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Background & Aims: Discomfort and pain are the principal conscious sensations that arise from the viscera, and both are increased in frequency and intensity in patients with a functional bowel disorder. Visceral receptors, perhaps sensitized, may contribute to these altered sensations. The aim of this study was to evaluate the effects of opioid receptor-selective agonists on afferent fibers innervating the colon., Methods: Mechanosensitive pelvic nerve afferent fibers were recorded from the decentralized S1 dorsal root in anesthetized rats. The effects of opioid agonists, given intra-arterially, were studied based on the fiber's responses to noxious colorectal distention (CRD) (80 mm Hg, 30 seconds)., Results: A total of 115 distention-sensitive fibers innervating the colon were studied, including 32 that were studied after colonic inflammation with 2.5% acetic acid. Neither mu-(morphine and fentanyl) nor delta- ([D-Pen2, D-Pen5]enkephalin- and SNC-80) opioid receptor agonists affected responses to CRD. In contrast, kappa- (U-50,488 and fedotozine) opioid receptor agonists dose-dependently attenuated responses to CRD. Acetic acid sensitized about half of the fibers studied, but neither the potency nor the efficacy of U-50, 488 or FDZ were changed after colonic inflammation., Conclusions: These results suggest a role for peripheral kappa-opioid receptors in the modulation of visceral nociception.

There is general agreement that activation of the N-methyl-D-aspartate receptor is involved in thermal hyperalgesia. However, there is less agreement on the specific intracellular events subsequent to receptor activation and the involvement of other excitatory amino acid receptors in thermal hyperalgesia. In the present study, we found that the intrathecal administration of N-methyl-D-aspartate produced a dose- (1 fmol-1 pmol) and time-dependent thermal hyperalgesia. In contrast, over the dose range tested, intrathecal administration of either alpha-amino-3-hydroxy-5-methylisoxazole-4-proprionate (AMPA; 10 fmol-100 pmol), 1,3-trans-1-aminocyclopentyl-1,3-dicarboxylate (10 fmol-100 pmol), quisqualate (10 pmol-5 nmol) or a 1:1 combination of AMPA and 1,3-trans-1-aminocyclopentyl-1,3-dicarboxylate (total dose 20 fmol-200 pmol) did not produce any evidence of thermal hyperalgesia; greater doses produced a caudally-directed biting and scratching behavior that precluded testing in the paradigm used. A fixed dose of 1,3-trans-1-aminocyclopentyl-1,3-dicarboxylate (100 pmol) did, however, potentiate the effects of N-methyl-D-aspartate (1-100 fmol). Thermal hyperalgesia produced by N-methyl-D-aspartate (1 pmol) was attenuated by intrathecal administration of the N-methyl-D-aspartate receptor-selective antagonist 2-amino-5-phosphonopentanoate (100 pmol), but not by the AMPA receptor-selective antagonist 6,7-dinitroquinoxaline-2,3-dione (1 nmol) or the metabotropic receptor antagonist 2-amino-3-phosphonoproprionate (10 nmol). In a second series of experiments, we examined the role of different signal transduction systems in acute N-methyl-D-aspartate-produced thermal hyperalgesia. N-Methyl-D-aspartate-produced thermal hyperalgesia (1 pmol) was attenuated by intrathecal hemoglobin (1-100 pmol) and dose-dependently by intrathecal N(G)-nitro-L-arginine methyl ester (10 pmol-l nmol), Methylene Blue (10 pmol-l nmol) and chelerythrine (1-100 pmol), suggesting that acute N-methyl-D-aspartate-mediated thermal hyperalgesia involves activation of nitric oxide synthase and protein kinase C. In contrast, N-methyl-D-aspartate-produced thermal hyperalgesia was unaffected by intrathecal administration of the phospholipase A2 inhibitor mepacrine (10 nmol) or the phospholipase C inhibitor neomycin (10 nmol). While prostaglandins and leukotrienes have been suggested to play a role in hyperalgesia, N-methyl-D-aspartate-produced thermal hyperalgesia (1 pmol) was unaffected by the non-selective eicosanoid inhibitor nordihydroguaiarate (1 nmol), the cyclo-oxygenase selective inhibitor indomethacin (10 nmol) or the lipoxygenase selective inhibitor baicalein (1 nmol). The results of the present study suggest that acute thermal hyperalgesia can be produced by activation of N-methyl-D-aspartate receptors. Activation of AMPA, metabotropic or co-activation of AMPA and metabotropic glutamate receptors, at the doses tested, did not produce an acute thermal hyperalgesia. The thermal hyperalgesia produced by N-methyl-D-aspartate is mediated by activation of nitric oxide synthase and protein kinase C, but not by phospholipase C, phospholipase A2, cyclo-oxygenase or lipoxygenase. Collectively, the results are consistent with a role for spinal N-methyl-D-aspartate receptors, nitric oxide and protein kinase C in thermal hyperalgesia.

We have previously reported that repetitive, noxious colorectal distention (CRD) induces c-Fos in the lumbosacral spinal cord. This study examined the effects of the analgesics morphine and tramadol on c-Fos expression resulting from noxious CRD in the rat. Pre-treatment (30 min or 1 min, i.v.) with morphine (1.25 mg/kg-5.0 mg/kg) or tramadol (1 mg/kg-20 mg/kg) dose-dependently attenuated c-Fos expression to CRD in all areas of the L6-S1 spinal gray matter. The highest dose of morphine was equipotent to the highest dose of tramadol. Repetitive dosing (1/4 of the greatest dose every 30 min) was as effective as a single bolus dose for both drugs. The visceromotor response to CRD was dose-dependently attenuated by tramadol and was reversed by naloxone. However, the dose of tramadol that eliminated the visceromotor response (7% of control) reduced the c-Fos expression to 47% of control. These results demonstrate that these two analgesics attenuate immediate-early gene expression and the visceromotor response to a noxious visceral stimulus and suggest that complete attenuation of c-Fos expression is not necessary for these compounds to produce analgesia to a noxious visceral stimulus.

Background: Amitriptyline and other tricyclic antidepressants exhibit high affinity binding to N-methyl-D-aspartate (NMDA) receptors in vitro and inhibit NMDA receptor activation-induced neuroplasticity in hippocampal slices. Because spinal NMDA receptor activation is believed to be central to generation and maintenance of hyperalgesic pain, the purpose of this study was to test whether intrathecal amitriptyline reduced inflammation-induced hyperalgesia in the rat., Methods: Rats were prepared with chronic lumbar intrathecal and femoral intravenous catheters and nociceptive threshold was assessed by hind paw withdrawal to a radiant heat stimulus. Rats received an injection of carrageenin in one hind paw followed by thermal paw withdrawal testing 3 hr later and intrathecal amitriptyline and/or intravenous morphine injection. In other rats, intrathecal NMDA injection was preceded by either intrathecal saline or 60 micrograms amitriptyline., Results: Intrathecal amitriptyline reversed thermal hyperalgesia in a dose-dependent manner, but had no effect on withdrawal latency of the contralateral, noninjected paw. Intrathecal phentolamine plus methysergide did not alter amitriptyline's effect, except at the lowest dose. Intravenous morphine increased paw withdrawal latency in both inflamed and control paws in a dose-dependent fashion, and morphine interacted additively with intrathecal amitriptyline to reverse hyperalgesia. Thermal hyperalgesia induced by NMDA was completely antagonized by intrathecal amitriptyline., Conclusions: Amitriptyline and other tricyclic antidepressants have been demonstrated to exhibit modest activity against clinical neuropathic pain after systemic administration. These data suggest that more profound pain relief might be obtained by intrathecal administration. Amitriptyline reverses hyperalgesia in rats by a mechanism unrelated to monoamine reuptake inhibition, and likely due to NMDA receptor antagonism.

Background: Systemically administered opioids induce analgesia in part by spinal noradrenergic, serotonergic, and cholinergic mechanisms. The current study tested whether antinociception from systemically administered opioids could therefore be enhanced by intrathecal injection of a monoamine reuptake inhibitor to potentiate the action of spinally released norepinephrine and serotonin (amitriptyline) and intrathecal injection of a cholinesterase inhibitor to potentiate the action of spinally released acetylcholine (neostigmine)., Methods: Rats were prepared with chronic lumbar intrathecal and femoral intravenous catheters and nociceptive threshold was assessed by hind paw withdrawal to a radiant heat stimulus. An isobolographic design was used to distinguish between additive and synergistic interactions., Results: Intravenous morphine and intrathecal neostigmine, but not intrathecal amitriptyline, caused dose-dependent antinociception alone. Combining any two of these three treatments yielded a synergistic interaction compared to each alone, whereas combining all three yielded an additive interaction compared to each two-way interaction. Intrathecal amitriptyline did not affect antinociception from intrathecal clonidine or intrathecal carbamylcholine., Conclusions: These data suggest that intrathecal doses of amitriptyline resulting in potentiation of intravenous morphine antinociception may not be adequate to block muscarinic receptors, because they did not affect carbamylcholine-induced antinociception. These results further support the relevance of spinal monoamine reuptake and cholinesterase inhibition to synergistically enhance analgesia from systemic opioids.

