Changes in galanin immunoreactivity in rat lumbosacral spinal cord and dorsal root ganglia after spinal cord injury

Alterations in the expression of the neuropeptide galanin were examined in micturition reflex pathways 6 weeks after complete spinal cord transection (T8). In control animals, galanin expression was present in specific regions of the gray matter in the rostral lumbar and caudal lumbosacral spinal cord, including: (1) the dorsal commissure; (2) the superficial dorsal horn; (3) the regions of the intermediolateral cell column (L1-L2) and the sacral parasympathetic nucleus (L6-S1); and (4) the lateral collateral pathway in lumbosacral spinal segments. Densitometry analysis demonstrated significant increases (P < or = 0.001) in galanin immunoreactivity (IR) in these regions of the S1 spinal cord after spinal cord injury (SCI). Changes in galanin-IR were not observed at the L4-L6 segments except for an increase in galanin-IR in the dorsal commissure in the L4 segment. In contrast, decreases in galanin-IR were observed in the L1 segment. The number of galanin-IR cells increased (P < or = 0.001) in the L1 and S1 dorsal root ganglia (DRG) after SCI. In all DRG examined (L1, L2, L6, and S1), the percentage of bladder afferent cells expressing galanin-IR significantly increased (4-19-fold) after chronic SCI. In contrast, galanin expression in nerve fibers in the urinary bladder detrusor and urothelium was decreased or eliminated after SCI. Expression of the neurotrophic factors nerve growth factor (NGF) and brain-derived neurotrophic factor (BDNF) was altered in the spinal cord after SCI. A significant increase in BDNF expression was present in spinal cord segments after SCI. In contrast, NGF expression was only increased in the spinal segments adjacent and rostral to the transection site (T7-T8), whereas spinal segments (T13-L1; L6-S1), distal to the transection site exhibited decreased NGF expression. Changes in galanin expression in micturition pathways after SCI may be mediated by changing neurotrophic factor expression, particularly BDNF. These changes may contribute to urinary bladder dysfunction after SCI.     

Prolonged sympathetic innervation of sensory neurons in rat thoracolumbar dorsal root ganglia during chronic colitis

Background Peripheral irritation‐induced sensory plasticity may involve catecholaminergic innervation of sensory neurons in the dorsal root ganglia (DRG). Methods Catecholaminergic fiber outgrowth in the thoracolumbar DRG (T13‐L2) was examined by tyrosine hydroxylase (TH) immunostaining, or by sucrose‐potassium phosphate‐glyoxylic acid histofluorescence method. TH level was examined by Western blot. Colonic afferent neurons were labeled by retrograde neuronal tracing. Colitis was induced by intracolonic instillation of tri‐nitrobenzene sulfonic acid (TNBS). Key Results The catecholaminergic fibers formed ‘basket‐like’ structures around the DRG cells. At 7 days following TNBS treatment, the number of DRG neurons surrounded by TH‐immunoreactive fibers and the protein levels of TH were significantly increased in T13, L1, and L2 DRGs (two‐ to threefold, P < 0.05). The DRG neurons that were surrounded by TH immunoreactivity were 200 kDa neurofilament‐positive, but not isolectin IB4‐positve or calcitonin gene‐related peptide‐positive. The TH‐immunoreactive fibers did not surround but adjoin the specifically labeled colonic afferent neurons, and was co‐localized with glial marker S‐100. Comparison of the level of TH and the severity of colonic inflammation showed that following TNBS treatment, the degree of colonic inflammation was most severe at day 3, subsided at day 7, and significantly recovered by day 21. However, the levels of TH in T13‐L2 DRGs were increased at both 3 days and 7 days post TNBS treatment and persisted up to 21 days (two‐ to fivefold increase, P < 0.05) as examined. Conclusions & Inferences Colonic inflammation induced prolonged catecholaminergic innervation of sensory neurons, which may have relevance to colitis‐induced chronic visceral hypersensitivity and/or referred pain.     

