Nociceptive Projection Neurons Research Articles

Contralateral Afferent Input to Lumbar Lamina I Neurons as a Neural Substrate for Mirror-Image Pain.

Mirror-image pain arises from pathologic alterations in the nociceptive processing network that controls functional lateralization of the primary afferent input. Although a number of clinical syndromes related to dysfunction of the lumbar afferent system are associated with the mirror-image pain, its morphophysiological substrate and mechanism of induction remain poorly understood. Therefore, we used ex vivo spinal cord preparation of young rats of both sexes to study organization and processing of the contralateral afferent input to the neurons in the major spinal nociceptive projection area Lamina I. We show that decussating primary afferent branches reach contralateral Lamina I, where 27% of neurons, including projection neurons, receive monosynaptic and/or polysynaptic excitatory drive from the contralateral Aδ-fibers and C-fibers. All these neurons also received ipsilateral input, implying their involvement in the bilateral information processing. Our data further show that the contralateral Aδ-fiber and C-fiber input is under diverse forms of inhibitory control. Attenuation of the afferent-driven presynaptic inhibition and/or disinhibition of the dorsal horn network increased the contralateral excitatory drive to Lamina I neurons and its ability to evoke action potentials. Furthermore, the contralateral Aβδ-fibers presynaptically control ipsilateral C-fiber input to Lamina I neurons. Thus, these results show that some lumbar Lamina I neurons are wired to the contralateral afferent system whose input, under normal conditions, is subject to inhibitory control. A pathologic disinhibition of the decussating pathways can open a gate controlling contralateral information flow to the nociceptive projection neurons and, thus, contribute to induction of hypersensitivity and mirror-image pain.SIGNIFICANCE STATEMENT We show that contralateral Aδ-afferents and C-afferents supply lumbar Lamina I neurons. The contralateral input is under diverse forms of inhibitory control and itself controls the ipsilateral input. Disinhibition of decussating pathways increases nociceptive drive to Lamina I neurons and may cause induction of contralateral hypersensitivity and mirror-image pain.

Netrin1 and reelin signaling are required for the migration of anterolateral system neurons in the embryonic spinal cord.

Projection neurons of the spinal cord dorsal horn which transmit pain, itch, and temperature information to the brain comprise the anterolateral system (AS). A recent molecular and genetic study showed that many developing AS neurons express the transcription factor Phox2a and provided insights into the mechanisms of their ontogeny and wiring of nociceptive neuronal circuits. Here, we show that the loss of the axonal guidance and neuronal migration signal netrin1 results in impaired migration of mouse Phox2a+ AS neurons into the spinal lamina I. Furthermore, we show that in the absence of Dab1, an intracellular transducer of the neuronal migration signal reelin, the migration of spinal lamina V and lateral spinal nucleus Phox2a+ AS neurons is impaired, in line with deficits in nociception seen in mice with a loss of reelin signaling. Together, these results provide evidence that netrin1 and reelin control the development of spinal nociceptive projection neurons, suggesting a mechanistic explanation for studies that link sequence variations in human genes encoding these neurodevelopmental signals and abnormal pain sensation.

Slow brushing reduces heat pain in humans.

