The spinal substantia gelatinosa (SG; Lamina II) is a major synaptic zone for unmyelinated (C) primary afferents. In spinal cord slices from a transgenic mouse in which certain GABAergic Lamina II neurones are labeled with green fluorescent protein (GFP), we compared primary afferent input with local efferent connections made by inhibitory SG neurones. Simultaneous whole-cell recordings from characterized neurones establish that inhibitory SG neurones receive monosynaptic input from a subset of unmyelinated primary afferents and connect to other Lamina II cells that have input from a different set of afferents, permitting interactions between distinctive afferent messages. Certain Lamina II inhibitory cells were found to connect to one another by reciprocal links. Inhibitory Lamina II connections appear arranged to modulate activity from different sets of peripheral unmyelinated fibres through neural circuitry that includes disinhibition.. 
Previous studies have shown sprouting of Abeta-fibres into Lamina II of the spinal cord dorsal horn after nerve injury and the formation of new synapses by these sprouts.
We found that NT5E was located on peptidergic and nonpeptidergic nociceptive neurons in dorsal root ganglia (DRG) and on axon terminals in Lamina II (the substantia gelatinosa) of spinal cord. Following nerve injury, NT5E protein and AMP histochemical staining were coordinately reduced in Lamina II.
Here we report that most interneurons in Lamina II are placed under phasic control by both GABAergic and glycinergic synapses, a number of which exhibit dual GABA/glycine co-release. A developmental switch is also apparent: a subpopulation of Lamina II interneurons controlled exclusively by either GABAergic or glycinergic synapses becomes detectable only after postnatal days 15 and 21, respectively. This allowed us to demonstrate that synapses containing GlyR alpha3 contribute in large part to synaptic inhibition in Lamina II. These findings explain the absence of any basal nociceptive hypersensitivity in Glra3 knockout mice, as GlyR alpha1 is still available for mediating synaptic inhibition at Lamina II synapses, but cannot be modulated by the prostaglandin-E-prostanoid type 2 (EP2) receptor-protein kinase A signalling cascade.
Additionally, this antibody detects PAP in mouse and rat small- to medium-diameter DRG neurons and axon terminals in Lamina II of spinal cord.
By using patch-clamp and confocal microscope calcium imaging techniques in rat spinal cord slices, we monitored the activity of dorsal horn Lamina II neurons following astrocyte activation. Results obtained revealed that stimuli that triggered Ca(2+) elevations in astrocytes, such as the purinergic receptor agonist BzATP and low extracellular Ca(2+), induce in Lamina II neurons slow inward currents (SICs).
OBJECTIVE: To investigate the Methionine Enkephalin (M-Enk) expression in Lamina II of cat spinal cord and the atypical complex terminal (ACT) after complete dorsal rhizotomy. The immuno-positive ACTs have been founded in the Lamina II of chronic operation side beside some M-Enk immunopositive simple terminals, they are round or ellipse in shape and usually form flat or convex two synapse with two post-compounds. CONCLUSION: M-Enk express in operation side after complete dorsal rhizotomy is mainly on the lateral side in Lamina II.
Unilateral intraplantar capsaicin injection in control animals evoked extracellular signal-regulated kinase 1/2 phosphorylation in a group of neurons in lamina I and Lamina II of the ipsilateral spinal dorsal horn in a somatotopically appropriate area.
METHODS: The effects of bupivacaine on the response to exogenous administration of N-methyl-D-aspartate (NMDA) receptor agonists were examined in Lamina II neurons of adult rat spinal cord slices using the whole-cell patch-clamp technique.
Injections of the cornea produced dense label in the interstitial islands in the ventral medullary dorsal horn (MDH), probably lamina I, and in neuropil in the ventromedial tip of the MDH, probably Lamina II.
METHODS:: The effects of bupivacaine on the response to exogenous administration of N-methyl-d-aspartate (NMDA) receptor agonists were examined in Lamina II neurons of adult rat spinal cord slices using the whole-cell patch-clamp technique.
BACKGROUND: Substantia gelatinosa (SG, Lamina II) is a spinal cord region where most unmyelinated primary afferents terminate and the central nociceptive processing begins.
Although it is known that C-tactile fibres terminate in the substantia gelatinosa (Lamina II) of the spinal cord, virtually all of the neurons in this region are interneurons, and currently it is not known how impulses in C-mechanoreceptors are transmitted to higher centres.
In the dorsal horn of the spinal cord, these afferents project to lamina I and the innermost layer of Lamina II, which has previously been implicated in persistent pain caused by injury.
Oxytocinergic axons originating from a subpopulation of paraventricular hypothalamic neurons establish synaptic contacts with Lamina II interneurons but little is known about the functional role of OT with respect to neuronal firing and excitability. RESULTS: Using the patch-clamp technique, we have recorded Lamina II interneurons in acute transverse lumbar spinal cord slices of rats (15 to 30 days old) and analyzed the OT effects on action potential firing ability. In the current clamp mode, we found that bath application of a selective OT-receptor agonist (TGOT) reduced firing in the majority of Lamina II interneurons exhibiting a bursting firing profile, but never in those exhibiting a single spike discharge upon depolarization. CONCLUSION: This effect of OT on the firing profile of Lamina II neurons is in good agreement with the antinociceptive and analgesic properties of OT described in vivo..
The Mas-related G-protein-coupled receptor D (Mrgprd) marks a distinct subset of sensory neurons that transmit polymodal nociceptive information from the skin epidermis to the substantia gelatinosa (SG, Lamina II) of the spinal cord.
Whole-cell patch-clamp recordings of Lamina II neurons were performed in spinal cord slices.
However, stalked cells were not the only Lamina II cells in direct contact with projection neurons.
Neurons with nNOS-like immunoreactivity (nNOS-LI) were concentrated mainly in the Lamina II of the Sp5C, showing no significant statistical difference during arthritis.
Synaptic glomeruli that involve tachykinin-containing primary afferent central terminals are numerous in Lamina II of the chicken spinal cord.
By contrast, PKCgamma interneurons of inner Lamina II only receive a myelinated afferent input.
In Lamina II, where mainly nociceptive afferents terminate, noradrenaline (20 microM) depolarised significantly more EGFP-labelled (41%) than non-EGFP-labelled GABAergic neurons (4%). In Lamina III, where low threshold afferents terminate, EGFP-labelled neurons were never depolarised but either hyperpolarised (25%) or not affected (75%) by noradrenaline. Depolarisations of EGFP-labelled Lamina II neurons were mimicked by the alpha(1)-adrenoceptor agonist phenylephrine (10-20 microM) and abolished by the alpha(1)-adrenoceptor antagonist prazosin (2 microM). These results show that noradrenaline directly excites inhibitory (GABAergic) Lamina II interneurons in addition to its inhibitory effect on (putatively excitatory) interneurons in superficial spinal dorsal horn.
In the superficial layers of the medullary dorsal horn, 12% of ERalpha-labeled cells, mainly located in Lamina II, also expressed noxious-induced Fos.
Moreover, the superficial c-kit(+) fibers originate from the dorsal root ganglion, and c-kit in Lamina II inner layer comes from intrinsic expression of the spinal cord.
To address this, we studied inhibitory synaptic transmission in Lamina II inhibitory interneurons identified under visual guidance in spinal slices taken from transgenic mice expressing enhanced green fluorescent protein (EGFP) under the control of the GAD promoter. These inhibitory inputs originated at least in part from local Lamina II interneurons as verified by simultaneous recordings from pairs of EGFP+ cells.
Excitatory interneuronal connectivity within Lamina II exhibited a pronounced sagittal orientation, in keeping with the somatotopic organization present in the pattern of primary afferent projections. Excitatory inputs to all classes of Lamina II neurons arose from a wider rostrocaudal area than inhibitory inputs, whereas both excitatory and inhibitory input zones were restricted mediolaterally. All cell types received excitatory input predominantly from positions ventral to that of their soma, but in lamina I neurons and Lamina II vertical cells this ventral displacement of the excitatory input zone was greater than in the other cell types, resulting in a more pronounced translaminar input pattern. A previously unknown excitatory input to the superficial dorsal horn from Lamina III-IV was identified in a subset of the vertical cell population.