Objective and Methods: Epicardial application of pharmacologic agonists has been used to study nociceptive and reflex responses to agents such as bradykinin. We utilized a model where intrapericardial bradykinin was administered in a closed-chest rat. The procedure allows for reproducible administration of microliter doses of pharmacologic agonists in both conscious and anesthetized animals., Results: Bradykinin (BK) has been shown to produce sympathoexcitatory reflexes when applied to the heart. BK typically produced a dose-dependent (0.001-10 micrograms) decrease in arterial blood pressure and tachycardia in pentobarbital-anesthetized rats. In contrast, in alpha-chloralose-anesthetized or awake rats, pericardial administration of BK produced a dose-dependent (0.001-10 micrograms) increase in arterial blood pressure and tachycardia. Maximal cardiovascular changes were produced by 1 microgram BK. The maximum change in arterial pressure was +33.6 +/- 9% in awake, +38.9 +/- 6% in chloralose-anesthetized, and -20 +/- 7% in pentobarbital-anesthetized rats. In alpha-chloralose-anesthetized rats, tachyphylaxis to pericardial administration of 1 microgram BK occurred at 5 and 15, but not at 30 min dosing intervals. Administration of the receptor selective B2-antagonist D-Arg,[Hyp3,Thi5,8 D-Phe7]-BK (200 micrograms) or the mixed B2/B1 antagonist [Thi5,8,D-Phe7]-BK (200 micrograms), produced similar attenuation of the pressor and tachycardia responses to BK. Bilateral transection of the cervical vagus nerve, bilateral removal of the stellate ganglion or ganglion blockade (hexamethonium), but not administration of indomethacin, reduced the magnitude of the tachycardia to BK. Only ganglionic blockade significantly reduced the pressor response to BK., Conclusions: These results demonstrate that pericardial administration of BK produces a tachycardia and pressor effect in awake and alpha-chloralose-anesthetized rats and a tachycardia and depressor effect in pentobarbital-anesthetized rats. These responses appear to be mediated through activation of BK (presumably B2) receptors on cardiac vagal and sympathetic afferents, and may include a direct action of BK on the heart. This model of pericardial administration of pharmacologic agonists may be useful in studies of cardiac pain and reflex responses.

It is widely appreciated that visceral pain differs significantly from pain that arises from cutaneous structures. Visceral pain is difficult for both the patient and physician to localize because it is diffuse in character and is typically referred to cutaneous structures. Further, there are differences between acute, post-operative visceral pain and the altered sensations associated with the so-called functional bowel disorders (e.g. non-ulcer dyspepsia, non-cardiac chest pain and irritable bowel syndrome). A variety of considerations suggests that sensory inputs from the fiscera, like nociceptive inputs from the skin, can be sensitized. Accordingly, inputs from the viscera that are not typically perceived may give rise to discomfort and pain if either visceral afferent fibres are sensitized or central neurones undergo a change in excitability ('central sensitization') after persistent visceral input. The anatomy and potential mechanisms associated with visceral hyperalgesia will be considered as will new information about opioid modulation of visceral inputs to the spinal cord.

Recent studies have suggested that glia might play a more active role in synaptic function than previously thought. Therefore, the present studies have evaluated the potential role of spinal cord glia in acute nociceptive processing and in the thermal and mechanical hyperalgesia produced by peripheral injury. In the present experiments, we found that: (1) selective inhibition of glia metabolism with intrathecal (i.t.) administration of fluorocitrate (1 nmol) results in a marked, but reversible, attenuation of the persistent thermal and mechanical hyperalgesia produced by intraplantar zymosan (5 mg); (2) selective inhibition of the inducible form of nitric oxide synthase (iNOS) with i.t. aminoguanidine (1 pmol-1 nmol) resulted in a dose-dependent inhibition of the persistent thermal, but not mechanical hyperalgesia produced by intraplantar zymosan (5 mg); (3) i.t. coadministration of interleukin 1 beta (IL1 beta; 10 ng) and interferon gamma (IFN; 1000 U) resulted in expression of the message for iNOS 8 hr after administration assessed using reverse-transcription polymerase chain reaction (RT-PCR) and Southern blot analysis; and (4) i.t. administration of lipopolysaccharide (LPS; 150 micrograms) produced a time-dependent thermal hyperalgesia compared with saline treated-rats (15 microliters). There was no change in mechanical withdrawal thresholds over time following any treatment, except fluorocitrate. We have previously shown that NO plays a significant role in mechanisms of hyperalgesia. In the present experiments we have extended these observations and have now shown a role for iNOS, expressed by glia, in mechanisms of hyperalgesia. These results suggest an unexplored avenue for the development of potential new and novel therapies for pain control.

Visceral organs receive dual innervation from primary afferents commonly referred to as 'sympathetic' and 'parasympathetic' afferents. We have previously reported a significantly greater induction of immediate-early genes in the viscerotopically appropriate spinal cord segments receiving 'parasympathetic' afferent innervation (pelvic nerve) compared with those receiving 'sympathetic' afferent innervation (hypogastric nerve) following noxious colorectal distention. In this study, the nucleus of the solitary tract (NTS) and viscerotopically appropriate spinal cord segments were labeled immunocytochemically for c-Fos following noxious gastric or esophageal distention to determine if the differential labeling following 'sympathetic' (spinal) and 'parasympathetic' (vagal) visceral afferent input is a general phenomenon of the gastrointestinal tract. Gastric distention and esophageal distention induced considerable c-Fos in the NTS and virtually none in the thoracic spinal segments. These data suggest that 'parasympathetic' visceral afferents may be different than 'sympathetic' visceral afferents with respect to their ability to induce c-Fos following noxious visceral distention.

Objective: The recently proposed animal model of lumbar radiculopathy was used to investigate behavioral consequences and histologic changes in spinal nerve roots, dorsal root ganglia, and spinal nerves after the L4, L5, and L6 nerve roots were loosely ligated with either silk or chromic gut sutures in an attempt to better understand the pathophysiologic mechanisms that give rise to pain associated with lumbar radiculopathy., Summary of Background Data: Little is known about the pathophysiologic mechanisms that give rise to pain associated with lumbar radiculopathy. The recently proposed animal model of unilateral lumbar radiculopathy, which demonstrated an association with motor paresis and thermal hyperalgesia of the affected hind limb and showed evidence of spontaneous pain has been demonstrated, may serve as a vehicle to allow direct investigation of the nature of the pathophysiological mechanisms associated with lumbar radiculopathy., Methods: Three distinct treatments of the nerve roots were initially investigated: 1) a sham intervention, where the surgery simply exposed the nerve roots and dorsal root ganglion followed by standard closing procedures; 2) 4-0 silk ligature, where two loose ligatures of 4-0 silk were placed around the nerve roots; and 3) 4-0 chromic gut 2, where four 0.3 cm pieces of 4-0 chromic gut were laid adjacent to the nerve roots and secured by two loose ligatures of 4-0 chromic gut., Study Design: ANOVA techniques were used to test for differential effects across time for the three treatment groups in terms of animal function. A qualitative analysis of the histology of the ipsilateral and contralateral nerve roots, dorsal root ganglia, and spinal nerves was done to correlate histologic changes with behavioral changes., Results: Behavioral results were consistent with the previous study. Rats treated with chromic gut, but not silk, reliably demonstrated a prolonged thermal hyperalgesia that was maximal 2 weeks after surgery and lasted for up to 12 weeks. These behavioral changes, however, were not correlated with histologic changes in myelinated fiber content in the L4, L5, and L6 nerve roots, dorsal root ganglia and spinal nerves, the ipsilateral spinal nerved, dorsal root ganglia, and nerve roots of rates ligated with silk or chromic gut showed similar, significant, decreased in the number of large diameter myelinated fibers., Conclusions: These results suggest that mechanical constriction of the L4, L5, and L6 spinal nerve roots, as evidenced by a loss of myelinated fibers, is not sufficient to produce the behavioral effects associated with this model of lumbar radiculopathy. It is hypothesized that chemical factors from the chromic gut play a role in the pathophysiology and development of the behavioral, but not histological, changes in this model of lumbar radiculopathy.

