperalgesia) on different days. Furthermore, we used an electrophysiological approach to analyze the effect of spinal 1nmol oxytocin on the activity of spinal dorsal horn wide dynamic range cells. In neuropathic rats, spinal or peripheral oxytocin partially restored the nociceptive threshold measured with the von Frey filaments (tactile allodynia), Hargreaves (thermal hyperalgesia) and Randall-Selitto (mechanical hyperalgesia) tests for 12 days. These results agree with electrophysiological data showing that spinal oxytocin diminishes the neuronal firing of the WDR neurons evoked by peripheral stimulation. This effect was associated with a decline in the activity of primary afferent Aδ- and C-fibers. The above findings show that repeated spinal or peripheral oxytocin administration attenuates the pain-like behavior in a well-established model of neuropathic pain. This study provides a basis for addressing the therapeutic relevance of oxytocin in chronic pain conditions.

Duloxetine Protects against Oxaliplatin-Induced Neuropathic Pain and Spinal Neuron Hyperexcitability in Rodents.

Oxaliplatin is a widely used chemotherapy agent, but induces serious peripheral neuropathy. Duloxetine is a dual reuptake inhibitor of serotonin and norepinephrine, and is shown to be effective against pain. However, whether and how duloxetine can attenuate oxaliplatin-induced allodynia in rodents is not clearly understood. A single injection of oxaliplatin (6 mg/kg, intraperitoneal; i.p.) induced a cold and mechanical allodynia, which was assessed by acetone and von Frey filament tests, respectively. When significant allodynic signs were observed, three different doses of duloxetine (10, 30, and 60 mg/kg, i.p.) were injected. Administration of 30 and 60 mg/kg of duloxetine significantly reduced the allodynia, whereas 10 mg/kg did not. By using an in vivo extracellular recording method, we further confirmed that 30 mg/kg of duloxetine could significantly inhibit the hyperexcitability of spinal wide dynamic range (WDR) cells. The anti-allodynic effect of duloxetine was completely blocked by an intrathecal injection of phentolamine (non-selective α-adrenergic receptor antagonist, 20 μg), or prazosin (α1-adrenergic receptor antagonists, 10 μg); however, idazoxan (α2-adrenergic receptor antagonist, 10 μg) did not block it. In conclusion, we suggest that duloxetine may have an effective protective action against oxaliplatin-induced neuropathic pain and spinal hyperexcitability, which is mediated by spinal α1-adrenergic receptors.

Differential effect of motor cortex stimulation on unit activities in the ventral posterior lateral thalamus in cats.

Although motor cortex stimulation (MCS) is being increasingly used to treat chronic refractory neuropathic pain in humans, its mechanisms of action remain elusive. Studies in animals have suggested the involvement of subcortical structures, in particular, the thalamus. Most of these studies have been performed in rats, a species presenting significant differences in thalamic anatomy and function relative to primates, in particular, a very limited number of thalamic GABA interneurons. The aim of this study was to investigate the effects of MCS on single-unit activities of the thalamic ventral posterior lateral (VPL) nucleus in cats, which contains substantial numbers of GABA interneurons. Spontaneous and evoked activities of VPL units were studied before and after MCS. Motor cortex stimulation induced significant depression of the wide-dynamic-range (WDR) cells' firing rate, concomitant with activity enhancement of nonnociceptive (NN) units. More than half of WDR cells showed a significant decrease in the firing rate, while a similar proportion of NN units exhibited the opposite after-effect. Maximal firing attenuation of WDR cells occurred when the MCS location matched somatotopically their receptive field. Repetition of MCS runs led to an accentuation of WDR depression. After peripheral stimulation, evoked activity in each cell showed MCS effects similar to those observed in spontaneous activity. These data demonstrate a selective top-down inhibition by MCS of nonspecific nociceptive (WDR) cells, enhanced by somatotopic concordance and stimulation repetition, in parallel to facilitation of NN cells. These 2 outcomes may play a role in the complex analgesic effect of MCS observed in neuropathic pain conditions.

