Calcitonin gene-related peptide-receptor component protein expression in the uterine cervix, lumbosacral spinal cord, and dorsal root ganglia

The neuropeptide calcitonin gene-related peptide (CGRP) may play a role in neurogenic inflammation, tissue remodeling of the uterine cervix, promoting vasodilation, parturition, and processing of sensory information in the spinal cord. CGRP-immunoreactive nerves of the cervix and spinal cord have been studied but cellular identification of the CGRP receptor has received little attention. CGRP-receptor component protein (CGRP-RCP) is a small protein associated with the CGRP receptor; thus, immunostaining for the CGRP-RCP can be used to identify sites of the CGRP receptor. We determined sites of CGRP-RCP immunoreactivity relative to the presence of CGRP-ir nerve fibers in the female rat uterine cervix, spinal cord, and dorsal root ganglia. CGRP-RCP immunoreactivity was expressed in the dorsal horn of the spinal cord, venules of the uterine cervix, and perikarya of sensory neurons in dorsal root ganglia. CGRP-immunoreactive fibers were adjacent to CGRP-RCP-immunoreactive vessels in the cervix and among CGRP-RCP-immunoreactive structures in the dorsal horn of the spinal cord. This suggests CGRP-RCP is associated with structures innervated by CGRP nerves and these interactions may be changed in tissues in response to an appropriate stimulus.     

Protein associated with Myc (PAM) is involved in spinal nociceptive processing

PAM (protein associated with Myc) is a potent inhibitor of adenylyl cyclases (ACs) which is primarily expressed in neurones. Here we describe that PAM is highly expressed in dorsal horn neurones and motoneuron of the spinal cord, as well as in neurones of dorsal root ganglia in adult rats. PAM mRNA expression is differentially regulated during development in both spinal cord and dorsal root ganglia of rats, being strongest during the major respective synaptogenic periods. In adult rats, PAM expression was up-regulated in the spinal cord after peripheral nociceptive stimulation using zymosan and formalin injection, suggesting a role for PAM in spinal nociceptive processing. Since PAM inhibited Galphas-stimulated AC activity in dorsal root ganglia as well as spinal cord lysates, we hypothesized that PAM may reduce spinal nociceptive processing by inhibition of cAMP-dependent signalling. Accordingly, intrathecal treatment with antisense but not sense oligonucleotides against PAM increased basal and Galphas-stimulated AC activity in the spinal cord and enhanced formalin-induced nociceptive behaviour in adult rats. Taken together our findings demonstrate that PAM is involved in spinal nociceptive processing.     

Reduced numbers of calcitonin gene-related peptide-(CGRP-) and tachykinin-immunoreactive sensory neurones associated with greater enkephalin immunoreactivity in the dorsal horn of a mutant rat with hereditary sensory neuropathy

Summary The mutilated foot rat is a mutant with autosomal recessive sensory neuropathy and frequent mutilation of the hindlimbs. Decreased numbers of dorsal root ganglion cells and diminished sensitivity to painful stimuli are characteristics of these animals. By use of immunocytochemistry, changes in the distributions of peptides involved in sensory and/or autonomic regulation, i.e. calcitonin generelated peptide (CGRP), tachykinins, enkephalin and neuropeptide Y in spinal cord, dorsal root ganglia and skin of these animals, were studied. In comparison with normal litter-mate controls, the dorsal horn of mutilated foot rats contained substantially fewer CGRP and tachykinin-immunoreactive fibres but more fibres immunoreactive for enkephalin. Many enkephalin-immunoreactive cell bodies were also found in the dorsal horn of the mutants, by contrast none were visible in control animals. Neuropeptide Y immunoreactivity was, however, unchanged in the spinal cord of the mutants. In the dorsal root ganglia of the mutants, the number of CGRPor tachykinin-immunoreactive cells and their proportion to total neuronal numbers were significantly less in comparison with normal controls. The diameter range of CGRP- and tachykinin-immunoreactive cells shifted from small (15–25 m) to medium size (25–45 m) as revealed by frequency distribution histograms. The skin from the affected foreand hindlimbs of the mutant rats, in keeping with fewer CGRP- and tachykinin-immunoreactive cells in the dorsal root ganglia, contained substantially less fibres immunoreactive for CGRP and tachykinins; a difference that was not seen in skin of unaffected areas (whiskers and snout). By contrast, neuropeptide Y-immunoreactive fibres showed a normal distribution around blood vessels and sweat glands of mutilated foot rats. The data suggest that diminished pain perception in the mutilated foot rat is related to loss of peptide-containing sensory neurones. Furthermore, the intraspinal increase of enkephalinergic neurones in the dorsal horn, concomitant with the decreased number of primary sensory neurones, may also play a contributory rôle in reducing pain thresholds.     

