Cholinesterases of rat sympathetic ganglia after immunosympathectomy, decentralization and axotomy

Abstract— Immunosympathectomy was produced in Sprague-Dawley rats by the subcutaneous injection of 300 units of nerve growth factor (NGF)-antiserum (1.56 mg of freeze-dried serum)/g/day for 6 days, the first dose being given 5–8 hr after birth. The immunosympathectomized rats and their control littermates were killed 2½ and 7 months after birth. Ganglionic acetylcholinesterase and pseudocholinesterase activities were measured by an adaption (Kungman , Kungman and Pouszczuk , 1968) of the colorimetric method of Ellman , Courtney , Andres and Featherstone (1961). Following immunosympathectomy the activities of these enzymes decreased significantly in superior cervical, stellate, thoracic chain, cardiac (abdominal), coeliac and superior mesenteric ganglia. The reduction of the acetylcholinesterase activity was greater than expected in a number of sympathetic ganglia, e.g. superior cervical, stellate, coeliac and cardiac ganglia, if one considered that only the postganglionic neurons were affected by immunosympathectomy. The activities of these enzymes were also reduced in the cervical sympathetic trunks from NGF-antiserum-treated rats. By means of decentralization and axotomy it was shown that 45 per cent of the total ganglionic acetylcholinesterase activity was associated with the preganglionic and 55 per cent with the postganglionic elements of the superior cervical ganglion from control rats. It was concluded that immunosympathectomy also affects the preganglionic sympathetic neurons. It is not known whether this is a primary effect of the NGF-antiserum or a secondary effect resulting from the absence of over 90 per cent of the postganglionic sympathetic cell bodies.     

Changes in the macrophage population of the rat superior cervical ganglion after postganglionic nerve injury

Following peripheral nerve transection, a series of biochemical changes occurs in axons and Schwann cells both at the site of lesion and distal to it. Macrophages differentiated from monocytes that invade the area in response to transection (elicited macrophages) and, perhaps, also macrophages normally present in the tissue (resident macrophages) play important roles in these changes. In addition, nerve transection produces changes in the cell bodies of axotomized neurons and their surrounding glial cells, located at some distance from the lesion. To determine whether macrophages might play a role in the changes occurring in the superior cervical ganglion (SCG) after axotomy, we examined the presence of macrophages before and after axonal damage. The monoclonal antibodies ED1, ED2, and OX6 were used, each of which recognizes a somewhat different population of macrophages. Ganglia from normal rats contained a population of resident cells that were ED2+ but very few that were ED1+. Within 2 days after the postganglionic nerves were transected, the number of ED1+ cells increased substantially, with little change in immunostaining for ED2. These data, in combination with published studies on other tissues, suggest that ED1 in the SCG is selective for elicited macrophages and ED2 for resident macrophages. OX6 immunostaining was prominent in normal ganglia but also increased significantly after axotomy, suggesting that it reflects both macrophage populations. Systemic administration of 6-hydroxydopamine, a neurotoxin that causes the destruction of sympathetic nerve endings, also produced an increase in ED1 immunostaining. Thus, the change in ED1 immunostaining in the SCG does not require surgery, with the attendant servering of local blood vessels and connective tissue, but rather only the disconnection of sympathetic neurons from their end organs. The time course of the invasion of monocytes after axotomy indicates that this process is not required to trigger the biochemical changes occurring in the ganglion within the first 24 h. On the other hand, the existence of a resident population of macrophages raises the possibility that changes in those cells might be involved. © 1995 John Wiley & Sons, Inc.     

Phenotypic alterations of neuropeptide Y and calcitonin gene-related peptide-containing neurons innervating the rat temporomandibular joint during carrageenan-induced arthritis

