Spinal Excitatory Dynorphinergic Interneurons Contribute to Burn Injury-Induced Nociception Mediated by Phosphorylated Histone 3 at Serine 10 in Rodents

The phosphorylation of serine 10 in histone 3 (p-S10H3) has recently been demonstrated to participate in spinal nociceptive processing. However, superficial dorsal horn (SDH) neurons involved in p-S10H3-mediated nociception have not been fully characterized. In the present work, we combined immunohistochemistry, in situ hybridization with the retrograde labeling of projection neurons to reveal the subset of dorsal horn neurons presenting an elevated level of p-S10H3 in response to noxious heat (60 °C), causing burn injury. Projection neurons only represented a small percentage (5%) of p-S10H3-positive cells, while the greater part of them belonged to excitatory SDH interneurons. The combined immunolabeling of p-S10H3 with markers of already established interneuronal classes of the SDH revealed that the largest subset of neurons with burn injury-induced p-S10H3 expression was dynorphin immunopositive in mice. Furthermore, the majority of p-S10H3-expressing dynorphinergic neurons proved to be excitatory, as they lacked Pax-2 and showed Lmx1b-immunopositivity. Thus, we showed that neurochemically heterogeneous SDH neurons exhibit the upregulation of p-S10H3 shortly after noxious heat-induced burn injury and consequential tissue damage, and that a dedicated subset of excitatory dynorphinergic neurons is likely a key player in the development of central sensitization via the p-S10H3 mediated pathway.

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The neurokinin-1 receptor is expressed with gastrin-releasing peptide receptor in spinal interneurons and modulates itch

10.1101/2020.07.14.199471 ◽

2020 ◽

Author(s):

Tayler D. Sheahan ◽

Charles A. Warwick ◽

Louis G. Fanien ◽

Sarah E. Ross

Keyword(s):

Dorsal Horn ◽

Projection Neurons ◽

Spinal Neurons ◽

Neurokinin 1 Receptor ◽

Peptide Receptor ◽

Gastrin Releasing Peptide ◽

Dorsal Horn Neurons ◽

Endogenous Expression ◽

Neurokinin 1

AbstractThe neurokinin-1 receptor (NK1R, encoded by Tacr1) is expressed in spinal dorsal horn neurons and has been suggested to mediate itch. However, previous studies relied heavily on neurotoxic ablation of NK1R spinal neurons, which limited further dissection of their function in spinal itch circuitry. Thus, we leveraged a newly developed Tacr1CreER mouse line to characterize the role of NK1R spinal neurons in itch. We show that pharmacological activation of spinal NK1R and chemogenetic activation of Tacr1CreER spinal neurons increases itch behavior, whereas pharmacological inhibition of spinal NK1R suppresses itch behavior. We use fluorescence in situ hybridization to characterize the endogenous expression of Tacr1 throughout the superficial and deeper dorsal horn, as well as the lateral spinal nucleus.Retrograde labeling studies from the parabrachial nucleus show that less than 20% of superficial Tacr1CreER dorsal horn neurons are spinal projection neurons, and thus the majority of Tacr1CreER are local interneurons. We then use a combination of in situ hybridization and ex vivo two-photon Ca2+ imaging of the spinal cord to establish that NK1R and the gastrin-releasing peptide receptor (GRPR) are coexpressed within a subpopulation of excitatory superficial dorsal horn neurons. These findings are the first to describe a role for NK1R interneurons in itch and extend our understanding of the complexities of spinal itch circuitry.

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Excitation of Mouse Superficial Dorsal Horn Neurons by Histamine and/or PAR-2 Agonist: Potential Role in Itch

Journal of Neurophysiology ◽

10.1152/jn.00463.2009 ◽

2009 ◽

Vol 102 (4) ◽

pp. 2176-2183 ◽

Cited By ~ 43

Author(s):

Tasuku Akiyama ◽

Mirela Iodi Carstens ◽

E. Carstens

Keyword(s):

