Nerve injury can produce hypersensitivity to noxious and normally innocuous stimulation. Injury-induced central (i.e. spinal) sensitization is thought to arise from enhanced afferent input to the spinal cord and to be critical for expression of behavioral hypersensitivity. Descending facilitatory influences from the rostral ventromedial medulla have been suggested to also be critical for the maintenance, though not the initiation, of experimental neuropathic pain. The possibility that descending facilitation from the rostral ventromedial medulla is required for the maintenance of central sensitization was examined by determining whether ablation of mu-opioid receptor-expressing cells within the rostral ventromedial medulla prevented the enhanced expression of repetitive touch-evoked FOS within the spinal cord of animals with spinal nerve ligation injury as well as nerve injury-induced behavioral hypersensitivity. Rats received a single microinjection of vehicle, saporin, dermorphin or dermorphin–saporin into the rostral ventromedial medulla and 28 days later, underwent either sham or spinal nerve ligation procedures. Animals receiving rostral ventromedial medulla pretreatment with vehicle, dermorphin or saporin that were subjected to spinal nerve ligation demonstrated both thermal and tactile hypersensitivity, and showed significantly increased expression of touch-evoked FOS in the dorsal horn ipsilateral to nerve injury compared with sham-operated controls at days 3, 5 or 10 post-spinal nerve ligation. In contrast, nerve-injured animals pretreated with dermorphin–saporin showed enhanced behaviors and touch-evoked FOS expression in the spinal dorsal horn at day 3, but not days 5 and 10, post-spinal nerve ligation when compared with sham-operated controls. These results indicate the presence of nerve injury-induced behavioral hypersensitivity associated with nerve injury-induced central sensitization. Further, the results demonstrate the novel concept that once initiated, maintenance of nerve injury-induced central sensitization in the spinal dorsal horn requires descending pain facilitation mechanisms arising from the rostral ventromedial medulla.

The calbindin-immunoreactivity of spinothalamic (STT) lamina I neurons and their ascending axons was examined in two experiments. In the first experiment, lamina I STT neurons in macaque monkeys were double-labeled for calbindin and for retrogradely transported WGA*HRP following large (n=2) or small (n=1) injections that included the posterior thalamus. Most, but not all (78%) of the contralateral retrogradely labeled lamina I STT cells were positive for calbindin. Calbindin-immunoreactivity was not selectively associated with any particular anatomical type of lamina I STT cell; 82% of the fusiform cells, 78% of the pyramidal cells and 67% of the multipolar cells were double-labeled. In the second experiment, oblique transverse sections from upper cervical spinal segments of three macaque monkeys, one squirrel monkey and five humans were stained for calbindin-immunoreactivity. In each case, a distinct bundle of fibers was densely stained in the middle of the lateral funiculus. This matches the location of anterogradely labeled ascending lamina I axons observed in prior work in cats and monkeys, and it matches the location of the classically described 'lateral spinothalamic tract' in humans. This bundle had variable shape across cases, an observation that might have clinical significance. These findings support the view that lamina I STT neurons are involved in spinal cordotomies that reduce pain, temperature and itch sensations.

Prevention of nerve injury-induced tactile, but not thermal, hypersensitivity is achieved by ipsilateral lesions of the dorsal columns or lidocaine microinjection into the nucleus gracilis (n. gracilis). These and other data support the possibility that tactile hyperresponsiveness after nerve injury may be selectively mediated by a low-threshold myelinated fiber pathway to the n. gracilis. Here we identify a transmitter that might selectively mediate such injury-induced tactile hypersensitivity. Neuropeptide Y (NPY), normally not detected in the dorsal root ganglion (DRG) or in the n. gracilis of rats, became markedly upregulated at both sites and in the spinal cord after spinal nerve injury. Injury-induced NPY-IR occurred predominately in large-diameter DRG cells, and the NPY-IR in the n. gracilis was blocked by dorsal rhizotomy or dorsal column lesion. NPY microinjection into the n. gracilis of uninjured rats elicited reversible tactile, but not thermal, hypersensitivity only in the ipsilateral hindpaw. Administration of anti-NPY antiserum, but not control serum or preabsorbed serum, into the n. gracilis ipsilateral to nerve injury reversed tactile, but not thermal, hypersensitivity. Similarly, microinjection of the NPY antagonists NPY18–36 and (R)-N-[[4-(aminocarbonylaminomethyl)-phenyl]methyl]-N2-(diphenylacetyl)-argininamide trifluoroacetate, into the n. gracilis ipsilateral to the injury reversed tactile, but not thermal, hypersensitivity. Antagonist administration into the contralateral n. gracilis had no effect on injury-induced hypersensitivity. These data suggest the selective mediation of nerve injury-induced tactile hypersensitivity by upregulated NPY via large fiber input to n. gracilis. Selective reversal of injury-induced tactile allodynia by NPY receptor antagonists would have significant implications for human neuropathic conditions.