C-tactile (CT) afferents are unmyelinated low-threshold mechanoreceptors optimized for signalling affective, gentle touch. In three separate psychophysical experiments, we examined the contribution of CT afferents to pain modulation. In total, 44 healthy volunteers experienced heat pain and CT optimal (slow brushing) and CT sub-optimal (fast brushing or vibration) stimuli. Three different experimental paradigms were used: Concurrent application of heat pain and tactile (slow brushing or vibration) stimulation; Slow brushing, applied for variable duration and intervals, preceding heat pain; Slow versus fast brushing preceding heat pain. Slow brushing was effective in reducing pain, whereas fast brushing or vibration was not. The reduction in pain was significant not only when the CT optimal touch was applied simultaneously with the painful stimulus but also when the two stimuli were separated in time. For subsequent stimulation, the pain reduction was more pronounced for a shorter time interval between brushing and pain. Likewise, the effect was more robust when pain was preceded by a longer duration of brush stimulation. Strong CT-related pain reduction was associated with low anxiety and high calmness scores obtained by a state anxiety questionnaire. Slow brushing - optimal for CT activation - is effective in reducing pain from cutaneous heating. The precise mechanisms for the pain relief are as yet unknown but possible mechanisms include inhibition of nociceptive projection neurons at the level of the dorsal horn as well as analgesia through cortical mechanisms. Slow brushing stimuli - optimal for activation of C-tactile fibres - can reduce pain from cutaneous heating. No such effect was seen with fast brushing or vibration. These observations indicate the role of C-tactile fibres in pain modulation.

Mechanism of GABA involvement in post-traumatic trigeminal neuropathic pain: activation of neuronal circuitry composed of PKCγ interneurons and pERK1/2 expressing neurons.

GABA disinhibition within the spinal dorsal horn has been implicated in pain hypersensitivity on injury in different neuropathic models. However, GABA alteration has been explored in only one study on trigeminal neuropathic pain. The present study investigated the implication of GABA in trigeminal dynamic mechanical allodynia (DMA) obtained after chronic constriction of the infraorbital nerve (CCI-IoN), and explored the cellular and molecular mechanisms by which GABA dysfunction induced DMA. Our data demonstrated a significant decrease in labelling in two GABA cell markers, glutamate acid decarboxylase (GAD67), and parvalbumin, in the medullary dorsal horn (MDH) of allodynic rats in comparison to sham rats. Increasing GABA by intracisternal injections of vigabatrin (VGB), a blocker of the catabolic enzyme GABA transaminase, alleviated pain behaviour and restored normal GABA cell marker expression in allodynic MDH. Interestingly, intracisternal VGB administration also significantly decreased PKCγ staining, i.e., of its phosphorylated active form and the number of pERK1/2 positive cells within the MDH. These two markers were highly expressed in allodynic MDH. The circuitry composed of PKCγ and pERK1/2 cells is silent under physiological conditions but is activated after CCI-IoN, therefore, switching touch stimuli to pain sensation. The decrease of GABA transmission constituted a key factor in the activation of this neuronal circuitry, which opens the gate for non-noxious stimuli to reach nociceptive projection neurons in lamina I.

Reduction in inhibitory control is sufficient to generate hyperalgesia in a spiking model of nociceptive integration in the superficial dorsal horn

The spinal cord’s dorsal horn is a major termination site for primary afferents carrying sensory information from the periphery. The superfic ial dorsal horn (SDH, laminae I and II), in particular, receives inputs from nociceptive C and Aδ fibers, and have an important role in relaying and processing nociceptive information. This work focuses on the pain condition named hyperalgesia, characterized by an increased response to a stimulus that is normally painful. Much is still unknown regarding the central mechanisms giving rise to this condition. Here we present a large network spiking model for hyperalgesia which includes significant functional components of the nociceptive circuit in the SDH. The network architecture (Figure 1A) was defined according to key experimental results, namely: the somatotopic mapping between body surface and the superficial dorsal horn [1]; the presence of an inhibitory control which, under normal conditions, leads to a proper perception of pain [2]; the existence of divergent/convergent fiber inputs to lamina I (LI) neurons [3]; and the presence of inhibitory interneurons, i.e. islet cells, with inputs/outputs within lamina II (LII) [4]. In the model, the connectivity profile between LII interneurons (excitatory, Exc and inhibitory, Inh) and LI projection neurons creates a mexican-hat input profile with a strong centered excitation flanked by a surrounding inhibition. Activity level control was provided by feed-forward inhibition from the inhibitory interneurons in LII. The network model was created and simulated using the simulation environment NeuralSyns [http://sourceforge. net/projects/neuralsyns/]. Neurons were represented as integrate-and-fire units with conductance based synapses. The full model was comprised of 2.600 neurons and 320.000 synapses. The analysis was performed by assessing the changes produced in the receptive fields of LI nociceptive specific projection neurons (NS), both in normal (control) conditions and after reduction in inhibitory control