The potential role of serotonergic mechanisms in the spinal oxytocin-induced antinociception

The role of oxytocin (OXT) in pain modulation has been suggested. Indeed, hypothalamic paraventricular nuclei (PVN) electrical stimuli reduce the nociceptive neuronal activity (i.e., neuronal discharge associated with activation of Aδ- and C-fibers) of the spinal dorsal horn wide dynamic range (WDR) cells and nociceptive behavior. Furthermore, raphe magnus nuclei lesion reduces the PVN-induced antinociception, suggesting a functional interaction between the OXT and the serotoninergic system. The present study investigated in Wistar rats the potential role of spinal serotonergic mechanisms in the OXT- and PVN-induced antinociception. In long-term secondary mechanical allodynia and hyperalgesia induced by formalin or extracellular unitary recordings of the WDR cells we evaluated the role of 5-hydroxytryptamine (5-HT) effect on the OXT-induced antinociception. All drugs were given intrathecally (i.t.). OXT (1×10−5–1×10−4nmol) or 5-HT (1×10−3–1×10−1nmol) prevented the formalin-induced sensitization, an effect mimicked by PVN stimulation. Moreover, administration of OXT (1×10−5nmol) plus 5-HT (1×10−3nmol) at ineffective doses, produced antinociception. This effect was antagonized by: (i) d(CH2)5[Tyr(Me)2,Thr4,Tyr-NH29]OVT (oxytocin receptor antagonist; 2×10−2nmol); or (ii) methiothepin (a non-specific 5-HT1/2/5/6/7 receptor antagonist; 80nmol). Similar results were obtained with PVN stimulation plus 5-HT (5×10−5nmol). In WDR cell recordings, the PVN-induced antinociception was enhanced by i.t. 5-HT and partly blocked when the spinal cord was pre-treated with methiothepin (80nmol). Taken together, these results suggest that serotonergic mechanisms at the spinal cord level are partly involved in the OXT-induced antinociception.

High-Frequency Stimulation of Dorsal Column Axons: Potential Underlying Mechanism of Paresthesia-Free Neuropathic Pain Relief

Spinal cord stimulation (SCS) treats neuropathic pain through retrograde stimulation of dorsal column axons and their inhibitory effects on wide dynamic range (WDR) neurons. Typical SCS uses frequencies from 50-100 Hz. Newer stimulation paradigms use high-frequency stimulation (HFS) up to 10 kHz and produce pain relief but without paresthesia. Our hypothesis is that HFS preferentially blocks larger diameter axons (12-15 µm) based on dynamics of ion channel gates and the electric potential gradient seen along the axon, resulting in inhibition of WDR cells without paresthesia. We input field potential values from a finite element model of SCS into an active axon model with ion channel subcomponents for fiber diameters 1-20 µm and simulated dynamics on a 0.001 msec time scale. Assuming some degree of wave rectification seen at the axon, action potential (AP) blockade occurs as hypothesized, preferentially in larger over smaller diameters with blockade in most medium and large diameters occurring between 4.5 and 10 kHz. Simulations show both ion channel gate and virtual anode dynamics are necessary. At clinical HFS frequencies and pulse widths, HFS preferentially blocks larger-diameter fibers and concomitantly recruits medium and smaller fibers. These effects are a result of interaction between ion gate dynamics and the "activating function" (AF) deriving from current distribution over the axon. The larger fibers that cause paresthesia in low-frequency simulation are blocked, while medium and smaller fibers are recruited, leading to paresthesia-free neuropathic pain relief by inhibiting WDR cells.

Immobilization stress sensitizes rat dorsal horn neurons having input from the low back.

Stress is known to promote several forms of muscle pain including non-specific low back pain. However, the question if stress alone activates nociceptive central neurons has not been studied systematically. Here, we investigated the influence of repeated immobilization stress on dorsal horn neurons and behaviour in the rat. The stress consisted of immobilization in a narrow tube for 1 h on 12 days. Single dorsal horn neurons were recorded with microelectrodes introduced into the spinal segment L2. In this segment, about 14% of the neurons responded to mechanical stimulation of the subcutaneous soft tissues of the low back in naïve rats. The neurons often behaved like wide dynamic range cells in that they had a low mechanical threshold and showed graded responses to noxious stimuli. The stress-induced changes in neuronal response behaviour were (1) appearance of new receptive fields in the deep tissues of the hindlimb, (2) increased input from deep soft tissues, but unchanged input from the skin and (3) significant increase in resting activity. Surprisingly, the pressure-pain threshold of the low back remained unchanged, although dorsal horn neurons were sensitized. In the open field test, the rats showed signs of increased anxiety. This study shows that stress alone is sufficient to sensitize dorsal horn neurons. The data may explain the enhanced pain low back patients report when they are under stress. The increased resting discharge may lead to spontaneous pain.

Intracisternal injection of palmitoylethanolamide inhibits the peripheral nociceptive evoked responses of dorsal horn wide dynamic range neurons.