The aim of this study was to identify immunoreactive neuropeptide Y (NPY) and calcitonin gene-related peptide (CGRP) neurons in the autonomic and sensory ganglia, specifically neurons that innervate the rat temporomandibular joint (TMJ). A possible variation between the percentages of these neurons in acute and chronic phases of carrageenan-induced arthritis was examined. Retrograde neuronal tracing was combined with indirect immunofluorescence to identify NPY-immunoreactive (NPY-IR) and CGRP- immunoreactive (CGRP-IR) neurons that send nerve fibers to the normal and arthritic temporomandibular joint. In normal joints, NPY-IR neurons constitute 78±3%, 77±6% and 10±4% of double-labeled nucleated neuronal profile originated from the superior cervical, stellate and otic ganglia, respectively. These percentages in the sympathetic ganglia were significantly decreased in acute (58±2% for superior cervical ganglion and 58±8% for stellate ganglion) and chronic (60±2% for superior cervical ganglion and 59±15% for stellate ganglion) phases of arthritis, while in the otic ganglion these percentages were significantly increased to 19±5% and 13±3%, respectively. In the trigeminal ganglion, CGRP-IR neurons innervating the joint significantly increased from 31±3% in normal animals to 54±2% and 49±3% in the acute and chronic phases of arthritis, respectively. It can be concluded that NPY neurons that send nerve fibers to the rat temporomandibular joint are located mainly in the superior cervical, stellate and otic ganglia. Acute and chronic phases of carrageenan-induced arthritis lead to an increase in the percentage of NPY-IR parasympathetic and CGRP-IR sensory neurons and to a decrease in the percentage of NPY-IR sympathetic neurons related to TMJ innervation.Key words: trigeminal ganglion, otic ganglion, superior cervical ganglion, arthritis, temporomandibular joint.     

Effects of Denervation and Axotomy on Nervous System-Specific Protein, Ornithine Decarboxylase, and Other Enzyme Activities in the Superior Cervical Sympathetic Ganglion of the Rat

The time courses of changes of three enolase isozymes (alpha alpha, alpha gamma, and gamma gamma), S-100 protein, 2',3'-cyclic nucleotide 3'-phosphodiesterase (CNPase), ornithine decarboxylase (ODC), beta-galactosidase, and glucose-6-phosphate dehydrogenase (G6PDH) were examined from 1 to 14 days after cutting of the preganglionic nerve (denervation) or the postganglionic nerve (axotomy) of the superior cervical sympathetic ganglion (SCG) of the rat. The wet weight and protein content in the axotomized SCG increased continuously, to nearly twice those of the denervated SCG for 1-2 weeks after the operations. Among enolase isozymes in the SCG, neuron-specific gamma gamma-enolase decreased rapidly after denervation and stayed at a low level for 2 weeks, whereas the isozyme remained almost unchanged after axotomy. On the contrary, ganglionic alpha alpha-enolase and the alpha gamma-hybrid form increased remarkably to reach a maximum at the second day after axotomy, and remained above control for 1 to 2 weeks; these two enolase isozymes showed little change after denervation. Denervation caused a much larger increase than did axotomy in the ganglionic S-100 protein, an astrocyte-specific protein, during the first week after the operation, while the protein content decreased after 2 weeks of either denervation or axotomy. CNPase, a myelin-associated enzyme, rose suddenly 2 days after axotomy, and remained at a rather high level compared with the denervated ganglion, which showed little variation.(ABSTRACT TRUNCATED AT 250 WORDS)     

Reduction in nerve growth factor availability leads to a conditioning lesion-like effect in sympathetic neurons

Axotomized peripheral neurons are capable of regeneration, and the rate of regeneration can be enhanced by a conditioning lesion (i.e., a lesion prior to the lesion after which neurite outgrowth is measured). A possible signal that could trigger the conditioning lesion effect is the reduction in availability of a target-derived factor resulting from the disconnection of a neuron from its target tissue. We tested this hypothesis with respect to nerve growth factor (NGF) and sympathetic neurons by administering an antiserum to NGF to adult mice for 7 days prior to explantation or dissociation of the superior cervical ganglion (SCG) and subsequently measuring neurite outgrowth. The antiserum treatment dramatically lowered the concentration of NGF in the SCG and increased the rate of neurite outgrowth in both explants and cell cultures. The increase in neurite outgrowth was similar in magnitude to that seen after a conditioning lesion. To determine if exogenous NGF could block the effect of a conditioning lesion, mice were injected with NGF or cytochrome C immediately prior to unilateral axotomy of the SCG, and for 7 days thereafter. A conditioning lesion effect of similar magnitude was seen in NGF-treated and control animals. While NGF treatment increased NGF levels in the contralateral control ganglion, it did not significantly elevate levels in the axotomized ganglion. The results suggest that the decreased availability of NGF after axotomy is a sufficient stimulus to induce the conditioning lesion effect in sympathetic neurons. While NGF administration did not prevent the conditioning lesion effect, this may be due to the markedly decreased ability of sympathetic neurons to accumulate the growth factor after axotomy.     

