Modulation of striatal dopaminergic function by local injection of 5'-N-ethylcarboxamide adenosine.

Rats rotated to the left when 5'-N-ethylcarboxamide adenosine (NECA) was injected into the left caudate nucleus and apomorphine was administered subcutaneously. The combination of NECA and apomorphine was more potent than L-(phenylisopropyl)adenosine and apomorphine in eliciting rotation, suggesting the involvement of adenosine receptors of the Ra type. The response was reduced when 2',5'-dideoxyadenosine was injected along with NECA into the caudate nucleus or when theorphylline was given intraperitoneally. Higher doses of apomorphine elicited a self-mutilatory response after the injection of NECA into the caudate nucleus. These results suggest that adenosine may be involved in the modulation of dopaminergic function in the striatum.

Neurotensin activation of the NTR1 on spinally-projecting serotonergic neurons in the rostral ventromedial medulla is antinociceptive.

Microinjection of neurotensin (NT) in the rostral ventromedial medulla (RVM) produces dose-dependent antinociception. The NTR1 (Neurotensin Receptor Subtype 1) may mediate part of this response, however definitive evidence is lacking, and the spinal mediators of NTR1-induced antinociception are unknown. In the present study, we used immunohistochemical techniques to show that the NTR1, but not the NTR2 is expressed by spinally projecting serotonergic neurons of the RVM. We also show that microinjection of NT or the NTR1-selective agonist PD149163 in the RVM both produce dose-dependent antinociception in the tail-flick test that is blocked by the NTR1-selective antagonist SR48692. The antinociception produced by NT or PD149163 is also blocked by intrathecal administration of the non-selective serotonergic receptor antagonist methysergide. The results of these experiments provide anatomical and behavioral evidence that activation of NTR1-expressing spinally projecting neurons in the RVM produces antinociception through release of serotonin in the spinal dorsal horn. These results support the conclusion that the NTR1 plays an important role in the central modulation of nociception.

Projections of neurons in the periaqueductal gray to pontine and medullary catecholamine cell groups involved in the modulation of nociception.

Stimulation of neurons in the periaqueductal gray (PAG) produces antinociception that is mediated in part by noradrenergic neurons that innervate the spinal cord dorsal horn. Because norepinephrine-containing neurons are not found in the PAG, noncatecholamine neurons in the PAG must project to, and activate, spinally projecting catecholamine neurons located in the pons or medulla. The present studies determined the projections of neurons in the ventrolateral PAG to the A5, A6 (locus coeruleus), and A7 catecholamine cell groups that are known to contain spinally projecting noradrenergic neurons. The anterograde tracer biotinylated dextran amine (BDA) was injected into the ventrolateral PAG, and labeled axon terminal profiles were identified near noradrenergic neurons that were visualized by processing tissue sections for tyrosine hydroxylase immunoreactivity. Highly varicose, anterogradely labeled terminal profiles were found apposed to the dendrites and somata of tyrosine-hydroxylase-immunoreactive neurons and non-tyrosine-hydroxylase-immunoreactive neurons in the dorsolateral and ventrolateral pontine tegmentum. These axon terminal profiles were more dense on the side ipsilateral to the BDA deposit, and both A7 and locus coeruleus neurons received a more dense innervation than did the A5 neurons. Although definitive evidence for a direct pathway from PAG neurons to spinally projecting A7 neurons requires ultrastructural studies, the results of the present studies provide presumptive evidence for direct projections from neurons in the PAG to noradrenergic A7 neurons that innervate the spinal cord dorsal horn and modulate pain perception. If neurons in the ventrolateral PAG do form synapses with noradrenergic A7 neurons, these spinally projecting catecholamine neurons may mediate part of the analgesic effect produced by systemic administration of morphine. In contrast, the projections of PAG neurons to the A5 cell group and the locus coeruleus may mediate the cardiovascular and motor effects produced by stimulation of sites in the ventrolateral PAG.

Periaqueductal gray neurons monosynaptically innervate extranuclear noradrenergic dendrites in the rat pericoerulear region.