Endogenous palmitoylethanolamide (PEA) has a key role in pain modulation. Central or peripheral PEA can reduce nociceptive behavior, but no study has yet reported a descending inhibitory effect on the neuronal nociceptive activity of Aδ- and C-fibers. This study shows that intracisternal PEA inhibits the peripheral nociceptive responses of dorsal horn wide dynamic range cells (i.e., inhibition of Aδ- and C-fibers), an effect blocked by spinal methiothepin. These results suggest that a descending analgesic mechanism mediated by the serotonergic system could be activated by central PEA.

Wide‐dynamic‐range neurons are heterogeneous in windup responsiveness to changes in stimulus intensity and isoflurane anesthesia level in mice

The windup phenomenon in wide-dynamic-range (WDR) neurons represents a short-term neuronal sensitization to repetitive noxious inputs that may share similar mechanisms with those that trigger the development of persistent pain and hyperalgesia. Some WDR cells are readily sensitized and express prominent windup (windup(+)), whereas others do not (windup(-)). We recorded extracellular single-unit activity of deep laminae WDR neurons (350-700 microm) in C57BL/6 mice to determine how changes in stimulus intensity (1x and >2x C-component threshold, n = 53) and concentrations of isoflurane anesthesia (2.0% and 1.0%, n = 30) might differently modulate windup responsiveness in windup(+) and windup(-) cells. Two principally different analysis methods [absolute windup (the number of action potentials) and relative windup (the percentage of action potentials evoked by the first stimulus of the train)] were used to interpret windup data. We observed that increasing the stimulus intensity and decreasing the isoflurane concentration: 1) facilitated windup generation at 0.2-Hz stimulation and significantly enhanced absolute windup at both 0.2-Hz and 0.5-Hz stimulation predominantly in windup(+) cells but did not confer windup capability on windup(-) cells and 2) significantly increased relative windup at 0.2-Hz, but not 0.5-Hz, stimulation in windup(+) cells. Our findings advance our understanding of the neurobiology of deep WDR neurons in mice and demonstrate that two populations of cells differ in their windup responsiveness to changes in experimental conditions. We also elucidate the usefulness and potential limitations of two widely used methods for calculating and presenting windup data.

GABA-mediated oxytocinergic inhibition in dorsal horn neurons by hypothalamic paraventricular nucleus stimulation

In anaesthetized rats, we tested whether the unit activity of dorsal horn neurons that receive nociceptive input is modulated by electrical stimulation of the hypothalamic paraventricular nucleus (PVN). An electrophysiological mapping of dorsal horn neurons at L3–L4 let us choose cells responding to a receptive field located in the toes region of the left hindpaw. Dorsal horn neurons were classified according to their response properties to peripheral stimulation. Wide Dynamic Range (WDR) cells responding to electrical stimulation of the peripheral receptive field and presenting synaptic input of Aδ, Aβ, and C-fibers were studied. Suspected interneurons that are typically silent and lack peripheral receptive field responses were also analyzed. PVN electrical stimulation inhibits Aδ (− 55.0 ± 10.2%), C-fiber (− 73.1 ± 6.7%), and post-discharge (− 75.0 ± 8.9%) peripheral activation in WDR cells, and silent interneurons were activated. So, this last type of interneuron was called a PVN-ON cell. In WDR cells, the inhibition of peripheral responses caused by PVN stimulation was blocked by intrathecal administration of a specific oxytocin antagonist or bicuculline. However, PVN-ON cell activation was blocked by the same specific oxytocin antagonist, but not by bicuculline. Our results suggest that PVN stimulation inhibits nociceptive peripheral-evoked responses in WDR neurons by a descending oxytocinergic pathway mediated by GABAergic PVN-ON cells. We discuss our observation that the PVN electrical stimulation selectively inhibits Aδ and C-fiber activity without affecting Aβ fibers. We conclude that Aδ and C-fibers receive a presynaptic inhibition mediated by GABA.