Hot receptors in the brain

Background

The complex neuronal circuitry of the dorsal horn of the spinal cord is as yet poorly understood. However, defining the circuits underlying the transmission of information from primary afferents to higher levels is critical to our understanding of sensory processing. In this study, we have examined phosphodiesterase 1C (Pde1c) BAC transgenic mice in which a green fluorescent protein (GFP) reporter gene reflects Pde1c expression in sensory neuron subpopulations in the dorsal root ganglia and spinal cord.

Results

Using double labeling immunofluorescence, we demonstrate GFP expression in specific subpopulations of primary sensory neurons and a distinct neuronal expression pattern within the spinal cord dorsal horn. In the dorsal root ganglia, their distribution is restricted to those subpopulations of primary sensory neurons that give rise to unmyelinated C fibers (neurofilament 200 negative). A small proportion of both non-peptidergic (IB4-binding) and peptidergic (CGRP immunoreactive) subclasses expressed GFP. However, GFP expression was more common in the non-peptidergic than the peptidergic subclass. GFP was also expressed in a subpopulation of the primary sensory neurons immunoreactive for the vanilloid receptor TRPV1 and the ATP-gated ion channel P2X₃. 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.

Conclusion

The expression of GFP in subclasses of nociceptors and also in dorsal horn regions densely innervated by nociceptors suggests that Pde1c marks a unique subpopulation of nociceptive sensory neurons.     

Dorsally derived netrin 1 provides an inhibitory cue and elaborates the 'waiting period' for primary sensory axons in the developing spinal cord

Dorsal root ganglion (DRG) neurons extend axons to specific targets in the gray matter of the spinal cord. During development, DRG axons grow into the dorsolateral margin of the spinal cord and projection into the dorsal mantle layer occurs after a ;waiting period' of a few days. Netrin 1 is a long-range diffusible factor expressed in the ventral midline of the developing neural tube, and has chemoattractive and chemorepulsive effects on growing axons. Netrin 1 is also expressed in the dorsal spinal cord. However, the roles of dorsally derived netrin 1 remain totally unknown. Here, we show that dorsal netrin 1 controls the correct guidance of primary sensory axons. During the waiting period, netrin 1 is transiently expressed or upregulated in the dorsal spinal cord, and the absence of netrin 1 results in the aberrant projection of sensory axons, including both cutaneous and proprioceptive afferents, into the dorsal mantle layer. Netrin 1 derived from the dorsal spinal cord, but not the floor plate, is involved in the correct projection of DRG axons. Furthermore, netrin 1 suppresses axon outgrowth from DRG in vitro. Unc5c(rcm) mutant shows abnormal invasion of DRG axons as observed in netrin 1 mutants. These results are the first direct evidence that netrin 1 in the dorsal spinal cord acts as an inhibitory cue for primary sensory axons and is a crucial signal for the formation of sensory afferent neural networks.     