Moreover, it was found that neuropathic animals with a higher degree of mechanical allodynia had a lower intensity of GFAP expression in Lamina II (substantia gelatinosa).
The upregulated IL-1 beta was mainly expressed in the neurons in the Lamina II approximately IV of the spinal cord.
Patch-clamp recordings in Lamina II neurons of isolated spinal cord slices showed that MCP-1 not only enhanced spontaneous EPSCs but also potentiated NMDA- and AMPA-induced currents.
Using whole-cell patch-clamp recording from rat transversal spinal cord slices, we investigated the actions of 50% N(2)O and 0.5% isoflurane (both 0.3 rat MAC; minimum alveolar concentration) on exogenously applied gamma-aminobutyric acid (GABA)- and glycine-induced currents in rat dorsal horn Lamina II neurons.
We have reported that <10% of AMPAr-containing synapses in Lamina II have the long form of GluR4, and that these are often arranged in dorsoventrally orientated clusters. All NK1r-positive Lamina III/IV neurons had numerous GluR2-immunoreactive puncta in their dendritic plasma membranes, and virtually all (97%) of the puncta tested were labelled (usually strongly) with the GluR4 antibody. These results show that synaptic AMPArs on the dendrites of the Lamina III/IV NK1r projection neurons contain GluR2, GluR3 and GluR4, but not GluR1 subunits..
To study the signaling mechanisms involved in this unique mAChR action, GABAergic spontaneous inhibitory postsynaptic currents (sIPSCs) of Lamina II neurons were recorded using whole-cell patch clamp techniques in rat spinal cord slices.
In this study, inhibitory and excitatory postsynaptic currents (IPSCs and EPSCs, respectively) were recorded from Lamina II neurons in rat spinal cord slices.
Whole-cell recording of Lamina II neurons was performed in spinal cord slices from control and nerve-ligated rats.
Capsaicin significantly increased the frequency of glutamatergic spontaneous excitatory post-synaptic currents and miniature excitatory post-synaptic currents in almost all the Lamina II neurons tested in control rats. In RTX-treated or in dorsal rhizotomized rats, capsaicin also produced an inward current in a subpopulation of Lamina II neurons. However, capsaicin had no effect on GABAergic and glycinergic spontaneous inhibitory post-synaptic currents of Lamina II neurons in RTX-treated or dorsal rhizotomized rats.
PVN electrical stimulation also activates a subpopulation of neurons in Lamina II.
Activated p38 MAPK was localized primarily to microglia and to a lesser extent in oligodendrocytes and Lamina II neurons.
Conversely, synapsin II knockout mice showed a stronger and longer-lasting increase of GABA in Lamina II of the dorsal horn after nerve injury than wild type mice.
Protein kinase C gamma (PKCgamma), which is concentrated in interneurons of the inner part of Lamina II of the dorsal horn, has been implicated in injury-induced allodynia, a condition wherein pain is produced by innocuous stimuli.
In this study, we determined the presynaptic and postsynaptic effects of the mu-opioid receptor agonist [ D-Ala(2),N-Me-Phe(4),Gly-ol(5)]-enkephalin (DAMGO) using whole-cell patch-clamp recordings of Lamina II neurons in rat spinal cord slices. However, the concentration response and the duration of the effects of DAMGO on G protein-coupled inwardly rectifying K+ currents in Lamina II neurons were not significantly different between vehicle- and RTX-treated groups.
Recent studies have unraveled some of the neuronal circuitries and mechanisms involved, highlighting the key role of synaptic glomeruli in Lamina II as the main sites for such a modulation..
We have used patch clamp recording of Lamina II neurons in spinal cord slices and immunocytochemistry in order to identify PVN-activated neurons in the superficial layers of the spinal cord and attempted to determine how this neuronal population may lead to OT-mediated antinociception. RESULTS: We show that OT released during PVN stimulation specifically activates a subpopulation of Lamina II glutamatergic interneurons which are localized in the most superficial layers of the dorsal horn of the spinal cord (lamina I-II). This OT-specific stimulation of glutamatergic neurons allows the recruitment of all GABAergic interneurons in Lamina II which produces a generalized elevation of local inhibition, a phenomenon which might explain the reduction of incoming Adelta and C primary afferent-mediated sensory messages. CONCLUSION: Our results obtained in Lamina II of the spinal cord provide the first clear evidence of a specific local neuronal network that is activated by OT release to induce antinociception.
In this study, we sought to establish the presence of adenosine A(2A) receptors in the Lamina II of the rat lumbar dorsal horn neurons and investigated whether the activation of A(2A) receptors is able to modulate NMDA receptor currents. The RT-PCR technique is also performed specifically in the Lamina II, and the presence of adenosine A(2A) receptor transcripts is assessed in neurons from the Lamina II with the single-cell RT-PCR technique. Finally, single cell RT-PCR revealed the presence of adenosine A(2A) receptor transcripts in a sample of Lamina II neurons.
Using patch-clamp recordings in Lamina II neurons of isolated spinal cord slices, we compared the effects of IL-1beta, IL-6, and TNFalpha on excitatory and inhibitory synaptic transmission.
The present study was undertaken to further examine the ultrastructural localization of SST(2A) receptor in Lamina II of the spinal dorsal horn and the role of SST(2A) receptor in thermal hyperalgesia following Complete Freund's Adjuvant (CFA)-induced inflammation. We found that SST(2A) receptors in Lamina II are located primarily in postsynaptic dendrites and soma, but not in axons or synaptic terminals. CFA-induced inflammation markedly increased SST(2A) receptor-like immunoreactivity in Lamina II.
p-ERK1/2 and p-p38 were expressed in neurons in distinct regions of the L4/L5 dorsal horn; p-ERK1/2 was mainly in lamina I, while p-p38 was mainly in Lamina II of the dorsal horn.
B(2) receptors are coexpressed in dorsal horn neurons with protein kinase A (PKA) and the delta isoform of protein kinase C (PKC), and we find that the augmentation by bradykinin of AMPA and NMDA receptor-mediated currents in Lamina II neurons requires coactivation of both PKC and PKA.
Fos-positive cells were mainly distributed within the isolectin B4-labeled region (inner aspect of Lamina II) after histamine injection.
EA treatment decreased the number of TUNEL-positive apoptotic cells in Lamina II of the L(3) and L(6) cord segments at 7 and 14 days post operation (dpo).
C-fos was not induced in inner Lamina II following c-fiber electrical stimulation of the intact or axotomized sciatic nerve, indicating no such plasticity at the spinal cord level.
The Nissl staining revealed that the thickness of lamina I (LI) and outer Lamina II remained mostly unchanged from birth until adulthood. CGRP afferents terminated mostly in LI and the outer two-thirds of Lamina II, whereas the termination area of fibers binding IB4 was centered on the middle one-third of Lamina II at all ages studied.
TSPO activity disappeared in laminas III-IV after P8 and was functionally downregulated in Lamina II after P15, resulting in a marked reduction of mIPSC duration in these laminas. TSPO-mediated synthesis of 3alpha5alpha-reduced steroids was spatially restricted, because, at P9-P15, when their production was maximal in Lamina II, no sign of spillover to laminas III-IV was apparent.
In the spinal cord, it was found that SSeCKS-immunoreactive axon collaterals terminate in the dorsal third of Lamina II outer in a region similar to that of unmyelinated C-, or small diameter myelinated Adelta-, fibers.
RESULTS: The main suppressive effects on lumbar c-fos expression of isoflurane were observed in the superficial Lamina II (P = 0.02), whereas fentanyl showed the strongest effects in lamina V (P = 0.05).
They project to outer Lamina II of spinal cord, and form synapse with the secondary sensory neurons.
Lumbar intrathecal Derm-sap (500 ng) produced (1) partial loss of Lamina II MOR-expressing dorsal horn neurons, (2) no effect on MOR-expressing dorsal root ganglion neurons, and (3) no change in baseline tail-flick and hotplate reflex nocifensive responses.
In laminae I-II, approximately 65% were GluR1-positive and approximately 60% were GluR3-positive, while in Lamina III the corresponding values were 34% (GluR1) and 80% (GluR3). Puncta stained with antibody against the C-terminus of GluR4 (which only detects the long form of this subunit) made up 23% of the AMPAr-containing puncta in lamina I, approximately 8% of those in Lamina II and 46% of those in Lamina III.