Dorsal Horn ◽

Potential Role ◽

Dynamic Range ◽

Intradermal Injection ◽

Mustard Oil ◽

Superficial Dorsal Horn ◽

Noxious Heat ◽

Dorsal Horn Neurons ◽

Sensory Pathways ◽

Transduction Mechanisms

Recent studies have suggested the existence of separate transduction mechanisms and sensory pathways for histamine and nonhistaminergic types of itch. We studied whether histamine and an agonist of the protease-activated receptor (PAR)-2, associated with nonhistaminergic itch, excite murine dorsal horn neurons. Single units were recorded in superficial lumbar dorsal horn of adult ICR mice anesthetized with pentobarbital. Unit activity was searched using a small intradermal hindpaw injection of histamine or the PAR-2 agonist SLIGRL-NH2. Isolated units were subsequently challenged with intradermal histamine followed by SLIGRL-NH2 (each 50 μg/1 μl) or reverse order, followed by mechanical, thermal, and algogenic stimuli. Forty-three units were classified as wide dynamic range (62%), nociceptive specific (22%), or mechano insensitive (16%). Twenty units gave prolonged (mean, 10 min) discharges to intradermal injection of histamine; 76% responded to subsequent SLIGRL-NH2, often more briefly. Units additionally responded to noxious heat (63%), cooling (43%), topical mustard oil (53%), and intradermal capsaicin (67%). Twenty-two other units gave prolonged (mean, 5 min) responses to initial intradermal injection of SLIGRL-NH2; 85% responded to subsequent intradermal histamine. They also responded to noxious heat (75%), mustard oil (93%), capsaicin (63%), and one to cooling. Most superficial dorsal horn neurons were excited by both histamine and the PAR-2 agonist, suggesting overlapping pathways for histamine- and non–histamine-mediated itch. Because the large majority of pruritogen-responsive neurons also responded to noxious stimuli, itch may be signaled at least partly by a population code.

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Intradermal endothelin-1 excites bombesin-responsive superficial dorsal horn neurons in the mouse

Journal of Neurophysiology ◽

10.1152/jn.00723.2015 ◽

2015 ◽

Vol 114 (4) ◽

pp. 2528-2534 ◽

Cited By ~ 4

Author(s):

T. Akiyama ◽

M. Nagamine ◽

A. Davoodi ◽

M. Iodi Carstens ◽

F. Cevikbas ◽

...

Keyword(s):

Dorsal Horn ◽

Dynamic Range ◽

Allyl Isothiocyanate ◽

High Threshold ◽

Mechanical Stimuli ◽

Superficial Dorsal Horn ◽

Noxious Heat ◽

Nociceptive Neurons ◽

Endothelin 1 ◽

Dorsal Horn Neurons

Endothelin-1 (ET-1) has been implicated in nonhistaminergic itch. Here we used electrophysiological methods to investigate whether mouse superficial dorsal horn neurons respond to intradermal (id) injection of ET-1 and whether ET-1-sensitive neurons additionally respond to other pruritic and algesic stimuli or spinal superfusion of bombesin, a homolog of gastrin-releasing peptide (GRP) that excites spinal itch-signaling neurons. Single-unit recordings were made from lumbar dorsal horn neurons in pentobarbital-anesthetized C57BL/6 mice. We searched for units that exhibited elevated firing after id injection of ET-1 (1 μg/μl). Responsive units were further tested with mechanical stimuli, bombesin (spinal superfusion, 200 μg·ml−1·min−1), heating, cooling, and additional chemicals [histamine, chloroquine, allyl isothiocyanate (AITC), capsaicin]. Of 40 ET-1-responsive units, 48% responded to brush and pinch [wide dynamic range (WDR)] and 52% to pinch only [high threshold (HT)]. Ninety-three percent responded to noxious heat, 50% to cooling, and >70% to histamine, chloroquine, AITC, and capsaicin. Fifty-seven percent responded to bombesin, suggesting that they participate in spinal itch transmission. That most ET-1-sensitive spinal neurons also responded to pruritic and algesic stimuli is consistent with previous studies of pruritogen-responsive dorsal horn neurons. We previously hypothesized that pruritogen-sensitive neurons signal itch. The observation that ET-1 activates nociceptive neurons suggests that both itch and pain signals may be generated by ET-1 to result in simultaneous sensations of itch and pain, consistent with observations that ET-1 elicits both itch- and pain-related behaviors in animals and burning itch sensations in humans.

