Position of spinothalamic tract axons in upper cervical spinal cord of monkeys.

Percutaneous upper cervical cordotomy continues to be performed on patients suffering from several types of severe chronic pain. It is believed that the operation is effective because it cuts the spinothalamic tract (STT), a primary pathway carrying nociceptive information from the spinal cord to the brain in humans. In recent years, there has been controversy regarding the location of STT axons within the spinal cord. The aim of this study was to determine the locations of STT axons within the spinal cord white matter of C2 segment in monkeys using methods of antidromic activation. Twenty lumbar STT cells were isolated. Eleven were classified as wide dynamic range neurons, six as high-threshold cells, and three as low-threshold cells. Eleven STT neurons were recorded in the deep dorsal horn and nine in superficial dorsal horn. The axons of the examined neurons were located at antidromic low-threshold points (<30 microA) within the contralateral lateral funiculus of C2. All low-threshold points were located ventral to the denticulate ligament, within the lateral half of the ventral lateral funiculus (VLF). None were found in the dorsal half of the lateral funiculus. The present findings support our previous suggestion that STT axons migrate ventrally as they ascend the length of the spinal cord. Also, the present findings indicate that surgical cordotomies that interrupt the VLF in C2 likely disrupt the entire lumbar STT.

Locations of spinothalamic tract axons in cervical and thoracic spinal cord white matter in monkeys.

The spinothalamic tract (STT) is the primary pathway carrying nociceptive information from the spinal cord to the brain in humans. The aim of this study was to understand better the organization of STT axons within the spinal cord white matter of monkeys. The location of STT axons was determined using method of antidromic activation. Twenty-six lumbar STT cells were isolated. Nineteen were classified as wide dynamic range neurons and seven as high-threshold cells. Fifteen STT neurons were recorded in the deep dorsal horn (DDH) and 11 in superficial dorsal horn (SDH). The axons of 26 STT neurons were located at 73 low-threshold points (<30 microA) within the lateral funiculus from T(9) to C(6). STT neurons in the SDH were activated from 33 low-threshold points, neurons in the DDH from 40 low-threshold points. In lower thoracic segments, SDH neurons were antidromically activated from low-threshold points at the dorsal-ventral level of the denticulate ligament. Neurons in the DDH were activated from points located slightly ventral, within the ventral lateral funiculus. At higher segmental levels, axons from SDH neurons continued in a position dorsal to those of neurons in the DDH. However, axons from neurons in both areas of the gray matter were activated from points located in more ventral positions within the lateral funiculus. Unlike the suggestions in several previous reports, the present findings indicate that STT axons originating in the lumbar cord shift into increasingly ventral positions as they ascend the length of the spinal cord.

Physiological studies of spinohypothalamic tract neurons in the lumbar enlargement of monkeys.

Recent anatomic results indicate that a large direct projection from the spinal cord to the hypothalamus exists in monkeys. The aim of this study was to determine whether the existence of this projection could be confirmed unambiguously using electrophysiological methods and, if so, to determine the response characteristics of primate spinohypothalamic tract (SHT) neurons. Fifteen neurons in the lumbar enlargement of macaque monkeys were antidromically activated using low-amplitude current pulses in the contralateral hypothalamus. The points at which antidromic activation thresholds were lowest were found in the supraoptic decussation (n = 13) or in the medial hypothalamus (n = 2). Recording points were located in the superficial dorsal horn (n = 1), deep dorsal horn (n = 10), and intermediate zone (n = 4). Each of the 12 examined neurons had cutaneous receptive fields on the ipsilateral hindlimb. All neurons responded exclusively or preferentially to noxious stimuli, suggesting that the transmission of nociceptive information is an important role of primate SHT axons. Twelve SHT neurons were also antidromically activated from the thalamus. In all cases, the antidromic latency from the thalamus was shorter than that from the hypothalamus, suggesting that the axons pass through the thalamus then enter the hypothalamus. These results confirm the existence of a SHT in primates and suggest that this projection may contribute to the production of autonomic, neuroendocrine, and emotional responses to noxious stimuli in primates, possibly including humans.

Cloning of rabbit TR4 and its bone cell-specific activity to suppress estrogen receptor-mediated transactivation.