The effects of N-methyl-D-aspartic acid (NMDA; 100 fmol-1 nmol) or quisqualic acid (QA; 10 pmol-10 nmol) on visceromotor and pressor responses to noxious colorectal distention (CRD; 40 mmHg, 20 s duration, interstimulus interval: 4 min) were studied in awake rats. Lesser doses of NMDA (100 fmol-1 pmol) administered intrathecally (i.t.) to the lumbar spinal cord produced a dose-dependent facilitation of visceromotor as well as pressor responses to CRD (maximum with 1 pmol NMDA at 1 min). The greatest dose tested (1 nmol) attenuated these responses (maximum at 1 min) and also produced a caudally-directed biting and scratching behavior accompanied by vocalizations. NMDA did not produce any of the above effects when administered i.t. to the thoracic spinal cord. I.t. pretreatment with the NMDA receptor antagonist, D-2-amino-5-phosphonovaleric acid (D-APV; 1 pmol), which produced no change in baseline activity or control responses, blocked all NMDA-produced effects in a reversible manner. QA produced dose-dependent inhibitory effects on visceromotor as well as pressor responses to noxious CRD when given i.t. to the lumbar spinal cord but not on administration to the thoracic spinal cord. Three nmol QA produced maximum inhibition at 2 min after administration and also produced caudally-directed biting and scratching. All of the QA-produced effects were reversibly blocked by i.t. pretreatment with the non-NMDA receptor antagonist, 6,7-dinitroquinoxaline-2,3-dione (DNQX; 3 nmol), which produced no change in baseline activity or control responses. We also examined the effects of NMDA and QA on responses to graded intensities of CRD. One pmol NMDA selectively facilitated visceromotor responses to CRD at distention pressures of 40 and 80 mmHg but not at 20 mmHg. In contrast, 3 nmol QA inhibited visceromotor responses to CRD at all intensities tested. In summary, these data suggest that activation of NMDA and non-NMDA receptors in the spinal cord differentially modulates visceral nociceptive input. Spinal segmental NMDA receptor activation produces selective facilitation of visceral nociceptive processing at noxious intensities of stimulation and may thereby contribute to central mechanisms underlying visceral hyperalgesia.

This report presents evidence that pericardial administration of a mixture of algogenic substances is a potentially useful model of cardiac nociception. In awake rats in which a looped silicone catheter had been placed in the pericardial sac at least 5 days previous, administration of a mixture containing equal concentrations of bradykinin (BK), acetylcholine (ACh), adenosine (ADEN), histamine (HIST), serotonin (5-HT) and prostaglandin E2 (PGE2) (total 25 nmol in 25 microliters) led to rapid acquisition of a passive avoidance behavior. In contrast, neither BK alone (5-25 nmol) nor the same mixture of ACh, ADEN, HIST, 5-HT and PGE2 without BK led to acquisition of the behavior or produced effects significantly different than produced by saline given into the pericardial sac in the same volume (25 microliters). Both BK and the mixture containing BK produced dose-dependent cardiovascular responses (pressor response and tachycardia) of similar magnitude. Neither saline nor the mixture without BK produced significant changes in mean arterial blood pressure or heart rate. In electrophysiological experiments in the same rats, thoracic spinal cord neurons responded dose-dependently to the mixture and, except for one neuron, responded also to BK in a dose-dependent manner. However, responses to BK, when compared to a similar dosage of BK contained in the mixture, were significantly less in magnitude and duration. All units received convergent somatic input from the thorax and all neurons also received convergent input from the esophagus. Balloon distension of the esophagus excited all units. Results of the behavioral characterization of algogenic substances administered into the pericardial sac of awake rats gave evidence of differences between the effects of BK and a mixture of six substances, including BK. BK in either of two dosages tested produced effects not different than saline while the mixture containing BK was aversive. In complementary electrophysiological studies, both BK and the mixture containing BK excited thoracic spinal cord neurons, suggesting that neuron responses to putative algogenic substances are not necessarily reliable measures of cardiac nociception.

Although physiological stimuli in the healthy gastrointestinal tract are generally not associated with conscious perception, chronic abdominal discomfort and pain are the most common symptoms resulting in patient visits with gastroenterologists. Symptoms may be associated with inflammatory conditions of the gut or occur in the form of so-called functional disorders. The majority of patients with functional disorders appear to primarily have inappropriate perception of physiological events and altered reflex responses in different gut regions. Recent breakthroughs in the neurophysiology of somatic and visceral sensation are providing a series of plausible mechanisms to explain the development of chronic hyperalgesia within the human gastrointestinal tract. A central concept to all these mechanisms is the development of hyperexcitability of neurons in the dorsal horn, which can develop either in response to peripheral tissue irritation or in response to descending influences originating in the brainstem. Taking clinical characteristics and the concept of central hyperexcitability into account, a model is proposed by which abdominal pain from chronic inflammatory conditions of the gut and functional bowel disorders such as noncardiac chest pain, nonulcer dyspepsia, and irritable bowel syndrome could develop by multiple mechanisms either alone or in combination.

1. Single-unit activity was recorded from S1 sacral dorsal root afferent fibers in the anesthetized rat. A total of 364 afferent fibers were identified by electrical stimulation of the pelvic nerve and subsequently tested for response to colorectal distension (CRD) and urinary bladder distension (UBD). Sixty-seven percent (n = 244) of the fibers were unmyelinated C-fibers and 33% (n = 120) were thinly myelinated A delta-fibers. 2. In three initial experiments, 35 fibers were identified by pelvic nerve stimulation and tested for response only to CRD; none of these fibers responded to CRD. In 20 subsequent experiments, 329 pelvic nerve afferent fibers were tested for response to CRD and UBD. Thirty-four percent (n = 112) of the 329 fibers were unresponsive to noxious CRD (80 mmHg) or to UBD (slow filling < or = 100 mmHg), 44% (n = 146) responded to UBD, 16% (n = 53) responded to CRD, and 6% (n = 18) responded to mechanical stimulation of the anal mucosa. 3. Of the total of 53 pelvic nerve afferent fibers that responded to CRD, 43 (81%) were C-fibers (mean: 1.5 m/s) and 10 (19%) were A delta-fibers (mean: 4.7 m/s). Fifteen of the CRD-sensitive fibers had no resting activity, whereas 38 fibers exhibited some resting activity (mean: 2.6 imp/s). 4. Reproducibility of responses to repeated CRD (80 mmHg, 30s, 10 trials at 4-min intervals) was tested in 17 fibers. In 16, responses to repeated distension were reproducible without evidence of facilitation or inhibition of subsequent responses. One fiber gave greater responses during the 9th and 10th trials. 5. Responses to graded CRD were studied in 44 fibers. All fibers exhibited monotonic, increasing stimulus-response functions < or = 80 mmHg of distension. Thresholds for response of the 44 fibers were determined after extrapolation of the least-squares linear-regression line to the ordinate and varied between 0 and 40 mmHg. Two populations of pelvic nerve afferent fibers in the colon were apparent: low threshold (LT) afferent fibers had a mean threshold of 2.9 mmHg (range: 0-10 mmHg; n = 34) and high threshold (HT) afferent fibers had a mean threshold of 32.6 mmHg (range: 28.0-40.0 mmHg; n = 10). 6. Chemosensitivity to bradykinin (BK) was tested in nine LT fibers. Seven fibers responded to BK (0.1 to 100 micrograms/kg ia) and two fibers did not respond up to 100 micrograms/kg of BK. Responses to BK tested in three fibers were dose dependent.(ABSTRACT TRUNCATED AT 400 WORDS)

Unilateral hindpaw inflammation produces several neurochemical changes in the ipsilateral spinal dorsal horn that have been interpreted as contributing to the associated hyperalgesia. NADPH-diaphorase histochemical staining was used to examine the response of a population of neurons 1, 2, 6 or 24 h following injection of 6 mg carrageenan into the left hindpaw of the rat. The resulting unilateral hindpaw inflammation produced a bilateral, time-dependent, reversible increase in the number of NADPH-diaphorase stained neurons in the lumbar spinal cord that peaked at 6 h. In laminae I-III, there was a significant increase in the number of NADPH-diaphorase stained neurons both ipsilateral (25.9 +/- 2.3) and contralateral (26.3 +/- 1.3) to the inflamed hindpaw relative to uninflamed, control animals (18.6 +/- 1.7, 17.4 +/- 1.7, respectively). A smaller but significant increase was observed in laminae IV-VII and X. Under dark field illumination, an increase in the number of densely stained neurons in laminae I-III was also observed to peak at 6 h. A greater percentage of the neurons observed under bright field illumination were visible under dark field illumination at 6 h (47%) compared to control (18%), suggesting an increase in the enzymatic activity of neurons in laminae I-III in addition to the increase in the number of neurons with threshold levels of NADPH-diaphorase activity. There was no consistent increase in this ratio over time in laminae IV-VII or X. Six hours following carrageenan, there was a bilateral 50% increase in the density of NADPH-diaphorase staining in the neuropil in the medial laminae I-III. Both spinal neurons and primary afferent axons contributed to this bilateral increase in staining as the number of NADPH-diaphorase stained dorsal root ganglion cell bodies increased 47% over control. In addition to the increase in staining in the lumbar spinal cord, at 6 h post carrageenan, there was a bilateral 23% increase over control in the number of stained neurons in the cervical dorsal spinal cord. For comparative purposes, the distribution of Fos-immunoreactive nuclei was compared to NADPH-diaphorase 6 h post carrageenan. The distribution of Fos-immunoreactive nuclei differed from the NADPH-diaphorase stained neurons, suggesting that two separate populations of neurons were stained. Unilateral hindpaw inflammation did not result in an increase in NADPH-diaphorase activity in the periaqueductal gray or the ventroposteriolateral thalamic nucleus. The relationship of NADPH-diaphorase to nitric oxide is discussed and it is concluded that NADPH-diaphorase,(ABSTRACT TRUNCATED AT 400 WORDS)