The role of neurotrophins in the maintenance of the spinal cord motor neurons and the dorsal root ganglia proprioceptive sensory neurons

The aim of this study was to approach the question of neuronal dependence on neurotrophins during embryonic development in mice in a way other than gene targeting. We employed amyogenic mouse embryos and fetuses that develop without any skeletal myoblasts or skeletal muscle and consequently lose motor and proprioceptive neurons. We hypothesized that if, in spite of the complete inability to maintain motor and proprioceptive neurons, the remaining spinal and dorsal root ganglia tissues of amyogenic fetuses still contain any of the neurotrophins, that particular neurotrophin alone is not sufficient for the maintenance of motor and proprioceptive neurons. Moreover, if the remaining spinal and dorsal root ganglia tissues still contain any of the neurotrophins, that particular neurotrophin alone may be sufficient for the maintenance of the remaining neurons (i.e., mostly non-muscle- and a few muscle-innervating neurons). To test the role of the spinal cord and dorsal root ganglia tissues in the maintenance of its neurons, we performed immunohistochemistry employing double-mutant and control tissues and antibodies against neurotrophins and their receptors. Our data suggested that: (a) during the peak of motor neuron cell death, the spinal cord and dorsal root ganglia distribution of neurotrophins was not altered; (b) the distribution of BDNF, NT-4/5, TrkB and TrkC, and not NT-3, was necessary for the maintenance of the spinal cord motor neurons; (c) the distribution of BDNF, NT-4/5 and TrkC, and not NT-3 and Trk B, was necessary for the maintenance of the DRG proprioceptive neurons; (d) NT-3 was responsible for the maintenance of the remaining neurons and glia in the spinal cord and dorsal root ganglia (possibly via TrkB).     

Differential expression of annexins I-VI in the rat dorsal root ganglia and spinal cord

The annexins are a family of Ca²⁻-dependent phospholipid-binding proteins. In the present study, the spatial expression patterns of annexins I-VI were evaluated in the rat dorsal root ganglia (DRG) and spinal cord (SC) by using indirect immunofluorescence. Annexin I is expressed in small sensory neurons of the DRG, by most neurons of the SC, and by ependymal cells lining the central canal. Annexin II is expressed by most sensory neurons of the DRG but is primarily expressed in the SC by glial cells. Annexin III is expressed by most sensory neurons, regardless of size, by endothelial cells lining the blood vessels, and by the perineurium. In the SC, annexin III is primarily expressed by astrocytes. In the DRG and the SC, annexin IV is primarily expressed by glial cells and at lower levels by neurons. In the DRG, annexin V is expressed in relatively high concentrations in small sensory neurons in contrast to the SC, where it is expressed mainly by ependymal cells and by small-diameter axons located in the superficial laminae of the dorsal horn areas. Annexin VI is differentially expressed by sensory neurons of the DRG, being more concentrated in small neurons. In the SC, annexin VI has the most striking distribution. It is concentrated subjacent to the plasma membrane of motor neurons and their processes. The differential localization pattern of annexins in cells of the SC and DRG could reflect their individual biological roles in Ca²⁻-signal transduction within the central nervous system. © 1996 Wiley-Liss, Inc.     

Pre- and post-natal ontogeny of serotonergic projections to the rat spinal cord

The development of 5-hydroxytryptamine (5-HT) innervation in the spinal cord was studied from embryonic day 14 (E14) to adulthood. Sprague-Dawley rats were fixed by perfusion with 5% glutaraldehyde in cacodylate-sodium metabisulfite buffer, and vibratome sections were processed for immunocytochemistry with a 5-HT antiserum. For electron microscopy, the sections were flat-embedded in araldite, and thin sectioning was performed. 5-HT neurons caudally directed from raphe nuclei invade the spinal cord at E14 and reach the caudalmost levels by E16-E17. In longitudinal sections, axons are seen by E15, at cervical and upper thoracic levels, to invade the presumptive gray matter from the anterior and lateral funiculi. The invasion process occurred either by sharp angulation of the axon or by branching of a collateral. By E16, at thoracic level the anterior horn and the intermediolateral columns are profusely innervated by very thin, varicose fibers; synapses are seen at E17 and E18 using EM. 5-HT immunoreactive boutons are involved here. After birth, 5-HT innervation of these two areas evolves progressively from a diffuse network to a more restricted pattern, especially at the thoracic level for the intermediolateral column and at cervical and lumbar levels for the anterior horn. The adult pattern is reached by postnatal day 21 (P21). The growth of axons toward the dorsal horn becomes noticeable by E19 at all spinal levels, when fibers invade the neck of the horn from the lateral funiculus, and innervation proceeds diffusely until P5. At P7, thin fibers course dorsally and laterally along the border of the gray matter and ramify profusely in layers I and II. The adult pattern is also reached in the dorsal horn by P21. These results are discussed in relation to the postnatal maturation of motor and sensory circuits and to the development of transplanted raphe neurons in the rat spinal cord.     