To clone a new nuclear receptor, we screened a rabbit heart complementary DNA (cDNA) library with degenerate oligonucleotide probes corresponding to the DNA-binding domain of nuclear receptors, which is highly conserved among receptors. One of the cDNA clones, clone 23, encodes a novel protein of 596 amino acids, and predicted molecular mass is 66 kDa. Homology search analysis identified this protein as rabbit TR4 (TR4-0). We also cloned the cDNA encoding a rabbit TR4 isoform (TR4-1), which lacks the putative C-terminal ligand-binding domain (350 amino acids) caused by a 23-bp exon deletion, which probably occurred during messenger RNA (mRNA) splicing. Northern blot analysis showed that TR4s are expressed with two kinds of mRNAs (9.0 kb and 2.8 kb), both of which are relatively abundant in brain, testis, and bone. RT-PCR analysis, using pairs of primers specific for each TR4, showed that both types of receptor express in various tissues. Furthermore, both are present in primary osteoblasts and bone marrow cells, though the mRNA levels of TR4-0 were much higher than those of TR4-1. A functional study, using a transient transfection assay, showed that both receptors suppressed retinoid X receptor (RXR)-retinoid acid receptor, RXR-TR, and RXR-VDR-mediated transactivation significantly in COS-1 and osteosarcoma cells (UMR-106, ROS17/2.8) and that TR4-0 was much more effective than TR4-1. Unexpectedly, we found that the TR4s effectively suppressed estrogen receptor-mediated transactivation in bone cells, but neither in kidney (COS-1) nor breast cancer cells (MCF-7, one of the major target cells of the estrogen action). Thus, the present study shows a novel property of the TR4 orphan receptor, acting as a bone cell-specific repressor in the estrogen receptor-mediated signaling pathway.

A method for improving the accuracy of stereotaxic procedures in monkeys using implanted fiducial markers in CT scans that also serve as anchor points in a stereotaxic frame.

We developed a relatively inexpensive method for stereotaxic placement of electrodes or needles in the brains of monkeys. Steel balls were affixed to the skulls of monkeys. These balls served as fiducial markers and were also used as points at which the monkey's skull was held in a modified stereotaxic apparatus. Computed tomography (CT) was used to establish the location of an injection target with respect to the fiducial markers. A computer program related the CT coordinates to stereotaxic coordinates. These were used to direct an electrode marker toward a target in the hypothalamus. With the marker left in place, the monkey was removed from the stereotaxic frame and a second CT scan was performed. Corrections for errors in marker placement were made and retrograde tracers were injected. This procedure was found to be more accurate and reliable than conventional stereotaxic procedures. The accuracy and repeatability of the technique were also established using a phantom model of a monkey's skull. Two important advantages of this method are that animals can be repeatedly placed into the stereotaxic frame in precisely the same position and that there are many opportunities during the procedure to check for and correct errors.

Distribution of four types of synapse on physiologically identified relay neurons in the ventral posterior thalamic nucleus of the cat.

This study was aimed at providing quantitative data on the thalamic circuitry that underlies the central processing of somatosensory information. Four physiologically identified thalamocortical relay neurons in the ventral posterior lateral nucleus (VPL) of the cat thalamus were injected with horseradish peroxidase and subjected to quantitative electron microscopy after pre- or postembedding immunostaining for gamma-aminobutyric acid to reveal synaptic terminals of thalamic inhibitory neurons. The four cells all had rapidly adapting responses to light mechanical stimuli applied to their receptive fields, which were situated on hairy or glabrous skin or related to a joint. Their dendritic architecture was typical of cells previously described as type I relay cells in VPL, and they lacked dendritic appendages. Terminals ending in synapses on the injected cells were categorized as RL (ascending afferent), F (inhibitory), PSD (presynaptic dendrite), and RS (mainly corticothalamic) types and were quantified in reconstructions of serial thin sections. RL and F terminals formed the majority of the synapses on proximal dendrites (approximately 50% each). The number of synapses formed by RL terminals declined on intermediate dendrites, but those formed by F terminals remained relatively high, declining to moderate levels (20-30%) on distal dendrites. RS terminals formed moderate numbers of the synapses on intermediate dendrites and the majority (> 60%) of the synapses on distal dendrites. Synapses formed by PSDs were concentrated on intermediate dendrites and were few in number (approximately 6%). They formed synaptic triads with F terminals and rarely with RL terminals. On somata, only a few synapses were found, all made by F terminals. The total number of synapses per cell was calculated to be 5,584-8,797, with a density of 0.6-0.9 per micrometer of dendritic length. Of the total, RL terminals constituted approximately 15%, F terminals approximately 35%, PSD terminals approximately 5%, and RS terminals approximately 50%. These results provide the first quantitative assessment of the synaptic architecture of thalamic somatic sensory relay neurons and show the basic organizational pattern exhibited by representatives of the physiological type of relay neurons most commonly encountered in the VPL nucleus.