Prolonged primary afferent induced alterations in dorsal horn neurones, an intracellular analysis in vivo and in vitro

1.) Peripheral tissues injury produces long lasting sensory and motor disturbances in man that present as the post-injury hypersensitivity syndrome with a reduction in the threshold required to elicit either pain or the flexion withdrawal reflex and an exaggeration of the normal response to suprathreshold stimuli. 2.) Two mechanisms contribute to these changes; sensitization of the peripheral terminals of high threshold primary afferents and an increase in the excitability of the spinal cord; a phenomenon known as central sensitization. 3.) Central sensitization has previously been shown by our laboratory to be the consequence of activity in unmyelinated primary afferents. Brief (20 s) C-fibre strength conditioning stimuli have the capacity to produce both a prolonged heterosynaptic facilitation of the flexion reflex and an alteration in the response properties of dorsal horn neurones, that long outlast the conditioning stimulus. 4.) In the adult decerebrate-spinal rat preparation we have, using intracellular recordings of dorsal horn neurones, examined the time course of the central effects of different types of orthodromic inputs. The hemisected spinal cord preparation isolated from 12-14 day rat pups has been used to see whether prolonged alterations in dorsal horn properties induced by orthodromic inputs can be studied in vitro. 5.) Single stimuli applied to a cutaneous nerve at graded strengths to successively recruit A beta, A delta and C-afferents produce, in the majority of neurones recorded in the deep dorsal horn in vivo, a series of post synaptic potentials that last from between ten and several hundred milliseconds. 6.) Repeated low frequency stimulation of C but not A-afferent fibres results in a pattern of progressive response increment or windup in a proportion of dorsal horn neurones. In some of the neurones the windup is associated with a depolarization that outlasts the stimulus period for tens of seconds. 7.) Application of the chemical irritant mustard oil to the skin activates chemosensitive C-afferent fibres for 1-3 minutes. Such a conditioning stimulus results however in an expansion in the size and an alteration in the response properties of the receptive fields of dorsal horn neurones that lasts for tens of minutes. 8.) In dorsal horn neurones recorded intracellularly in the isolated hemisected spinal cord, both intrinsic membrane properties and the orthodromic responses to primary afferent input can be studied. Repeated stimulation of a dorsal root produces in some neurones a prolonged heterosynaptic facilitation with both an augmentation of the response to the conditioning root (homosynaptic potentiation) and to adjacent test roots (heterosynaptic potentiation).(ABSTRACT TRUNCATED AT 400 WORDS)     

Role of mitochondria in intracellular calcium signaling in primary and secondary sensory neurones of rats

The participation of different calcium-regulated mechanisms in the generation of cytosolic Ca(2+) transients during neuronal excitation has been compared in isolated large and small primary (dorsal root ganglia (DRG)) and secondary (spinal dorsal horn (DH)) rat sensory neurones. As it was shown before in murine primary sensory neurones the application of mitochondrial protonophore CCCP by itself induced only small elevation of [Ca(2+)](i). However, its preceding application substantially increased the peak amplitude of depolarization-induced transients. Application of CCCP immediately after termination of the depolarizing pulse induced in both types of primary neurones a massive release of Ca(2+) from mitochondria into the cytosol. In secondary neurones application of CCCP by itself induced a substantial release of Ca(2+) from the mitochondria, but its preceding application resulted in only an insignificant increase in the peak amplitude of depolarization-triggered calcium transients. Application of CCCP immediately after termination of depolarization elicited a small release of Ca(2+), which became more pronounced when the application was delayed. Preceding application of CCCP increased the amplitude of the transients induced by caffeine-triggered Ca(2+) release from the endoplasmic reticulum in secondary neurones and did not affect those in large primary neurones. These findings may be explained by substantial differences in the density and distribution of mitochondria in the cytosol of primary and secondary sensory neurones. This suggestion was confirmed electronmicroscopically, showing a much lower density of mitochondria near plasmalemma in secondary sensory neurones and predominant clustered location of mitochondria beneath the plasmalemma in the primary cells. The possible functional importance of these differences is discussed.     

E-cadherin expression in a particular subset of sensory neurons.

We found that the dorsal root ganglia (DRG) and trigeminal ganglia of mouse embryos express the E-cadherin cell-cell adhesion molecule and analyzed its expression profile. E-cadherin expression began around Embryonic Day 12 (E12) in these ganglia, thereafter increased, and persisted to the adult stage. This cadherin was expressed by 10 and 30% of DRG neurons in E17 and postnatal animals, respectively, as well as by satellite cells and some Schwann cells. E-cadherin-positive primary sensory fibers terminated only in a narrow region of the dorsal horn of the spinal cord, which was identified as part of lamina II by double-staining for E-cadherin and substance P or somatostatin. This E-cadherin expressing area of the spinal cord extended to part of the trigeminal nucleus in the medulla. These results showed that E-cadherin is expressed in a particular subset of primary sensory neurons which may have specific functional properties. We suggest that this adhesion molecule may play a role in the selective adhesion of sensory neuronal fibers.     