Windup in Dorsal Horn Neurons Is Modulated by Endogenous Spinal μ-Opioid Mechanisms

The mu-opioid receptor (MOR) plays a critical role in morphine analgesia and nociceptive transmission. However, the physiological roles for endogenous MOR mechanisms in modulating spinal nociceptive transmission, and particularly in the enhanced excitability of spinal nociceptive neurons after repeated noxious inputs, are less well understood. Using a MOR gene knock-out (-/-) approach and an MOR-preferring antagonist, we investigated the roles of endogenous MOR mechanisms in processing of acute noxious input and in neuronal sensitization during windup-inducing stimuli in wide dynamic range (WDR) neurons. Extracellular single-unit activity of WDR neurons was recorded in isoflurane-anesthetized MOR(-/-) and wild-type C57BL/6 mice. There were no significant differences between the genotypes in the responses of deep WDR cells to acute mechanical stimuli, graded electrical stimuli, and noxious chemical stimuli applied to the receptive field. Intracutaneous electrical stimulation at 1.0 Hz produced similar levels of windup in both genotypes. In contrast, 0.2 Hz stimulation induced significantly higher levels of windup in MOR(-/-) mice compared with the wild-type group. In wild-type mice, spinal superfusion with naloxone hydrochloride (10 mM, 30 microl) significantly enhanced windup to 0.2 Hz stimulation in both deep and superficial WDR cells. A trend toward facilitation of windup was also observed during 1.0 Hz stimulation after naloxone treatment. These results suggest that endogenous MOR mechanisms are not essential in the processing of acute noxious mechanical and electrical stimuli by WDR neurons. However, MORs may play an important role in endogenous inhibitory mechanisms that regulate the development of spinal neuronal sensitization.

Differential electrophysiological effects of brain-derived neurotrophic factor on dorsal horn neurons following chronic spinal cord hemisection injury in the rat

To assess the role of brain-derived neurotrophic factor (BDNF) in nociceptive processing after chronic lateral spinal cord hemisection injury (SCI) at T13, we studied the effects of BDNF on evoked activity of dorsal horn wide dynamic range (WDR) neurons. Evoked responses of WDR cells ( n=34 total) at L3–L5 were characterized electrophysiologically after spinal administration of vehicle, or BDNF (10 μg). In hemisected animals, application of BDNF to the surface of the cord resulted in reductions in evoked activity in 24 of 32 cells (75%), and enhancement of evoked activity in eight of 32 (25%) cells. Phosphate-buffered saline-receiving animals demonstrated evoked response rates of between 75 and 93 Hz, while BDNF(−) cells had evoked rates from between 20 and 41 Hz, and BDNF(+) activities were between 80 and 119 Hz, significant changes of 76 and 124%, respectively. Effects were bilateral and differences in sidedness were not observed. These results further implicate BDNF in nociceptive processing, but suggest a complex role after chronic SCI.

Reduction in inflammation-induced sensitization of dorsal horn neurons by transcutaneous electrical nerve stimulation in anesthetized rats.

Transcutaneous electrical nerve stimulation (TENS) is utilized to treat a variety of painful conditions. Inflamed animals present with an increased response to noxious stimuli, i.e., hyperalgesia, at the site of injury (primary hyperalgesia) and outside the site of injury (secondary hyperalgesia). Further, following acute inflammation, dorsal horn neurons show an increased responsiveness to peripherally applied stimuli, which has been termed sensitization. Previous studies demonstrate a reduction in dorsal horn neuron activity following TENS treatment in normal animals and a reduction in primary and secondary hyperalgesia in acutely inflamed animals. The purpose of this study was to examine the effects of TENS on dorsal horn neurons sensitized by acute inflammation. Extracellular recordings from wide dynamic range (WDR), high threshold (HT) and low threshold (LT) dorsal horn neurons in anesthetized rats were assessed for spontaneous activity, responses to innocuous and noxious mechanical stimulation and receptive field size. Responses were measured before and 3 h after induction of inflammation, and immediately and 1 h after application of either high (100 Hz) or low (4 Hz) frequency TENS (motor intensity, pulse duration = 100 microseconds). TENS was applied to the inflamed paw to encompass the receptive field of the neuron for 20 min. WDR and HT dorsal horn neurons sensitized to mechanical stimulation after induction of inflammation. Application of either high or low frequency TENS to the inflamed paw reduced both innocuous and noxious evoked responses of WDR and HT dorsal horn neurons immediately and 1 h after treatment with TENS. Comparison of responses after TENS with baseline responses showed that the evoked responses in the majority of WDR and HT cells returned to or fell below baseline responses. TENS had no effect on responses of LT neurons. In summary, central neuron sensitization is reduced by TENS and may underlie the reduction in hyperalgesia observed after treatment with TENS.