Anti-allergic effect of bryonolic acid from Luffa cylindrica cell suspension cultures.

The anti-allergic activity of bryonolic acid (1) isolated from the cultured cells of Luffa cylindrica L. (Cucurbitaceae) was compared with that of glycyrrhetinic acid (2), the aglycone of glycyrrhizin from licorice. Compound 1, when administered to rats intraperitoneally at a dose of 600 mg/kg, inhibited homologous passive cutaneous anaphylaxis more strongly than 2 at the same dose. Compound 1 also significantly inhibited delayed hypersensitivity in mice which could not be inhibited by 2. In contrast to 2, 1 showed not only little toxicity but no visible side effects on mice, without impairing the activity of the hepatic enzyme (4,5 beta-dihydrocortisone:NADP+ delta 4-oxidoreductase) involved in steroid catabolism.

Electrophysiological study of spinothalamic inputs to ventrolateral and adjacent thalamic nuclei of the cat.

1. Extracellular and intracellular methods were used to record from fibers and neurons in the ventral lateral (VL) and adjacent nuclei of the cat thalamus. The receptive fields of the recorded units were analyzed and the units tested for inputs from the medial lemniscus (ML) and spinothalamic tract (STT) by electrical stimulation of the dorsal columns (DC) and ventrolateral funiculus (VLF) at the C2-3 spinal level. 2. Thirty-eight STT fibers were isolated in the thalamus. Their conduction velocities ranged from 15 to 75 m/s (mode 36 m/s). Adequate stimuli were found for 23 of these fibers. Seventeen were low-threshold (LT), 3 were wide-dynamic-range (WDR), and 3 were high-threshold (HT) units. 3. Five STT fibers were intra-axonally injected. Three were sufficiently well filled for analysis of their terminal fields. An intermediate-velocity STT fiber (conduction velocity 38 m/s) had a 4.3-microns axon and a single large terminal field in the central lateral nucleus (CL). The other two STT fibers were smaller, with diameters of 2.5 and 2.3 microns, conduction velocities of 15 and 19 m/s, and terminal fields made up of a few small boutons at the borders of the ventral posterior lateral nucleus (VPL). 4. Of 319 neurons isolated, 14 out of 129 (10.8%) in VL, 14 out of 76 (18.4%) in the VPL or ventral posterior medial (VPM) nucleus, 27 out of 64 (42.2%) in the CL nucleus, and 5 out of 50 (10%) in the reticular nucleus (R) responded at latencies less than 50 ms to VLF stimuli. A train of three pulses was more effective in driving VLF-responding neurons in all these nuclei than a single pulse. VLF-responding cells were widely dispersed in VL, concentrated in a focus in CL, and distributed around the borders of VPL. Most of those in VL and a small number in CL could be antidromically activated by stimulation of motor cortex. 5. Latencies of presynaptic responses (STT fibers) to VLF stimulation were short and varied from 0.8 to 3.9 ms (mode 1.6 ms). Despite this, very few fast-responding neurons were found. These were six VPL neurons (2.5 to 4 ms), one VL neuron (3 ms), and four CL neurons (3-4 ms). The initial spike latencies of the majority of thalamic neurons responding to VLF stimulation appeared in two peaks, one between 6 and 8 ms and the other at 10-15 ms.(ABSTRACT TRUNCATED AT 400 WORDS)

Descending adrenergic input to the primate spinal cord and its possible role in modulation of spinothalamic cells.

