Temporal and spatial profiles of pontine-evoked monoamine release in the rat's spinal cord

In the spinal cord, the monoamine neurotransmitter norepinephrine, which is released mainly from fibers descending from the dorsal pons, has major modulatory effects on nociception and locomotor rhythms. To map the spatial and temporal patterns of this release, changes in monoamine level were examined in laminae I-VIII of lumbar segments L3-L6 of halothane-anesthetized rats during pontine stimulation. The changes were measured through a carbon fiber microelectrode at 0.5-s intervals by fast cyclic voltammetry, which presently is the method of best spatiotemporal resolution. When different pontine sites were tested with 20-s pulse trains (50-to 200-microA amplitude, 0.5-ms pulse width, and 50-Hz frequency) during measurement in the dorsal horn (lamina IV), the largest consistent increases were produced by the locus ceruleus, although effective pontine sites extended 1.5 mm dorsally and ventral from the locus ceruleus. When the locus ceruleus stimulus was used to map the spinal cord, increased levels were always seen in lamina I and laminae IV-VIII, whereas 50% of sites in laminae II and III showed substantial decreases and the rest showed increases. These increases typically had short latencies [4.5 +/- 0.4 (SE) s] and variable decay times (5-200 s), with peaks occurring during the stimulus train (mean rise-time: 12.0 +/- 0.6 s). The mean peak level was 544 +/- 82 nM as estimated from postexperimental calibration with norepinephrine. Other significant laminar differences included higher mean peak concentrations (805 nM) and rise times (14.9 s) in lamina I and shorter latencies in lamina VI (3.2 s). Peak concentrations were inversely correlated with latency. When stimulation frequency was varied, increases were disproportionately larger with faster frequencies (> or =50 Hz), hence extrajunctional overflow probably contributed most of the signal. We conclude, generally, that pontine noradrenergic control is exerted on widespread spinal laminae with a significant component of paracrine transmission after several seconds of sustained activity. Relatively stronger effects prevail where nociceptive transmission (lamina I) and locomotor rhythm generation (lamina VI) occur.     

Quantitative comparison of inhibition in spinal cord of nociceptive information by stimulation in periaqueductal gray or nucleus raphe magnus of the cat

The descending inhibition of spinal neuronal responses by focal electrical stimulation in the periaqueductal gray (PAG) or nucleus raphe magnus (NRM) was quantitatively studied and compared in the anesthetized, paralyzed cat. All 60 dorsal horn neurons studied were driven by electrical stimulation of hindlimb cutaneous nerves at strengths supramaximal for activation of A-alpha,delta- and C-fibers, and 52 also responded to noxious radiant heating (50 degrees C, 10 s) of the skin of the foot- or toepads; 8 units had receptive fields in the hairy skin of the hindlimb. All neurons studied also responded to mechanical stimuli; recording sites were located in laminae I-VI of the dorsal horn. The inhibition of spinal neuronal heat-evoked responses by stimulation in the PAG or NRM differed quantitatively when examined on the same spinal neurons. Inhibition of heat-evoked spinal neuronal responses occurred at a lower threshold of stimulation in the NRM than in the PAG. The mean intensity of stimulation in the NRM producing an attenuation to 50% of the control 50 degrees C heat-evoked response was significantly lower than the mean intensity of stimulation in the PAG producing a 50% attenuation of the same spinal units. The mean magnitude of inhibition produced by stimulation in the NRM was significantly greater than that produced on the same spinal units by the same intensity of stimulation in the PAG. However, stimulation in the NRM and PAG produced the same mean percent change in inhibition per 100-microA increase in the intensity of stimulation. Thus, the slopes of the recruitment of descending inhibition from the PAG and the NRM as a function of increasing intensities of stimulation are the same; the lines of recruitment of inhibition are parallel. When examined on the same dorsal horn units, stimulation in the PAG influenced their intensity coding to graded noxious heating of the skin differently than did stimulation in the NRM. The responses of the class 2 and class 3 spinal units examined to increasing temperatures of heat applied to the skin was a monotonic linear function throughout the temperature range studied (42-50 degrees C). Stimulation in the PAG decreased the slope of the stimulus-response function (SRF) without affecting unit thresholds of response, thus influencing the gain control of nociceptive transmission in the dorsal horn. Stimulation in the NRM produced a parallel shift to the right of the SRF, influencing the set point and threshold of response.(ABSTRACT TRUNCATED AT 400 WORDS)     