Stimulation-produced modulation from the rostral, medial medulla (RMM) on the spinal nociceptive tail-flick (TF) reflex and on lumbar spinal dorsal horn neuron responses to noxious cutaneous stimuli was studied in adult rats treated as neonates with capsaicin or vehicle. In vehicle-treated rats (n = 7), both descending facilitatory and inhibitory influences on the TF reflex were produced from the RMM. At 11/23 sites in the RMM, electrical stimulation produced biphasic modulatory effects. Electrical stimulation facilitated the spinal nociceptive TF reflex at low intensities (5-25 microA) and inhibited the TF reflex at greater intensities (50-200 microA). The mean threshold intensity of stimulation to inhibit the TF reflex (cut-off time = 7.0 s) was 66 microA (n = 11). At 11 of 23 sites, electrical stimulation only inhibited the TF reflex; the mean threshold intensity of stimulation to inhibit the TF reflex was 50 microA (n = 11). At one stimulation site, electrical stimulation only facilitated the TF reflex at the intensities tested (5-100 microA). In capsaicin-treated rats (n = 6), the proportion of sites from which electrical stimulation only inhibited the TF reflex was significantly less (3/27 sites = 11%) than in vehicle-treated rats (11/23 = 48%). The threshold intensity of stimulation to inhibit the TF reflex from these three sites was 50 microA. The number of sites in RMM from which electrical stimulation only facilitated the TF reflex was significantly greater in capsaicin-treated rats (15/27 = 56%) than in vehicle-treated rats (1/23 = 4%). Neither the number of sites in RMM from which electrical stimulation produced biphasic modulatory effects on the TF reflex (48% and 33%, respectively) nor the intensities of stimulation or magnitudes of facilitation or inhibition of the TF reflex significantly differed between vehicle- and capsaicin-treated rats. In electrophysiological experiments, all units studied responded to non-noxious and noxious intensities of mechanical stimulation applied to the glabrous skin of the plantar surface of the ipsilateral hind foot and also to noxious heating of the skin (50 degrees C). The number of sites where electrical stimulation produced only facilitatory effects on responses of spinal dorsal horn neurons to noxious stimulation (thermal or mechanical) of the skin was significantly increased from 13% of the total sites in vehicle-treated rats to 40% in capsaicin-treated rats.(ABSTRACT TRUNCATED AT 400 WORDS)

Background: Combinations of opioids and nonsteroidal antiinflammatory drugs (NSAIDs) are commonly used to control pain in the perioperative period, yet there are no quantitative evaluations of the interaction between opioids and nonsteroidal antiinflammatory drugs during visceral nociception. This study evaluated the interaction between morphine and ketorolac during visceral nociception in the rat., Methods: The pressor response to noxious colorectal distention (80 mmHg, 20 s) was evaluated in 29 male Sprague-Dawley rats and dose-response curves were determined for intravenous morphine, ketorolac and the mixture of morphine and ketorolac. The data were interpreted using an isobolographic analysis to establish the nature of the interaction., Results: Intravenous ketorolac alone (8-32 mg/kg) did not have a significant antinociceptive effect, whereas morphine alone (1-4 mg/kg) produced significant antinociception during noxious colorectal distention (dose yielding a 50% reduction in nociceptive response relative to baseline pressor response = 1.7 +/- 0.6 mg/kg). Isobolographic analysis of the antinociceptive interaction demonstrated a highly significant, naloxone-reversible potentiation of intravenous morphine by ketorolac in the rat during visceral nociception (P < 0.001)., Conclusions: Ketorolac is a powerful potentiator of morphine antinociception during visceral nociception in the rat. However, intravenous ketorolac alone did not demonstrate antinociceptive properties during colorectal distention, a model of acute visceral nociception without a major inflammatory component. These data suggest that ketorolac may have a central modulatory effect on opioid pharmacology and the synergistic effect may be separate from its peripheral antiinflammatory properties. This study encourages further basic as well as clinical evaluations of the improved antinociception provided by combination therapy of opioids and nonsteroidal antiinflammatory drugs.

Activation of the N-methyl-D-aspartate (NMDA) receptor has been reported to be involved in the mechanisms that underlie thermal hyperalgesia produced by the intraplantar injection of carrageenan. As NMDA-mediated thermal hyperalgesia produced in models of acute and persistent pain have been reported to involve production of nitric oxide, we examined the role of nitric oxide in both the development and maintenance of the thermal hyperalgesia produced by the intraplantar injection of carrageenan. In addition, we examined the role of nitric oxide in the maintenance of the mechanical hyperalgesia produced by intraplantar injection of carrageenan. Prior to the intraplantar injection of carrageenan (2 mg in 100 microliters) there was no significant difference in thermal withdrawal latencies or mechanical withdrawal thresholds between the left and right hindpaws. Three hours after injection of carrageenan into the left hindpaw, rats showed evidence of a significantly faster thermal withdrawal latency and lower mechanical withdrawal threshold of the left hindpaw compared to the right hindpaw. In addition, the left hindpaw was significantly increased in size (diameter) compared with the right hindpaw. In these same rats, the intrathecal administration of saline, NG-nitro-L-arginine methyl ester (L-NAME; 2-200 nmol) or the inactive enantiomer, NG-nitro-D-arginine methyl ester (D-NAME; 200 nmol) did not produce any significant change in thermal nociceptive withdrawal latencies in the non-injected paw. However, administration of L-NAME (2-20 nmol), but not saline or D-NAME produced a dose dependent and reversible block of the thermal hyperalgesia for a period of up to 3 h.(ABSTRACT TRUNCATED AT 250 WORDS)

Noxious stimuli applied to the skin can produce long-lasting, C-fiber-dependent, secondary hyperalgesia that is mediated by central mechanisms. NMDA receptor antagonists and low doses of morphine can preferentially block the development of hyperalgesia without significantly altering unpotentiated responses to nociceptive stimuli. The aim of our study was to determine if low doses of pentobarbital can also preferentially alter either hyperalgesic or unpotentiated responses to nociceptive heat stimuli in spinalized and intact rats. Our results demonstrate the following. (1) Mustard oil applied above the ankle joint or electrical stimulation of the sciatic nerve at C-fiber intensity in spinalized, unanesthetized rats decreased the latency to withdrawal of the foot from water maintained at 47-49 degrees C. This secondary hyperalgesia to thermal stimulation persisted for at least 1 h and was most likely mediated by central mechanisms. (2) Pentobarbital in both spinalized and spinal cord-intact rats prevented the development of the late component (42-120 min) but only partially decreased the early (2-6 min) component of hyperalgesia. In contrast, pentobarbital had relatively minimal effects on unpotentiated withdrawal responses. Thus, pentobarbital is similar to morphine in its ability to prevent hyperalgesia, but may differ from the anesthetic isoflurane, which does not interfere with the development of hyperalgesia.

Neuronal plasticity associated with altered sensations arising from tissue damage involves both established (e.g. substance P and excitatory amino acids) and novel (e.g. nitric oxide and metabolites of arachidonic acid) mediators released from terminals of primary afferent neurons or synthesised in the spinal cord. These and other mediators lead to activity-dependent synaptic plasticity and enhanced sensitivity to noxious stimuli (hyperalgesia). Activation of the N-methyl-D-aspartate (NMDA) receptor results in a calcium-dependent production of nitric oxide, while activation of alpha-amino-3-hydroxy-5-methylisoxazole-5-propionate (AMPA)-and 1,3- trans-1-amino-cyclopentyl-1,3-dicarboxylate (trans-ACPD)-sensitive glutamate receptors results in a phospholipase A2 (PLA2)-mediated production of different intracellular mediators, including arachidonic acid. Thermal hyperalgesia requires NMDA receptor activation and is primarily mediated by production of nitric oxide. Mechanical hyperalgesia requires AMPA and metabotropic glutamate receptor coactivation, and is primarily mediated by cyclo-oxygenase products of arachidonic acid metabolism.