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Superficial Dorsal Horn Neurons Identified by Intracutaneous Histamine: Chemonociceptive Responses and Modulation by Morphine

Journal of Neurophysiology ◽

10.1152/jn.2000.84.2.616 ◽

2000 ◽

Vol 84 (2) ◽

pp. 616-627 ◽

Cited By ~ 50

Author(s):

Steven L. Jinks ◽

E. Carstens

Keyword(s):

Dorsal Horn ◽

Dynamic Range ◽

Mustard Oil ◽

Mechanical Stimuli ◽

Superficial Dorsal Horn ◽

Neuronal Responses ◽

Noxious Heat ◽

Morphine Concentration ◽

Dorsal Horn Neurons ◽

Low Concentrations

We have investigated whether neurons in superficial laminae of the spinal dorsal horn respond to intracutaneous (ic) delivery of histamine and other irritant chemicals, and thus might be involved in signaling sensations of itch or chemogenic pain. Single-unit recordings were made from superficial lumbar dorsal horn neurons in pentobarbital sodium–anesthetized rats. Chemoresponsive units were identified using ic microinjection of histamine (3%, 1 μl) into the hindpaw as a search stimulus. All superficial units so identified [9 nociceptive-specific (NS), 26 wide-dynamic-range (WDR)] responded to subsequent ic histamine. A comparison group of histamine-responsive deep dorsal horn neurons ( n = 16) was similarly identified. The mean histamine-evoked discharge decayed to 50% of the maximal rate significantly more slowly for the superficial (92.2 s ± 65.5, mean ± SD) compared with deep dorsal horn neurons (28.2 s ± 11.6). In addition to responding to histamine, most superficial dorsal horn neurons were also excited by ic nicotine (22/25 units), capsaicin (21/22), topical mustard oil (5/6), noxious heat (26/30), and noxious and/or innocuous mechanical stimuli (except for 1 unit that did not have a mechanosensitive receptive field). Application of a brief noxious heat stimulus during the response to ic histamine evoked an additive response in all but two cases, followed by transient depression of firing in 11/20 units. Intrathecal (IT) administration of morphine had mixed effects on superficial dorsal horn neuronal responses to ic histamine and noxious heat. Low morphine concentrations (100 nM to 1 μM) facilitated histamine-evoked responses (to >130% of control) in 9/24 units, depressed the responses (by >70%) in 11/24, and had no effect in 4. Naloxone reversed morphine-induced effects in some but not all cases. A higher morphine concentration (10 μM) had a largely depressant, naloxone-reversible effect on histamine responses. Responses of the same superficial neurons to noxious heat were facilitated (15/25), reduced (8/25), or unaffected (2/25) by low morphine concentrations and were depressed by the higher morphine concentration. In contrast, deep dorsal horn neuronal responses to both histamine and noxious heat were primarily depressed by low concentrations of morphine in a naloxone-reversible manner. These results indicate that superficial dorsal horn neurons respond to both pruritic and algesic chemical stimuli and thus might participate in transmitting sensations of itch and/or chemogenic pain. The facilitation of superficial neuronal responses to histamine by low concentrations of morphine, coupled with inhibition of deep dorsal horn neurons, might underlie the development of pruritis that is often observed after epidural morphine.

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Altered synaptic input and GABABreceptor function in spinal superficial dorsal horn neurons in rats with diabetic neuropathy

The Journal of Physiology ◽

10.1113/jphysiol.2006.126102 ◽

2007 ◽

Vol 579 (3) ◽

pp. 849-861 ◽

Cited By ~ 66

Author(s):

Xiu-Li Wang ◽

Hong-Mei Zhang ◽

Shao-Rui Chen ◽

Hui-Lin Pan

Keyword(s):

Diabetic Neuropathy ◽

Dorsal Horn ◽

Synaptic Input ◽

Superficial Dorsal Horn ◽

Dorsal Horn Neurons

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Developmental Changes in the Fidelity and Short-Term Plasticity of GABAergic Synapses in the Neonatal Rat Dorsal Horn

Journal of Neurophysiology ◽

10.1152/jn.01342.2007 ◽

2008 ◽

Vol 99 (6) ◽

pp. 3144-3150 ◽

Cited By ~ 18

Author(s):

Rachel A. Ingram ◽

Maria Fitzgerald ◽

Mark L. Baccei

Keyword(s):