The laminar organization of dorsal horn cells responding to peripheral C fibre stimulation

Summary Cat dorsal horn was searched for all detectable units that responded to peripheral C fibre input. Fifty-seven such units were examined in detail. They were located in two main areas. One group was in the superficial laminae 1, 2, and possibly dorsal 3 (n = 29), and the other group was much deeper in laminae 5 and 6 (n = 24). Only four units were situated in the region of lamina 4.Differences were found in the responses to C fibre stimulation of these two groups, both in the optimum stimulus and in the timing of responses to repeated stimulation. Superficial units often did not respond to C fibre stimulation unless a train of two or more stimuli (10 ms apart) were applied, but when responses did occur they were usually very even and regular, with precise onset latencies on repeated stimulation. Deep units tended to need only one peripheral C fibre stimulus for excitation, but the responses were irregular with latencies fluctuating with each stimulus. Some superficial and deep units showed a steady increase in latency of the late C response on repeated stimulation. Increases of up to 80 ms after 30 s of stimulation at 1 Hz were observed.The results are discussed in terms of the neuronal connections in the dorsal horn.The work was supported by the Medical Research Council and the National Institutes of HealthM. Fitzgerald is a Medical Research Council Training Fellow     

The influence of stimulus temperature rise rate, adapting temperature, and stimulus duration on suprathreshold responses evoked by noxious heat in the glabrous skin of the limb Comparison of neuronal discharge in the rat spinal dorsal horn with human sensations

 The influence of stimulus temperature rise rate (2.5oC/s, 5.0oC/s, and 10.0oC/s), adapting (baseline) temperature (25oC, 30oC, and 35oC), and duration of peak stimulus temperature (1.0 s, 2.5 s, 5.0 s, and 10.0 s) on responses evoked by noxious heat stimuli of suprathreshold intensity was studied in wide dynamic range (WDR) neurons of the rat spinal dorsal horn. The spinal neuronal responses were compared with human psychophysical data obtained using the same stimuli. Noxious heat stimuli with a peak temperature of 54oC were applied with a contact thermostimulator to the glabrous skin of the hindfoot in rats or to the palmar skin in humans. With the highest ramp rate and the highest adapting temperature, the sensory and spinal neuronal response latencies were decreased more than expected on the basis of the change in physical parameters of the stimulus. The magnitudes of sensory and spinal neuronal response were independent of the stimulus ramp rate, whereas pain magnitude estimates and spinal neuronal impulse counts evoked by the same peak stimulus temperature were increased with an increase in the adapting stimulus temperature. The onset latencies of pain reactions and spinal neuronal responses were independent of the peak stimulus duration, whereas the latency of the maximum discharge in spinal neurons increased with prolongation of the peak stimulus. The sensory magnitude estimate of pain and the neuronal impulse count were increased with increase in stimulus duration. Following spinalization, the spinal neuronal responses were stronger and the stimulus duration-dependent increase in the impulse count developed faster. Moreover, the peak frequency of spinal neuronal response increased significantly with prolongation of the heat stimuli after spinalization, but not in animals with an intact spinal cord. The results indicate that stimulus rise rate, stimulus duration, and the adapting temperature are important factors in determining the sensory and spinal neuronal responses to high-intensity heat stimuli. The changes in the total impulse counts evoked by varying supraliminal heat stimuli in spinal dorsal horn WDR neurons corresponded well with the changes in pain magnitude estimates in humans. Also, the changes in spinal neuronal response onset latencies were accompanied by corresponding changes in onset latencies of human pain reactions but not with pain magnitude estimates. The effect of spinalization indicated that descending pathways control not only the response magnitude in the spinal dorsal horn WDR neurons but also the temporal characteristics of the spinal neuronal response. Received: 24 August 1998 / Accepted: 25 January 1999     

Course and mode of action of descending system conveying nucleus raphe magnus induced inhibition of flexion reflex in rats