Insulin-like growth factor-II/Mannose-6-phosphate receptor in the spinal cord and dorsal root ganglia of the adult rat

The insulin-like growth factor-II/mannose-6-phosphate (IGF-II/M6P) receptor is a multifunctional transmembrane glycoprotein, which interacts with a number of molecules, including IGF-II and M6P-containing lysosomal enzymes. The receptor is widely distributed throughout the brain and is known to be involved in lysosomal enzyme trafficking, cell growth, internalization and degradation of IGF-II. In the present study, using autoradiographic, Western blotting and immunocytochemical methods, we provide the first report that IGF-II/M6P receptors are discretely distributed at all major segmental levels of the spinal cord and dorsal root ganglia of the adult rat. In the spinal cord, a high density of [(125)I]IGF-II binding sites was evident in the ventral horn (lamina IX) and in areas around the central canal (lamina X), whereas intermediate grey matter and dorsal horn were associated with moderate receptor levels. The dorsal root ganglia exhibited rather high density of [(125)I]IGF-II binding sites. Interestingly, meninges present around the spinal cord displayed highest density of [(125)I]IGF-II binding compared to any given region of the spinal grey matter or the dorsal root ganglia. Western blot results indicated the presence of the IGF-II/M6P receptor at all major levels of spinal cord and dorsal root ganglia, with little segmental variation. At the cellular level, spinal motorneurons demonstrated the most intense IGF-II/M6P receptor immunoreactivity, followed by interneurons in the intermediate region and deeper dorsal horn. Some scattered IGF-II/M6P immunoreactive fibers were found in the superficial laminae of the dorsal horn and dorsolateral funiculus. The meninges of the spinal cord also seemed to express IGF-II receptor immunoreactivity. In the dorsal root ganglia, receptor immunoreactivity was evident primarily in a subset of neurons of all diameters. These results, taken together, provide anatomical evidence of a role for the IGF-II/M6P receptor in general cellular functions such as transport of lysosomal enzymes and/or internalization followed by clearance of IGF-II in the spinal cord and dorsal root ganglia.     

Immunocytochemical localization of somatostatin receptor sst2A in the rat spinal cord and dorsal root ganglia

Intrathecal administration of octreotide, a stable somatostatin analogue, provides pain relief in patients, and locally applied somatostatin inhibits firing of nociceptive dorsal horn neurons. In the present study, we have raised polyclonal antibodies that specifically detect the somatostatin receptor sst_2A and used these antisera for immunocytochemical localization of the receptor protein in the rat spinal cord and dorsal root ganglia. In the superficial layers of the dorsal horn, sst_2A-like immunoreactivity (Li) formed a dense network consisting of neuronal perikarya and dendrites which were often closely apposed by, but not co-contained within, somatostatin-14-immunoreactive nerve fibres and terminals. sst_2A-Li was resistant to dorsal rhizotomy and did not colocalize with either substance P or calcitonin gene-related peptide suggesting that sst_2A-Li was not located to primary afferents, but rather confined to second-order spinal neurons. The position of sst_2A-Li perikarya and dendrites in the dorsal horn appeared to be similar to those containing μ-opioid receptor-Li; however, double labelling experiments revealed no instances of coexistence of these two receptors. sst_2A-Li was also observed in the dorsal root ganglia predominantly targeted to the somatic plasmalemma of medium size neurons distinct from those expressing somatostatin-14 or δ-opioid receptors. Thus, the present results not only provide a morphological substrate for spinal octreotide analgesia but also show that somatostatin and opioids are poised to modulate nociceptive transmission by distinct anatomical systems.     