Previous studies in the macaque monkey have identified a thalamic nucleus, the posterior portion of the ventral medial nucleus (VMpo), as a dedicated lamina I spinothalamocortical relay for pain and temperature sensation. The dense plexus of calbindin-immunoreactive fibres that characterizes VMpo in primates enables its homologue to be identified in the human thalamus by immunohistochemical labelling for calbindin. We have now analysed in detail the cytoarchitectonic characteristics of VMpo and its relationship with immunoreactivity for calbindin, substance P and calcitonin gene-related peptide (CGRP) in the human thalamus. The area in the posterolateral thalamus in which dense calbindin-immunoreactive fibre terminations are present coincides nearly completely with a distinct region that contains small to medium-sized cells with round or oval shapes that are aggregated in clusters separated by cell sparse areas. This region, which we identify as VMpo, is located posteromedial to the ventral posterior lateral (VPL) and ventral posterior medial (VPM) nuclei, ventral to the anterior pulvinar and centre médian nuclei, lateral to the limitans and parafascicular nuclei and dorsal to the medial geniculate nucleus. Calbindin-immunoreactive fibres enter VMpo from the spinal lemniscus and form large patches of dense terminal-like staining over clusters of VMpo neurons. A few of these clusters also display terminal-like substance P labelling. Small bursts of CGRP staining are intercalated between the calbindin-labelled clusters, but there is little or no overlap between these two markers. CGRP immunoreactivity is also present over small, non-clustered neurons in the calbindin-negative area that separates VMpo from the VPL and VPM nuclei, which we denote as the posterior nucleus (Po). These observations provide a concise description of VMpo in the human thalamus. Further, they suggest that the lamina I spinothalamic tract fibres (represented by calbindin and probably also substance P immunoreactivity) and vagal-solitary-parabrachial afferents (represented by CGRP immunoreactivity) form closely related, but separate, termination fields that can be considered to represent different aspects of enteroceptive information regarding the physiological status of the tissues and organs of the body. The location of VMpo and the adjacent Po fits with clinical descriptions of the thalamic area from which pain, temperature and visceral sensations can be evoked by microstimulation, and where nociceptive and thermoreceptive neurons have been recorded in humans. It also corresponds to the area in which infarcts cause analgesia and thermoanaesthesia and can lead to the paradoxical development of central pain.

The nonopioid actions of spinal dynorphin may promote aspects of abnormal pain after nerve injury. Mechanistic similarities have been suggested between opioid tolerance and neuropathic pain. Here, the hypothesis that spinal dynorphin might mediate effects of sustained spinal opioids was explored. Possible abnormal pain and spinal antinociceptive tolerance were evaluated after intrathecal administration of [d-Ala2,N-Me-Phe4, Gly-ol5]enkephalin (DAMGO), an opioid μ agonist. Rats infused with DAMGO, but not saline, demonstrated tactile allodynia and thermal hyperalgesia of the hindpaws (during the DAMGO infusion) and a decrease in antinociceptive potency and efficacy of spinal opioids (tolerance), signs also characteristic of nerve injury. Spinal DAMGO elicited an increase in lumbar dynorphin content and a decrease in the μ receptor immunoreactivity in the spinal dorsal horn, signs also seen in the postnerve-injury state. Intrathecal administration of dynorphin A(1–17) antiserum blocked tactile allodynia and reversed thermal hyperalgesia to above baseline levels (i.e., antinociception). Spinal dynorphin antiserum, but not control serum, also reestablished the antinociceptive potency and efficacy of spinal morphine. Neither dynorphin antiserum nor control serum administration altered baseline non-noxious or noxious thresholds or affected the intrathecal morphine antinociceptive response in saline-infused rats. These data suggest that spinal dynorphin promotes abnormal pain and acts to reduce the antinociceptive efficacy of spinal opioids (i.e., tolerance). The data also identify a possible mechanism for previously unexplained clinical observations and offer a novel approach for the development of strategies that could improve the long-term use of opioids for pain.