In lightly anaesthetized rats, the latencies and magnitudes of heat-evoked withdrawal reflexes from the hindlimb were measured electromyographically. Low-intensity (20-50 microA) stimulation of the nucleus raphe magnus (NRM) resulted in an inhibition of the flexion reflex (commonly referred to as analgesia) as evidenced by increased latency and decreased amplitude. The effect lasted for several minutes after the stimulation was terminated. Following lesions of the dorsolateral funiculus (DLF) at the neural thoracic levels 7-8, baseline latencies were reduced and the effect of the NRM stimulation was abolished. Lesions of the DLF at lumbar level 1 resulted in unaltered baseline latencies with persistence of the inhibitory effect of NRM stimulation. The results of the present experiment show that pathways exerting a tonic inhibition of the withdrawal reflex, and mediating the effect of electrical stimulation from the NRM, descend in the DLF at the thoracic level of the spinal cord. At the lumbar level, there is a shift away from the DLF. The antinociceptive effect of 20 microA NRM stimulation was partly reduced by pretreatment with the serotonin synthesis inhibitor para-chlorophenylalanine (PCPA) or the opiate naloxone (1 mg kg-1 i.v.). In animals pretreated with PCPA, naloxone (1 mg kg-1 i.v.) completely abolished the analgesic effect of the stimulation. Thus, both serotonergic and opioid systems may be implicated in mediating the analgesia. With 50 microA stimulation the same treatment only partly attenuated the NRM-induced analgesia, indicating an additional non-serotonergic and non-opioid mechanism which requires a higher current intensity for its activation.     

Serotonin is found in myelinated axons of the dorsolateral funiculus in monkeys.

Physiological measurements suggest that the inhibition of primate spinothalamic tract cells by serotonin is mediated by myelinated axons. Previous morphologic studies emphasize that most serotonin-containing axons in the spinal cord are unmyelinated. Accordingly, the possibility that some serotonin-containing axons in the primate dorsolateral funiculus of the spinal cord are myelinated was investigated. Macaque monkeys were given L-tryptophan and the monoamine oxidase inhibitor, nialamide, intraperitoneally 1 h prior to sacrifice to increase axonal stores of serotonin. The animals were perfused (0.05 or 0.5% glutaraldehyde, 4% paraformaldehyde), and transverse sections of the thoracic cord were reacted with antibody against serotonin and then prepared for electron microscopy. Many of the immunostained axons in the dorsolateral funiculus included fine, myelinated fibers with diameters of 0.7-2.2 microns. Unmyelinated serotonin-containing axons were also observed. The observation of myelinated serotonin-containing axons in the white matter of the monkey dorsolateral funiculus contradicts the view that the descending serotoninergic projection consists entirely of unmyelinated fibers, particularly since the conduction velocity of the fine fibers would be too slow to account for the earliest latency of descending inhibition following stimulation in the brainstem. The presence of myelinated serotoninergic axons presumably accounts for the latencies reported for the inhibition of primate spinothalamic cells following stimulation of the periaqueductal gray, an inhibition that can be blocked with serotonin antagonists and that is associated with the release of serotonin in the dorsal horn.     

Oscillatory interaction between dorsal root excitability and dorsal root potentials in the spinal cord of the turtle.

The response to dorsal root stimulation, at one to two times threshold, was investigated in the isolated cervical enlargement of the turtle spinal cord. At frequencies near 10 Hz the synaptic response in motoneurons and the cord dorsum potential, after an initial lag time, oscillated in amplitude with a period of more than 1 s. The mono- and polysynaptyic postsynaptic response in motoneurons, the pre- and postsynaptic component of the cord dorsum potential and the dorsal root potential oscillated in synchrony. These oscillations were only observed with stimulus frequencies in the range 9-11 Hz. The oscillating response could only be evoked from stimulus sites to which dorsal root potentials were conducted from the spinal cord (2-3 mm). At more distant stimulus sites cyclic variations in amplitude of the cord dorsum potential and the synaptic response in motoneurons were not observed. During an oscillating spinal response to a stimulus train in one dorsal root filament, the response evoked by a stimulus in another short filament (2-3 mm) from the same root varied in amplitude with the induced oscillation. The spinal response to a stimulus in a longer filament (i.e. more than 3 mm) did not oscillate. It is argued that the oscillating responses described rely on interactions between distributed elements rather than on unit oscillators. We also show that primary afferent transmission is unaffected by the substantial variations in dorsal root potentials during oscillations.     