Sensory pathways in amphioxus larvae II. Dorsal tracts and translumenal cells

Serial and interval EM series were used to examine the dorsal nerve tracts in the anterior nerve cord of a 12.5 day larva of Branchiostoma floridae. Fibres within the tracts derive from peripheral sensory cells and a class of intramedullary sensory neurones known as dorsal (Retzius) bipolar cells. Both form repeated synapses of similar type, apparently with the same targets. The synapses occur at points where, at intervals, the tracts expand to form large synaptic zones. The target dendrites, which form complex tangles, belong chiefly to dorsal translumenal cells, a class of neurone distinguished by their apical processes. The latter range from short extensions of the cell apex that contact the opposite side of the cord via junctions, but go no further, to elongate processes with slender branches that project to the contralateral dorsal tract. The morphology indicates that translumenal cells play the same role in amphioxus as internuncial neurones in vertebrate spinal cord. Their axons can be ipsilateral or contralateral; some synapse with motoneurones directly while others innervate other interneurones, including other translumenal cells. From the circuitry, the cells appear to be chiefly involved in integrating sensory input from peripheral mechanoreceptors. This could include acting as a filter that amplifies some input patterns over others, or that normalizes input, so that CNS circuits are not overloaded as new sensory cells differentiate during development. The functional importance of the translumenal system to the organism is reflected in a massive increase in size and cell numbers during the larval phase. The anterior, brain-like integrative centres of the cerebral vesicle, in contrast, are initially small and change very little.     

Effects of dorsal root section and occlusion of dorsal spinal artery on the neurotransmitter candidates in rat spinal cord

In order to obtain further evidence of putative neurotransmitters in primary sensory neurons and interneurons in the dorsal spinal cord, we have studied the effects of unilateral section of dorsal roots and unilateral occlusion of the dorsal spinal artery on cholinergic enzyme activity and on selected amino acid levels in the spinal cord. One week after sectioning dorsal roots from caudal cervical (C₇) to cranial thoracic (T₂) levels, the specific activity of choline acetyltransferase (ChAT) was significantly decreased and acetylcholinesterase (AChE) showed a tendency to decrease in the dorsal quadrant on the operated side of the spinal cord. Dorsal root sectioning had little effect on the levels of free glutamic acid or other amino acids in the dorsal spinal cord. These results suggest that primary sensory neurons may include some cholinergic axons, and that levels of putative amino acid transmitters are not regulated by materials supplied by axonal transport from the dorsal root ganglia. By contrast, one week following unilateral occlusion of the dorsal spinal artery, the activities of ChAT and AChE were unchanged in the operated quadrant of the spinal cord, while decreases of Asp, Glu, and GABA, and an increase in Tau were detected. These findings are consistent with the proposals that such amino acids, but not ACh, may function as neurotransmitter candidates in interneurons of the dorsal spinal cord.Abbreviation used ACh acetylcholine - AChE acetylcholinesterase - Asp aspartic acid - ChAT choline acetyltransferase - GABA -aminobutyric acid - Glu glutamic acid - Gly glycine - SP substance P - Tau taurine     

Analysis of receptive fields revealed by in vivo patch-clamp recordings from dorsal horn neurons and in situ intracellular recordings from dorsal root ganglion neurons