We previously localized full-length trkB receptors on their terminals within Lamina II. In this preparation, BDNF (100-500 ng/mL) enhances the release of sensory neurotransmitters (glutamate, substance P, CGRP) in Lamina II by acting on trkB receptors expressed by primary afferent fibers of the peptidergic nociceptive type (PN-PAFs). A pre-synaptic mechanism was demonstrated after (i) patch-clamp recordings where the neurotrophin induced a significant increase in frequency, but not amplitude, of AMPA-mediated mEPSCs, (ii) real time calcium imaging, where sustained application of BDNF evoked an intense response in up to 57% Lamina II neurons with a significant frequency rise. This is the first demonstration that trkB receptors expressed by PN-PAF terminals in Lamina II are functional during postnatal development.
The results show that Ca(V)1.3-like immunoreactivity is widely distributed in all segments of the spinal cord but that the distribution in the different laminae of the spinal gray matter varies, with the highest density of labeled neurons in lamina IX and the lowest in Lamina II.
Nerve fibers immunolabelled for GFRalpha1 (GDNF receptor), GFRalpha2 (neurturin receptor), or GFRalpha3 (artemin receptor) showed distinct distribution patterns in the spinal cord, suggesting separate populations of sensory fibers with different functions: GFRalpha1-labeled fibers were in outer Lamina II and the lateral-collateral pathway and associated with autonomic interneurons and preganglionic neurons; GFRalpha2-labeled fibers were only in inner Lamina II; GFRalpha3-labeled fibers were in lamina I, the lateral-collateral pathway, and areas surrounding dorsal groups of preganglionic neurons and associated interneurons.
In this study, we performed whole-cell recordings in rat Lamina II neurons to record glycinergic spontaneous inhibitory postsynaptic currents (sIPSCs). The potentiating effect of capsaicin on sIPSCs was blocked by ionotropic glutamate receptor antagonists or tetrodotoxin in most Lamina II neurons examined.
In the TBSN, P2X(3) immunoreactivity was dispersed in the rostral TBSN but was dense in the superficial laminae of Vc, especially in the inner Lamina II.
In this study, we investigated the effects of EA on CNTF expression in the spared L(6) dorsal root ganglion (DRG), and spinal Lamina II at spinal segments L(3) and L(6) as well as nucleus dorsalis (ND) of L(3) spinal segment following removal of L(1)-L(5) and L(7)-S(2) (DRG) in the cat. After EA, their number significantly increased as early as 7 dpo in Lamina II of L(6) segment, and as late as 14 dpo in ND of L(3) segment.
Using whole-cell patch-clamp recordings from spinal cord slices of young (10-15 days old) rats, we have characterized and compared the properties of inhibitory synaptic transmission in Lamina II and laminae III-IV of the dorsal horn, which are involved in the processing of nociceptive and non-nociceptive sensory information, respectively. All (100%) of laminae III-IV neurons, but only 55% of Lamina II neurons, received both gamma-aminobutyric acid (GABA)ergic and glycinergic inputs. The remaining 45% of Lamina II neurons received only GABAergic synapses. Among the 55% of Lamina II neurons receiving both GABAergic and glycinergic inputs, all displayed a small proportion (approximately 10%) of mixed miniature inhibitory postsynaptic currents (mIPSCs), indicating the presence of a functional GABA/glycine co-transmission at a subset of synapses. The presence of mixed mIPSCs and the differences in decay kinetics of GABAA-type receptor mIPSCs between Lamina II and laminae III-IV were due to the endogenous tonic production of 3alpha5alpha-reduced steroids (3alpha5alpha-RS) in Lamina II. Our results indicate that, compared with Lamina II, inhibitory synaptic transmission in laminae III-IV is characterized by a dominant role of glycinergic inhibition and the absence of a functional GABA/glycine co-transmission..
The expression of RXRbeta in Lamina II neurons in the dorsal horn of transgenic and wild type (WT) animals was associated with extensive astrocyte staining in end-stage lumbar spinal cord from transgenic rats.
To determine their role, glycinergic spontaneous inhibitory postsynaptic currents (sIPSCs) were recorded in Lamina II neurons by using whole-cell recordings in spinal cord slices of wild-type (WT) and mAChR subtype knockout (KO) mice.
The activity of GABAergic inhibitory interneurones located in Lamina II of the spinal cord is of fundamental importance for the processing of peripheral nociceptive messages. Using patch-clamp recordings of Lamina II interneurones in the spinal cord of 15-20-day-old rats, we showed that synaptic inhibition mediated by GABA(A)Rs and its modulation by 3alpha5alphaNS in Lamina II of the spinal cord largely depend on activation of PKC. These findings confirmed that there was a significant production of endogenous 3alpha5alphaNS in Lamina II of the spinal cord but also indicated that PKC-dependent phosphorylation processes were tonically activated to control GABA(A)R-mediated inhibition under resting conditions.
It was noted that these inhibitory cells had axonal projections confined to Lamina II whereas excitatory vertical cells projected to lamina I and II.
Large neurons of the L6 DRG at 7 days post operation (dpo), and small to medium-sized neurons at 14 dpo, as well as in the Lamina II of the L6 spinal cord at 14 dpo was observed.
NT-3 immunoreactivity increased at 3 days post-operation (dpo), but decreased at 10 dpo in spinal Lamina II after ganglionectomy of L1-L5 and L7-S2 (leaving L6 DRG intact). It is possible that increased NT-3 in spinal Lamina II is derived from anterograde transport from small- and medium-sized neurons of L6 DRG, whereas decreased NT-3 immunoreactivity in the nucleus dorsalis is due to decreased transport of NT-3 from large neurons in the DRG at this time.
In the spinal cords of 10-month-old WT mice, ERbeta-positive cells were localized in Lamina II, whereas ERalpha-positive cells were mainly localized in lamina I.
In this study, we have compared the actions of 5alpha3alpha and minaxolone upon inhibitory transmission mediated by both GABA(A) and strychnine-sensitive GlyRs in Lamina II neurones of juvenile (P15-21) rats.
In addition, there was increased expression of protein kinase C gamma (PKCgamma) in cells outside of the inner region of Lamina II (IIi) in both strains after spinal contusion injury.
Here, we report that, in contrast to the nuclear localization of other transcription factors, DREAM showed a punctate staining pattern in rat spinal dorsal horn in immunofluorescent analysis, with a membrane localization profile in some neurons and its expression accumulated in the inner zone of Lamina II.
Although A-beta terminals centrally sprouted into Lamina II of the dorsal horn of the spinal cord, the peripheral A-beta fibers in the skin retracted from the epidermis to deeper layers of the dermis.
To determine whether the neurotrophin is effective in regulating the spontaneous release of the two neurotransmitters, we have recorded miniature inhibitory postsynaptic currents (mIPSCs) in Lamina II of post-natal rats. Our results thus describe a yet uncharacterized effect of BDNF in Lamina II, giving further strength to the notion that the neurotrophin plays an important role in pain neurotransmission..
The effects of 50% N2O on electrically evoked and spontaneous excitatory glutamatergic transmission and on the response to exogenous administration of N-methyl-d-aspartate (NMDA) and alpha-amino-3-hydroxy-5-methyl-4-isoxazole-4-propionic acid (AMPA) receptor agonists were examined in Lamina II neurons of adult rat spinal cord slices using the whole-cell patch-clamp technique.
Vertical, radial and tonic central Lamina II cells consistently expressed outward current to both NA and 5HT, but transient central and Substance P (SP)-insensitive lamina I cells were unaffected directly by either NA or 5HT.
This study evaluated the effect of EA on expression of neurotrophins in the Lamina II of the spinal cord, in cats subjected to dorsal rhizotomy. Significantly greater numbers of nerve growth factor (NGF) and neurotrophin-3 (NT-3) positive neurons, brain-derived neurotrophic factor (BDNF) immunoreactive varicosities and NT-3 positive neurons and glial cells were observed in Lamina II on the acupunctured (left) side, compared to the non-acupunctured, contralateral side.
We previously reported that the pronociceptive neurotrophin brain-derived neurotrophic factor (BDNF) induces facilitation of C-fiber evoked EPSCs and NMDA currents in Lamina II neurons of rats up to P40.