Effects of midbrain and medullary stimulation on spinomesencephalic tract cells in the cat

1. The effects of electrical stimulation at different rostrocaudal levels of the midbrain, and at sites in the rostral medulla ipsilateral and contralateral to spinal recording sites, were evaluated against the responses of 46 cells belonging to the cat spinomesencephalic tract (SMT). 2. Inhibitory and/or excitatory effects of brain stem stimulation were observed on SMT cells that responded best (26 cells) or exclusively (12 cells) to noxious mechanical or thermal stimuli, as well as on 7 cells responding only to tap and/or stimulation of deep tissues. Recording sites for 32 cells were located in laminae V-VIII (27 cells) and laminae I-III (5 cells). 3. Midbrain stimulation sites were located in the superior colliculus, central gray (CG), red nucleus, and the midbrain reticular formation. Both inhibitory-only and excitatory-only effects were observed, although the most common effect of midbrain stimulation was excitation followed by inhibition (mixed effects). The effects of stimulation at different midbrain levels were determined for each cell. Stimulation in the caudal, middle, or rostral midbrain was often found to exert different effects on the same SMT cell. 4. Stimulation in the rostral medulla at sites located in nucleus raphe magnus (NRM), nucleus reticularis gigantocellularis, and nucleus reticularis magnocellularis produced the same complement of effects observed with midbrain stimulation. Excitation followed by inhibition was the most common effect observed. 5. Stimulus intensities required to produce excitatory or inhibitory effects from midbrain were 114 +/- 85 (SD) microA and 210 +/- 91 microA, respectively. Stimulus currents required to produce excitatory or inhibitory effects from medullary stimulation sites were 124 +/- 56 microA and 70 +/- 60 microA, respectively. The mean currents required to produce mixed effects were 221 +/- 120 microA (midbrain) and 127 +/- 71 microA (medulla). Increasing the stimulus intensity used to evaluate brain stem effects increased the magnitude and duration of effects for 33 cells. Mixed effects were observed on 11 cells at stimulus intensities greater than those required to produce inhibitory-only effects. 6. Significant differences were found between the latencies of excitation and inhibition produced from different brain stem levels. These differences suggest that midbrain and medullary stimulation activate descending pathways with a wide range of conduction velocities and/or supraspinal and spinal connectivities. 7. The spinal trajectory of pathways contributing to the varied effects of brain stem stimulation as well as the complex receptive fields (RFs) of SMT cells were evaluated by the placement of lesions in the cervical spinal cord.(ABSTRACT TRUNCATED AT 400 WORDS)     

The effects of focal stimulation in nucleus raphe magnus and periaqueductal gray on intracellularly recorded neurons in spinal laminae I and II

Single neurons in spinal laminae I and II of cats were recorded intracellularly while stimulating in nucleus raphe magnus (NRM) and periaqueductal gray (PAG) with monopolar tungsten microelectrodes. Brain stem stimulation inhibited about one-half of the nociceptive-specific neurons, whereas the other half was unaffected. Brain stem stimulation inhibited about one-half of the multireceptive neurons, but the other half was excited and then inhibited. Brain stem stimulation inhibited about one-third of the low-threshold neurons, one-half was excited then inhibited, and one-fifth showed no effect. In all classes of neurons, the inhibition was produced by an inhibitory postsynaptic potential (IPSP) that began with a latency of approximately 25 ms and lasted approximately 400 ms following a single stimulus. The IPSP occurred with a small conductance increase and was reversed by hyperpolarizing currents applied to the cell. These data indicate that NRM and PAG modulated laminae I and II neurons via a postsynaptic mechanism. The conduction velocity of this descending pathway was calculated to range from 6.1 to 66.6 m/s with an average of 13.8 m/s. These data also indicate heterogeneity in the pathway, since some neurons were inhibited, whereas other neurons were excited then inhibited by descending stimulation. Finally, these data indicate specificity in these descending pathways since nearly one-half of neurons that had low-threshold inputs were excited by brain stem stimulation, whereas nearly all nociceptive-specific neurons were either inhibited or unaffected.     

Electrical stimulation of visceral afferent pathways in the pelvic nerve increases c-fos in the rat lumbosacral spinal cord.

Electrical stimulation (20-35 Hz, 2-5 V, 1.5 h) of the pelvic nerve in urethane-anesthetized rats increased the expression of c-fos protein-immunoreactivity primarily in neurons in the L6-S1 segments of the spinal cord. The neurons were localized to areas receiving afferent input from the pelvic viscera including the superficial dorsal horn, the dorsal commissure, and lateral laminae V-VII in the region of the sacral parasympathetic nucleus. These experiments indicate that (1) electrical stimulation of abdominal nerves following surgical exposure is a useful method for tracing visceral afferent pathways and (2) afferent information from the pelvic viscera is received by neurons in specific areas of the dorsal horn.     