Previous reports using light microscopy have provided anatomical evidence that neurons in the ventrolateral periaqueductal gray (PAG) innervate the medial pericoerulear dendrites of noradrenergic neurons in the nucleus locus coeruleus (LC). The present study used anterograde tracing and electron microscopic analysis to provide more definitive evidence that neurons in the ventrolateral PAG form synapses with the somata or dendrites of noradrenergic LC neurons. Deposits of either biotinylated dextran amine or Phaseolus vulgaris leucoagglutinin into the rat ventrolateral PAG labeled a moderate to high number of axons in the region of the medial pericoerulear region and Barrington's nucleus, but a relatively low number were labeled in the nuclear core of the LC. Ultrastructural analysis of anterogradely labeled terminals at the levels of the rostral (n = 233) and caudal (n = 272) subdivisions of the LC indicated that approximately 20% of these form synapses with tyrosine hydroxylase-immunoreactive dendrites; most of these were located in the medial pericoerulear region. In rostral sections, about 12% of these were symmetric synapses, 9% were asymmetric synapses, and 79% were membrane appositions without clear synaptic specializations. In caudal sections, about 30% were symmetric synapses, 11% were asymmetric synapses, and 59% were appositions. In both rostral and caudal sections, 60% of the anterogradely labeled terminals formed synapses with noncatecholamine dendrites, and 20% formed axoaxonic synapses. These results provide direct evidence for monosynaptic projections from neurons in the ventrolateral PAG to the extranuclear dendrites of noradrenergic LC neurons. This monosynaptic pathway may mediate in part the analgesia, reduced responsiveness to external stimuli, and decreased excitability of somatic motoneurons produced by stimulation of neurons in the ventrolateral PAG.

Hypoalgesia induced by the local injection of phentolamine in the nucleus raphe magnus: blockade by depletion of spinal cord monoamines.

Blockade of the noradrenergic input to the nucleus raphe magnus (NRM) by the injection of alpha-adrenergic antagonists produces hypoalgesia. Previous studies show that this hypoalgesia is blocked by the intrathecal injection of either phentolamine or methysergide. The present study demonstrates that depletion of spinal cord serotonin, norepinephrine, or both also blocks this hypoalgesia. Together these studies suggest that the hypoalgesia produced by microinjection of noradrenergic antagonists in the NRM is mediated by the activation of both raphe-spinal serotonergic neurons and bulbospinal noradrenergic neurons.

Synergistic interactions between two alpha(2)-adrenoceptor agonists, dexmedetomidine and ST-91, in two substrains of Sprague-Dawley rats.

Several lines of evidence indicate that the antinociception produced by intrathecal administration of the alpha(2)-adrenoceptor agonists dexmedetomidine or ST-91 is mediated by different subtypes of the alpha(2)-adrenoceptor. We recently provided additional pharmacologic evidence for this idea, as well as for differences in the function of these receptors between Harlan and Sasco rats, two widely-used outbred substrains of Sprague-Dawley rat. The present study used isobolographic analysis to further characterize the receptors at which intrathecally administered ST-91 and dexmedetomidine act in these two substrains. The rationale for these studies derives from the assumption that if dexmedetomidine and ST-91 act as agonists at the same receptor then they should interact in an additive manner. However, if they interact in a supra-additive manner, then they must act at different subtypes of the alpha(2)-adrenoceptor. In the tail-flick test, the dose-effect relationship for a 1:3 mixture of dexmedetomidine and ST-91 was shifted significantly to the left of the theoretical dose-additive line in both Harlan and Sasco Sprague-Dawley rats. A similar finding was made in the hot-plate test despite the fact that the dose-response characteristics of the agonists were different in this test. Thus, in Harlan rats, in which ST-91 is a full agonist and dexmedetomidine is essentially inactive, the dose-effect relationship for the mixture of dexmedetomidine and ST-91 was shifted far to the left of the dose-additive line. Similarly, in Sasco rats, in which ST-91 is a partial agonist and dexmedetomidine is inactive, co-administration of the two agonists also shifted the dose-response relationship to the left of the dose-additive line. The consistent finding that these two alpha(2)-adrenoceptor agonists interact in a supra-additive manner provides strong evidence that dexmedetomidine and ST-91 produce antinociception by acting at different alpha(2)-adrenoceptor subtypes in the spinal cord. This conclusion is consistent with the earlier proposal that dexmedetomidine acts predominantly at alpha(2A)-adrenoceptors whereas ST-91 acts predominantly at non-alpha(2A)-adrenoceptors. Recent anatomical evidence indicates that these non-alpha(2A) adrenoceptors may be of the alpha(2C) type. The synergistic combination of an alpha(2A)- and an alpha(2C)-adrenoceptor agonist may provide a unique and highly effective drug combination for the treatment of pain without the sedation produced by an equianalgesic dose of a single alpha(2)-adrenoceptor agonist.