Functional differences between neurochemically defined populations of inhibitory interneurons in the rat spinal dorsal horn

In order to understand how nociceptive information is processed in the spinal dorsal horn we need to unravel the complex synaptic circuits involving interneurons, which constitute the vast majority of the neurons in laminae I–III. The main limitation has been the difficulty in defining functional populations among these cells. We have recently identified 4 non-overlapping classes of inhibitory interneuron, defined by expression of galanin, neuropeptide Y (NPY), neuronal nitric oxide synthase (nNOS) and parvalbumin, in the rat spinal cord. In this study we demonstrate that these form distinct functional populations that differ in terms of sst2A receptor expression and in their responses to painful stimulation. The sst2A receptor was expressed by nearly all of the nNOS- and galanin-containing inhibitory interneurons but by few of those with NPY and none of the parvalbumin cells. Many galanin- and NPY-containing cells exhibited phosphorylated extracellular signal-regulated kinases (pERK) after mechanical, thermal or chemical noxious stimuli, but very few nNOS-containing cells expressed pERK after any of these stimuli. However, many nNOS-positive inhibitory interneurons up-regulated Fos after noxious thermal stimulation or injection of formalin, but not after capsaicin injection. Parvalbumin cells did not express either activity-dependent marker following any of these stimuli. These results suggest that interneurons belonging to the NPY, nNOS and galanin populations are involved in attenuating pain, and for NPY and nNOS cells this is likely to result from direct inhibition of nociceptive projection neurons. They also suggest that the nociceptive inputs to the nNOS cells differ from those to the galanin and NPY populations.

Presynaptic CaV3.2 Channels Regulate Excitatory Neurotransmission in Nociceptive Dorsal Horn Neurons

It is generally accepted that presynaptic transmitter release is mainly regulated by subtypes of neuronal high-voltage-activated Ca(2+) channels. Here for the first time, we examined the role of T-type Ca(2+) channels (T-channels) in synaptic transmission in the dorsal horn (DH) of the spinal cord using patch-clamp recordings from acute spinal cord preparations from both rat and mouse. We found that selective pharmacological antagonism of T-channels inhibited spontaneous synaptic release of glutamate in superficial laminae I-II of the DH, while GABA release was spared. We found similar effect in identified nociceptive projection neurons of lamina I of the DH, but not in inhibitory DH interneurons. In comparison, antagonism of T-channels did not affect excitatory transmission in deeper non-nociceptive DH laminae. Furthermore, we used isoform-specific agents, knock-out mice and immunohistochemistry to specifically implicate presynaptic Ca(V)3.2 channels. We also used an animal model of painful diabetic neuropathy to demonstrate that blocking T-channels in superficial DH neurons suppressed spontaneous excitatory synaptic transmission in diabetic rats in greater degree than in healthy age-matched animals. These studies provide previously unknown information regarding the role of presynaptic T-channels in nociceptive signaling in the spinal cord.