Substantia gelatinosa (SG, Lamina II) is a spinal cord region where most unmyelinated primary afferents terminate and the central nociceptive processing begins.
Using immunoelectron microscopy, we found that 3 weeks after unilateral sciatic nerve transection, the number of galanin-containing afferents was increased in ipsilateral Lamina II of monkey spinal cord. These results suggest that peripheral nerve injury induces an expansion of the central projection of galanin-containing afferents in Lamina II of the monkey spinal cord, not only by increasing galanin levels in primary afferents but also by triggering afferent branching..
By patch clamp recordings and correlative immunocytochemistry, we studied here the effect of 2 microM capsaicin-induced vanilloid receptor-1 (TRPV1) activation on IPSCs in spinal Lamina II neurons from post-natal mice. The peptide then excites inhibitory neurons in laminae I, III and IV, leading to an increased release of GABA/glycine in Lamina II via a parallel alternative pathway to glutamate..
The substantia gelatinosa (Lamina II) of the spinal dorsal horn contains inhibitory and excitatory interneurons that are thought to play a critical role in the modulation of nociception. However, the organization of the intrinsic circuitry within Lamina II remains poorly understood. We used glutamate uncaging by laser scanning photostimulation to map the location of neurons that give rise to local synaptic inputs to islet cells, a major class of inhibitory interneuron in Lamina II. Local synaptic inputs to islet cells arose almost entirely from within Lamina II, and these local inputs included both excitatory and inhibitory components.
To determine the changes in synaptic input to dorsal horn neurons and the GABAB)receptor function in streptozotocin-induced diabetes, we performed whole-cell recording (GDP-beta-S included in the internal solution) on Lamina II neurons in rat spinal cord slices.
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.
Single-pulse stimulation to the dorsal root elicited membrane excitation in Lamina II, and high-frequency pulse-train stimulation evoked long-lasting excitation that expanded widely in the dorsal horn.
Finally, application of fractalkine to spinal slices did not produce acute facilitation of excitatory synaptic transmission in Lamina II dorsal horn neurons, arguing against a direct action of fractalkine on spinal neurons.
The present study investigated whether morphine can promote regeneration and synaptic reconstruction of the terminals of injured primary afferent fibers in Lamina II of the spinal cord in rats following sciatic nerve injury. Fluoride-resistant acid phosphatase (FRAP)-positive terminals in Lamina II of the L4 spinal segment after sciatic nerve injury were assessed after treatment with vehicle, morphine, and naloxone plus morphine. Under the electron microscope, types I and II complex terminals of unmyelinated afferent fibers from the dorsal root, simple terminals of interneuronal axons, and terminals of descending axons at Lamina II of the L4 spinal segment were documented in the different groups after injury. FRAP-positive terminals in Lamina II were depleted after sciatic nerve injury in the vehicle group. The present study demonstrates that morphine may promote the regeneration and synaptic reconstruction of the terminals of injured primary unmyelinated afferent fibers in Lamina II of spinal cord, by a process mediated by mu-opioid receptors..
In dorsal horn, CGRP-positive fibers with and without H3R-LI ramified extensively in Lamina II; many of the former formed a plexus in lamina V.
In contrast to phasic inhibitory currents, using patch-clamp recording technique on spinal cord slices prepared from adult mice we revealed that tonic inhibitory currents were mediated by GABAA receptors but not by glycine receptors in dorsal horn Lamina II region.
Some of this input is relayed directly to supraspinal sites by projection neurons, whereas much of the input impinges on a heterogeneous population of interneurons in Lamina II. Previously, we demonstrated that G-protein-gated inwardly rectifying potassium (GIRK) channels are expressed in Lamina II of the mouse spinal cord and that pharmacologic ablation of spinal GIRK channels selectively blunts the analgesic effect of high but not lower doses of intrathecal mu-opioid receptor (MOR) agonists. Here, we report that GIRK channels formed by GIRK1 and GIRK2 subunits are found in two large populations of Lamina II excitatory interneurons. Ultrastructural analysis revealed that GIRK subunits preferentially label type I synaptic glomeruli, suggesting that GIRK-containing Lamina II interneurons receive prominent input from C fibers, while receiving little input from A delta fibers. Thus, excitatory interneurons in Lamina II of the mouse spinal cord can be subdivided into different populations based on the neurotransmitter system coupled to GIRK channels.
Enkephalin immunolabeling was detected within a single morphological subpopulation of nonpyramidal neurons located predominantly in Lamina II/III, 30% (33/109) of which were also GABA immunopositive. Axons immunolabeled for enkephalin were also abundant in Lamina II/III.
In this study, we determined how increased nociceptive inflow affects GABAergic spontaneous inhibitory postsynaptic currents (sIPSCs) of Lamina II neurons by using whole cell recordings in rat spinal cord slices. Additionally, bradykinin significantly inhibited sIPSCs in a population of Lamina II neurons and this inhibitory effect was also abolished by LY341495 and CPPG.
By applying 3-dimensional analysis of Golgi-Cox-stained material, we now demonstrate that corticosteroids can exert differential effects on dendritic arborizations of pyramidal neurons in Lamina II/III of the mPFC. Treatment with the glucocorticoid receptor-selective agonist dexamethasone and with the natural adrenosteroid, corticosterone (CORT), results in significant reductions in the total length of apical dendrites in the pyramidal neurons in Lamina II/III of the anterior cingulate/prelimbic and infralimbic cortices.
To test these hypotheses, we quantitatively compared active and passive membrane properties, firing patterns in response to depolarizing current steps and synaptic input of GABAergic neurons in spinal dorsal horn Lamina II of neuropathic and of control animals. Thus, a reduced membrane excitability, altered firing patterns or changes in synaptic input of this group of GABAergic neurons in Lamina II of the spinal cord dorsal horn are unlikely causes for neuropathic pain..
The numbers of Fos immunoreactive neurons in the spinal dorsal horn and in the medulla oblongata were significantly correlated mainly immediately after stimulation: lamina I correlated with the VLMlat, LRt, Sol and RVM; Lamina II correlated with the VLMlat, LRt and Sol; and laminae IV-V correlated with the VLMlat and LRt.
To investigate the mAChR subtypes involved in the inhibitory effect of mAChR agonists on glutamate release, evoked excitatory postsynaptic currents (eEPSCs) were recorded in Lamina II neurons using whole cell recordings in rat spinal cord slices.
The new procedure was applied successfully to a sample of 51 interneurons from Lamina II/III of the spinal dorsal horn..
Gal(1) protein, which could not be demonstrated in Gal(1) knockout mice, colocalized with VGLUT2 protein, but not with glutamate decarboxylase, in many nerve endings in Lamina II.
After 12-24 h, a significant increase in the number of small COX-2-containing neurons was observed in Lamina II on the injected side compared with the contralateral side. Furthermore, using fluorescent double-labeling for COX-2 and SP, an increase in the number of small COX-2-containing neurons in contact with SP-containing elements was observed ipsilaterally (1.4-1.6-fold compared with the contralateral side) in Lamina II. Using electron microscopy, two types of SP-containing axon terminals were found to form synapses with COX-2-containing neurons in Lamina II.
Peripheral nerve injury leads to the establishment of a novel synaptic connection between afferent Abeta-fiber and Lamina II neurons in spinal dorsal horn, which is hypothesized to underlie mechanical allodynia. In the present study, the role of protein tyrosine kinases (PTKs) in Abeta-fiber-evoked excitatory postsynaptic currents (EPSCs) recorded in Lamina II neurons in transverse spinal cord slices of rats was investigated using the whole-cell patch-clamp recording technique.
Co-localization of CB and CR was present in 0.3% of the cells and these cells were localized in Lamina II.
More than 90% of the dually labeled neurons were distributed in lamina I (marginal zone), less than 10% of them were located in Lamina II (substantia gelatinosa), and only a few (about 1%) were found in Lamina III (magnocellular zone).
In the spinal dorsal horn, synaptophysin-immunolabeling was weak in the afferent fibers in lamina I, outer Lamina II and the dorsal part of inner Lamina II, but strong in the afferent fibers in laminae III-IV. However, a subpopulation of isolectin B4-positive small dorsal root ganglion neurons expressed both synaptoporin and synaptophysin, and their afferent fibers were mainly distributed in the ventral part of inner Lamina II.