The quantitative approach to the study of descending inhibition of spinal nociceptive transmission was initiated in Heidelberg through the use of natural, noxious stimulation and examination of the modulation of the encoding properties of spinal dorsal horn neurons. This important approach required control of the noxious stimulus, which had previously been inadequately considered, and the parametric assessment of modulatory influences on the encoding properties of spinal dorsal horn neurons. As a consequence, descending inhibition of spinal nociceptive transmission was found not to be homogeneous throughout the brainstem, but rather to be significantly different from different brainstem nuclei. The early contributions from Zimmermann's laboratory and subsequent contributions, mostly by previous coworkers, led to a clearer understanding of how these inhibitory systems were organized in the brainstem. For example, it was established in the early 1980s that the nucleus raphe magnus was not the requisite or the only bulbar relay between the midbrain and the spinal cord important to the endogenous pain control system. The same quantitative approach has proved useful in studies of nociceptive reflexes evoked either by noxious thermal or visceral stimuli and in electrophysiological studies of neuronal responses to noxious stimulation of hollow visceral organs. Most recently, the same approach has been profitably applied to studies that have focused onfacilitatory influences descending from many of the same brainstem sites. As a consequence, it has been proposed that there exists an endogenous pain facilitating system analogous to the well-accepted endogenous pain inhibitory system. While the function of the facilitatory system remains unknown, it is proposed that it may be important to long-lasting pain conditions that exist in the absence of pathology.

The effects of N-methyl-D-aspartate (100 amol-1 nmol) on the nociceptive tail-flick reflex were studied in awake rats. Lesser doses of N-methyl-D-aspartate (100 amol-10 pmol) administered intrathecally to the lumbar spinal cord produced a dose-dependent facilitation of the tail-flick reflex (maximum at 0.5-1 min). The greatest dose tested (1 nmol) inhibited the tail-flick reflex (maximum at 2-5 min) and produced a caudally directed scratching and biting behavior accompanied by vocalizations. Intrathecal pretreatment with the N-methyl-D-aspartate receptor antagonist, D-2-amino-5-phosphonovaleric acid (1 fmol-1 pmol), which produced no change in baseline tail-flick latency, blocked all N-methyl-D-aspartate produced effects in a dose-dependent manner (100 fmol D-2-amino-5-phosphonovaleric acid produced maximum blockage for about 40 min). The magnitude and duration of N-methyl-D-aspartate-produced biphasic effects on tail-flick latency were similar in awake and lightly pentobarbital-anesthetized rats, but caudally directed biting and scratching behavior was not produced in lightly anesthetized rats. Reversible spinalization at T8-T10 in lightly anesthetized rats (produced by cold-block) completely abolished inhibition of the tail-flick reflex produced by 1 nmol N-methyl-D-aspartate whereas facilitation produced by 10 pmol N-methyl-D-aspartate remained unchanged, indicating that N-methyl-D-aspartate-produced facilitation is a local, segmental effect and that N-methyl-D-aspartate-produced inhibition requires a supraspinal loop. To examine the nature of the supraspinal loop, potential contributions of descending noradrenergic and serotonergic systems were studied. Intrathecal pretreatment with 100 nmol phentolamine completely blocked N-methyl-D-aspartate-produced inhibition of the tail-flick reflex, while N-methyl-D-aspartate-produced facilitation and caudally directed biting and scratching behavior remained unchanged. Intrathecal pretreatment with 50 nmol methysergide reversed the inhibitory effect of 1 nmol N-methyl-D-aspartate, resulting in a potent and prolonged facilitation which could be blocked by D-2-amino-5-phosphonovaleric acid. (1 pmol). Intrathecal pretreatment with an alternate substrate for nitric oxide synthase, NG-nitro-L-arginine methyl ester (100 nmol), completely blocked N-methyl-D-aspartate-produced facilitation of the tail-flick reflex, whereas N-methyl-D-aspartate-produced inhibition and caudally directed biting and scratching behavior were unaffected.(ABSTRACT TRUNCATED AT 400 WORDS)

Dorsal root ganglion (DRG) neurons express receptors for bradykinin and capsaicin, both algesic substances. Administration of bradykinin or capsaicin to neurons cultured from embryonic rat DRG stimulated the production of cyclic GMP but did not affect the production of cyclic GMP in nonneuronal DRG cultures. Bradykinin-evoked cyclic GMP production was mediated by B2 receptors and was unaltered by indomethacin. Both bradykinin- and capsaicin-stimulated cyclic GMP production required Ca2+ and was inhibited by methylene blue. Furthermore, methylene blue attenuated basal cyclic GMP production in DRG neurons, suggesting tonic cyclic GMP production in these cells. L-NG-monomethyl arginine inhibited both bradykinin- and capsaicin-stimulated cyclic GMP production as well as basal cyclic GMP production. These findings suggest the involvement of a nitrosyl compound in bradykinin- and capsaicin-stimulated cyclic GMP production and in tonic cyclic GMP production in DRG neurons.

Thoracolumbar and lumbosacral spinal segments that receive afferent input from the descending colon and rectum were stained immunocytochemically for c-Fos- and c-Jun-like proteins following repetitive, noxious colorectal distention (CRD). Noxious CRD (80 mmHg) resulted in significantly more c-Fos- and c-Jun-like immunoreactivity in the sacral dorsal horn than in the thoracic dorsal horn. In both regions of the spinal cord the increase in c-Fos-like immunoreactivity was at least twice that of c-Jun-like immunoreactivity. Basal levels of c-Jun but not c-Fos were observed in the thoracic intermediolateral nucleus (IML) and the sacral parasympathetic nucleus (SPN). Noxious CRD induced both c-Fos and c-Jun in the SPN, but not the IML.

Several recent reports document that activation of the NMDA receptor is required for the development and maintenance of thermal hyperalgesia. In contrast, the receptor subtype(s) involved in mechanisms that underlie mechanical hyperalgesia are at present unknown. We report here that acute mechanical hyperalgesia in the rat is not produced by NMDA receptor agonists, but instead requires coactivation of ionotropic AMPA and metabotropic glutamate receptor subtypes. Collectively, the results are consistent with a role for activation of spinal cord neurons using NMDA receptor agonists in mechanisms of thermal hyperalgesia and for coactivation of AMPA and metabotropic glutamate receptors on spinal cord neurons in mechanisms of mechanical hyperalgesia.

Recently, it has been reported that loosely constrictive chromic gut ligatures around the sciatic nerve produce behavioral evidence of neuropathic pain in rats. It has been shown that axonal swelling after ligation results in a constriction injury associated with a decrease in the number of both large-diameter myelinated and small-diameter unmyelinated axons, but the mechanism(s) producing spontaneous pain and thermal hyperalgesia remain largely unknown. The present study systematically evaluated potential mechanisms involved in development of the behavioral changes produced by chromic gut ligatures loosely tied around the sciatic nerve. Four ligatures of either silk (4-0), plain gut (4-0), or chromic gut (4-0, 3-0, or 2-0) were placed loosely around the left sciatic nerve of male Sprague-Dawley rats. An additional group of rats had 8 x 0.5 cm sections of 4-0 chromic gut laid adjacent to the left sciatic nerve. The right sciatic nerve was exposed in all rats for sham surgery. The posture and gait of all rats was qualitatively assessed before (day 0) and for 20-30 days after surgery. Rats were tested for evidence of thermal and mechanical hyperalgesia prior to surgery, and on postoperative days 3, 5, 10, 20 and, in some groups, on day 30. Chromic gut, but not plain gut or silk, ligatures placed around or laid next to the sciatic nerve produced an alteration in the posture of rats such that most of the pressure was placed on the heel and medial aspect of the left (ligated) hind paw with the toes held together and plantar-flexed while pressure appeared to be evenly distributed on the right (sham) hind paw. As a result, a pronounced limp was evident, often with the left hind paw held in the air for prolonged periods of time during the first few days after surgery. These postural changes were most pronounced in the 2-0 and 3-0 chromic gut-treated rats. Chromic gut sutures (4-0, 3-0, or 2-0) tied loosely around the left sciatic nerve also produced a 'dose-dependent' decrease in thermal withdrawal latency that was maximal on postoperative day 3 (25%, 39%, and 41%, respectively). The magnitude of the thermal hyperalgesia declined over time such that a return to baseline was observed by postoperative day 20 in 4-0 and 3-0 chromic gut-treated rats.(ABSTRACT TRUNCATED AT 400 WORDS)