It has been thought that spinal dorsal horn neurons receive convergent inputs from not only somatosensory but also visceral pathways. For instance, the referred pain is presumed to be due to the convergence of sensory inputs from cardiac and shoulder receptive fields. However, precise investigation has not been made from dorsal horn neurons yet, because of difficulty in studying the pathways from those regions by means of conventional electrophysiology. The purpose of this study is to clarify the convergent inputs to single dorsal horn neurons from wide receptive fields using an in vivo patch-clamp recording technique from the superficial spinal dorsal horn and an intracellular recording from dorsal root ganglion neurons that keep physiological connections with the peripheral sites. Identified dorsal root ganglion neurons received an input from a quite small area, about 1 x 1 mm in width of the skin. In contrast, substantia gelatinosa neurons in the spinal cord received inputs from an unexpectedly wide area of the skin. Previous extracellular recordings have, however, revealed that substantia gelatinosa neurons have small receptive field. This discrepancy is probably due mainly to an availability of the in vivo patch-clamp method to analyze sub-threshold synaptic responses. In contrast, the extracellular recording technique allows us to analyze predominantly the firing frequency of neurons. Thus, the in vivo patch-clamp recordings from dorsal horn neurons and the intracellular recordings from DRG neurons will be useful for well understanding the sensory processing in the spinal cord.     

THE EFFECT OF CHRONIC DORSAL ROOT SECTION ON THE CONCENTRATION OF FREE AMINO ACIDS IN THE RABBIT SPINAL CORD

Abstract— Amino acids may be involved in primary afferent excitatory neurotransmission in the spinal cord. To test this possibility the effect of chronic dorsal root section on amino acid levels of the rabbit spinal cord has been investigated. Dorsal roots L6-S2 were sectioned under anaesthesia. Control animals were subjected to similar surgical procedures but the dorsal roots were left intact. Electromyogram recordings taken 6 days after surgery confirmed the absence of sensory input to the lower lumbosacral cord of dorsal root sectioned animals although motor function was retained. In contrast to this control animals exhibited normal reflex activity. The spinal cord was removed from each animal and extracted in trichloracetic acid for subsequent analysts of amino acids on an autoanalyser. Sections of cord were retained for histological determination of neuronal degeneration. Comparison of amino acid levels in dorsal root sectioned and control animals revealed that the only excitatory amino acid to be significantly reduced by dorsal root section wasaspartic acid (–50 percent X although glutamic acid was also reduced (– 30 per cent). Two inhibitory amino acids, cystathionine and GABA, were also significantly depleted (– 50 and - 35 per cent). The possible involvement of these amino acids in spinal cord neurotransmission is discussed.     

Nerve growth factor in glia and inflammatory cells of the injured rat spinal cord

Nerve growth factor (NGF) is crucial for the development of sympathetic and small-diameter sensory neurons and for maintenance of their mature phenotype. Its role in generating neuronal pathophysiology is less well understood. After spinal cord injury, central processes of primary afferent fibers sprout into the dorsal horn, contributing to the development of autonomic dysfunctions and pain. NGF may promote these states as it stimulates sprouting of small-diameter afferent fibers and its concentration in the spinal cord increases after cord injury. The cells responsible for this increase must be identified to develop a strategy to prevent the afferent sprouting. Using immunocytochemistry, we identified cells containing NGF in spinal cord sections from intact rats and from rats 1 and 2 weeks after high thoracic cord transection. In intact rats, this neurotrophin was present in a few ramified microglia and in putative Schwann cells in the dorsal root. Within and close to the lesion of cord-injured rats, NGF was in many activated, ramified microglia, in a subset of astrocytes, and in small, round cells that were neither glia nor macrophages. NGF-immunoreactive putative Schwann cells were prevalent throughout the thoracolumbar cord in the dorsal roots and the dorsal root entry zones. Oligodendrocytes were never immunoreactive for this protein. Therapeutic strategies targeting spinal cord cells that produce NGF may prevent primary afferent sprouting and resulting clinical disorders after cord injury.     