Antinociception induced by microinjection of substance P into the A7 catecholamine cell group in the rat.

Stimulation of neurons in the ventromedial medulla produces antinociception that is mediated in part by indirect activation of pontospinal noradrenergic neurons. Substance P-containing neurons located in the ventromedial medulla project to the A7 catecholamine cell group and may serve as an excitatory link between these two cell groups. Thus, the antinociception induced by stimulation of the neurons in ventromedial medulla may be mediated by substance P released from these projections which activates spinally projecting noradrenergic neurons in the A7 cell group. This hypothesis was tested by determining whether microinjection of various doses of substance P into the A7 cell group of the rat could induce antinociception. The results indicated that substance P induced dose-dependent antinociception that was more pronounced in the hindlimb ipsilateral to the microinjections. This observation is consistent with anatomical observations that noradrenergic A7 neurons project predominantly to the ipsilateral spinal cord dorsal horn. Moreover, the antinociceptive effects of substance P microinjection appear to be mediated at least in part by activation of spinally projecting noradrenergic neurons in the A7 cell group, because intrathecal injections of the alpha-2 noradrenergic antagonists yohimbine and idazoxan blocked these antinociceptive effects. The results of these experiments support the hypothesis that the antinociception induced by stimulation of neurons in the ventromedial medulla is mediated in part by activation of substance P-containing neurons that project to, and activate, spinally projecting noradrenergic neurons located in the A7 catecholamine cell group.

Bidirectional modulation of nociception by GABA neurons in the dorsolateral pontine tegmentum that tonically inhibit spinally projecting noradrenergic A7 neurons.

The A7 catecholamine cell group in the dorsolateral pontine tegmentum constitutes an important part of the descending pathways that modulate nociception. Evidence from immunocytochemical studies demonstrate that noradrenergic A7 neurons are densely innervated by GABA terminals arising from GABA neurons that are located in the dorsolateral pontine tegmentum medial to the A7 cell group. GABA(A) receptors are also located on the somata and dendrites of noradrenergic A7 neurons. These findings suggest that noradrenergic neurons in the A7 cell group may be under tonic inhibitory control by GABA neurons. To test this hypothesis, the GABA(A) antagonist bicuculline methiodide in doses of 0.2 or 1.0nmol was microinjected into sites located dorsal to the A7 cell group and the resulting effects on tail flick and nociceptive foot withdrawal responses were measured. Both doses of bicuculline produced significant increases in tail flick latencies and small, but significant, increases in foot withdrawal latencies. Intrathecal injection of the alpha(2)-adrenoceptor antagonist yohimbine, in a dose of 76.7nmol (30microg), attenuated the antinociceptive effect of bicuculline on both the tail and the feet. In contrast, the alpha(1)-adrenoceptor antagonist WB4101, in a nearly equimolar dose of 78.6nmol (30microg), increased the antinociceptive effect of bicuculline on both the tail and the feet. Intrathecal injection of the antagonists alone did not consistently alter nociceptive responses of either the feet or the tail. These findings suggest that noradrenergic neurons in the A7 cell group are tonically inhibited by local GABA neurons. Furthermore, these findings suggest that inhibition of GABA(A) receptors located on spinally-projecting A7 noradrenergic neurons disinhibits, or activates, two populations of A7 neurons that have opposing effects on nociception. One of these populations facilitates nociception by an action mediated by alpha(1)-adrenoceptors in the spinal cord dorsal horn and the other population inhibits nociception by an action mediated by alpha(2)-adrenoceptors.

Enkephalin neurons that project to the A7 catecholamine cell group are located in nuclei that modulate nociception: ventromedial medulla.