Recent studies have suggested that a spinal cholinergic system is important in spinal nociceptive modulation. In the present study, the role of a nitric oxide (NO)-generating system in spinal cholinergic modulation of nociception was examined in awake rats. Intrathecal (i.t.) administration of the cholinergic muscarinic receptor antagonist atropine produced a dose-dependent (1.4-14.4 nmol) decrease in the mechanical threshold for tail withdrawal, which was reversed rapidly (2-3 min) by subsequent i.t. administration of the NO precursor, L-arginine (10 pmol to 10 nmol). Intrathecal administration of L-arginine alone (10 pmol to 10 nmol) produced a dose-dependent increase in the mechanical nociceptive withdrawal threshold of the tail. The reflexive withdrawal of the tail produced by noxious heat was not significantly affected by i.t. administration of either atropine or L-arginine. Inhibition of the NO-cGMP pathway by i.t. administration of either Nw-nitro-L-arginine methyl ester (L-NAME, 10 nmol) or methylene blue (10 nmol) significantly enhanced the magnitude and prolonged the time course of the decrease in the mechanical threshold for tail withdrawal produced by i.t. pretreatment with atropine (1.4 nmol). Neither L-NAME nor methylene blue affected mechanical withdrawal thresholds in rats pretreated with saline. These data suggest that the production of endogenous NO is required for tonic inhibition of spinal nociceptive mechanical transmission. Moreover, the present data support the speculation that there exists in the lumbar spinal cord a tonic, cholinergic modulation of NO synthase.

Ischemia of visceral organs, especially the heart, is often a painful and potentially life-threatening condition. However, in at least 75% of all cases myocardial ischemia may be "silent" (i.e., without pain or sensation). Yet, the mechanisms responsible for silent ischemia are not well understood. As such, many different theories have been advanced to explain silent ischemia; however, none have been able to adequately explain all of the experimental and clinical findings. This paper proposes a hypothetical mechanism that may help to understand mechanisms of silent ischemia.

Recent observations using acute and persistent pain models have suggested that activation of the N-methyl-D-aspartate (NMDA) receptor is required for mechanisms that underly the development and maintenance of thermal hyperalgesia. The present results document that both NMDA-mediated thermal hyperalgesia produced after acute intrathecal NMDA administration and NMDA-mediated thermal hyperalgesia produced in a model of neuropathic pain are dose-dependently and reversibly attenuated by intrathecal administration of ethanol (0.5-1.0%; total dose, 106-213 nmol, i.t.). This is consistent with recent reports that ethanol may function as a selective NMDA receptor antagonist at low concentrations and further extends the evidence that thermal hyperalgesia is mediated by NMDA receptors.

1. The effects of electrical stimulation of cervical vagal afferent fibers on the nociceptive tail-flick (TF) reflex and responses of spinal dorsal horn neurons to noxious cutaneous stimulation were studied in adult rats treated as neonates with either capsaicin or vehicle. 2. Vagal afferent stimulation (VAS) produced biphasic, intensity-dependent effects on the TF reflex in vehicle-treated and untreated control rats. The TF reflex was facilitated in both groups of rats at lesser intensities of VAS (2.5-50 microA) and fully inhibited at greater intensities of VAS (50-100 microA). In contrast, biphasic effects of VAS on the TF reflex generally were not produced in rats treated as neonates with capsaicin. Facilitation of the TF reflex was produced in these rats by lesser intensities of VAS as well as by typically "inhibitory" intensities of VAS; the TF reflex was never inhibited in 6/12 rats, even at the greatest intensity of VAS tested (1,000 microA). When the TF reflex was inhibited by VAS in capsaicin-treated rats, the intensities required were significantly greater than those required in vehicle-treated or untreated rats. 3. In electrophysiological experiments, 77 neurons were recorded in the lumbar spinal dorsal horn of pentobarbital sodium-anesthetized, paralyzed rats treated as neonates with either vehicle or capsaicin. The neurons had receptive fields on the glabrous skin of the plantar surface of the ipsilateral hind foot, and all responded to mechanical stimuli of both nonnoxious and noxious intensities; 16/77 neurons also responded to noxious thermal stimulation. In vehicle-treated rats, nociceptive responses of 50% of 30 units studied were biphasically modulated by VAS, 33% were only inhibited, and 17% were only facilitated by VAS at the intensities tested (5-500 microA). In capsaicin-treated rats, nociceptive responses of 32% of 47 units studied were biphasically modulated by VAS, 15% were only inhibited, and 34% were only facilitated by VAS at the intensities tested (5-500 microA). In addition, nociceptive responses of neurons facilitated at lesser intensities of VAS and not affected at greater intensities of VAS were observed in capsaicin-treated rats (19% of the 47-unit sample). Overall, the proportion of the neuronal sample inhibited by VAS was less, and the proportion of the sample facilitated by VAS was greater in capsaicin-treated rats compared with vehicle-treated rats. 4. The efficacy of the capsaicin treatment was evaluated immunocytochemically.(ABSTRACT TRUNCATED AT 400 WORDS)

1. Three hundred fifty neurons in the T2-T4 spinal segments of 38 intact, pentobarbital sodium-anesthetized, pancuronium-paralyzed male rats were examined for somatic receptive fields and responses to midthoracic esophageal distension (ED). Recordings were made at a depth of 0.1-1.45 mm from the dorsal spinal cord surface and from the midline to approximately 1.0 mm lateral. 2. Fifty-six of the 350 total neurons (16%) responded to ED, produced by air inflation of a latex balloon (0.5-1.5 ml). Most of these 56 neurons (84%) were excited by ED, and all except one were excited at a short latency (< 2 s) to stimulus onset. The response to ED in about one-half of all excited neurons terminated abruptly with termination of the stimulus; the other neurons exhibited an afterdischarge of 5 to > 80 s. Repeated ED at a constant intensity (1.25 ml, 30 s every 6 min) produced stable and reproducible responses of neurons excited by ED. Twenty-one percent of neurons that responded to ED were antidromically invaded from the spinomedullary junction. 3. Graded ED (0.5-1.5 ml, 30 s every 6 min) produced linear and accelerating stimulus-response functions in the 29 neurons tested. The mean threshold for distension, determined with a least-squares regression analysis, was extrapolated to near 0.5 ml of distending volume, and no difference in response threshold was found between neuronal groups with or without after-discharge. 4. The spontaneous activity of 7 of the 56 neurons (12.5%) that responded to ED was inhibited by the stimulus. Stimulus-response functions for four neurons inhibited by ED were intensity dependent. The spontaneous activity of these neurons was inhibited to a mean of 24.5% of the prestimulus control by 1.25 ml ED. 5. Two neurons of the total sample of 56 (3.5%) responded to ED (1.50 ml) in a biphasic excitatory-inhibitory manner. The excitatory component of excitatory-inhibitory neurons encoded the intensity of ED; the inhibitory component during the second half of ED was apparent only at greater distending volumes (1.25-1.5 ml). 6. Somatic receptive fields were found for 303/350 neurons, and 98% were located on the thorax and proximal forearm (all ipsilateral). Five neurons in T2-T4 spinal segments had their cutaneous receptive fields located on caudal parts of the body (tail, hindleg, scrotum).(ABSTRACT TRUNCATED AT 400 WORDS)

There is considerable evidence to implicate N-methyl-D-aspartate (NMDA) receptor activation in the mechanisms that underly thermal hyperalgesia in the spinal cord. As many of the effects of NMDA receptor activation appear to be ultimately mediated through production of nitric oxide (NO), recent reports have begun to define the role of NO in spinal nociceptive processing. From this evidence, it is likely that NO, produced in neurons in the spinal cord that contain NO synthase, like NMDA, plays a pivotal role in multisynaptic local circuit nociceptive processing in the spinal cord. Collectively, these reports suggest that the reflex withdrawal response to noxious heat is not mediated through activation of NMDA receptors and subsequent production of NO and cGMP, but that the acute NMDA-produced facilitation of thermal reflexes is NMDA-, NO- and cGMP-mediated and that a sustained production of NO and subsequent activation of soluble guanylate cyclase (GC-S) in the lumbar spinal cord appears to be required for maintenance of the thermal hyperalgesia produced in persistent pain models. As our knowledge and understanding of the new and intriguing class of neurotransmitters typified by NO emerges, it is likely that the next few years of pain and analgesia research will focus on the cellular events underlying mechanisms of chronic pain.