GABAB receptor protein and mRNA distribution in rat spinal cord and dorsal root ganglia

The presence of metabotropic receptors for GABA, GABAB, on primary afferent terminals in mammalian spinal cord has been previously reported. In this study we provide further evidence to support this in the rat and show that the GABAB receptor subunits GABAB1 and GABAB2 mRNA and the corresponding subunit proteins are present in the spinal cord and dorsal root ganglion. We also show that the predominant GABAB1 receptor subunit mRNA present in the afferent fibre cell body appears to be the 1a form. In frozen sections of lumbar spinal cord and dorsal root ganglia (DRG) GABAB receptors were labelled with [3H]CGP 62349 or the sections postfixed with paraformaldehyde and subjected to in situ hybridization using oligonucleotides designed to selectively hybridize with the mRNA for GABAB(1a), GABAB(1b) or GABAB2. For immunocytochemistry (ICC), sections were obtained from rats anaesthetized and perfused-fixed with paraformaldehyde. The distribution of binding sites for [3H]CGP 62349 mirrored that previously observed with [3H]GABA at GABAB sites. The density of binding sites was high in the dorsal horn but much lower in the ventral regions. By contrast, the density of mRNA (pan) was more evenly distributed across the laminae of the spinal cord. The density of mRNA detected with the pan probe was high in the DRG and distributed over the neuron cell bodies. This would accord with GABAB receptor protein being formed in the sensory neurons and transported to the primary afferent terminals. Of the GABAB1 mRNA in the DRG, approximately 90% was of the GABAB(1a) form and approximately 10% in the GABAB(1b) form. This would suggest that GABAB(1a) mRNA may be responsible for encoding presynaptic GABAB receptors on primary afferent terminals in a manner similar to that we have previously observed in the cerebellar cortex. GABAB2 mRNA was also evenly distributed across the spinal cord laminae at densities equivalent to those of GABAB1 in the dorsal horn. GABAB2 mRNA was also detected to the same degree within the DRG. Immunocytochemical analysis revealed that GABAB(1a), GABAB(1b) and GABAB2 were all present in the spinal cord. GABAB(1a) labelling appeared to be more dense than GABAB(1b) and within the superficial dorsal horn GABAB(1a) was present in the neuropil whereas GABAB(1b) was associated with cell bodies in this region. Both 1a and 1b immunoreactivity was expressed in motor neurons in lamina IX. GABAB2 immunoreactivity was expressed throughout the spinal cord and was evident within the neuropil of the superficial laminae.     

Ontogeny of enkephalin- and VIP-containing neurons in dissociated cultures of embryonic mouse spinal cord and dorsal root ganglia

The ontogeny of vasoactive intestinal polypeptide (VIP), and Met-enkephalin in primary cultures of spinal cord/dorsal root ganglia from 12-day mouse embryos was examined by radioimmunoassay and immunohistochemistry. Met-enkephalin levels rose from less than 5 to 700 pg/culture over 26 days and were half maximal by day 16-18 in culture. VIP levels rose from less than 1 to 30 pg/culture over the same period, but were already half maximal by day 9. Met-enkephalin immunoreactivity was localized in multipolar medium sized neurons while VIP immunoreactivity was visualized both in neurons with extensively branched processes and in bipolar cells some of which appeared to be dorsal root ganglion cells. Tetrodotoxin (TTX)-sensitive spontaneous release of both peptides developed in parallel with the ability to stimulate peptide release with elevated potassium. Factors affecting the ontogeny of neuropeptide expression in, and release from, spinal cord neurons can now be examined in vitro in a strictly defined neurochemical environment.     

Sympathetic sprouting in the dorsal root ganglia of the injured peripheral nerve in a rat neuropathic pain model