Spinal Cord ◽

Dorsal Horn ◽

Receptive Fields ◽

Neuronal Firing ◽

Neonatal Rat ◽

Superficial Dorsal Horn ◽

Short Term ◽

Dorsal Horn Neurons ◽

Gabaergic Synapses

The lower thresholds and increased excitability of dorsal horn neurons in the neonatal rat suggest that inhibitory processing is less efficient in the immature spinal cord. This is unlikely to be explained by an absence of functional GABAergic inhibition because antagonism of γ-aminobutyric acid (GABA) type A receptors augments neuronal firing in vivo from the first days of life. However, it is possible that more subtle deficits in GABAergic signaling exist in the neonate, such as decreased reliability of transmission or greater depression during repetitive stimulation, both of which could influence the relative excitability of the immature spinal cord. To address this issue we examined monosynaptic GABAergic inputs onto superficial dorsal horn neurons using whole cell patch-clamp recordings made in spinal cord slices at a range of postnatal ages (P3, P10, and P21). The amplitudes of evoked inhibitory postsynaptic currents (IPSCs) were significantly lower and showed greater variability in younger animals, suggesting a lower fidelity of GABAergic signaling at early postnatal ages. Paired-pulse ratios were similar throughout the postnatal period, whereas trains of stimuli (1, 5, 10, and 20 Hz) revealed frequency-dependent short-term depression (STD) of IPSCs at all ages. Although the magnitude of STD did not differ between ages, the recovery from depression was significantly slower at immature GABAergic synapses. These properties may affect the integration of synaptic inputs within developing superficial dorsal horn neurons and thus contribute to their larger receptive fields and enhanced afterdischarge.

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Five Sources of a Dorsal Root Potential: Their Interactions and Origins in the Superficial Dorsal Horn

Journal of Neurophysiology ◽

10.1152/jn.1997.78.2.860 ◽

1997 ◽

Vol 78 (2) ◽

pp. 860-871 ◽

Cited By ~ 28

Author(s):

Patrick D. Wall ◽

Malcolm Lidierth

Keyword(s):

Motor Cortex ◽

Dorsal Horn ◽

Dorsal Root ◽

Repetitive Stimulation ◽

Superficial Dorsal Horn ◽

Dorsal Horn Neurons ◽

Dorsal Roots ◽

Myelinated Fibers ◽

Root Potential ◽

Stimulation Of

Wall, Patrick D. and Malcolm Lidierth. Five sources of a dorsal root potential: their interactions and origins in the superficial dorsal horn. J. Neurophysiol. 78: 860–871, 1997. The dorsal root potential (DRP) was measured on the lumbar dorsal roots of urethan anesthetized rats and evoked by stimulation of five separate inputs. In some experiments, the dorsal cord potential was recorded simultaneously. Stimulation of the L3 dorsal root produced a DRP on the L2 dorsal root containing the six components observed in the cat including the prolonged negative wave (DRP V of Lloyd 1952 ). A single shock to the myelinated fibers in the sural nerve produced a DRP on the L6 dorsal root after the arrival in the cord of the afferent volley. The shape of this DRP was similar to that produced by dorsal root stimulation. Repetitive stimulation of the myelinated fibers in the gastrocnemius nerve also produced a prolonged negative DRP on the L6 dorsal root. When a single stimulus (<5 μA; 200 μs) was applied through a microelectrode to the superficial Lissauer Tract (LT) at the border of the L2 and L3 spinal segments, a characteristic prolonged negative DRP (LT-DRP) began on the L2 dorsal root after some 15 ms. Stimulation of the LT evoked DRPs bilaterally. Recordings on nearby dorsal roots showed this DRP to be unaccompanied by stimulation of afferent fibers in those roots. The LT-DRP was unaffected by neonatal capsaicin treatment that destroyed most unmyelinated fibers. Measurements of myelinated fiber terminal excitability to microstimulation showed that the LT-DRP was accompanied by primary afferent depolarization. Repetitive stimulation through a microelectrode in sensorimotor cortex provoked a prolonged and delayed negative DRP (recorded L2–L4). Stimulation in the cortical arm area and recording on cervical dorsal roots showed that the DRP was evoked more from motor areas than sensory areas of cortex. Interactions were observed between the LT-DRP and that evoked from the sural or gastrocnemius nerves or motor cortex. The LT-DRP was inhibited by preceding stimulation of the other three sources but LT stimulation did not inhibit DRPs evoked from sural or gastrocnemius nerves on the L6 dorsal root or from motor cortex on the L3 root. However, LT stimulation did inhibit the DRP evoked by a subsequent Lissaeur tract stimulus. Recordings were made from superficial dorsal horn neurons. Covergence of input from LT sural, and gastrocnemius nerves and cortex was observed. Spike-triggered averaging was used to examine the relationship between the ongoing discharge of superficial dorsal horn neurons and the spontaneous DRP. The discharge of 81% of LT responsive cells was correlated with the DRP.

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