Galanin receptor-expressing dorsal horn neurons: Role in nociception

Galanin, along with enkephalins and neuropeptide Y, has been hypothesized to negatively modulate nociception in the superficial dorsal horn of the spinal cord. In the present study, we sought to determine the role of presumably excitatory dorsal horn galanin receptor-expressing neurons in nociception by selectively destroying GalR1-expressing superficial dorsal horn interneurons using lumbar intrathecal injections of the targeted cytotoxin, galanin-saporin (Gal-sap). Lumbar intrathecal injection of Gal-sap (500 ng) reduced immunoperoxidase staining for GalR1 in the superficial dorsal horn without affecting primary afferent neurons in lumbar dorsal root ganglia. Lumbar intrathecal Gal-sap also: 1--reduced nocifensive reflex responding on the thermal plate at 0.3 °C, 44 °C, and 47 °C; 2--increased hot side occupancy in a thermal preference task (15 °C vs 45 °C); and, 3--decreased escape from 44 °C and 47 °C, but not 20 °C. Thus, similar to lesions of mu opiate receptor-expressing dorsal horn interneurons, selective destruction of GalR1-expressing superficial dorsal horn neurons produces heat hypo-algesia, likely due to loss of GalR1-expressing excitatory interneurons leading to reduced activation of nociceptive projection neurons in response to aversive heat. These results are different than those seen with intrathecal neuropeptide Y-saporin and suggest the potential value of selectively targeting GalR1-expressing dorsal horn neurons to control pain.

Distribution of RET immunoreactivity in the rodent spinal cord and changes after nerve injury

RET (for "rearranged during transfection") is a transmembrane tyrosine kinase signaling receptor for members of the glial cell line-derived neurotrophic factor (GDNF) family of ligands. We used RET immunohistochemistry (IHC), double-labeling immunofluorescence (IF), and in situ hybridization (ISH) in adult naïve and nerve-injured rats to study the distribution of RET in the spinal cord. In the dorsal horn, strong RET-immunoreactive (-ir) fibers were abundant in lamina II-inner (II(i)), although this labeling was preferentially observed after an antigen-unmasking procedure. After dorsal rhizotomy, RET-ir fibers in lamina II(i) completely disappeared from the dorsal horn, indicating that they were all primary afferents. After peripheral axotomy, RET-ir in primary afferents decreased in lamina II(i) and appeared to increase slightly in laminae III and IV. RET-ir was also observed in neurons and dendrites throughout the dorsal horn. Some RET-ir neurons in lamina I had the morphological appearance of nociceptive projection neurons, which was confirmed by the finding that 53% of RET-ir neurons in lamina I colocalized with neurokinin-1. GDNF-ir terminals were in close proximity to RET-ir neurons in the superficial dorsal horn. In the ventral horn, RET-ir was strongly expressed by motoneurons, with the strongest staining in small, presumably gamma-motoneurons. Increased RET expression following peripheral axotomy was most pronounced in alpha-motoneurons. The expression and regulation pattern of RET in the spinal cord are in line with its involvement in regenerative processes following nerve injury. The presence of RET in dorsal horn neurons, including nociceptive projection neurons, suggests that RET also has a role in signal transduction at the spinal level. This role may include mediating the effects of GDNF released from nociceptive afferent fibers.

Substance P-Driven Feed-Forward Inhibitory Activity in the Mammalian Spinal Cord

In mammals, somatosensory input activates feedback and feed-forward inhibitory circuits within the spinal cord dorsal horn to modulate sensory processing and thereby affecting sensory perception by the brain. Conventionally, feedback and feed-forward inhibitory activity evoked by somatosensory input to the dorsal horn is believed to be driven by glutamate, the principle excitatory neurotransmitter in primary afferent fibers. Substance P (SP), the prototypic neuropeptide released from primary afferent fibers to the dorsal horn, is regarded as a pain substance in the mammalian somatosensory system due to its action on nociceptive projection neurons. Here we report that endogenous SP drives a novel form of feed-forward inhibitory activity in the dorsal horn. The SP-driven feed-forward inhibitory activity is long-lasting and has a temporal phase distinct from glutamate-driven feed-forward inhibitory activity. Compromising SP-driven feed-forward inhibitory activity results in behavioral sensitization. Our findings reveal a fundamental role of SP in recruiting inhibitory activity for sensory processing, which may have important therapeutic implications in treating pathological pain conditions using SP receptors as targets.

Association of serotonin-like immunoreactive axons with nociceptive projection neurons in the caudal spinal trigeminal nucleus of the rat.