Different increase in C-fibre evoked responses after nociceptive conditioning stimulation in sham-operated and neuropathic rats

Mechanisms of neuropathic pain are not clear. Recently we showed long-term potentiation (LTP) of wide dynamic range (WDR) neurones after electrical conditioning stimulation of the sciatic nerve in normal rats. In this study we investigated the effects of the same conditioning on both the evoked responses of WDR cells and on vital signs in neuropathic rats. Nerve injury was produced by tight ligation of the L5 and L6 spinal nerves and a control group received sham surgery. The electrical conditioning gave a significantly smaller LTP in neuropathic rats as compared to controls. Despite giving a smaller LTP in neuropathic rats, the rise in mean arterial blood pressure and heart rate induced by the conditioning was exactly the same as in the control group. The threshold for activating C-fibres was significantly lower and the baseline tended to be higher in neuropathic rats. Thus, it is suggested that the nerve injury itself is likely to induce an LTP-like state in the neurones studied. Further studies using the LTP-inducing model can provide new information on the mechanisms of neuropathic pain.

Interactions between brainstem and trigeminal neurons detected by cross-spectral analysis

Cells in the nucleus raphe magnus that are inhibited by noxious skin stimuli (off-cells) have been postulated to suppress pain by continuously inhibiting spinal and trigeminal nociceptive neurons. To test this hypothesis, spontaneous activity was simultaneously recorded from off-cells ( n=15) and wide-dynamic range cells ( n=27) of the trigeminal complex (subnucleus interpolaris), in rats anesthetized with pentobarbital. Most off-cells ( n=14) had rhythmic interspike intervals, their modes averaging 106 ms. No trigeminal cell fired rhythmically. Rhythmic firing was defined quantitatively: the autospectrum's peak power had to exceed 1.75 times its asymptote. This formula was obtained by generalizing from a natural cut-off in the theoretical autospectrum for serially uncorrelated, gamma-distributed intervals, whose firing can be varied from Poissonian to highly regular by adjusting one parameter. It encompasses the qualitative judgement of autocorrelograms commonly made in neurophysiology. Cross-correlograms ( n=29) appeared noisy and otherwise featureless. However, their power spectra (cross-periodograms) sometimes showed significant peaks, compared with simulated non-interactive distributions. The latter were generated by interchanging the raphe interval sequences at one random point (as in cutting a deck of cards), thus retaining most of their serial correlation. Of 29 cross-periodograms, 21 were significant at 1 to 13 frequencies (100 points, 0.4 to 39 Hz). These frequencies were often near the peak raphe power, and sometimes near its harmonics too. Furthermore, cross-spectral phase angles at peak power were non-uniform, most falling between 0 and 180 degrees (unit vector sum 60°, n=20). To understand why the frequency domain gave better detection, cross-spectra and cross-correlations were modeled theoretically by convolving idealized input autocorrelations and synaptic response functions. This demonstrated that rhythmic firing is insufficient for better frequency-domain detection, and that serially correlated input intervals or non-additive synaptic responses are necessary. The conclusion was confirmed by stochastic simulation of a simple non-additive synapse, that required successful input spikes to fall within a specified interval of the preceding spike. Experimentally, serial correlation was found in 12 of the 15 raphe cells, and in 20 of the 27 trigeminal cells. It is proposed that the weak experimental cross-correlograms arise because many asynchronous raphe inputs converge on each trigeminal cell, possibly to optimize the resting suppression of pain. The distribution of cross-spectral phase angles at peak raphe power suggested that raphe spikes arriving at the synapses' preferred interval cause a fall in trigeminal activity. In general, cross-spectral analysis can sometimes uncover influences hidden in cross-correlograms, but the firing of one neuron must be rhythmic and non-renewal, or else certain input intervals must be favored in synaptic transmission.

Incision-induced Changes in Receptive Field Properties of Rat Dorsal Horn Neurons

To learn more about pain mechanisms produced by surgery, responses of wide dynamic range (WDR) and high threshold (HT) dorsal horn neurons were studied before and after an incision. For this study, an incision was made in a mechanically insensitive area of the receptive field (RF) of the dorsal horn neuron in the plantar aspect of the foot and changes in mechanical response properties were studied. Action potentials from single dorsal horn neurons were recorded in halothane anesthetized rats and these neurons were characterized as WDR or HT. Changes in background activity and responses to a variety of mechanical stimuli adjacent to the incision, distant to the injury, and in areas throughout the hindquarters were recorded. Fifty neurons were recorded (29 WDR, 21 HT cells); only nine of these had a sustained increase in background activity after incision. Marked decreases in threshold to von Frey filaments applied adjacent to the wound occurred in 9 of 28 WDR neurons but in none of 21 HT cells. Von Frey filament thresholds distant to the incision were largely not changed. A blunt mechanical stimulus activated 18 of 22 WDR neurons when applied directly on the incision. HT cells were largely not excited by this mechanical stimulus after incision. The RF to pinch was enlarged in 31 neurons to include areas outside the injury. Pinch RFs of both WDR and HT cells expanded. These results suggest that incisions in mechanically insensitive areas of the RF of dorsal horn neurons produced little change in background activity; expansion of pinch RFs outside the injury was common. Changing a mechanically insensitive area of the RF of WDR neurons to a mechanically sensitive area by an incision could contribute to pain behaviors that indicate primary mechanical hyperalgesia in behavioral studies.