The present study focuses on 3 different aspects of the descending adrenergic system in the primate: (1) the distribution of adrenergic fibers and terminals in the spinal cord, (2) the source of this input and (3) the possible physiological effects of this system on spinal nociceptive processing. Antibodies to the enzyme phenylethanolamine-N-methyltransferase (PNMT) were employed to map the distribution of epinephrine-containing axonal profiles in the primate spinal cord. Smooth longitudinally oriented fibers were localized to the outer edge of the lateral funiculus. PNMT-containing axonal enlargements were distributed to the superficial dorsal horn, intermediate gray matter and the region surrounding the central canal at all spinal cord levels. PNMT-immunostained profiles were also observed in the intermediolateral cell column. A double labeling study employing retrograde transport of HRP from the spinal cord and PNMT immunohistochemistry identified a small population of HRP-PNMT-labeled neurons in the 'C1' region at the levels of the medulla and ponto-medullary junction. Thus, these cells are a probable source of adrenergic input to the spinal cord. Electrophysiological studies demonstrated that iontophoresis of epinephrine onto identified primate spinothalamic tract neurons in the lumbar dorsal horn resulted in inhibition of the glutamate-induced firing of these cells. The data from these studies support the hypothesis that adrenergic (PNMT-containing) cells in the caudal brainstem project to all levels of the cord and may contribute to descending modulation of nociceptive processing at these levels.

Natural groupings of primate spinothalamic neurons based on cutaneous stimulation. Physiological and anatomical features.

1. Two hundred and twenty-one spinothalamic tract (STT) neurons in the lumbar spinal cord of anesthetized monkeys were studied. The majority of the recordings were in laminae IV-VI. Thirteen of these neurons were intracellularly injected with horseradish peroxidase and histologically reconstructed. 2. A standard series of four mechanical cutaneous stimuli, which ranged in intensity from innocuous brushing to tissue-damaging pinching, were used to test the mechanical responsiveness of STT neurons. The mean alterations in discharge rate produced by these test stimuli when delivered to a neuron's excitatory receptive field were used as response measures. 3. Univariate and bivariate analyses of these response measures failed to reveal natural groupings of STT neurons. To assess whether natural groupings dependent upon shared multivariate response patterns were present, a k-means cluster analysis of the responses was performed. 4. Because an assumption about the type of coding used by the STT system had to be made prior to clustering, two independent analyses were performed. One approach assumed a labeled line coding model; response magnitudes were determined within the context of the neuron under study (within-neuron analysis). The other approach assumed a population coding model; response magnitudes were determined within the context of the STT population (across-neuron analysis). 5. The within-neuron analysis suggested that the STT sample could be partitioned into four groups. The smallest group (n = 18, 8%) responded primarily to brushing but often had a convergent nociceptive input; this group was referred to as type I. A second group (n = 31, 14%) had strong responses to low-intensity stimuli, particularly pressure, and modestly larger responses to noxious stimuli; this group was referred to as type II. The clustering in these two groups was relatively weak, reflecting some heterogeneity in response pattern. 6. The largest within-neuron group (n = 108, 49%) was most responsive to noxious stimuli but had a saturating response function; because of their apparent role in coding intermediate intensity stimuli, this group was referred to as type III. The fourth group (n = 64, 29%) responded best to the most intense stimulus used; this group was referred to as type IV. 7. The across-neuron analysis also suggested that the STT sample could be partitioned into four groups. The largest group (n = 122, 55%) had relatively weak responses to all the cutaneous stimuli; this group was referred to as type A. 8. All of the remaining across-neuron groups had mean responses at or above the mean for all cutaneous stimuli.(ABSTRACT TRUNCATED AT 400 WORDS)

Classification of primate spinothalamic and somatosensory thalamic neurons based on cluster analysis.