Effects of electroacupuncture with different frequencies on spinal ionotropic glutamate receptor expression in complete Freund's adjuvant-injected rat

We investigated expressional changes of spinal glutamate receptors by electroacupuncture (EA) in an inflammatory animal model. Inflammation was induced by an intraplantar injection of complete Freund's adjuvant (CFA) into the hindpaw of male Sprague-Dawley rats. Bilateral EA stimulation at 2, 15 and 120 Hz was applied at those acupoints corresponding to Zusanli and Sanyinjiao in man using needles with 3-day intervals for 30 days. Paw edema and mechanical thresholds were measured by a water displacement plethysmometer and Analgesy-Meter, respectively. Edema and mechanical sensitivity of the hindpaw induced by CFA-injection were strongly inhibited by EA stimulation. At 30 days after CFA-injection, effects of EA on ionotropic glutamate receptor (NR-1, NR-2A, GluR-1 and GluR-2/3) expression in association with c-fos and calcitonin gene-related peptide (CGRP) expression were observed in the dorsal horn of the spinal cord using immunohistochemistry. The number of c-fos-like immunostained cells was decreased significantly in the superficial laminae of the dorsal horn by 2Hz EA, but CGRP expression also showed a marked decrease in the same region using the other types of EA stimulation. N-methyl-D-aspartate receptor (NR-1 and NR-2A) expression was attenuated in all regions of the dorsal horn by all types of EA. Of the alpha-amino-3-hydroxy-5-methylisoxazole-4-propionic acid receptor (GluR-1 and -2/3), only GluR-1 expression was prevented by EA treatment in the superficial laminae and the neck of the dorsal horn. It is concluded that EA treatment can attenuate inflammatory edema and mechanical thresholds in CFA-injected rats through modulating expression of ionotropic glutamate receptors, and especially N-methyl-D-aspartate receptors, in the dorsal horn of the spinal cord.     

Bicuculline and strychnine suppress the mesencephalic locomotor region-induced inhibition of group III muscle afferent input to the dorsal horn

We examined the effect of iontophoretic application of bicuculline methiodide and strychnine hydrochloride on the mesencephalic locomotor region (MLR)-induced inhibition of dorsal horn cells in paralyzed cats. The activity of 60 dorsal horn cells was recorded extracellularly in laminae I, II, V-VII of spinal segments L7-S1. Each of the cells was shown to receive group III muscle afferent input as demonstrated by their responses to electrical stimulation of the tibial nerve (mean latency and threshold of activation: 20.1+/-6.4 ms and 15.2+/-1.4 times motor threshold, respectively). Electrical stimulation of the MLR suppressed transmission in group III muscle afferent pathways to dorsal horn cells. Specifically the average number of impulses generated by the dorsal horn neurons in response to a single pulse applied to the tibial nerve was decreased by 78+/-2.8% (n=60) during the MLR stimulation. Iontophoretic application (10-50 nA) of bicuculline and strychnine (5-10 mM) suppressed the MLR-induced inhibition of transmission of group III afferent input to laminae I and II cells by 69+/-5% (n=10) and 29+/-7% (n=7), respectively. Likewise, bicuculline and strychnine suppressed the MLR-induced inhibition of transmission of group III afferent input to lamina V cells by 59+/-13% (n=14) and 39+/-11% (n=10), respectively.Our findings raise the possibility that GABA and glycine release onto dorsal horn neurons in the spinal cord may play an important role in the suppression by central motor command of thin fiber muscle afferent-reflex pathways.     

Pathways mediating descending control of spinal nociceptive transmission from the nuclei locus coeruleus (LC) and raphe magnus (NRM) in the cat