Serotoninergic projections to the spinal dorsal horn are implicated in the modulation of nociceptive transmission. However, morphological evidence indicating that serotoninergic projection fibers make synapses on nociceptive neurons in the medullary dorsal horn is still meager. Thus, we examined whether axonal varicosities with serotonin (5-HT)-like immunoreactivity (5-HT-LI) might make synapses on nociceptive projection neurons in the caudal spinal trigeminal nucleus (Vc) of the rat. Projection neurons were retrogradely labeled with tetramethylrhodamine-dextran amine (TMR-DA) or wheat germ agglutinin-horseradish peroxidase (WGA-HRP) that was injected into the parabrachial or thalamic region. Vc neurons in which c-fos protein-like immunoreactivity (Fos-LI) was induced by subcutaneous injection of formalin into the lip were considered nociceptive. Vc neurons in direct contact with axonal varicosities that bind isolectin I-B4 were also considered nociceptive. Triple labeling for 5-HT, TMR-DA, and Fos as well as that for 5-HT, TMR-DA, and I-B4 were done by using the immunofluorescence and fluorescence histochemical techniques. Confocal laser-scanning microscopy revealed that axonal varicosities with 5-HT-LI were in close apposition to TMR-DA-labeled neurons showing Fos-LI in lamina I and the outer part of lamina II (lamina IIo), and that both axonal varicosities with 5-HT-LI and those binding I-B4 were in close apposition to single neuronal profiles labeled with TMR-DA. The presumed nociceptive neuronal profiles in close apposition to axon terminals with 5-HT-LI were mainly those of laminae I and II neurons as well as dendrites of lamina III neurons. Electron microscopy confirmed that axon terminals with 5-HT-LI and those with I-B4 binding activity in laminae I and II made synapses on somatic and dendritic profiles that were labeled with WGA-HRP. The results indicate that serotoninergic neurons project directly on nociceptive projection neurons in the Vc.

Is there a pathway in the posterior funiculus that signals visceral pain?

The present report provides evidence that axons in the medial part of the posterior column at T10 convey ascending nociceptive signals from pelvic visceral organs. This evidence was obtained from human surgical case studies and histological verification of the lesion in one of these cases, along with neuroanatomical and neurophysiological findings in animal experiments. A restricted lesion in this area can virtually eliminate pelvic pain due to cancer. The results remain excellent even in cases in which somatic structures of the pelvic body wall are involved. Following this procedure, neurological testing reveals no additional neurological deficit. There is no analgesia to pinprick stimuli applied to the body surface, despite the relief of the visceral pain. Since it is reasonable to attribute the favorable results of limited midline myelotomies to the interruption of axons of visceral nociceptive projection neurons in the posterior column, we have performed experiments in rats to test this hypothesis. The results in rats indicate that the dorsal column does indeed include a nociceptive component that signals pelvic visceral pain. The pathway includes neurons of the postsynaptic dorsal column pathway at the L6-S1 segmental level, axons of these neurons in the fasciculus gracilis, and neurons of the nucleus gracilis and the ventral posterolateral nucleus of the thalamus.

Raphe magnus stimulation-induced antinociception in the cat is associated with release of amino acids as well as serotonin in the lumbar dorsal horn

Stimulation in the nucleus raphe magnus (NRM) inhibits transmissionof nociceptive information within the spinal cord through activation of bulbospinal pathways. This study used microdialysis in combination with high pressure liquid chromatography to measure the release of serotonin (5HT) and several amino acids, including glutamate, aspartate and glycine, from the lumbar dorsal horn during electrical stimulation within the NRM in the α-chloralose anesthetized cat. Observed release of putative neurotransmitters was correlated with inhibition of nociceptive projection neurons recorded from sites within 800 μm rostral or caudal to the dialysis fiber. NRM stimulus parameters considered to preferentially activate myelinated fibers caused inhibition of nociceptive evoked activity, and increased the release of excitatory amino acids and glycine within the spinal cord, with no detectable release of 5HT. When pulse widths were lengthened and unmyelinated fibers were also activated, increases in 5HT in the spinal dialysate were observed as well. Strychnine administered through the dialysis fiber (0.02-1 mM) antagonized NRM-induced inhibition when 5HT release was not detected. Inhibition produced by stimulation that increased 5HT concentrations was relatively strychnine resistant. These results point to a raphe-spinal inhibitory pathway that is not dependent on 5HT, the activation of which results in the spinal release of glycine.