An immunohistochemically distinct zone was identified in the superficial aspect of trigeminal nucleus caudalis of the New World owl monkey that is not immunoreactive for substance P or serotonin, in stark contrast to the dense staining present in the surrounding laminae I and II. Thionin-stained sections in different planes showed that this is a subregion of lamina I containing clusters of neurons that appear to have pyramidal or polygonal somata. Extracellular microelectrode recordings in this region revealed clusters of thermoreceptive-specific (COLD) cells with nasal or labial receptive fields, whereas nociceptive neurons were found in the adjacent portions of lamina I. Anterograde tracer injections in this region produced trigeminothalamic terminal labeling in the site homologous to the lamina I spino-thalamo-cortical relay nucleus identified previously in the Old World macaque monkey and in humans. Retrograde tracer injections involving this thalamic site, where recordings of trigeminal COLD-like neurons were obtained, produced clusters of retrogradely labeled trigeminothalamic neurons in this immunohistochemically distinct subregion of lamina I, nearly all of which are pyramidal neurons. We conclude that the nocturnal owl monkey has a specialized perinasal thermoreceptive trigeminothalamic sensory pathway that is probably of behavioral significance during olfactory sniffing. In addition, these observations corroborate other findings that have indicated that lamina I COLD cells are pyramidal neurons and are not physiologically modulated by substance P or serotonin, in contrast to nociceptive neurons.

In cat and monkey, lamina I cells can be classified into three basic morphological types (fusiform, pyramidal, and multipolar), and recent intracellular labeling evidence in the cat indicates that fusiform and multipolar lamina I cells are two different types of nociceptive cells, whereas pyramidal cells are innocuous thermoreceptive-specific. Because earlier observations indicated that only nociceptive dorsal horn neurons respond to substance P (SP), we examined which morphological types of lamina I neurons express receptors for SP (NK-1r). We categorized NK-1r-immunoreactive (IR) lamina I neurons in serial horizontal sections from the cervical and lumbar enlargements of four monkeys. Consistent results were obtained by two independent teams of observers. Nearly all NK-1r-IR cells were fusiform (42%) or multipolar (43%), but only 6% were pyramidal (with 9% unclassified). We obtained similar findings in three monkeys in which we used double-labeling immunocytochemistry to identify NK-1r-IR and spinothalamic lamina I neurons retrogradely labeled with cholera toxin subunit b from the thalamus; most NK-1r-IR lamina I spinothalamic neurons were fusiform (48%) or multipolar (33%), and only 10% were pyramidal. In contrast, most (approximately 75%) pyramidal and some (approximately 25%) fusiform and multipolar lamina I spinothalamic neurons did not display NK-1r immunoreactivity. These data indicate that most fusiform and multipolar lamina I neurons in the monkey can express NK-1r, consistent with the idea that both types are nociceptive, whereas only a small proportion of lamina I pyramidal cells express this receptor, consistent with the previous finding that they are non-nociceptive. However, these findings also indicate that not all nociceptive lamina I neurons express receptors for SP.

Pain and temperature stimuli activate neurons of lamina I within the dorsal horn of the spinal cord, and although these neurons can be classified into three basic morphological types and three major physiological classes, earlier studies did not establish a structure/function correlation between their morphology and their physiological responses. We recorded and intracellularly labeled 38 cat lamina I neurons. All 12 fusiform cells were nociceptive-specific, responsive only to pinch and/or heat. All 11 pyramidal cells were thermoreceptive-specific, responsive only to innocuous cooling. Of ten multipolar cells, six were polymodal, responsive to heat, pinch and cold, and four were nociceptive-specific. Five unclassified cells had features consistent with this pattern. These results support the view that central pain and temperature pathways contain anatomically discrete sets of modality-selective neurons.