The location of methionine enkephalin neurons in the medulla oblongata that project to the dorsolateral pontine tegmentum was investigated using anterograde and retrograde tract tracing combined with immunocytochemical neurotransmitter identification. The results of these experiments demonstrate that enkephalinergic neurons from areas known to modulate nociception project to the region of the A7 catecholamine cell group in the dorsolateral pontine tegmentum. The medullary nuclei that contain these enkephalinergic neurons include the nucleus raphe magnus and the nucleus reticularis gigantocellularis pars alpha in the ventromedial medulla. While some of these enkephalinergic axons appose the somata and dendrites of A7 neurons, the majority of these axons appear to contact non-catecholamine neurons in the dorsolateral pontine tegmentum. Unidentified neurons located in the nucleus raphe magnus, the nucleus reticularis gigantocellularis pars alpha, and the nucleus reticularis gigantocellularis also project to the A7 area. Many of the neurons in the nucleus reticularis gigantocellularis pars alpha appear to contact both noradrenergic A7 neurons and non-catecholamine neurons in the dorsolateral pontine tegmentum, whereas most of those in the nucleus raphe magnus appear to contact non-catecholamine neurons. The anatomical findings described in this report and the results of preliminary behavioral studies provide evidence to support a model in which activation of the enkephalin-containing neurons in the ventromedial medulla facilitates nociception, while the non-enkephalin neurons mediate part of the antinociception produced by stimulating sites in the ventromedial medulla.

Ultrastructural evidence that substance P neurons form synapses with noradrenergic neurons in the A7 catecholamine cell group that modulate nociception.

Potent antinociception can be produced by electrical stimulation of spinally projecting noradrenergic neurons in the A7 catecholamine cell group and this effect is blocked by intrathecal injection of alpha2-adrenoceptor antagonists. Microinjection of substance P near A7 neurons also produces antinociception that is blocked by intrathecal injection of alpha2-adrenoceptor antagonists. These observations suggest that substance P produces antinociception by activating noradrenergic A7 neurons. However, it is not known whether this effect of substance P is produced by a direct or an indirect action on A7 neurons. Although light microscopic studies have demonstrated the existence of both substance P-containing axon terminals and neurokinin-1 receptors in the region of the A7 cell group, it is not known whether substance P terminals form synapses with noradrenergic A7 neurons. These experiments used double-labeling immunocytochemical methods and electron microscopic analysis to determine whether substance P-containing axons form synapses with noradrenergic neurons in the A7 cell group. Pre-embedding immunocytochemistry, combined with light and electron microscopic analysis, was used to provide ultrastructural evidence for synaptic connections between substance P-immunoreactive terminals labeled with immunoperoxidase and tyrosine hydroxylase-immunoreactive A7 neurons labeled with silver-enhanced immunogold. Tyrosine hydroxylase labeling was found in perikarya and dendrites in the A7 region, and substance P labeling was found in axons and synaptic terminals. Substance P-labeled terminals formed asymmetric synapses with tyrosine hydroxylase-labeled dendrites, but only a few of these were present on tyrosine hydroxylase-labeled somata. Substance P-labeled terminals also formed asymmetric synapses with unlabeled dendrites, and many unlabeled terminals formed both symmetric and asymmetric synapses with tyrosine hydroxylase-labeled dendrites. These results demonstrate that substance P neurons form a significant number of synapses with the dendrites of noradrenergic A7 neurons and support the conclusion that microinjection of substance P in the A7 cell group produces antinociception by direct activation of spinally projecting noradrenergic neurons.

Microinjection of morphine in the A7 catecholamine cell group produces opposing effects on nociception that are mediated by alpha1- and alpha2-adrenoceptors.

Stimulation of neurons in the ventromedial medulla produces antinociception in part by inhibiting nociceptive dorsal horn neurons. This antinociceptive effect is mediated in part by spinally projecting noradrenergic neurons located in the A7 catecholamine cell group. Methionine-enkephalin-immunoreactive neurons in the ventromedial medulla project to an area that includes the A7 cell group, and these enkephalin neurons may mediate part of the antinociception produced by stimulation of sites in the ventromedial medulla. This possibility was tested by determining the effects of microinjecting morphine near the A7 cell group on nociceptive foot and tail responses. Microinjection of a 3.75 nmol dose of morphine in the A7 region did not alter nociceptive responses, but a higher dose of 7.5 nmol facilitated these responses. In contrast, a higher dose of 15 nmol of morphine did not alter nociceptive responses. Selective alpha-adrenoceptor antagonists were injected intrathecally to determine whether the hyperalgesia produced by morphine is mediated by spinally projecting noradrenergic A7 neurons. Intrathecal injection of the alpha2-adrenoceptor antagonist yohimbine did not alter the hyperalgesic effect produced by the 7.5 nmol dose of morphine, but the alpha1 antagonist WB4101 reversed the hyperalgesia and produced antinociception that lasted for nearly 30 min. Although the 15 nmol dose of morphine did not alter nociceptive responses, intrathecal injection of yohimbine after the microinjection of morphine produced a significant facilitation of nociception, and intrathecal injection of WB401 produced a significant antinociceptive effect. Intrathecal injection of the antagonists alone did not consistently alter nociception. These findings, and those of published reports, suggest that morphine indirectly activates two populations of spinally projecting A7 noradrenergic neurons that have opposing effects on nociception. One of these populations facilitates nociception by an action mediated by alpha1-adrenoceptors in the spinal cord dorsal horn and the other population inhibits nociception by an action mediated by alpha2-adrenoceptors. These results suggest that some of the methionine-enkephalin neurons located in the ventromedial medulla that project to the A7 cell group can exert bidirectional control of nociceptive responses.