Temporal regulation of neuropilin‐1 expression and sensitivity to semaphorin 3A in NGF‐ and NT3‐responsive chick sensory neurons

The extracellular molecule semaphorin 3A (Sema3A) is proposed to be a negative guidance cue that participates in patterning DRG sensory axons in the developing chick spinal cord. During development Sema3A is first expressed throughout the spinal cord gray matter, but Sema3A expression later disappears from the dorsal horn, where small‐caliber cutaneous afferents terminate. Sema3A expression remains in the ventral horn, where large‐muscle proprioceptive afferents terminate. It has been proposed that temporal changes in the sensitivity of different classes of sensory afferents to Sema3A contribute to the different pathfinding of these sensory afferents. This study compared the expression of the semaphorin 3A receptor subunit, neuropilin‐1, and the collapse response of growth cones to semaphorin 3A for NGF (cutaneous)‐ and NT3 (proprioceptive)‐dependent sensory axons extended from E6‐E10 chick embryos. Growth cones extended from E6 DRGs in NT3‐containing medium expressed neuropilin‐1 and collapsed in response to Sema3A. From E7 until E10 NT3‐responsive growth cones expressed progressively lower levels of neuropilin‐1, and were less sensitive to Sema3A. On the other hand, growth cones extended from DRGs in NGF‐containing medium expressed progressively higher levels of neuropilin‐1 and higher levels of collapse response to Sema3A over the period from E6–E10. Thus, developmental patterning of sensory terminals in the chick spinal cord may arise from changes in both Sema3A expression in the developing spinal cord and accompanying changes in neuronal expression of the Sema3A receptor subunit, neuropilin‐1. © 2002 Wiley Periodicals, Inc. J Neurobiol 51: 43–53, 2002     

Calcitonin gene-related peptide coexists with substance P in capsaicin sensitive neurons and sensory ganglia of the rat

Immunohistochemical and radioimmunoassay studies revealed that both CGRP- and SP-like immunoreactivity in the caudal spinal trigeminal nucleus and tract, the substantia gelatinosa and the dorsal cervical spinal cord as well as in cell bodies of the trigeminal ganglion and the spinal dorsal root ganglion is markedly depleted by capsaicin which is known to cause degeneration of a certain number of primary sensory neurons. Higher brain areas and the ventral spinal cord were not affected by capsaicin treatment. Furthermore CGRP and substance P-like immunoreactivity were shown to be colocalized in the above areas and to coexist in cell bodies of the trigeminal ganglion and the spinal dorsal root ganglia. It is suggested that CGRP, like substance P, may have a neuromodulatory role on nociception and peripheral cardiovascular reflexes.     

Tracing of sensory neurones and spinal motoneurones of the pigeon by injection of fluorescent dyes into peripheral nerves

The projection of peripheral sensory and motor nerves was investigated in the pigeon (Columba livia) by means of retrogradely transported fluorescent dyes. Two combinations of fluorescent tracers were used that could be identified within the same cell when excited by light of 405 nm: 1) Propidium iodide and Bisbenzimide, which label the cytoplasm orange and the nucleus blue, respectively; 2) Fast Blue, which labels the cytoplasm blue, and Nuclear Yellow, which labels the nucleus (especially the nucleolar ring) yellow. The presence of the tracers in a given cell was confirmed microspectrophotometrically. Following injection of the tracers into peripheral nerves, labelled sensory neurones were seen in the dorsal root ganglia and motoneurones of the spinal cord. The peroneal and tibial nerves projected to L2-L5 and L2-L7, respectively, whereas the median and ulnar nerves projected to C12-Th2 and C13-Th1. Double-labelled sensory neurones were observed when both peroneal and tibial, or median and ulnar nerves were injected with different tracers. This indicates that some sensory neurones possess peripheral processes that dichotomize to pass down two different peripheral nerves. Double labelling was never seen in motoneurones, or in sensory neurones after tracer injection into the sciatic and femoral nerves.     

Monoaminergic Modulation of Spinal Viscero-Sympathetic Function in the Neonatal Mouse Thoracic Spinal Cord