The extent of the sprouting of sympathetic postganglionic fibers in the dorsal root ganglion (DRG) and the peripheral nerves was examined in neuropathic rats at different postoperative times. After the L5 and L6 spinal nerves were ligated on one side, three different pain behavior tests (representing mechanical allodynia, cold allodynia, ongoing pain exacerbated by cold stress) were performed at various time intervals. The sympathetic postganglionic fibers were visualized by immunostaining with antibodies to tyrosine hydroxylase (TH). In the neuropathic rats, all three pain behaviors were fully developed within 3 days after the surgery, maintained up to 2 weeks, and then started to decline gradually afterward. At 20 weeks after neuropathic surgery, pain behaviors were reduced significantly compared to the peak response, but were still higher than the presurgery levels. Sympathectomy, performed 4 days after neuropathic surgery, almost completely abolished the signs of mechanical allodynia and ongoing pain behaviors, and it reduced the behaviors of cold allodynia to approximately half. The numerical density of sympathetic fibers in the DRG of an injured segment was significantly higher at 1, 4, and 20 weeks after neuropathic surgery as compared to the normal, suggesting that there is sprouting of sympathetic fibers in the DRG after peripheral nerve injury. Sprouting of sympathetic fibers in the DRG was extensive as early as 2 days after the spinal nerve ligation, and the sprouted fibers were almost completely eliminated after sympathectomy. The data suggest that sympathetic innervation of the DRG may play an important role in the development and maintenance of sympathetically maintained neuropathic pain. © 1996 Wiley-Liss, Inc.     

Distribution of calretinin mRNA in rat spinal cord and dorsal root ganglia: a study using non-radioactive in situ hybridization histochemistry

Using non-radioactive in situ hybridization calretinin mRNA was detected in numerous small neurons within lamina II and IV of the dorsal horn. Many labelled cells are distributed over the whole ventral horn; however, no motorneurons contained the mRNA. In dorsal root ganglia4.9 ± 1.7% (mean ± S.D., n = 5 animals) of the primary afferent neurons contained calretinin mRNA. Labelled cells were of intermediate and large size with diameters ranging from 36 to 68 μm indicating that calretinin is synthesized in neurons with myelinated afferetn fibers and presumably a corpuscular ending.     

Differences in developmental cell death between somatic and autonomic motor neurons of rat spinal cord

Considerable knowledge concerning developmental cell death has come from the study of somatic motor neurons (SMNs), but a related set of spinal neurons, the autonomic motor neurons (AMNs), have been studied less extensively in this respect. In the present study, we used three different approaches to determine the amount of AMN cell death during normal development in the rat. First, target dependency was studied in organotypic slice cultures, and it was found that AMNs survived for at least 12 days after removal of their postsynaptic targets. No factors were added to the serum-free medium to substitute for the ablated targets, indicating that AMNs were able to survive without target-derived trophic factors. Such target-independent survival is not characteristic of neurons that undergo typical developmental cell death. Second, AMNs were counted in double-stained choline acetyltransferase immunocytochemical and NADPH diaphorase histochemical preparations at ages (postnatal days 4–22) encompassing the period when AMN postsynaptic target cells undergo developmental death. Neuron numbers were essentially identical at all ages examined, indicating that no AMN cell death occurred postnatally. Finally, from embryonic day 13 to postnatal day 22, animals were analyzed by using terminal transferase-mediated nick-end labeling to identify dying cells. Many fewer labeled cells were observed among AMNs than among SMNs. Thus, all three approaches indicated that there is a significant SMN/AMN difference in developmental cell death. The phenotypic trait(s) that underlies this difference may also be important in the relative resistance of AMNs to pathological conditions that induce death of SMNs, e.g., those involved in amyotrophic lateral sclerosis and excitotoxicity. J. Comp. Neurol. 396:483–492, 1998. © 1998 Wiley-Liss, Inc.     

Morphine analgesia and newly synthesized 5-hydroxytryptamine in the dorsal and the ventral halves of the spinal cord of the rat

In the rat, morphine (5 mg/kg, s.c.) induced an increase in 5-hydroxytryptamine (5-HT) synthesis in the spinal cord. These effects appeared with a shorter latency and are much more marked in the dorsal half than in the ventral half. Although the increase in the dorsal half was slightly delayed by comparison with the onset of the analgesic effect the maxima for both phenomena were simultaneous. The role of the bulbo-spinal serotonergic system in morphine analgesia is discussed.     