Ultrastructural analysis of ventrolateral periaqueductal gray projections to the A7 catecholamine cell group.

Stimulation of neurons in the ventrolateral periaqueductal gray produces antinociception that is mediated in part by pontine noradrenergic neurons. Previous light microscopic analysis provided suggestive evidence for a direct projection from neurons in the ventrolateral periaqueductal gray to noradrenergic neurons in the A7 cell group that innervate the spinal cord dorsal horn. Therefore, the present ultrastructural study used anterograde tracing combined with tyrosine hydroxylase immunoreactivity to provide definitive evidence that neurons in the ventrolateral periaqueductal gray form synapses with the somata and dendrites of noradrenergic neurons of the A7 cell group. Injections of the anterograde tracers biotinylated dextran amine or Phaseolus vulgaris leucoagglutinin into the ventrolateral periaqueductal gray of Sasco Sprague-Dawley rats yielded a dense innervation in the region of the lateral pons containing the A7 cell group. Electron microscopic analysis of anterogradely labeled terminals (n=401) in the region of the A7 cell group indicated that approximately 10% of these formed plasmalemmal appositions to tyrosine hydroxylase-immunoreactive dendrites with no intervening astrocytic processes. About 23% of these were asymmetric synapses, 10% were symmetric synapses, and 67% did not exhibit clearly differentiated synaptic specializations. The majority of anterogradely labeled terminals (60%) formed plasmalemmal appositions with dendrites and somata that lacked detectable tyrosine hydroxylase immunoreactivity. About 35% of these were symmetric synapses, 9% were asymmetric synapses and 56% did not form synaptic specializations. Approximately 30% of all anterogradely labeled terminals displayed features characteristic of axo-axonic synapses.The present results provide direct ultrastructural evidence to support the hypothesis that the analgesia produced by stimulation of neurons in the ventrolateral periaqueductal gray is mediated, in part, by activation of spinally projecting noradrenergic neurons in the A7 catecholamine cell group.

The projections of locus coeruleus neurons to the spinal cord.

Spinally projecting noradrenergic neurons located in the locus coeruleus/subcoeruleus (LC/SC) are a major source of the noradrenergic innervation of the spinal cord. However, the specific terminations of these neurons have not been clearly defined. The purpose of this chapter is to describe the results of experiments that used the anterograde tracer Phaseolus vulgaris leucoagglutinin in combination with dopamine-beta-hydroxylase immunocytochemistry to more precisely determine the spinal cord terminations of neurons located in the LC/SC. The results of these experiments indicate that the axons of LC neurons are located primarily in the ipsilateral ventral funiculus and terminate most heavily in the medial part of laminae VII and VIII, the motoneuron pool of lamina IX, and lamina X. LC neurons provide a moderately dense innervation of the ventral part of the dorsal horn, but only a very sparse innervation of the superficial dorsal horn. The SC projects ipsilaterally in the ventrolateral funiculus and terminates diffusely in the intermediate and ventral laminae of the spinal cord. Finally, the results of preliminary experiments indicate that different rat substrains may have LC neurons that exhibit qualitatively different termination patterns in the spinal cord. More specifically, LC neurons in some rat substrains innervate the dorsal horn, while those in other substrains primarily innervate the ventral horn and intermediate zone.

A low-priced computer-assisted densitometer and analysis software for quantitative autoradiography.