Descending serotonergic, noradrenergic, and dopaminergic systems project diffusely to sensory, motor and autonomic spinal cord regions. Using neonatal mice, this study examined monoaminergic modulation of visceral sensory input and sympathetic preganglionic output. Whole-cell recordings from sympathetic preganglionic neurons (SPNs) in spinal cord slice demonstrated that serotonin, noradrenaline, and dopamine modulated SPN excitability. Serotonin depolarized all, while noradrenaline and dopamine depolarized most SPNs. Serotonin and noradrenaline also increased SPN current-evoked firing frequency, while both increases and decreases were seen with dopamine. In an in vitro thoracolumbar spinal cord/sympathetic chain preparation, stimulation of splanchnic nerve visceral afferents evoked reflexes and subthreshold population synaptic potentials in thoracic ventral roots that were dose-dependently depressed by the monoamines. Visceral afferent stimulation also evoked bicuculline-sensitive dorsal root potentials thought to reflect presynaptic inhibition via primary afferent depolarization. These dorsal root potentials were likewise dose-dependently depressed by the monoamines. Concomitant monoaminergic depression of population afferent synaptic transmission recorded as dorsal horn field potentials was also seen. Collectively, serotonin, norepinephrine and dopamine were shown to exert broad and comparable modulatory regulation of viscero-sympathetic function. The general facilitation of SPN efferent excitability with simultaneous depression of visceral afferent-evoked motor output suggests that descending monoaminergic systems reconfigure spinal cord autonomic function away from visceral sensory influence. Coincident monoaminergic reductions in dorsal horn responses support a multifaceted modulatory shift in the encoding of spinal visceral afferent activity. Similar monoamine-induced changes have been observed for somatic sensorimotor function, suggesting an integrative modulatory response on spinal autonomic and somatic function.     

cGMP-mediated signaling via cGKIalpha is required for the guidance and connectivity of sensory axons

Previous in vitro studies using cGMP or cAMP revealed a cross-talk between signaling mechanisms activated by axonal guidance receptors. However, the molecular elements modulated by cyclic nucleotides in growth cones are not well understood. cGMP is a second messenger with several distinct targets including cGMP-dependent protein kinase I (cGKI). Our studies indicated that the alpha isoform of cGKI is predominantly expressed by sensory axons during developmental stages, whereas most spinal cord neurons are negative for cGKI. Analysis of the trajectories of axons within the spinal cord showed a longitudinal guidance defect of sensory axons within the developing dorsal root entry zone in the absence of cGKI. Consequently, in cGKI-deficient mice, fewer axons grow within the dorsal funiculus of the spinal cord, and lamina-specific innervation, especially by nociceptive sensory neurons, is strongly reduced as deduced from anti-trkA staining. These axon guidance defects in cGKI-deficient mice lead to a substantial impairment in nociceptive flexion reflexes, shown using electrophysiology. In vitro studies revealed that activation of cGKI in embryonic dorsal root ganglia counteracts semaphorin 3A-induced growth cone collapse. Our studies therefore reveal that cGMP signaling is important for axonal growth in vivo and in vitro.     

ErbB transmembrane tyrosine kinase receptors are expressed by sensory and motor neurons projecting into sciatic nerve.

Adult spinal cord motor and dorsal root ganglion (DRG) sensory neurons express multiple neuregulin-1 (NRG-1) isoforms that act as axon-associated factors promoting neuromuscular junction formation and Schwann cell proliferation and differentiation. NRG-1 isoforms are also expressed by muscle and Schwann cells, suggesting that motor and sensory neurons are themselves acted on by NRG-1 isoforms produced by their peripheral targets. To test this hypothesis, we examined the expression of the NRG-1 receptor subunits erbB2, erbB3, and erbB4 in rat lumbar DRG and spinal cord. All three erbB receptors are expressed in these tissues. Sciatic nerve transection, an injury that induces Schwann cell expression of NRG-1, alters erbB expression in DRG and cord. Virtually all DRG neurons are erbB2- and erbB3-immunoreactive, with erbB4 also detectable in many neurons. In spinal cord white matter, erbB2 and erbB4 antibodies produce dense punctate staining, whereas the erbB3 antibody primarily labels glial cell bodies. Spinal cord dorsal and ventral horn neurons, including alpha-motor neurons, exhibit erbB2, erbB3, and erbB4 immunoreactivity. Spinal cord ventral horn also contains a population of small erbB3+/S100beta+/GFAP- cells (GFAP-negative astrocytes or oligodendrocytes). We conclude that sensory and motor neurons projecting into sciatic nerve express multiple erbB receptors and are potentially NRG-1 responsive.