Met5-enkephalin-Arg6-Gly7-Leu8-immunoreactive nerve fibers in the major salivary glands of the rat: evidence for both sympathetic and parasympathetic origin

Summary The localization of the proenkephalin A-derived octapeptide, Met⁵-enkephalin-Arg⁶-Gly⁷-Leu⁸ (MEAGL), was studied in the major salivary glands of Sprague-Dawley and Wistar rats with the indirect immunofluorescence method. MEAGL-immunoreactive nerve fibers were found around the acini, along intra-and interlobular salivary ducts and in close contact with blood vessels. In the parotid and submandibular glands tyrosine hydroxylase (TH) immunoreactivity was observed in nerve fibers around the acini, in association with intra- and interlobular salivary ducts and around blood vessels, while in the sublingual gland TH-immunoreactive nerve fibers were only seen around blood vessels. Parasympathetic neurons in submandibular ganglia contained MEAGL immunoreactivity. Moderate TH immunoreactivity was seen in some neurons of the submandibular ganglia. A subpopulation of sympathetic principal neurons in the superior cervical ganglion were immunoreactive for both MEAGL and TH. In the trigeminal ganglion, no MEAGL-immunoreactive sensory neurons or nerve fibers were observed. Superior cervical ganglionectomies resulted in a complete disappearance of TH-immunoreactive nerve fibers, while MEAGL-immunoreative nerve fibers were still present in the glands. The presence of MEAGL immunoreactivity in neurons of both sympathetic superior cervical ganglia and parasympathetic submandibular ganglia and the results of superior cervical ganglionectomies suggest, that MEAGL-immunoreactive nerve fibers in the major salivary glands of the rat have both sympathetic and parasympathetic origin.     

Calcium-Dependent Transglutaminase of Rat Sympathetic Ganglion in Development and After Nerve Injury

The activity of transglutaminase (TG) was examined in the rat superior cervical ganglion (SCG) during development and after postganglionic nerve crush. During postnatal development the enzyme activity is increased by sevenfold in parallel to protein content of the ganglion and reaches adult levels by day 35 after birth. The endogenous activity (enzyme activity assayed in the absence of the exogenous substrate) during development is transiently elevated with a peak at day 21 postnatal. In the adult ganglion the enzyme specific activity is evenly distributed in all subcellular compartments, but most of it is contained in the cytosol. Within the first hour after axotomy TG activity is rapidly and transiently elevated. The peak value, 80% above control levels, is attained by 30 min postoperative. At this time the activity is increased in all subcellular fractions, but the endogenous activity is selectively increased in the fraction containing nuclei. The enhanced TG activity after axotomy can be prevented by topical treatments with verapamil, an inhibitor of voltage-dependent calcium fluxes across excitable membranes, or with the calcium chelator EGTA. The results show that intracellular TG activity is present in the SCG and that it increases with postnatal growth of the ganglion. After axotomy the enzyme activity is rapidly and transiently increased in the ganglion and this elevation critically depends on calcium fluxes.     

Neuroanatomy of morphine-modulating peptides

Antisera against two mammalian peptides related to the molluscan cardioexcitatory peptide Phe-Met-Arg-Phe-NH2 were used to locate immunoreactive neurons in the rat brain, nerve fibres and terminals in the spinal cord, sympathetic ganglion cells and adrenal chromaffin cells. Immunoreactivity for the newly characterised octa- and octadecapeptide was detected in nerve cell bodies in the hypothalamic area, including parts of the dorsomedial, periventricular and paraventricular nuclei, and in the nucleus tractus solitarii. Nerve terminals in the superficial laminae of the spinal cord were also immunoreactive for these peptides, while the sensory ganglia were nonreactive. Some principal ganglion cells in the superior cervical ganglia exhibited bright immunofluorescence for the peptides, and a few adrenal medullary cells were immunoreactive. The presence of these peptides in the substantia gelatinosa of the spinal cord suggests that they may be involved in sensory neurotransmission, especially in the mechanisms mediating pain. In the hypothalamo-hypophysial system these peptides may be involved in the regulation of hormonal systems. They may also act as co-transmitters in the sympathetic nervous system.     