A computer-assisted densitometer which consists of a darkroom enlarger, a black-and-white exposure meter, an amplifier, an analog-to-digital converter, and a microcomputer with a monitor and a graphic printer can be utilized for the quantitative analysis of autoradiograms. QUANTAR, a computer program written in BASIC, records the density measurements and stores the data in a file that can be easily retrieved by commercially available spreadsheet software. A spreadsheet template was designed to convert the digital readings into concentration of ligand in tissue. A variety of spreadsheet templates can be created to analyze data for a standard curve, a Scatchard plot, or a binding competition curve. This system, which is easy to assemble, may be useful to the frugal investigator with a modest equipment budget.

The projections of noradrenergic neurons in the A5 catecholamine cell group to the spinal cord in the rat: anatomical evidence that A5 neurons modulate nociception.

Brainstem noradrenergic neurons located in the A5, A6, and A7 catecholamine cell groups provide the entire noradrenergic innervation of the spinal cord. We have previously demonstrated that noradrenergic neurons in the A6 and A7 cell groups innervate the ventral and dorsal horns, respectively. Since the specific spinal cord terminations of the A5 cell group have not been clearly delineated, the present experiments were designed to trace the projections from this noradrenergic cell group to the spinal cord, using the anterograde tracer Phaseolus vulgaris-leucoagglutinin (PHA-L) in combination with dopamine-beta-hydroxylase immunocytochemistry. The results of these experiments indicate that A5 noradrenergic neurons project ipsilaterally through the dorsolateral funiculus in cervical, thoracic, and lumbar segments. In cervical segments, these axons terminate primarily in the ipsilateral deep dorsal horn (laminae IV-VI) and the intermediate zone (lamina VII). In thoracic segments, the intermediolateral cell column is heavily innervated by A5 axons. In lumbar segments, the concentration of A5 axons is more diffuse and more widely distributed than that in cervical and thoracic segments. Although there is a higher density of axons in the deep dorsal horn and the intermediate zone, there are also scattered axons in the dorsal and ventral horns. The innervation of these regions of the spinal cord by A5 neurons provides anatomical support for the conclusion that these noradrenergic neurons are involved in modulating cardiovascular reflexes and nociceptive transmission in the spinal cord.

The projection of noradrenergic neurons in the A7 catecholamine cell group to the spinal cord in the rat demonstrated by anterograde tracing combined with immunocytochemistry.

Noradrenergic neurons located in the A5, A7 and locus coeruleus/subcoeruleus (LC/SC) catecholamine cell groups innervate all levels of the spinal cord. However, the specific spinal cord terminations of these neurons have not been clearly delineated. This study determined the spinal cord terminations of the A7 catecholamine cell group using the anterograde tracer Phaseolus vulgaris leucoagglutinin (PHA-L) in combination with dopamine-beta-hydroxylase (DBH) immunocytochemistry. In addition, the spinal cord projections of A7 neurons were examined by measuring the reduction in the density of DBH-immunoreactive axons in specific regions of the spinal cord after a unilateral electrolytic lesion of the A7 cell group. The results of these experiments indicate that noradrenergic neurons in the A7 cell group project primarily in the ipsilateral dorsolateral funiculus and terminate most heavily in the dorsal horn (laminae I-IV).

The projection of locus coeruleus neurons to the spinal cord in the rat determined by anterograde tracing combined with immunocytochemistry.

Pontospinal noradrenergic neurons located in the A5, A7 and locus coeruleus/subcoeruleus (LC/SC) nuclei are the major source of the noradrenergic innervation of the spinal cord. However, the specific terminations of spinally-projecting noradrenergic neurons located in these nuclei have not been clearly defined. The purpose of the experiments described in this report was to more precisely define the spinal terminations of neurons located in the LC/SC using the anterograde tracer phaseolus vulgaris-leucoagglutinin in combination with dopamine-beta-hydroxylase (D beta H) immunocytochemistry. In addition, the spinal cord regions in which LC/SC neurons terminate was assessed by measuring the reduction in the density of D beta H-immunoreactive axon terminals in specific spinal cord regions after a unilateral electrolytic lesion that included LC/SC neurons. The results of these experiments indicate that the axons of LC neurons are located primarily in the ipsilateral ventral funiculus and terminate most heavily in the medial part of laminae VII and VIII, the motoneuron pool of lamina IX, and lamina X. LC neurons provide a moderately dense innervation of the ventral part of the dorsal horn, but only a very sparse innervation of the superficial dorsal horn. The SC projects ipsilaterally in the ventrolateral funiculus and terminates diffusely in the intermediate and ventral laminae of the spinal cord.