Summary We have previously reported that electrical stimulation in LC or NRM when tested on the activity of a multireceptive neurone in the spinal cord produced similar inhibitory actions. The present study aimed to define the pathways that mediate this descending inhibitory action in the spinal cord by pharmacological means and by making surgical lesions in the spinal cord or NRM. Attempts to differentiate pathways pharmacologically did not succeed since the i.v. administration of the 5-HT antagonists, methysergide and cinnanserin failed to antagonise descending inhibition evoked from either NRM or LC. Lesions involving a part or whole of the ipsilateral ventral quadrant reduced the inhibition produced from LC to a greater extent than that from NRM in 24 multireceptive neurones. In seven of these neurones stimulation in LC was without any effect after the lesion. In 23 multireceptive neurones recorded after making lesions that spared the ipsilateral ventral quadrant the effects of LC stimulation were unchanged. NRM effectiveness was reduced by an ipsilateral dorsolateral funiculus (DLF) lesion but required a bilateral DLF lesion for an almost complete abolition. Similar results were obtained when the effect of the various lesions were studied on the dorsal root potentials (DRPs) generated from LC or NRM. Lesions in the midline raphe complex, that included NRM, did not block the inhibitory action of LC stimulation. The inhibition produced from both these nuclei was additive whereas excitation was not. We conclude that LC actions in the spinal cord are mediated primarily through a pathway in the ipsilateral ventral quadrant whereas those from NRM are mediated through bilateral projections in DLF. Furthermore, although NRM plays no part in mediating LC actions and separate and independent pathways mediate their spinal action yet these apparently independent pathways have plenty of scope for interaction in the dorsal horn of the spinal cord itself.     

Spinal pathways mediating tonic or stimulation-produced descending inhibition from the periaqueductal gray or nucleus raphe magnus are separate in the cat

1. The spinal pathways for tonic and stimulation-produced descending inhibition of spinal nociceptive neurons were investigated in anesthetized paralyzed cats. Reversible circumscribed blocks were produced at various depths in the lateral funiculi (LF) at L1-L2 using the microinjection of the local anesthetic lidocaine. The total amount of tonic descending inhibition in the absence of LF blocks was evaluated by monitoring the spinal neuronal activity during reversible spinalization by cold block and compared with the activity of the same neuron during LF blocks. Stimulation-induced descending inhibition of neuronal responses to noxious skin heating was produced by bipolar focal electrical stimulation in the periaqueductal gray (PAG) or nucleus raphe magnus (NRM) and compared with the inhibition of the same neurons during LF blocks. The relative significance of ipsi- and contralateral pathways in the dorsal, medial, or ventral aspects of the lateral funiculi for these types of descending inhibition are quantitatively described. 2. All 35 lumbar spinal dorsal horn neurons studied responded to noxious and innocuous mechanical and noxious thermal stimuli applied within the receptive fields on the glabrous skin of the hindlimb. Responses to noxious skin stimuli (50 degrees C, 10 s at 3-min intervals) were constant over time and served as a parameter to evaluate tonic and stimulation-produced descending inhibition. All neurons also responded to electrical stimulation of hindlimb cutaneous nerves supramaximal for the activation of A-beta-, delta-, and C-fibers. Neurons were located in laminae I-VI of the dorsal horn at L5-L7 levels. LF blocks were produced by the microinjection of 1 microliter lidocaine at each of one to six sites in the ipsilateral and/or contralateral LF 500, 1,500, and/or 2,500 microns below cord surface. 3. LF blocks ipsilateral to the recording sites in the cord significantly reduced tonic inhibition, with blocks in the dorsal part of the LF [i.e., the dorsolateral funiculus (DLF)] being equally effective to complete LF blocks. Stimulation-produced inhibition from PAG or NRM was, however, not significantly affected by ipsilateral LF blocks. 4. Contralateral LF blocks significantly reduced stimulation-produced descending inhibition and failed to affect tonic descending inhibition. Ventral LF blocks attenuated inhibition from the PAG but not from NRM, whereas DLF blocks were more effective on inhibition from the NRM. 5. Bilateral LF blocks significantly reduced tonic as well as stimulation-produced descending inhibition.(ABSTRACT TRUNCATED AT 400 WORDS)     

Quantitative characterization of ceruleospinal inhibition of nociceptive transmission in the rat