Lamina I spinothalamic tract (STT) neurons were identified by retrograde labeling with cholera toxin subunit b (CTb) in monkeys. On the basis of the criteria of somatal shape and dendritic orientation in horizontal sections used in prior work in the cat, three distinct morphological types were recognized: fusiform (F) cells with spindle-shaped somata and two main longitudinal dendritic arbors; pyramidal (P) cells with triangular somata and three main dendrites oriented primarily longitudinally; and multipolar (M) cells with polygonal somata and four or more dendrites directed longitudinally and mediolaterally. Some cells had transitional shapes, but cells with indeterminate shapes and a few with small round, unipolar, or eccentric somata were grouped as unclassified (U). Greater variation appeared in the monkey than had been seen in the cat, and more subtypes were noted. The overall proportions of these cell types were: 47% F, 27% P, 22% M, and 5% U. Differential longitudinal distributions were found over the length of the spinal cord (from the second cervical through the first coccygeal segments). Pyramidal and multipolar cells together predominated in the enlargements, whereas fusiform cells predominated in thoracic segments. We conclude that three distinct morphological types of lamina I STT cells are present in the monkey as in the cat. Considered with other recent findings, the present results support the possibility that these three cell types may correspond to distinct physiological classes of nociceptive and thermoreceptive lamina I STT cells.

We examined the morphology and distribution of retrogradely labeled spinothalamic tract (STT) neurons in lamina I (the marginal zone) of the spinal dorsal horn after large injections of cholera toxin subunit B (CTb) or Fast Blue (FB) into the contralateral thalamus of cats. Based on the shape and orientation of the somata and proximal dendrites in horizontal sections, three distinct cell types were identified: (1) fusiform cells with small, spindle-shaped somata and bipolar, longitudinal dendritic arbors; (2) pyramidal cells with triangular somata and three main dendritic origins with primarily longitudinal arborizations; and (3) multipolar cells with larger, multiangular somata and four or more radiating dendritic arbors directed both longitudinally and mediolaterally. These three morphological types differed significantly in the number of primary dendrites and the size of the somata. Subclasses of multipolar cells were noted. Nearly all cells could be categorized into these three classes consistently in horizontal sections. A small number of cells with transitional shapes or with small, round somata were unclassified.The proportional distributions of these cell types were found to vary over the length of the spinal cord (from the third cervical through the coccygeal segments) in three cats. The overall proportions of cell types were 34% fusiform, 36% pyramidal, 25% multipolar, and 5% unclassified. The proportions of pyramidal and multipolar cells were strikingly higher within the C7-8 and L6-7 segments and lowest in the thoracic segments. In contrast, fusiform cells formed about 20% of the labeled lamina I STT population in the C7-8 and L6-7 segments but more than 60% in thoracic segments. Across all nine cats, the proportions were similar within the cervical (C5-8) and lumbosacral (L5-S1) enlargements, although considerable interanimal variability was noted.These distinct morphological types of lamina I STT cells with differential longitudinal distributions probably have different functional roles. They may correspond to the three main physiological classes of lamina I STT cells. (C) 1996 Wiley-Liss, Inc.

Division of Neurobiology Barrow Neurological Institute 350 West Thomas Road Phoenix, AZ 85013 USA Centre de Recherche en Sciences Neurologiques and Département de Stomatologie Faculté de Médicine Dentaire Université de Montreal Montreal, Quebec H3C 3J7 Canada Dept. of Cell Biology Faculty of Health Sciences University of Linköping Linköping S-581 85 Sweden

THE existence of a posterolateral thalamic relay nucleus for pain and temperature sensation was postulated in 1911, on the basis of the stroke-induced analgesia and thermanaesthesia found paradox-ically in patients with thalamic pain syndrome1. Pain or tempera-ture sensations can be evoked in humans by electrical stimulation in a vaguely defined region of the posterolateral thalamus2'3. Here we use anterograde tracing and single unit recordings to demon-strate that there is a distinct nucleus in the posterior thalamus of the macaque monkey that receives a dense, topographic input from spinothalamic lamina I neurons and in which almost all neurons are nociceptive- or thermoreceptive-specific. Immunohistochemical staining showed that this nucleus is defined by a dense calbindin-positive fibre plexus in the macaque, so we applied the same stain-ing method to sections of human thalamus. We found a nearly identical fibre plexus localized within a distinct nucleus that is cytoarchitectonically homologous to the lamina I relay nucleus in the macaque thalamus. The stereotaxic coordinates of this nucleus and its location relative to the main somatosensory representation fit clinical descriptions of the pain-producing region in humans. We conclude that this is a specific thalamic nucleus for pain and temperature sensation in both monkey and human.