This study evaluated the plastic changes of c-jun and c-fos in the right sixth lumbar dorsal root ganglion (L6 DRG), Rexed's Lamina II in representative spinal segments L3, L5, and L6 and in the nucleus dorsalis (ND) at L3 segments after electro-acupuncture (EA) in cats subjected to removal of L1-L5 and L7-S2 DRG. Following dorsal root ganglionectomy, there was a significant increase in the density of c-jun immunoreactivity in the neurons and glia in spinal Lamina II and in the ND; there was also marked elevation in the expression of c-fos in ND. After EA in the operated animals, there was an up-regulation in the expression of c-jun in the L6 DRG and the associated spinal Lamina II; however, increased c-fos expression was detected only in the L6 DRG.
neurons in the olfactory epithelium and vomeronasal organ, olfactory glomeruli, possibly amacrine cells, neurons in the ganglion cell layer and three bands in the inner plexiform layer of the retina, axons in the optic nerve and tract up to the superior colliculus, inner and outer hair cells and the spiral ganglion cells in the cochlea, vestibulocochlear nerve bundle, spinal trigeminal tract, and Lamina II of the spinal cord.
In the spinal cord dorsal horn, GFP positive neurons were largely restricted to lamina I and to a lesser extent Lamina II, but surprisingly did not coexpress markers for key neuronal populations present in the superficial dorsal horn.
We present some of our recent results concerning inhibitory synaptic transmission in Lamina II of the spinal cord and show that endogenous 5alpha-reduced neurosteroids are produced locally in Lamina II and modulate synaptic gamma-aminobutyric acid A(GABAA) receptor function during development, as well as during inflammatory pain.
Rexed's Lamina II was densely immunostained for calbindin D-28k, whereas, in the anterior horn, calbindin-D-28k-positive small neurons were barely dispersed in a scattered pattern.
At all levels of the spinal cord dorsal horn, HCN1 immunoreactivity (HCN1-IR) was predominantly absent from laminae I and II, while a dense band of punctate labeling was visible in Lamina III. Labeled neurons were identified in close vicinity to the central canal, in the lateral spinal nucleus, in the ventral horn and occasionally in Lamina II and III.
We show that AMPA receptor activation enhances spontaneous release of inhibitory amino acids in the presence of tetrodotoxin onto both Lamina II neurons and NK1 receptor-expressing (NK1R+) lamina I neurons.
At least seven distinct Y1 receptor-positive populations could tentatively be recognized: Type 1) abundant small, fusiform Y1 receptor-positive neurons in laminae I-II, producing a profuse neuropil; Type 2) Y1 receptor-positive projection neurons in lamina I; Type 3) small Y1 receptor-positive neurons in Lamina III, similar to Type 1 neurons, but less densely packed; Type 4) a number of large, multipolar Y1 receptor-positive neurons in the border area between laminae III-IV, with dendrites projecting toward laminae I-II; Type 5) a considerable number of large, multipolar Y1 receptor-positive neurons in laminae V-VI; Type 6) many large Y1 receptor-positive neurons around the central canal (area X); and Type 7) a small number of large Y1 receptor-positive neurons in the medial aspect of the ventral horns (lamina VIII). The Lamina II neurons express somatostatin [ The neuropeptide Y Y1 receptor is a somatic receptor on dorsal root ganglion neurons and a postsynaptic receptor on somatostatin dorsal horn neurons.
We found that preprotachykinin B-immunoreactive neurons were present throughout laminae I-III, constituting 10-11% of the neuronal population in laminae I-II, and 4% of that in Lamina III. They formed a prominent band in the ventral half of Lamina II (where they made up 16% of the population) and the dorsalmost part of Lamina III. By examining expression of Fos protein in response to intraplantar injection of formaldehyde we provide evidence that many of the preprotachykinin B cells in lamina I and the outer part of Lamina II respond to noxious stimulation..
METHODS: The effects of midazolam on electrically evoked and spontaneous excitatory transmission were examined in Lamina II neurons of adult rat spinal cord slices using the whole cell patch clamp technique.
Whole-cell voltage-clamp recordings were performed on Lamina II neurones in the rat spinal cord slices.
To determine the role of individual mAChR subtypes in control of synaptic GABA release, spontaneous inhibitory postsynaptic currents (sIPSCs) and miniature IPSCs (mIPSCs) were recorded in Lamina II neurons using whole-cell recordings in spinal cord slices of wild-type and mAChR subtype knockout (KO) mice.
Here, we show that during the development of thermal hyperalgesia and mechanical allodynia associated with inflammatory pain, synaptic inhibition mediated by GABAA receptors in Lamina II of the DH was in fact markedly increased.
In the spinal cord, IB4 and CGRP colocalized in fibers and terminals in the inner part of Lamina II, the lateral collateral path, and the sacral parasympathetic nucleus (SPN). After injection of IB4 into the bladder wall, immunoreaction for IB4 was detected in SPN, but not in Lamina II.
Spinal neurons are the main source of trkB in Lamina II (substantia gelatinosa). Ultrastructurally, full-length trkB (fl-trkB) receptors were present at somato-dendritic membranes of Lamina II neurons (rat: 66.8%; mouse: 73.8%) and at axon terminals (rat: 33.2%; mouse: 26.2%). This is the first ultrastructural description of fl-trkB localization at synapses between first- and second-order sensory neurons in Lamina II, and suggests that BDNF may be released by fl-trkB-immunopositive PAFs to modulate nociceptive input in this lamina of dorsal horn..
OBJECTIVE: To explore the changes of the expression of NT-4 in spared dorsal root ganglia (DRG,L6) on both the operation/Acup side and the nonoperation/non-Acup side as well as in the spinal Lamina II (L3, L5, L6) and Clarke' nucleus (L3) of the normal adult cats, partial dorsal rhizotomy cats, and Acup spared DRG cats so as to disclose the relation between NT-4 and the plasticity of spinal cord as well as the Acup promoting spinal cord plasticity. The distribution and the number of NT-4 immunoreactive neurons in bilateral spared DRG (L6) on the operation/Acup side and the nonoperation/Acup side as well as in the, spinal Lamina II (L3, L5, L6) and Clarke' nucleus (L3) of each cat were oberserved and counted. In addition, Acup increased NT-4 expression in L5, L6 spinal Lamina II.
Immunoperoxidase labeling with ENT1 antibodies produced specific staining in dorsal horn which was concentrated over superficial laminae, especially the substantia gelatinosa (Lamina II). Electron microscopy analysis of ENT1 expression in Lamina II indicated its presence within pre- and post-synaptic elements, although a number of other structures, including myelinated and unmyelinated, axons were also labeled.
Pharmacological activation of group I mGluRs leads to long-term depression (LTD) of synaptic strength between Adelta-fibers and neurons in Lamina II of spinal dorsal horn of the rat. Synaptic strength between Adelta-fibers and Lamina II neurons was assessed by perforated whole-cell patch-clamp recordings in a spinal cord-dorsal root slice preparation of young rats.
Using immunohistochemistry, we found that Kv 4.2 and Kv 4.3 immunoreactivity was concentrated in the superficial dorsal horn, mainly in Lamina II.
In an attempt to clarify the mechanism underlying the regulation of the release of substance P (SP) from the central axon terminals of the synaptic glomeruli in Lamina II of the dorsal horn, we examined the expression patterns of delta and mu opioid receptors (DOR and MOR) in relation to those of enkephalin (ENK) and SP in the synaptic glomeruli. DOR, MOR, ENK and SP immunoreactivities in Lamina II of the dorsal horn in the chicken were examined by confocal laser scanning and electron microscopies.
We found that interneurons of Lamina II are at the origin of the major ascending circuits targeted by the nonpeptide nociceptors.
The two nerve injury models also reduce the postsynaptic potassium channel opening action of DAMGO on Lamina II spinal cord neurons, but again only in segments receiving injured afferent input.
GABAergic inhibitory interneurons are among the neurons lost, and a marked decrease in inhibitory postsynaptic currents of Lamina II neurons coincides with the induction of apoptosis. Preventing nerve injury-induced apoptosis of dorsal horn neurons by blocking caspase activity maintains inhibitory transmission in Lamina II and reduces pain hypersensitivity..