Release of endogenous monoamines into spinal cord superfusates following the microinjection of phentolamine into the nucleus raphe magnus.

Previous studies have suggested that raphe-spinal neurons located in the nucleus raphe magnus (NRM) are tonically inhibited by noradrenergic neurons. Furthermore, blockade of the inhibitory noradrenergic input to the NRM induces antinociception which appears to be mediated by the release of both serotonin and norepinephrine in the spinal cord. The present experiments were designed to directly measure the release of endogenous serotonin and norepinephrine into spinal cord superfusates before and after the microinjection of the alpha-adrenergic antagonist phentolamine into the NRM. High-performance liquid chromatography with electrochemical detection was used to quantitate the monoamines. The injection of phentolamine into the NRM induced a significant increase in the amount of both norepinephrine and serotonin released in the spinal cord. This enhanced release was not observed following either the injection of phentolamine into sites outside the NRM or the injection of saline vehicle into the NRM. These results support the proposal that the antinociception induced by the blockade of the inhibitory noradrenergic input to the NRM is mediated by the activation of spinally-projecting serotonergic and noradrenergic neurons.

Antinociception induced by local injections of carbachol into the nucleus raphe magnus in rats: alteration by intrathecal injection of monoaminergic antagonists.

Electrical stimulation of neurons located in the nucleus raphe magnus (NRM) produces antinociception which appears to result from inhibition of spinothalamic tract neurons located in the spinal cord dorsal horn. Iontophoretic application of acetylcholine also activates NRM neurons and microinjection of cholinergic agonists such as carbachol into the NRM produces a profound, long-lasting antinociception. Since the antinociception induced by electrical stimulation of NRM neurons is mediated, at least in part, by bulbospinal serotonergic and noradrenergic neurons, the role of these monoaminergic neurons in mediating the antinociception induced by microinjecting carbachol in the NRM was examined in the present study. To this end, various antagonists of serotonin and norepinephrine were injected into the spinal cord subarachnoid space following the induction of antinociception by the local injection of carbachol into the NRM. The serotonergic antagonist methysergide had no effect on carbachol-induced antinociception. However, the alpha 2-noradrenergic antagonist yohimbine attenuated, while the alpha 1-noradrenergic antagonists prazosin and WB4101 increased the effects of carbachol. The non-selective noradrenergic antagonist phentolamine also attenuated the effects of carbachol. These results lead to the suggestion that the antinociception induced by the local injection of carbachol into the NRM is mediated by selective activation of bulbospinal noradrenergic neurons. Furthermore, the antinociception resulting from the activation of these descending noradrenergic neurons appears to be mediated by alpha 2-noradrenergic receptors located in the spinal cord dorsal horn. Finally, the local injection of carbachol into the NRM also appears to activate another population of noradrenergic neurons which produces hyperalgesia mediated by alpha 1-noradrenergic receptors.

Alterations in nociception following lesions of the A5 catecholamine nucleus.

Neurons located in the nucleus raphe magnus (NRM), a region important in the control of nociception, appear to be tonically inhibited by noradrenergic (NA) neurons. Anatomical studies have suggested that the A5 catecholamine nucleus may be the primary source of noradrenergic neurons whose terminals are located in the NRM. The purpose of the present study was to examine the role of A5 neurons in the modulation of nociception. Bilateral electrolytic lesions of the A5 nuclei produced a marked and long lasting antinociception as assessed by both the tail-flick and hot-plate tests. Unilateral A5 lesions also produced a long-lasting elevation in hot-plate latency, but the elevation of tail-flick latency was smaller in magnitude and was only observed one day following the lesion. This finding is consistent with previous studies which have shown that blockade of the NA input to the NRM by the microinjection of NA antagonists also produces antinociception. These data indicate that neurons located in the A5 nucleus may be the origin of this NA projection to the NRM. The elevation in tail-flick latency observed following A5 lesions was significantly attenuated by the intrathecal injection of either the NA antagonist phentolamine or the serotonergic antagonist methysergide. However, the elevation in hot-plate latency was not significantly altered by these monoaminergic antagonists. Similarly, previous studies have shown that the elevation in tail-flick, but not hot-plate latency, produced by the microinjection of NA antagonists in the NRM is attenuated by the intrathecal injection of either phentolamine or methysergide.(ABSTRACT TRUNCATED AT 250 WORDS)