Tissue injury results in several changes in spinal cord neurons that contribute to hyperalgesia arising from the injured tissue. In models of unilateral hindpaw inflammation, changes in the neurochemistry and electrophysiology of dorsal horn neurons ipsilateral, and to a much lesser extent contralateral, to the inflamed paw have been reported. For example, the excitability of dorsal horn neurons increases, receptive field size increases, and the content of various proteins and neuropeptides in the dorsal horn (e.g. FOS, dynorphin, enkephalin) are affected following peripheral inflammatory insult. These changes are typically interpreted on the basis of their relevance to nociception.

Loose ligation of the sciatic nerve with 4-0 chromic gut sutures in rats produces behavioral evidence of neuropathic pain. In the present experiments we examined the involvement of capsaicin-sensitive afferents in mediating the thermal hyperalgesia produced by this model. Male Sprague-Dawley rats, treated as neonates (within 48 h of birth) with capsaicin (50 mg/kg, s.c.) or vehicle, were used at 16-18 weeks of age. Chromic gut sutures (4-0) were tied around the left sciatic nerve and withdrawal latencies of both hind paws to radiant heat were determined on postoperative days 3, 5, 10 and 20. Whereas there was a pronounced thermal hyperalgesia which lasted for up to 20 days in vehicle-treated rats, there was no evidence of thermal hyperalgesia in capsaicin-treated rats. There was no difference in baseline (pre-surgery) withdrawal latencies between the two groups. Radioimmunoassay revealed that there was a significant depletion of substance P (43.8%) and calcitonin-gene-related peptide (72.6%) in the lumbar spinal cord of neonatal capsaicin-treated rats compared to vehicle-treated rats. These results demonstrate that the chromic gut-induced thermal hyperalgesia is mediated by capsaicin-sensitive afferents and suggest that central mechanisms which process and control the reflex response to heat are different than mechanisms involved in thermal hyperalgesia.

The intravenous (i.v.) administration of serotonin (5-HT) to rats is a noxious visceral stimulus which produces distinct vagal afferent-mediated pseudaffective responses, a passive avoidance behavior, a vagal afferent-mediated inhibition of the tail-flick (TF) reflex and a complex cardiovascular response. In the present study, we examined the effects of age (10 or 16 weeks), strain (Sprague-Dawley, SD; Wistar-Kyoto, WKY; spontaneously hypertensive rats, SHR) and anesthetic (conscious or lightly pentobarbital-anesthetized) on nociceptive (TF reflex and pseudaffective responses) and cardiovascular responses produced by 5-HT (3-288 micrograms/kg, i.v.). There were no age-related differences in baseline TF latencies in the 3 strains. Further, latencies were generally not significantly different whether rats were tested conscious or lightly anesthetized. There were, however, strain differences. Both conscious or lightly pentobarbital-anesthetized SHR and WKY rats at 10 weeks of age had significantly faster response latencies than 10 week old SD rats. At 16 weeks of age, only the lightly pentobarbital-anesthetized WKY and SHR showed faster response latencies than SD rats. The WKY and SHR, but not the SD rats, were more sensitive to the nociceptive effect of i.v. 5-HT at 16 weeks of age compared to 10 weeks of age. At both ages, WKY and SHR, but not SD rats, showed an anesthetic-dependent increase in nociceptive sensitivity to i.v. 5-HT. In addition, at both ages, regardless of the presence of anesthetic, the order of sensitivity to the nociceptive effects of i.v. 5-HT was SD greater than WKY much much greater than SHR.(ABSTRACT TRUNCATED AT 250 WORDS)

The present study examined the spinal pathway and receptors that mediate nocigenic inhibition of the tail-flick (TF) reflex produced by conditioning colorectal distension (CRD). Conditioning CRD (80 mmHg; 30 s) inhibited the TF reflex in all rats studied (n = 29). In 19 rats where intensity-dependent effects of CRD were studied, conditioning CRD in 7 rats facilitated the TF reflex at lesser, non-noxious intensities (mean 7.9 +/- 2.1 mmHg) and inhibited the TF reflex at greater, noxious intensities (40-100 mmHg); conditioning CRD at all intensities tested only inhibited the TF reflex in the other 12 rats. Inhibition of the TF reflex produced by 30 s CRD was short-lasting, repeatable and graded with the intensity of CRD. The mean threshold of CRD for inhibition of the TF reflex to cut off (10 s) was 61.4 +/- 3.3 mmHg (n = 29). Intrathecal pretreatment with atropine or methysergide significantly attenuated the inhibitory effect of CRD on the TF reflex; the effects were time- and dose-related. Intrathecal pretreatment with mecamylamine, phentolamine or naloxone was without effect. Intrathecal administration of physostigmine, an acetylcholinesterase inhibitor, significantly reduced the threshold intensity of conditioning CRD necessary to inhibit the TF reflex to cut off (mean 36.0 +/- 4.0 mmHg; n = 5). Bilateral transections of the spinal dorsolateral funiculi (DLF) did not affect the inhibitory effect of CRD in 4/7 rats and attenuated the inhibitory effect of CRD in the other 3 rats. The antagonistic effect of methysergide on CRD-produced inhibition of the TF reflex was abolished following the DLF transections, while scopolamine retained its efficacy in rats with bilateral DLF transections. These findings provide evidence for involvement of spinal cholinergic interneurons as well as a descending serotoninergic pathway traveling in the DLF in CRD-produced inhibition of the TF reflex.

In the rat, intravenous (i.v.) serotonin (5-HT) is a noxious stimulus which produces distinct vagal afferent-mediated pseudoaffective responses, a passive avoidance behavior, a vagal afferent-mediated inhibition of the nociceptive tail-flick (TF) reflex and a complex triad of cardiovascular responses. In the present study, we have used a variety of 5-HT receptor antagonists to characterize the receptor subtype(s) in the rat that mediate (1) inhibition of the TF reflex and (2) the cardiovascular responses produced by i.v. 5-HT. 5-HT produced a dose-dependent (3-72 micrograms/kg, i.v.) inhibition of the TF reflex (ED50 = 15.3 +/- 0.7 micrograms/kg). Following administration of the 5-HT2 receptor-selective antagonists ketanserin (50-250 micrograms/kg, i.v.) or xylamidine (10-100 micrograms/kg, i.v.), or the 5-HT3 receptor-selective antagonists ICS 205-930 (50-250 micrograms/kg, i.v.) or MDL 72222 (25-250 micrograms/kg, i.v.), there appeared to be a parallel shift of the 5-HT dose-response curve to the right. Following co-administration of xylamidine (50 micrograms/kg, i.v.) with ICS 205-930 (100 micrograms/kg, i.v.), the 5-HT-induced inhibition of the TF reflex was completely abolished at all doses of 5-HT tested (3-288 micrograms/kg, i.v.). In contrast, administration of the centrally acting 5-HT2 receptor-selective antagonist LY 53857 (10-100 micrograms/kg, i.v.) or the non-specific receptor antagonist methysergide (25-500 micrograms/kg, i.v.) resulted in a dose-dependent, but not parallel shift of the 5-HT dose-response curve to the right. The maximal doses of LY 53857 and methysergide tested (250 micrograms/kg and 500 micrograms/kg, respectively) completely abolished the effects of 5-HT (3-288 micrograms/kg, i.v.). Administration of the alpha 1-adrenoceptor antagonist prazosin (25-100 micrograms/kg, i.v.) failed to alter the 5-HT dose-response curve, indicating that the effects of ketanserin were due to blockage of 5-HT2 receptors rather than alpha 1 receptors. Administration of each of the antagonists also produced marked, but selective effects on components of the complex cardiovascular response to i.v. 5-HT. Each of the 5-HT3 receptor selective antagonists (ICS 205-930 or MDL 72222) produced a dose-dependent attenuation of the Bezold-Jarisch reflex-mediated hypotension and bradycardia, and each of the 5-HT2 receptor selective antagonists (xylamidine, ketanserin or LY 53857) produced a dose-dependent attenuation of the pressor response. The non-specific 5-HT receptor antagonist methysergide produced a dose-dependent attenuation of the 5-HT-induced pressor response.(ABSTRACT TRUNCATED AT 400 WORDS)