Electron microscopic examination of the dorsal horn revealed three main localizations: 1) a postsynaptic localization on peptidergic cell bodies in laminae I-III and in numerous dendrites; 2) a presynaptic localization on unmyelinated and thin myelinated peptidergic fibers (two types of axon terminals are observed, large ones, presumably of primary afferent origin, and smaller ones partially from intrinsic cells; this presynaptic labelling represents 60% and 22% of total labelling in laminae I and II, respectively); and 3) 16.9% of labelling in lamina I and 19.8% in Lamina II are observed in astrocytes.
Under chronic conditions of neuropathic pain, nociceptive C terminals are lost from their target region in spinal Lamina II, leading to reduced thermal hyperalgesia.
This procedure resulted in more than an 80% reduction in the expression of both NR1 mRNA and protein and a corresponding loss of NMDA, but not AMPA currents, in the Lamina II neurons in the injected area.
Moreover, dense reaction product was found in the most medial aspect of Lamina II, especially Lamina II inner part, and less in Lamina III and IV of levels L3-L5.
In the current study, the authors investigated the effects of remifentanil hydrochloride, with and without its vehicle, glycine, on the activation of NMDA receptors and the modulation of NMDA-induced current on neurons inside the Lamina II from the dorsal horn of rat spinal cord.
Antiserum against the P2X3-receptor subtype showed P2X3- receptor immunoreactivity in axon-like structures of the cervix, in small and medium-sized neurons in the L6/S1 DRG, and in Lamina II of the L6/S1 spinal cord segments.
We previously reported that brain-derived neurotrophic factor (BDNF), a pronociceptive neurotransmitter, induces synaptic facilitation of excitatory postsynaptic current (EPSC) in Lamina II neurons of neonatal rats up to P14 in a N-methyl-d-aspartate (NMDA) receptor-dependent manner. After contusion injury, BDNF was unable to facilitate dorsal root-evoked EPSCs in Lamina II neurons despite the finding that NMDA-evoked currents were only slightly smaller than those observed in age-matched uninjured animals.
Lamina II central neurons, with dorsal root (DR) C-fiber input, monosynaptically excited Lamina II vertical neurons with DR Adelta input. Lamina II outer vertical neurons with DR Adelta input monosynaptically excited lamina I neurons. Together, these observations indicate that the neural circuitry in the SDH, including its substantia gelatinosa (Lamina II), has an explicit organization in which particular combinations of neurons comprise modules arranged to modify and transmit sensory information arriving from Adelta and C primary afferent fibers..
At the light microscopic level, CB immunoreactivity was observed most intensely in the Lamina II using the avidin-biotinylated peroxidase complex (ABC) and immunofluorescence methods.
OBJECTIVE: To explore the changes in expression of Substance P (SP) in spinal Lamina II at different time following hemisected spinal cord injury (hSCI). The expression of SP was measured by immunohistochemical ABC method, and the number of SP positive varicosities in Lamina II was counted. RESULTS: SP positive varicosities were observed in spinal Lamina II.
In the dorsal horn, these fibers ascended into and then terminated in Lamina II. Some terminals made synaptic contact with dendritic profiles in Lamina II.
OBJECTIVE: This study was conducted to detect the expression levels of NT-4 in the spared dorsal root ganglia (DRG, L6) on the operated side and un-operated side, in the spinal Lamina II (L3, L5, L6) and Clarke's nucleus (L3) of the normal adult cats and the cats submitted to partial dorsal rhizotomy. The distribution and the number of NT-4 positive neurons in bilateral spared DRG (L6) in the operated/un-operated side, spinal Lamina II (L3, L5, L6) and Clarke's nucleus (L3) of each animal were observed and counted. (2) There was no difference in respect to the number of NT-4 positive neuron in L3, L5, L6 spinal Lamina II and L3 Clarke's nucleus in the operated side between the normal group, 7-day operation group and 14-day operation group (P>0.05).
OBJECTIVE: To explore the temporospatial changes of IGF-I expression in the spared dorsal root ganglia (DRG, L6) on the operated side and un-operated side, in the spinal Lamina II (L3, L5, L6) and Clarke's nucleus (L3) of the adult cats that have undergone partial dorsal rhizotomy, and compare them against those of the normal adult cats so as to unveil the relation between IGF-I and the plasticity of spinal cord. (2) There was no difference in number of IGF-I positive neuron in L3, L5, L6 spinal Lamina II between normal group, 7th day post-operation group and 14th day post-operation group (P>0.05).
AMPA was found as the most potent agonist tested producing market production of c-Fos particularly in neurons of Lamina II at doses of 10 pM per 10-microl injection.
METHODS: The effects of a clinically relevant concentration (1 rat minimum alveolar concentration [ MAC]) of isoflurane on electrically evoked and spontaneous excitatory/inhibitory transmission and on the response to exogenous administration of the gamma-aminobutyric acid type A receptor agonist muscimol were examined in Lamina II neurons of adult rat spinal cord slices using the whole cell patch clamp technique.
To determine the role of receptor subtypes in the muscarinic agonist-induced synaptic GABA release, spontaneous inhibitory postsynaptic currents (sIPSCs) were recorded in Lamina II neurons using whole-cell voltage-clamp recordings in rat spinal cord slices.
Previous work suggested that neurons in the inner part of Lamina II (IIi), onto which IB(4)-positive sensory neurons project, facilitate nociceptive transmission following tissue or nerve injury.
For both picrotoxin and strychnine, the increase in Fos-like immunoreactivity peaked in lamina V (at 3579+/-319 and 3649+/-375% of control, respectively; mean+/-SEM) but for picrotoxin an additional peak was observed in the outer part of Lamina II (1959+/-196%).
Substance P (SP) is a well-established pain messenger in the spinal cord, although its role in substantia gelatinosa (Lamina II) still remains elusive. We carried out patch-clamp recordings on Lamina II neurons from transverse mouse spinal cord slices (P8-12), using the selective NK1 receptor agonist [ Sar9,Met(O2)11]-SP (SM-SP, 3-5 microM) in the presence of NBQX.
Within the nociceptive afferent pathway to Lamina II, nonpeptidergic C-fiber synapses in the deeper half of Lamina II (IIi) contain E-cadherin but mostly lack N-cadherin, whereas the majority of the peptidergic C-fiber synapses in the outer half of Lamina II (IIo) contain N-cadherin but lack E-cadherin. Approximately one-half of the Abeta-fiber terminations in Lamina III contain N-cadherin; none contain E-cadherin.
We found extensive sprouting of uninjured WGA-HRP-labeled afferents into the central termination field in Lamina II of dorsal horn normally occupied by L5 afferents whose peripheral axons had been ligated distal to the dorsal root ganglion.
The staining for TRPV1 was found in axon collaterals in the dorsal parts of Vp, Vo, and Vi and in terminals and fibers throughout lamina I and the outer zone of Lamina II (IIo) of Vc.
Immunohistochemistry showed that EGFP-expressing neurones accounted for about one-third of the GABAergic neurones in Lamina II of the spinal dorsal horn. The whole-cell patch-clamp technique was used to intracellularly label and physiologically characterize EGFP- and non-EGFP-expressing Lamina II neurones in spinal cord slices. In conclusion, EGFP expression defined a substantial but, with respect to the measured parameters, rather inhomogeneous subgroup of GABAergic neurones in spinal Lamina II. These results provide a base to elucidate the functional roles of this subgroup of GABAergic Lamina II neurones, e.g.
Colchicine pretreatment allows the staining of numerous cell bodies in Lamina II.
Pyramidal neurons in Lamina II-III of medial prefrontal cortex were drawn in three dimensions, and the morphology of apical and basilar arbors was quantified.
Three weeks after the treatment, the area in Lamina II of the L6 spinal cord stained with IB4 was significantly reduced compared with the area stained in control rats. These results indicate that intrathecal treatment with the IB4-saporin conjugate at the level of L6-S1 spinal cord, which reduces IB4 afferent nerve terminal staining in Lamina II of the L6 spinal cord as well as the number of IB4-binding neurons in L6 DRG, suppressed bladder overactivity induced by bladder irritation without affecting normal micturition.