Chronic immobilization stress: evidence for decreases of 5-hydroxy-tryptamine immunoreactivity and for increases of glucocorticoid receptor immunoreactivity in various brain regions of the male rat

Summary Male rats were exposed to severe 14 day immobilization stress. Body weight, body temperature, food and water intake, behavioral parameters, and serum corticosterone levels were measured during and after the stress period. On the 7th day after cessation of stress the experimental animals together with the control rats were taken to immunocytochemical analysis involving morphometry and microdensitometry of tyrosine hydroxylase (TH), 5-hydroxytryptamine (5-HT), various neuropeptide, and glucocorticoid receptor (GR) immunoreactivities (IRs) in a large number of regions of the central nervous system. In addition, adrenocorticotropic hormone (ACTH) IR was analyzed in the pituitary gland.Seven days following cessation of the chronic stress food intake, total locomotion and forward locomotion had been restored to normal. Serum corticosterone levels appeared to remain increased even 6 days following cessation of the chronic immobilization stress, probably caused by increased release of ACTH. Paraventricular corticotropin releasing hormone (CRF) IR was negatively correlated with the pituitary ACTH IR, indicating that the increase in ACTH release was produced by an increased release of CRF from the hypothalamus.The major immunocytochemical change observed 7 days after cessation of stress was a disappearance of 5-HT IR in the 5-HT cell groups B 1, B 2, B 3, and B 7. 5-HT IR in nerve terminals was only affected in the dorsal horn, where 5-HT IR was increased in the substantia gelatinosa. GR IR was found to be significantly increaed in monoaminergic cell groups: serotoninergic B 7, dopaminergic A 12, and noradrenergic A 1, A 2, and A 6. A trend for a reduction of TH IR was observed in nigral DA cells associated with significant reductions in TH IR in striatal DA nerve terminals. Finally, increases in 5-HT and substance P (SP) IR were found in the nerve terminals of the substantia gelatinosa of the cervical spinal cord in the stress group.In the present experimental model evidence has been obtained for a maintained activation of the hypothalamic-pituitary-adrenal axis as evaluated 7 days after cessation of severe chronic immobilization stress. The reduction of 5-HT IR in various 5-HT cell groups indicates a reduction of 5-HT synthesis, which may also be associated with reduced 5-HT release from the nerve terminals, since no depletion was observed in terminal regions and in one case an increase in 5-HT IR was noted (substantia gelatinosa). The increase in GR IR, demonstrated in the NA and 5-HT cell groups in the presence of a maintained hypersecretion of corticosterone may represent signs of an upregulation of GR synthesis and/or increased translocation, which take place in the presence of maintained hypersecretion of corticosterone. Thus, 5-HT and NA neurons may respond more effectively to circulating glucocorticoids after severe chronic stress. In this way glucocorticoids may protect against stress-induced exhaustion of neurons leading to impairment of transmission. Studies on TH IR suggest a deficit in the DA transmission line of the nigrostriatal DA neurons, but of no other CA neurons studied. Such effects may contribute to behavioral suppression. Finally, the stress-induced increases in 5-HT and SP IR in the substantia gelatinosa may in part underlie the phenomenon of stress-induced analgesia.     

Peptidylglycine alpha-amidating monooxygenase activity in spinal cord, dorsal roots, and dorsal root ganglia of Macaca fascicularis

Several peptides synthesized by primary sensory neurons are alpha-amidated at the C-terminal residue, including vasoactive intestinal polypeptide, neurokinin A and substance P, which is also abundant in spinal cord. In pituitary and other tissues, the C-terminal amidation is catalyzed by peptidylglycine alpha-amidating monooxygenase (PAM). In the present study, soluble PAM activity in spinal cord and in primary sensory neurons is quantified and characterized as to cofactor and cosubstrate requirements and substrate specificity.     

VGLUT1 and VGLUT2 innervation in autonomic regions of intact and transected rat spinal cord