Data analyzed in this study were derived from the responses of 128 spinothalamic tract (STT) cells and 110 thalamic neurons recorded in 75 anesthetized monkeys. A k-means cluster analysis, a nonhierarchical clustering technique, was performed using the relative magnitudes of responses to a graded series of innocuous and noxious mechanical stimuli applied to the receptive field. For comparison, a parallel analysis was performed based on definitions of low-threshold (LT), wide dynamic range (WDR), and high-threshold (HT) cells used by our laboratory. For 128 STT cells, a classification scheme with three clusters was found statistically to be the best. This yielded groups of 22, 57, and 49 cells in clusters 1, 2, and 3, respectively. Cluster 1 cells were activated best by low-intensity mechanical stimuli, whereas cluster 3 cells were activated primarily by nociceptive stimuli. Cluster 2 cells had intermediate characteristics. When the classification scheme based on the cluster analysis was compared with the classification of the same neurons as LT, WDR, and HT cells, cluster 1 cells were divided into LT and WDR cells, whereas cluster 2 and 3 cells included WDR and HT cells. For 110 thalamic neurons, a classification scheme with five clusters was found statistically to be the best. Clusters 1-5 contained 25, 34, 17, 10, and 24 cells, respectively. Response characteristics of cells in each group indicated a gradual change in sensitivity to higher intensities of peripheral input from cluster 1 to 5. When this classification scheme was compared with the classification scheme previously used by our laboratory, cluster 1 cells belonged to the LT group, clusters 2 and 3 split into LT and WDR cells, and clusters 4 and 5 included WDR and HT cells. It is concluded that a classification scheme based on a cluster analysis of the responses of neurons to standardized stimuli may provide an objective and functionally meaningful way to categorize somatosensory neurons.

Responses of primate spinothalamic neurons to noxious thermal stimulation of glabrous and hairy skin.

Extracellular recordings were made from 81 primate spinothalamic (STT) neurons in the L7-S1 segments of the spinal cord. The majority of the sample was recorded from within laminae IV-V. The responses of STT neurons to noxious thermal stimulation of glabrous and hairy skin were studied in an attempt to identify a neural substrate for the differences in thermal sensation evoked by noxious stimulation of these two types of skin. In addition, the responses to graded mechanical stimuli were examined for evidence of differential sensitivity. Thermal intensity-response functions were constructed from the alteration in the mean discharge rate produced by a 30-s thermal pulse of 43-55 degrees C. Generally, the functions derived from stimulation of both hairy and glabrous skin were either linear or positively accelerating. Deceleration in the response functions was occasionally observed above 53 degrees C. The population mean discharge rate derived from glabrous skin stimulation was significantly greater than that derived from hairy skin stimulation above 49 degrees C. Cluster analysis was used to assess whether the STT population could be partitioned into functionally relevant subgroups. No clustering was evident on the basis of the alteration in discharge rate during stimulation alone. Analysis of the alteration in mean discharge rate during and following thermal stimulation identified four groups; these were referred to as the Amnr, Bmnr, Cmnr, and Dmnr classes. The clustering was not dependent on differences in the responses evoked from hairy and glabrous skin. The mechanical and thermal sensitivities of each thermal class covaried. The capacity of the STT population to code the quality of noxious thermal stimuli, as judged by changes in the across-neuron discharge pattern, was assessed with a multidimensional scaling technique (MDS). The results suggest that the population discharge could be used to order stimuli correctly from 45 to 55 degrees C. Also, it was found that a substantial change in the population's discharge pattern occurred to stimuli between 47 and 49 degrees C when delivered to hairy skin. A similar alteration in the population's discharge pattern occurred to glabrous skin stimuli near 51 degrees C. These alterations in population behavior may underly the alterations in sensory quality in humans that occur in these temperature ranges when stimulating hairy and glabrous skin. The possible roles of the thermally and mechanically based classes in thermal intensity and quality coding were examined. Within the lower intensity ranges (less than 49-51 degrees C), the Cmnr and Dmnr classes appeared to be best suited to intensity coding.(ABSTRACT TRUNCATED AT 400 WORDS)

Temporal features of the responses of primate spinothalamic neurons to noxious thermal stimulation of hairy and glabrous skin.