Light and electron microscopic analysis of the spinal cord revealed transganglionically transported lectin in unmyelinated axons in the dorsolateral funiculus and axon terminals concentrated mainly within Lamina II of the dorsal horn.
CTb-labeling was detected in regions known to receive input from myelinated sciatic afferents: lamina I and a band extending from the inner part of Lamina II (IIi) to lamina V in the L3-5 segments, and the deepest part of the dorsal horn in L2. Importantly, no CTb-labeling was detected in the outer part of Lamina II (IIo) in the denervated areas. Again, CTb-immunoreactivity showed the normal sciatic pattern in L4-L5, with no labeling detected in Lamina IIo in the denervated region. These results do not support the suggestion that the central terminals of intact myelinated afferents sprout into regions of Lamina II occupied by adjacent nerves that have been axotomized peripherally..
In the dorsal horn of the spinal cord, HCN-1, HCN-2, and HCN-4 were all expressed in laminae I-IV, although HCN-1 was not detectable in Lamina II.
Ten days after nerve ligation, there was a 2.46+/-0.38 fold ( p<0.05) bilateral increase in nociceptin immunoreactivity in the lamina superficiales (I and II), with a notable increase in the inner Lamina II at the level of L4.
Aside from somatic motor neurons and autonomic preganglionic neurons choline acetyltransferase-immunoreactivity was found throughout the spinal cord in Lamina III of the superficial dorsal horn and near the central canal. A dense band of NADPH diaphorase staining was found in Lamina II and in centrally located neurons of all segments.
Here we show that in rats rendered tolerant to the analgesic action of morphine, cross-tolerance to the analgesic action of intrathecally administered [ d-Ala2,N-MePhe4,Gly-ol5]enkephalin (DAMGO) can be produced without changes in the magnitude of DAMGO-induced internalization of the MOR in Lamina II neurons of the rat spinal cord.
Previous in vitro studies have shown that muscarinic agonists increase GABA release and reduce the glutamatergic synaptic input to Lamina II interneurons through GABAB receptors in the spinal cord.
RESULTS: CB-, CR-, PV-, GABA- and glycine-like immunoreactive (LI) neurons were differentially observed in all layers of the MDH, but particularly in Lamina II. Neurons that exhibited immunoreactivity for both CaBPs and GABA or glycine were also observed mainly in Lamina II.
Rofecoxib (100 microM) did not affect spontaneous excitatory postsynaptic currents or alpha-amino-3-hydroxy-5-methyl-4-isoxazole propanoic acid (AMPA) and N-methyl-d-aspartate (NMDA) responses in Lamina II neurons from spinal cords of animals with SNI indicating no detectable action on transmitter release or postsynaptic activity.
Morphological changes in the ipsilateral L4-L5 Lamina II consisted of: (i) cell loss (38 +/- 5%), (ii) increased TUNEL-positive profiles, (iii) decreased SP-immunoreactive primary afferents, and (iv) reactive gliosis. This study shows that: (a) spinal cord neuron loss may be triggered by a p53- and E2F1-independent apoptosis in Lamina II with the participation of glutamate mGlu5 receptors, (b) these receptors seem to be involved transiently, as their blockade was no longer protective by 7 days CCI, and (c) this delayed cell death occurred in the absence of Bax activation, suggesting the involvement of an alternative death pathway..
The densest VRL-1 immunoreactivity in the spinal cord was found in lamina I, inner Lamina II, and laminae III/IV.
epsilonPKC expression in dorsal root ganglion neurons and gammaPKC expression in Lamina II of the spinal cord increased from the first to the third postnatal week.
In Lamina II of the spinal dorsal horn, synaptic inhibition mediated by ionotropic GABA(A) and glycine receptors contributes to the integration of peripheral nociceptive messages. Whole-cell patch-clamp recordings were performed from Lamina II neurons in spinal cord slices to study the properties of miniature IPSCs (mIPSCs) mediated by activation of GABA(A) and glycine receptors in immature (<30 d) and adult rats.
Spinal Lamina II (substantia gelatinosa) neurons play an important role in processing of nociceptive information from primary afferent nerves. Anatomical studies suggest that neurons in the outer (Lamina II(o)) and inner (Lamina II(i)) zone of Lamina II receive distinct afferent inputs. The functional significance of this preferential afferent termination in Lamina II remains unclear. In this study, we examined the differential synaptic inputs to neurons in Lamina II(o) and II(i) in response to primary afferent stimulation. Whole cell voltage-clamp recordings were performed on neurons in Lamina II(o) and II(i) of the rat spinal cord slice under visual guidance. Capsaicin (1 microM) significantly increased the frequency of glutamatergic miniature excitatory postsynaptic currents (mEPSCs) in all 27 Lamina II(o) neurons and significantly increased the amplitude of mEPSCs in 12 of 27 Lamina II(o) neurons. However, capsaicin only significantly increased the frequency of mEPSCs in 9 of 22 (40.9%) Lamina II(i) neurons and increased the amplitude of mEPSCs in 6 of these 9 neurons. Furthermore, the peak amplitude of EPSCs, evoked by electrical stimulation of the attached dorsal root, in 40 Lamina II(o) neurons was significantly greater than that [ 160.5 +/- 16.7 vs. 87.0 +/- 10.4 (SE) pA] in 37 Lamina II(i) neurons. On the other hand, the peak amplitude of evoked inhibitory postsynaptic currents (IPSCs) in 40 Lamina II(o) neurons was significantly smaller than that (103.1 +/- 11.6 vs. 258.4 +/- 24.4 pA) in 37 Lamina II(i) neurons. In addition, the peak amplitudes of both EPSCs and IPSCs, evoked by direct stimulation of Lamina II, were similar in Lamina II(o) and II(i) neurons. This study provides new information that stimulation of primary afferents differentially potentiates synaptic inputs to neurons in Lamina II(o) and II(i). The quantitative difference in excitatory and inhibitory synaptic inputs to Lamina II(o) and II(i) neurons may be important for integration of sensory information from primary afferent nerves..
Intraneural injection of cholera toxin B subunit, a transganglionic tracer, into the injured sciatic nerve of wild-type mice revealed that the myelinated Abeta-fiber primary afferents sprouted from laminae III-VI of the dorsal horn of the spinal cord and invaded Lamina II.
We found that each of the sciatic branches targets a distinct mediolateral location in inner Lamina II and that each of the spared nerve injury models produced a more reproducible pattern of thiamine monophosphatase staining loss than did partial tight ligation.
Here, we investigate the effect of BDNF on dorsal root-evoked synaptic transmission in Lamina II neurons. A significant minority of cells was minimally affected by BDNF and consistent with this, not all neurons in Lamina II were immunoreactive for the tyrosine kinase (trk) B receptor.
We examined the effects of corticotropin-releasing factor (CRF) on plasticity of optically recorded neuronal activity in the substantia gelatinosa (Lamina II) of 12-18-day-old rat spinal cord slices stained with a voltage-sensitive dye. Single-pulse test stimulation to the dorsal root that activated A and C fibres evoked prolonged (>100 ms) light-absorption change in the Lamina II. Although interneuronal actions might contribute, these results suggest that CRF may have dual effects on excitatory synaptic transmission within the Lamina II depending upon cellular conditions: a conversion from the induction of long-term depression to long-term potentiation (LTP), and inhibition of LTP induction.
In the lumbar cord dorsal horn, dense immunoreactivity was seen in the inner part of Lamina II. The results indicate that most of the adenosine A1 receptors in the dorsal horn are located in inner Lamina II postsynaptic neuronal cell bodies and processes whose functional and neurochemical identity is so far unknown.
The central arborizations of large myelinated cutaneous afferents normally extend as far dorsally as the ventral part of Lamina II in rat spinal cord. (1992) reported that after nerve injury some of these afferents sprouted into lamina I and the dorsal part of Lamina II, and it has been suggested that this could contribute to allodynia associated with neuropathic pain. Both normal and axotomized populations included axons with hair follicle afferent-like morphology and arbors that entered the ventral half of Lamina II; however, none of these extended farther dorsally. We observed that both ipsilateral and contralateral to the sectioned nerve, arbors of axons with hair follicle afferent-like morphology in the sciatic territory extended only as far as the ventral half of Lamina II.