1. Descending influences produced by focal electrical stimulation in the nuclei reticularis gigantocellularis (NGC) and gigantocellularis pars alpha (NGC alpha) on spinal nociceptive transmission and the dorsoventral region of spinal white matter mediating stimulation-produced modulation were examined in pentobarbital sodium-anesthetized, paralyzed rats. Spinal units studied responded to mechanical stimuli and noxious heating (50 degrees C) of cutaneous receptive fields confined to the glabrous skin of the ipsilateral hind foot. Recording sites were located in laminae I-VI of the L3-L5 spinal segments. 2. Electrical stimulation in the NGC or NGC alpha produced both facilitation and inhibition of responses of spinal units to noxious heating of the skin. At 33 of 57 stimulation sites in the NGC and NGC alpha, electrical stimulation produced biphasic effects, facilitating responses at lesser intensities (5-25 microA) and inhibiting responses at greater intensities (50-100 microA). At 21 other sites in the NGC and NGC alpha, electrical stimulation (5-100 microA) only inhibited, and at 3 sites stimulation (5-100 microA) only facilitated responses of spinal units to noxious heating of the skin. 3. Electrical stimulation in the NGC or NGC alpha contralateral to the spinal recording site produced the same magnitude of facilitation/inhibition or inhibition of spinal nociceptive transmission as did stimulation in the ipsilateral NGC and NGC alpha. 4. The latencies to descending facilitation and inhibition of spinal nociceptive transmission from the NGC and NGC alpha were estimated by a cumulative sum technique to be 232 and 80 ms, respectively. 5. Responses of spinal units to graded heating (42-50 degrees C) of the skin exhibited positively accelerating stimulus-response functions (SRF) throughout the temperature range tested. Electrical stimulation at lesser, "facilitating" intensities produced a parallel, leftward shift of the SRF, whereas stimulation at greater, "inhibitory" intensities significantly decreased the slope of the SRF without affecting the threshold for response. 6. To determine whether activation of cell bodies in the NGC or NGC alpha were capable of replicating the effects of electrical stimulation, L-glutamate was microinjected into sites where electrical stimulation facilitated at lesser and inhibited at greater intensities the responses of spinal units to 50 degrees C heating of the skin. L-Glutamate (5 nmol) produced a rapid onset, short-lasting and reproducible facilitation of nociceptive transmission; glutamate microinjection into the same site at a greater dosage (50 nmol) inhibited responses of the same spinal units.(ABSTRACT TRUNCATED AT 400 WORDS)

The rat lumbosacral spinal cord was immunocytochemically stained for Fos-like immunoreactivity following repetitive colorectal distention (CRD) to 20, 40 or 80 mm Hg. Following all 3 distention pressures, Fos-like immunoreactive (Fos-ir) nuclei were observed primarily in laminae I-II, V-VII and X, although some labeled nuclei were observed in laminae III-IV. Eighty mm Hg CRD resulted in significantly more Fos-ir nuclei than 20 or 40 mm Hg CRD. Morphological examination of the colon revealed clear signs of inflammation following 80 but not 20 mm Hg CRD. Acute 20 mm Hg CRD is a non-noxious stimulus, suggesting that both noxious and non-noxious visceral stimuli can induce Fos-like immunoreactivity in the spinal cord. It is suggested that activation of neuropeptide-containing small-diameter primary afferents is necessary, although not necessarily sufficient, to induce c-fos in the rat spinal cord.

Chemical, electrical or physiological activation of cardiopulmonary vagal (cervical, thoracic or cardiac), diaphragmatic vagal (DVAG) or subdiaphragmatic vagal (SDVAG) afferents can result in either facilitation or inhibition of nociception in some species. In the rat, these effects depend upon vagal afferent input to the NTS and subsequent CNS relays, primarily in the NRM and ventral LC/SC, although specific relay nuclei vary as a function of the vagal challenge stimulus. Spinal pathways and neurotransmitters have been identified for vagally mediated effects on nociception and consistently implicate the involvement of descending 5-HT and noradrenergic systems, as well as intrinsic spinal opioid receptors. Species differences may exist with respect to both the effects of DVAG and SDVAG afferents on nociception and the efficacy of vagal afferents to modulate nociception. However, it is also possible that such differences reflect the modality of noxious input (e.g., visceral versus cutaneous), the type of neuronal activity investigated (e.g., resting versus noxious-evoked), spinal location of recording (e.g., thoracic versus lumbosacral) and/or parameters of stimulation. It is also possible that activation of some vagal afferents is aversive, but whether this contributes to changes in nociception produced by vagal activation has not clearly been established. Finally, the vagal-nociceptive networks described in this review provide a fertile area for future study. These networks can provide an understanding of physiological and pathophysiological peripheral events that affect nociception.

Bradykinin (BK) is a potent algesic compound. Therefore, we hypothesized that BK, acting as a peripheral noxious stimulus, would attenuate or inhibit responses to another noxious stimulus. When administered intravenously (i.v.) to rats lightly anesthesized with pentobarbital, BK produced a dose-dependent (12-144 micrograms/kg) inhibition of the nociceptive tail-flick (TF) reflex. BK also produced a dose-dependent decrease in mean arterial blood pressure, a subsequent increase in heart rate and an increase in the rate of respiration. The latency to the maximal effect of BK on the TF reflex was 10 s and was occasionally preceded by pseudoaffective responses at doses greater than 48 micrograms/kg BK. Neonatal treatment with capsaicin (50 mg/kg, subcutaneous) significantly attenuated BK-produced inhibition of the TF reflex indicating that BK was acting via peripheral afferents to inhibit the TF reflex. Reversible cold block or complete spinal transection at a low thoracic level (T9-12), but not reversible cold block at a high cervical level (C1-2), significantly attenuated BK-produced inhibition of the TF reflex, suggesting that BK activates afferents which enter the spinal cord between C2 and T9. Pretreatment with intrathecally administered phentolamine (30 micrograms), an alpha-adrenoceptor antagonist, or methysergide (30 micrograms), a non-selective serotonin receptor antagonist, did not alter BK-produced inhibition of the TF reflex, further supporting the absence of activation of descending systems from the brainstem by i.v. BK. While coadministration of PGE2 and BK significantly potentiated BK-produced inhibition of the TF reflex, neither bilateral removal of the stellate ganglion nor bilateral cervical vagotomy significantly affected the inhibitory action of i.v. BK on the TF reflex. These results suggest that i.v. BK inhibits the nociceptive TF reflex by activation of capsaicin-sensitive visceral afferents entering the spinal cord between C2 and T9.

The intrathecal (i.t.) administration of either N-methyl-D-aspartate (NMDA, 10 fmol to 10 pmol) or L-arginine (1 pmol to 10 nmol), but not D-arginine (1 pmol to 10 nmol), produced a rapid, transient, dose-dependent facilitation (maximal response of 30.9 +/- 6.0% and 33.7 +/- 1.5%, respectively) of the nociceptive tail-flick reflex (ED50 = 47.8 +/- 15.4 fmol and 11.4 +/- 2.7 pmol, respectively). Maximal NMDA-produced facilitation of the tail-flick reflex (1 pmol i.t.) was completely abolished by prior treatment (10 min prior) with either N omega-nitro-L-arginine methyl ester (L-NAME, 10 nmol i.t.), methylene blue (10 nmol i.t.) or DL-5-aminophosphonovaleric acid (AP5, 100 pmol i.t.). NMDA-produced facilitation was completely recovered 40 min after L-NAME, 50 min after methylene blue and 30 min after AP5. L-NAME, methylene blue or AP5 did not significantly alter baseline tail-flick latency. These results suggest that NMDA-produced facilitation of a thermal nociceptive reflex is mediated through activation of an NMDA receptor that results in an increase in endogenous nitric oxide and activation of soluble guanylate cyclase in lumbar spinal cord.

1. In pentobarbital sodium-anesthetized rats, we evaluated changes in cortical evoked potentials (EPs) associated with electrical and chemical stimulation of nucleus raphe magnus (NRM). A condition-test (C-T) paradigm was used. Cortical EPs were produced by test stimuli delivered to a hindpaw or the thalamic ventral posterior lateral nucleus (VPL; electrical stimulation), or by photic stimulation of the eyes or electrical stimulation of contralateral homotopical cortex (transcallosal EPs). These test stimuli were then preceded by electrical or chemical conditioning stimulation (CS) delivered to NRM through a stereotaxically implanted electrode or injection cannula, respectively. Effects of CS on EPs produced by the test stimuli were characterized. 2. Electrical CS preceding a test stimulus delivered to the foot reduced the amplitude of EPs at thresholds as low as 10-25 microA. The magnitude of EP reduction was dependent on CS intensity, frequency, and the C-T interval. Optimal parameters were trains of 10 pulses (400 Hz) delivered at a C-T interval of 5-10 ms. Injection of glutamate and lidocaine into NRM demonstrated that these effects were due to activation of NRM neurons and not to current spread to medial lemniscus (ML). NRM CS also reduced cortical EPs produced by test stimulation in VPL but did not alter EPs from visual stimulation or from electrical stimulation of contralateral homotopical cortex. 3. These findings suggest that NRM CS attenuates EPs by inhibiting thalamic or thalamocortical afferent activity. Because NRM CS affected all components of the cortical EPs, the effect appears to involve alteration of general sensory activity and is not nociception specific.(ABSTRACT TRUNCATED AT 250 WORDS)