Fast excitatory neurotransmission to sympathetic and parasympathetic preganglionic neurons (SPN and PPN) is glutamatergic. To characterize this innervation in spinal autonomic regions, we localized immunoreactivity for vesicular glutamate transporters (VGLUTs) 1 and 2 in intact cords and after upper thoracic complete transections. Preganglionic neurons were retrogradely labeled by intraperitoneal Fluoro-Gold or with cholera toxin B (CTB) from superior cervical, celiac, or major pelvic ganglia or adrenal medulla. Glutamatergic somata were localized with in situ hybridization for VGLUT mRNA. In intact cords, all autonomic areas contained abundant VGLUT2-immunoreactive axons and synapses. CTB-immunoreactive SPN and PPN received many close appositions from VGLUT2-immunoreactive axons. VGLUT2-immunoreactive synapses occurred on Fluoro-Gold-labeled SPN. Somata with VGLUT2 mRNA occurred throughout the spinal gray matter. VGLUT2 immunoreactivity was not noticeably affected caudal to a transection. In contrast, in intact cords, VGLUT1-immunoreactive axons were sparse in the intermediolateral cell column (IML) and lumbosacral parasympathetic nucleus but moderately dense above the central canal. VGLUT1-immunoreactive close appositions were rare on SPN in the IML and the central autonomic area and on PPN. Transection reduced the density of VGLUT1-immunoreactive axons in sympathetic subnuclei but increased their density in the parasympathetic nucleus. Neuronal cell bodies with VGLUT1 mRNA occurred only in Clarke's column. These data indicate that SPN and PPN are densely innervated by VGLUT2-immunoreactive axons, some of which arise from spinal neurons. In contrast, the VGLUT1-immunoreactive innervation of spinal preganglionic neurons is sparse, and some may arise from supraspinal sources. Increased VGLUT1 immunoreactivity after transection may correlate with increased glutamatergic transmission to PPN.     

Increased brain-derived neurotrophic factor immunoreactivity in rat dorsal root ganglia and spinal cord following peripheral inflammation

Our recent study showed that peripheral inflammation induced an increased expression of brain-derived neurotrophic factor (BDNF) mRNA which was mediated by nerve growth factor (NGF) in the dorsal root ganglion (DRG). In the present study, we evaluated the change of BDNF immunoreactivity in the DRG and spinal cord following peripheral inflammation by means of immunohistochemistry. Significant increases in the percentage of BDNF-immunoreactive (IR) neuron profiles in the L5 DRG and marked elevation in the expression of BDNF-IR terminals in the spinal dorsal horn were observed following peripheral tissue inflammation produced by an intraplantar injection of Freund's adjuvant into the rat paws. These findings suggest that peripheral tissue inflammation induces an increased BDNF synthesis in the DRG and an elevated anterograde transport of BDNF to the spinal dorsal horn. The functional role of this increased BDNF was discussed briefly.     

Ontological study of calbindin-D28k-like and parvalbumin-like immunoreactivities in rat spinal cord and dorsal root ganglia

The calcium ion plays an important role in some critical developmental events in the nervous system, such as neurulation and neurite elongation. Therefore, as the intracellular calcium-binding proteins calbindin-D28k (CaB) and parvalbumin (PV) may be expressed in these developmental events. Accordingly, the ontological expression of CaB and PV was examined immunocytochemically in the spinal cord and dorsal root ganglia (DRG) of the rat, in order to evaluate the relationship between CaB and PV expression, and other important developmental events. During the ontogenesis of the spinal cord, the CaB-like immunoreactivity was mainly observed in the cell somata. The immunoreactive cells in the ventral horn of the cervical and thoracic, lumbar, and sacral segments first appeared at embryonic day (E)-12, E-13, and E-14, respectively. However, these cells were not detected in the intermediate gray matter of the same segments at E-14, E-15, and E-16, respectively, and in the dorsal horn at E-14-E-15, E-16, and E-17, respectively. The peak of immunoreactive cells, both as to number and intensity, occurred in the perinatal period. However, from postnatal day (P)-14 on, the number and intensity of the positive cells decreased, the adult levels being reached at P-35. The PV-like immunoreactivity was mainly detected in the fibers and punctata during the ontogenesis of the spinal cord. The immunoreactive fibers first appeared on the surface of the dorsal horn in the cervical and thoracic segments at E-14, then entered the dorsal horn at E-15, and reached the intermediate gray matter and ventral horn at E-16. The first appearance of these fibers in the same areas of the lumbar and sacral segments occurred 1 day later than in the cervical and thoracic segments. During the perinatal period, the maximum content of PV-like immunoreactive fibers, together with many punctata, was seen in the gray matter. However, between P-14 and P-17, most of them lost immunoreactivity rapidly, with the exception of the medial region of the intermediate gray matter, where the PV-immunoreactive punctata remained up to the adult stage. In DRG neurons, both CaB and PV was expressed, but in different neurons. Neurons labeled with anti-CaB and anti-PV sera were first detected at E-16 and E-14, respectively. These neurons were large or medium-sized in the prenatal period.(ABSTRACT TRUNCATED AT 400 WORDS)