A total of 51 dorsal horn units responsive to heat were isolated and their receptive fields characterized (i.e., response properties and adequate stimuli determined) in pentobarbital-anesthetized, paralyzed rats. In 39 of the 51 units, the descending inhibition of heat-evoked activity produced by focal electrical stimulation in the locus ceruleus/subceruleus (LC/SC) was examined. All units studied responded to mechanical stimulation, to electrical stimulation of the ipsilateral tibial nerve at intensities supramaximal to activate A-alpha, delta- and C-fibers, and to noxious heating (50 degrees C) of the footpad. The cutaneous receptive fields of all units were confined to the glabrous skin of the toes and footpad. All neurons examined were located in the dorsal horn of the spinal cord in laminae I-VI. Tracking experiments established that inhibition of heat-evoked dorsal horn unit activity could be reliably produced by focal electrical stimulation in both the contralateral and ipsilateral LC/SC. The inhibition produced by electrical stimulation in the LC/SC was intensity-, pulse duration-, and frequency-dependent. In six experiments, the efficacy of LC/SC stimulation-produced inhibition of heat-evoked activity was compared using two pulse durations (100 and 400 microseconds); greater inhibition of heat-evoked activity was produced at lower intensities of stimulation at the 400-microseconds pulse duration. In 10 experiments, the frequency of stimulation was varied (25-200 Hz); stimulation at a frequency of 100 Hz resulted in maximal inhibition of heat-evoked activity for stimulation sites both inside (n = 7) and outside (n = 3) the LC/SC. Inhibition of heat-evoked dorsal horn unit activity could be reliably produced by focal electrical stimulation in sites inside the LC/SC (n = 18). Significant descending inhibition of noxious heat-evoked spinal neuronal activity could also be produced by stimulation in pontine sites located outside the LC/SC, however, not as reliably. Systematic electrode tracks were made through the pons, using a searching stimulus of 100 microA, to locate sites medial, lateral, and ventral to the LC/SC from which significant descending inhibition could be produced. Stimulation in 156 sites outside the LC/SC at 100 microA produced inhibition of heat-evoked spinal unit activity to 50% of control or less in only 37 sites. Descending inhibition was characterized quantitatively from 14 of these 37 sites; the mean intensities of stimulation to inhibit heat-evoked activity to 50% of control were experimentally determined, and the mean thresholds of stimulation for inhibition and the mean recruitment indices were calculated.(ABSTRACT TRUNCATED AT 400 WORDS)     

Mechanisms mediating the brain stem control of somatosensory transmission in the dorsal horn of the cat's spinal cord: an intracellular analysis

Summary The effect of brainstem stimulation was studied on neurones recorded intracellularly in the superficial and deeper laminae of the lumbosacral dorsal horn of the spinal cord in anaesthetised cats. Stimulation in the nucleus locus coeruleus (LC) produced a hyperpolarisation in 4/13 multireceptive neurones and produced a biphasic action consisting of a hyperpolarisation which was followed by a depolarisation in 3/13 neurones. These actions were produced irrespective of whether the multireceptive neurone was located in the superficial or deeper laminae of the dorsal horn. Stimulation failed to produce postsynaptic potentials in the remaining 6/13 multireceptive neurones. The amplitude of hyperpolarisation was increased by the passage of depolarising pulses through the recording microelectrode and decreased by hyperpolarising pulses. Stimulation in other brainstem areas such as, the lateral (FTL), paralemniscal (FTP) and central (FTC) divisions of the tegmental field and the nuclei raphe magnus (NRM) and reticularis magnocellularis (RMc) also hyperpolarised neurones in the dorsal horn. The polarity of hyperpolarisation evoked from some brainstem areas (FTP, FTC, RMc) could be reversed to depolarisation by the passive diffusion of ions from the recording microelectrode containing 3M-KCl. Brainstem (LC, NRM, FTP, FTL) stimulation generated long lasting (700 ms) hyperpolarisation on 4/4 selectively nocireceptive neurones of lamina I. There was, however, no effect on the activity of 5/5 neurones recorded in laminae I/II which in addition to receiving excitatory cutaneous inputs were inhibited by heat stimuli. Stimulation in LC also produced dorsal root potentials (DRPs) and reduced the amplitude of simultaneously recorded excitatory postsynaptic potentials (EPSPs) generated by the activation of primary afferent fibres in 3 multireceptive neurones. It is concluded that inhibition of nociceptive transmission in the spinal cord from LC and other brainstem areas may involve both pre- and postsynaptic mechanisms.     

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)     

Substance P release and neurokinin 1 receptor activation in the rat spinal cord increase with the firing frequency of C-fibers