Extracellular recordings were made from 81 primate spinothalamic (STT) neurons in the L7-S1 segments of the spinal cord. The majority of the sample was recorded from within laminae IV-V. The temporal features of the responses to noxious thermal stimulation of glabrous and hairy skin were studied in an attempt to determine whether natural groupings of STT neurons could be identified on the basis of response time course alone and whether these groups were skin type dependent. The relationship between these groups and those based on static response features (37) was also explored in an attempt to define more fully their potential functional roles. In most STT neurons, the thermally evoked responses typically appeared to have two response components, particularly at stimulus temperatures above 49 degrees C. The first response phase typically peaked within 1-12 s of stimulus onset and then adapted. The second phase slowly rose to a maximum, typically 15-30 s following stimulus onset. The existence of natural groupings of STT neurons based upon the characteristics of these two response components was assessed with a k-means cluster analysis. On the basis of the onset and early peak latencies, two well-defined short and long latency neuronal clusters were found in the responses evoked from both glabrous and hairy skin; these were referred to as the SP1 and LP1 classes, respectively. The glabrous and hairy skin SP1 classes did not differ significantly in either onset or early peak latency for stimuli of 47-55 degrees C. However, the hairy skin LP1 class had significantly shorter onset latencies than the glabrous skin LP1 class for stimuli of 49-53 degrees C, as well as shorter peak latencies for stimuli of 49 and 51 degrees C. The SP1 class constituted 62% of the hairy skin subset, whereas the LP1 class constituted 57% of the glabrous skin subset. A cluster analysis of the late-peak latencies also revealed two subgroups. In the responses evoked from both glabrous and hairy skin, the longer latency classes (LP2) constituted more than 80% of the samples. With one exception, no dependence upon the type of skin that was stimulated was found in the latencies of either the LP2 class or the shorter latency SP2 class. Prior conditioning of the skin with a 30-s thermal pulse of 51-55 degrees C led to a suppression of the early response phase and an enhancement of the late phase in nearly all cases examined (n = 11). This pattern was independent of skin type.(ABSTRACT TRUNCATED AT 400 WORDS)

Neurons in ventrobasal region of cat thalamus selectively responsive to noxious mechanical stimulation.

1. A survey was made of neurons located in the ventral posterior lateral nucleus of the cat thalamus and its immediate vicinity for elements with specifically nociceptive properties. 2. Pipette microelectrodes filled with a dye solution were used to obtain extracellular recordings of unitary activity in 34 animals anesthetized with chloralose. 3. The great majority of the over 1,000 different single units responding to sciatic nerve stimulation noted in this series of experiments could also be excited by innocuous mechanical stimulation of skin or subcutaneous tissues. An infrequent but consistently noted group of units excited by A-alpha beta delta sciatic nerve volleys did not respond to innocuous mechanical manipulation or A-alpha beta sciatic nerve volleys; they were excited only by either noxious levels of mechanical stimulation or when volleys included the activity of more slowly conducting myelinated fibers. The latencies of such "high-threshold" units to sciatic volleys were longer than those of the other units. 4. Histologically identified recording sites marked by dye were recovered for 17 high-threshold units. Twelve of the 17 could be excited by noxious manipulations of restricted parts of the contralateral hindlimb. Nine of the 12 had cutaneous receptive fields, whereas 3 responded only to stimulation of subcutaneous tissues. None of the 17 high-threshold units evidenced additional discharges that could be correlated with the C-fiber component of sciatic nerve volleys. 5. The high-threshold units typically exhibited a low level of irregular background activity, which increased on repeated noxious stimulation of the peripheral receptive fields. Tactile units of the same or adjacent penetrations usually had a much greater degree of ongoing activity, often marked by bursts at a relatively high frequency. 6. The recording sites for the 17 high-threshold neurons were located dorsal and ventrolateral to the core of the ventrobasal nuclei and were not found in the midst of the low-threshold, cutaneous, mechanoreceptive population. During vertical stereotaxic penetrations, high-threshold units were noted dorsal or ventral to the location of ventrobasal tactile units in a pattern consistent with the core's somatotopic arrangement. 7. These results support the concept that the cat ventrolateral thalamus receives a small but distinct selectively nociceptive projection. The nociceptive neurons appear to be located in a shell that surrounds the main tactile projection to the ventral posterior lateral nucleus and that retains at least part of the topographic arrangement characteristic of the tactile core. Presumably, this projection is part of an organization identifying and localizing noxious stimulation.