Responses of primate spinomesencephalic tract cells to intradermal capsaicin

The responses of 32 spinomesencephalic tract cells to intradermal capsaicin were examined in anesthetized monkeys. Wide dynamic range ( n=20) and nociceptive specific ( n=6) cells showed two types of excitatory responses to intradermal injection of capsaicin. The first excitatory response shown by the majority of wide dynamic range ( n=13) and nociceptive specific ( n=4) cells was consistent with their sensitization by capsaicin. The cells showed an acute and prolonged increase in ongoing activity with capsaicin injection. Responses to mechanical stimuli were substantially increased after capsaicin and an expansion of receptive field was often observed. The responses of the same cells to excitatory amino acid agonists applied locally by iontophoresis also increased. All cells showing sensitization were antidromically activated from periaqueductal gray regions dorsal to the sulcus limitans. Electrical stimulation at these sites did not affect the ongoing or evoked discharges of these cells. The second excitatory response of wide dynamic range ( n=5) and nociceptive specific ( n=1) cells was a novel pattern not consistent with sensitization. These cells nevertheless showed an acute and prolonged increase in background activity after capsaicin injection. Yet, there was no change or a decrease in responses to cutaneous stimuli, no evidence for change in receptive field size and no increase in responses to locally released excitatory amino acids. These cells projected to regions in the periaqueductal gray ventral to the sulcus limitans. Electrical stimulation at these sites produced a decrease in spontaneous activity of the same cell. Low threshold mesencephalic-projecting neurons ( n=6) showed a single inhibitory pattern ( n=4) of responses to capsaicin. The injection produced an acute decrease in spontaneous activity that was sustained for at least 30 min after injection. The responses to cutaneous stimuli and to excitatory amino acids were also substantially reduced. Low threshold cells were found that projected to both dorsal-lateral and ventral-lateral regions of the periaqueductal gray. In summary, three patterns of responses shown by primate spinomesencephalic tract cells to intradermal capsaicin appear dependent on the functional regions of the periaqueductal gray to which they project. These results suggest that inputs of spinomesencephalic tract neurons to the periaqueductal gray may evoke important components of the systemic response to the neurogenic hyperalgesia produced by intradermal capsaicin.

Nitric oxide-mediated spinal disinhibition contributes to the sensitization of primate spinothalamic tract neurons.

This study concentrated on whether an increase in spinal nitric oxide (NO) diminishes inhibition of spinothalamic tract (STT) cells induced by activating the periaqueductal gray (PAG) or spinal glycinergic and GABAergic receptors, thus contributing to the sensitization of STT neurons. A reduction in inhibition of the responses to cutaneous mechanical stimuli induced by PAG stimulation was seen in wide dynamic range (WDR) STT cells located in the deep layers of the dorsal horn when these neurons were sensitized during administration of a NO donor, 3-morpholinosydnonimine (SIN-1), into the dorsal horn by microdialysis. In contrast, PAG-induced inhibition of the responses of high-threshold (HT) and superficial WDR STT cells was not significantly changed by spinal infusion of SIN-1. A reduction in PAG inhibition when STT cells were sensitized after intradermal injection of capsaicin could be nearly completely blocked by pretreatment of the dorsal horn with a NO synthase inhibitor, 7-nitroindazole. Moreover, spinal inhibition of nociceptive activity of deep WDR STT neurons elicited by iontophoretic release of glycine and GABA agonists was attenuated by administration of SIN-1. This change paralleled the change in PAG-induced inhibition. However, the inhibition of HT and superficial WDR cells induced by glycine and GABA release did not show a significant change when SIN-1 was administered spinally. Combined with our recent results, these data show that the effectiveness of spinal inhibition can be reduced by the NO/cGMP pathway. Thus disinhibition may constitute one mechanism underlying central sensitization.