Both the firing frequency of primary afferents and neurokinin 1 receptor (NK1R) internalization in dorsal horn neurons increase with the intensity of noxious stimulus. Accordingly, we studied how the pattern of firing of primary afferent influences NK1R internalization. In rat spinal cord slices, electrical stimulation of the dorsal root evoked NK1R internalization in lamina I neurons by inducing substance P release from primary afferents. The stimulation frequency had pronounced effects on NK1R internalization, which increased up to 100 Hz and then diminished abruptly at 200 Hz. Peptidase inhibitors increased NK1R internalization at frequencies below 30 Hz, indicating that peptidases limit the access of substance P to the receptor at moderate firing rates. NK1R internalization increased with number of pulses at all frequencies, but maximal internalization was substantially lower at 1–10 Hz than at 30 Hz. Pulses organized into bursts produced the same NK1R internalization as sustained 30 Hz stimulation. To determine whether substance P release induced at high stimulation frequencies was from C-fibers, we recorded compound action potentials in the sciatic nerve of anesthetized rats. We observed substantial NK1R internalization when stimulating at intensities evoking a C-elevation, but not at intensities evoking only an Aδ-elevation. Each pulse in trains at frequencies up to 100 Hz evoked a C-elevation, demonstrating that C-fibers can follow these high frequencies. C-elevation amplitudes declined progressively with increasing stimulation frequency, which was likely caused by a combination of factors including temporal dispersion. In conclusion, the instantaneous firing frequency in C-fibers determines the amount of substance P released by noxious stimuli.     

Changes in intracellular free Ca ion concentration evoked by electrical activity in cat spinal neurons in situ

In cats under allobarbitone anaesthesia, Ca2+-sensitive microelectrodes were inserted into the lumbosacral spinal neurons to measure intracellular free Ca2+ concentration [Ca]i. In 72 resting motoneurons, the global mean [Ca]i was 7.9 microM (SD +/- 25.9). In the 36 "best" cells (with resting and action potentials better than 60 mV), mean [Ca]i was 1.6 microM (SD +/- 1.64). Activation of motoneurons by antidromic or direct stimulation evoked mean increases in [Ca]i of about 90 nM when stimulating for 30 s at 10 Hz, and 170 nM at 20 Hz. The mean time to half-recovery was 23 s (SD +/- 14.5). Orthodromic stimulation consistently produced smaller increases in [Ca]i. Measurements in motor axons showed a comparable resting level of [Ca]i, but only minimal changes during stimulation, even at 100 Hz. Sensory axons (also recorded within the spinal cord) similarly failed to show any increase in [Ca]i during high frequency stimulation. In some interneurons, however, particularly large and rapid increases in [Ca]i could be evoked by dorsal root stimulation at 1-5 Hz. Unresponsive cells (presumably neuroglia), with a typically high and stable resting potential, had a variable [Ca]i giving a mean of 32 microM (SD +/- 63.0). A tentative theoretical analysis of the magnitude and time course of delta [Ca]i evoked in motoneurons by tetanic stimulation is consistent with remarkably slow apparent diffusion of intracellular Ca2+ (1/250 of rate of diffusion in water), such as might be expected in the presence of very efficient mechanisms of Ca2+ sequestration.     

Inhibition of the responses of cat dorsal horn neurons to noxious skin heating by stimulation in medial or lateral medullary reticular formation

Summary Responses of single lumbar dorsal horn units to noxious radiant heating (50° C, 10 s) of glabrous footpad skin were recorded in cats anesthetized with sodium pentobarbital and 70% nitrous oxide. The heat-evoked responses of 37/40 units were reduced during electrical stimulation (100 ms trains, 100 Hz, 3/s, 25–600 A) in the medullary nucleus raphe magnus (NRM) and/or in laterally adjacent regions of the medullary reticular formation (MRF). Inhibition was elicited by stimulation in widespread areas of the medulla, but with greatest efficacy at ventrolateral sites. The magnitude of inhibition increased with graded increases in medullary stimulation intensity. Mean current intensities at threshold for inhibition or to produce 50% inhibition were higher for NRM than for MRF sites. Units' responses to graded noxious heat stimuli increased linearly from threshold (42–43° C) to 52° C. During NRM (5 units) or ipsilateral MRF stimulation (7 units), responses were inhibited such that the mean temperature-response functions were shifted toward higher temperatures with increased thresholds (1.5° and 1° C, respectively) and reduced slopes (to 60% of control). Contralateral MRF stimulation had a similar effect in 4 units. Inhibitory effects of NRM and MRF stimulation were reduced (by >25%) or abolished in 4/6 and 5/12 units, respectively, following systemic administration of the serotonin antagonist methysergide. Inhibitory effects from NRM, ipsi- and contralateral MRF were reduced or abolished in 2/9, 4/8 and 6/9 cases, respectively, following systemic administration of the noradrenergic antagonist phentolamine. These results confirm and extend previous studies of medullospinal inhibition and the role of monoamines, and are discussed in terms of analgesic mechanisms.