Substance P-like and serotonin-like immunoreactivity in the lateral cervical nucleus of the raccoon.

The distribution of substance P and serotonin in the lateral cervical nucleus (LCN) of the raccoon was examined by light microscopic immunohistochemistry. Substance P-immunoreactive fibers were found to be clustered in the ventromedial part of the LCN, whereas only few such fibers appeared in the dorsolateral part of the nucleus. This organization is closely similar to that previously observed in the cat, and provides further evidence for an anatomic and functional segregation along the transverse axis of the LCN in carnivores. In some sections, substance P-positive fibers were found primarily in areas of the ventromedial LCN containing small neurons, indicating that such fibers may be involved in functions of the LCN associated with nociceptive projection neurons and/or local circuit neurons. The raccoon LCN also received a relatively sparse innervation of serotonin-positive fibers that were distributed throughout the nucleus, an organization similar to that previously observed in the cat. The functional role of the serotonergic fibers is unclear. However, their presence suggests that descending influences on transmission in the spinocervicothalamic pathway, in addition to the well-documented descending control of spinocervical tract neurons, may be present also at the level of the LCN.

Neurotransmitters in Nociceptive Modulatory Circuits

Significant advances have been made in our understanding of nociceptive modulation from RVM. Among the most useful conceptually has been the discovery that there are two classes of modulatory neurons in the RVM that are likely to have opposing actions on nociception: on-cells, which may facilitate nociceptive transmission, and off-cells, which probably have a net inhibitory effect on nociception. The similarity in response properties among the members of each class, their large, somatic "receptive fields," and the wide distribution of the terminal fields of axons of individual neurons to the trigeminal sensory complex and to multiple spinal segments indicate that these neurons exert a global influence over nociceptive responsiveness. Drug microinjections into the RVM presumably shift the balance between states of on- or off-cell firing and also produce measurable changes in the threshold for nocifensor reflexes. The meaningful unit of function in the RVM nociceptive modulatory system therefore probably consists of large ensembles of physiologically and pharmacologically similar neurons. The strong coordination of activity of the two classes of RVM neuron may depend largely upon intranuclear projections from RVM off-cells that excite other off-cells and inhibit on-cells. The off-cell pause is GABA-mediated, and it is likely that there is a subset of GABA-containing RVM on-cells that directly inhibit off-cells. Furthermore, the available evidence indicates that exogenous opiates activate off-cells by inhibiting GABAergic release. Presumably, enkephalinergic cells in the RVM disinhibit off-cells in a similar way. Although non-serotonin-containing off-cells certainly exist, we propose that some off-cells contain serotonin. Other possible connections are based on more limited data; however, ACh, neurotensin, NE, and EAAs are present in neurons that project to the RVM, and each of these compounds, when microinjected into the RVM, has a modulating effect on nociceptive transmission. The local circuits in the RVM that underlie these actions remain to be elucidated. At the level of the dorsal horn, there is good evidence for each of three inhibitory mechanisms: direct inhibition of nociceptive projection neurons, inhibition of excitatory relay interneurons, and excitation of an inhibitory interneuron. The relative contribution made by each of these circuits is unknown.(ABSTRACT TRUNCATED AT 400 WORDS)

Effects of excitatory amino acids (EAA) and μ opioid agonists on nociceptive projection neurons in rat

Effects of excitatory amino acids (EAA) and μ opioid agonists on nociceptive projection neurons in rat