PURPOSE: To clarify the relationship between allodynia and the sprouting of myelinated fibers, we examined whether the administration of nerve growth factor (NGF) affected the paw withdrawal response to non-noxious mechanical stimuli and the sprouting of myelinated fibers into Lamina II of the spinal dorsal horn, using a chronic constriction injury model of the sciatic nerve. RESULTS: With vehicle infusion, significantly increased responsiveness to mechanical stimuli was observed on postoperative days (PODs) 5, 7, and 14 after ligation, compared with before surgery, and B-HRP-positive fibers were newly localized in Lamina II on PODs 7 and 14. Infusion of NGF reduced the responsiveness to mechanical stimuli on 5, 7, and 14 PODs and B-HRP-positive fibers in Lamina II on PODs 7 and 14. CONCLUSION: We propose that the suppression of the increased responsiveness to mechanical stimuli produced by NGF could be related to the disappearance of B-HRP-positive fibers in Lamina II..
The 25 microl CFA group was also the only group that displayed a significant expansion of the sciatic and saphenous nerve terminal field in Lamina II of the dorsal horn at 8 weeks, using wheat-germ agglutinin-horse radish peroxidase transganglionic labelling. Lower dose CFA caused an acute, reversible expansion of terminal fields in Lamina II in neonatal animals, while CFA did not produce this effect in adults. The expansion of afferent terminals in Lamina II following neonatal CFA inflammation is maintained into adulthood if the inflammation is also maintained, as seen following 25 microl CFA.
The immunohistochemistry results indicated that GABA receptor/channel rho 1 subunits were expressed in mouse spinal cord superficial dorsal horn (lamina I and Lamina II) and in DRG.
Punctate labelling resembling that of axonal fibres and terminals was evident in Lamina II of the dorsal horn and throughout the cord.
A role of ovarian hormones was also suggested after immunostaining of oestrogen receptors in the Lamina II of Caudalis subnucleus of the trigeminal sensory complex and cervical (C1-C2) spinal dorsal horn.
NR1 subunit mRNA and dendritic protein are reduced by 80% in the area of the virus injection, and NMDA currents, but not AMPA currents, are reduced 86-88% in Lamina II neurons.
These neurons projected to the inner portion of Lamina II in the dorsal horn of spinal cord and the dermis of skin.
Using spinal cord slice preparations and patch-clamp recordings in Lamina II and lamina V regions, we tested a hypothesis that P2X receptor subtypes differentially modulate glutamate release from primary afferent terminals innervating different sensory regions. In Lamina II recordings, >70% of the modulation was transient. Transient modulation was not observed in the presence of 1 microM trinitrophenyl-ATP (TNP-ATP), a subtype-selective P2X receptor antagonist, suggesting that homomeric P2X3 receptors may be involved in the transient modulation in Lamina II.
Segmental distribution and rostrocaudal central level of dorsal root ganglion (DRG) neurons innervating reference points were examined and DiI-induced fluorescent areas were mapped in the horizontal plane through Lamina II of the dorsal horn.
In OFC of the perinatal human brain, indication of prominent anastomosis formation in the upper layers (Lamina II and III) is observed.
Vacuolation in area V1 was most severe in Lamina III and the glial cell reaction in lamina V or VI. Surviving neurons were most abundant in Lamina II or III, whereas prion protein deposition either affected all laminae equally or was maximal in Lamina II or III.
In RTX-treated rats, IB(4)-labeled terminals in the dorsal horn were significantly reduced, and CTB-labeled terminals appeared to sprout into Lamina II of the spinal dorsal horn. The delayed tactile allodynia induced by RTX is likely attributable to damage to myelinated afferent fibers and their abnormal sprouting in Lamina II of the spinal dorsal horn.
The ventral or inner region of spinal substantia gelatinosa (SG; Lamina II(i)) is a heterogeneous sublamina important for the generation and maintenance of hyperalgesia and neuropathic pain.
Nitrobenzylthioinosine (NBMPR), a potent non-transportable inhibitor of rENT1, attenuated synaptically evoked EPSCs onto Lamina II neurons in a concentration-dependent manner.
The numbers of positive neuron for NT3 and it's mRNA in large, medium, small neuron of L6 DRG and the numbers of positive neurons and glia cells for NT3 in Lamina II were counted respectively. RESULTS: The numbers of positive large, small neurons for NT3 and its mRNA in DRG and the number of positive neurons and glia cells for NT3 in Lamina II on the acupuncture side increased apparently than those on the non-acupuncture side (P < 0.05). However, the positive signal of NT3 mRNA in Lamina II was not seen in our study. CONCLUSION: The results indicate that acupuncture promoting the plasticity of spinal cord involves both the increase in expression of NT3 in large and small neurons of spared DRG and the increase in number of NT3 positive neurons and glia cells in spinal Lamina II.
Neuronal cell bodies exhibiting PPE-like immunoreactivity were present in all laminae of the Vc, with a higher concentration in Lamina II. Most of the CB-, CR- and PV-like immunoreactive neurons were located in Lamina II, and some of them were also found in laminae I and III of the Vc. CB/PPE, CR/PPE and PV/PPE double-labelled neurons were mainly observed in Lamina II.
Using the blind whole cell patch-clamp recording technique, we investigated peripheral nerve injury-induced changes in excitatory synaptic transmission to neurones in Lamina II of the dorsal horn. sciatic nerve transection (SNT)) peripheral nerve injury altered the mean threshold intensity for eliciting A fibre-mediated EPSCs in Lamina II neurones. In spinal cord slices from naive rats, polysynaptic A beta fibre-evoked EPSCs were observed in 24 % of Lamina II neurones, monosynaptic A delta fibre EPSCs were observed in 34 % and polysynaptic A delta fibre EPSCs were observed in 7 %.
CTb labelling was seen in the same areas on the lesioned side, but with a dramatic increase in Lamina II.
By using ABC immunohistochemistry and in situ hybridization techniques, the dynamic changes of the above three factors and their mRNA in spinal Lamina II of different segments were analysed.
The released NA might take part in the modulation of nociceptive transmission through the following pathways: (1) inhibits the glutamate and substance P release from primary afferent terminals; (2) increases the release of inhibitory neurotransmitters from Lamina II (substantia gelatinosa) neurons.
VGluT1 immunoreactivity (IR) was intense in the inner part of Lamina II but weak in lamina I and the outer part of Lamina II. VGluT2-IR was most intense in lamina I and the outer part of Lamina II. The dual immunofluorescence histochemistry indicated expression of VGluTs in substance P (SP)-containing axon terminals in lamina I and the outer part of Lamina II. It was further observed that isolectin IB4, a marker for unmyelinated axons, often bound with VGluT2-immunopositive structures but rarely with VGluT1-immunopositive structures in Lamina II. Thus, the results indicated in laminae I and II of the medullary and spinal dorsal horns that both VGluT1 and VGluT2 were expressed in axon terminals of primary afferent fibers, including SP-containing nociceptive fibers and that VGluT in unmyelinated primary afferent fibers terminating in Lamina II was primarily VGluT2..
Therefore, we have used an in vitro spinal cord slice preparation to study the effects of GBP on Lamina II neurons.
The most prominent alpha(1B)-subunit upregulation was found in the outer as well as the inner part of Lamina II (II(o), II(i)), extending from the medial toward the lateral region of the L4 and L5 spinal segments.
Previous studies have shown sprouting of Abeta-fibres into Lamina II of the spinal cord dorsal horn after nerve injury and the formation of new synapses by these sprouts.
Injured neurons with C-fibers exhibit transganglionic degeneration of their terminations within Lamina II of the spinal cord dorsal horn, while peripheral nerve injury of medium to large neurons induces collateral sprouting of myelinated A-fibers from lamina I and III/IV into Lamina II in rats, cats, and primates. In all animals, 2-5 weeks after nerve transection (treated or otherwise), IB4- and SP-ir is absent from Lamina II. Animals without nerve cap treatment exhibited robust fiber sprouting into Lamina II at 2 weeks. In sharp contrast, animals treated with silicone caps did not exhibit betaHRP-ir fibers in Lamina II at 2 weeks.
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