Reduction in nerve growth factor availability leads to a conditioning lesion‐like effect in sympathetic neurons

Axotomized peripheral neurons are capable of regeneration, and the rate of regeneration can be enhanced by a conditioning lesion (i.e., a lesion prior to the lesion after which neurite outgrowth is measured). A possible signal that could trigger the conditioning lesion effect is the reduction in availability of a target‐derived factor resulting from the disconnection of a neuron from its target tissue. We tested this hypothesis with respect to nerve growth factor (NGF) and sympathetic neurons by administering an antiserum to NGF to adult mice for 7 days prior to explantation or dissociation of the superior cervical ganglion (SCG) and subsequently measuring neurite outgrowth. The antiserum treatment dramatically lowered the concentration of NGF in the SCG and increased the rate of neurite outgrowth in both explants and cell cultures. The increase in neurite outgrowth was similar in magnitude to that seen after a conditioning lesion. To determine if exogenous NGF could block the effect of a conditioning lesion, mice were injected with NGF or cytochrome C immediately prior to unilateral axotomy of the SCG, and for 7 days thereafter. A conditioning lesion effect of similar magnitude was seen in NGF‐treated and control animals. While NGF treatment increased NGF levels in the contralateral control ganglion, it did not significantly elevate levels in the axotomized ganglion. The results suggest that the decreased availability of NGF after axotomy is a sufficient stimulus to induce the conditioning lesion effect in sympathetic neurons. While NGF administration did not prevent the conditioning lesion effect, this may be due to the markedly decreased ability of sympathetic neurons to accumulate the growth factor after axotomy. © 2006 Wiley Periodicals, Inc. J Neurobiol, 2006     

大鼠脊神经节内交感神经支配的荧光组织化学与HRP示踪观察

本研究应用乙醛酸诱发儿茶酚胺（ＣＡ）荧光技术观察大鼠肾上腺素（ＮＡ）能神经在脊神经节内的分布;并应用ＨＲＰ顺、逆行追踪技术对脊神经节内ＮＡ能神经纤维的起源及其与脊神经节神经元的关系进行了探讨。荧光组织化学观察发现、有些神经节神经元胞体周围分布有带膨体的ＮＡ能神经末梢;有的紧密围绕脊神经节细胞——卫星细胞复合体。颈上交感神经节内注射霍乱毒素Ｂ亚单位结合ＨＲＰ（ＣＢ┐ＨＲＰ）,在同侧Ｃ３～６节段脊神经节内可见标记的点状纤维末梢紧邻于节细胞旁。Ｔ１１～Ｌ２节段脊神经节内注射ＨＲＰ后,在同侧椎旁交感链（Ｔ９～Ｌ１）内可见标记的交感节后神经元胞体。上述实验结果表明,交感节后神经元发出节后纤维可直接到达脊神经节内,与节细胞发生接触。本研究提示、交感神经在脊神经节水平可能参与躯体初级传入信息的调制     

Localization of fibroblast growth factor 2 (FGF-2) protein and the receptors FGFR 1–4 in normal human seminiferous epithelium

 Fibroblast growth factor 2 (FGF-2), which occurs in various isoforms both species and tissue specifically, regulates cell proliferation and differentiation via a dual receptor system consisting of heparan sulphate proteoglycans and receptor tyrosine kinases (FGFRs). This study demonstrates for the first time the distribution pattern of FGF-2 and the receptors FGFR 1–4 in the normal seminiferous epithelium of adult men. In western blot analyses, the polyclonal antibody, anti-FGF-2, shows two immunoreactive bands at 18 and 24 kDa. On paraffin sections, positive immunoreaction occurs within the cytoplasm of spermatogonia. The distribution pattern of the polyclonal anti-FGFR 1–4 antibodies is as follows: anti-FGFR-1 (one 68-kDa band) stains nuclei and cytoplasm of spermatogonia; anti-FGFR-3 (five bands at 68, 78, 105, 125 and 145 kDa) stains the nuclei of all germ cells except those of elongated spermatids; and anti-FGFR-4 (one 48-kDa band) stains the cytoplasm of primary pachytene spermatocytes. We were unable to demonstrate FGFR-2 immunoreactivity either in western blot analysis or on paraffin sections. This distribution pattern suggests that FGF-2 in spermatogonia is involved in the autocrine and paracrine regulation of the proliferation and differentiation of spermatogonia and spermatocytes via the receptors FGFR-1, FGFR-3 and FGFR-4. Accepted: 23 December 1997     

Cellular reactions to axotomy in rat superior cervical ganglia includes apoptotic cell death

The cellular response to axonal injury in the superior cervical ganglion was examined by immunofluoresence at intervals from 6 h to 14 days after transection of the internal and external carotid nerves. GAP-43-immunoreactivity (IR) appeared in some neurons in the ganglia 1 day after axotomy, while neurons in control ganglia were GAP-43 negative. In 3 days axotomized ganglia GAP-43-IR structures were increased in number and intensity in nerve fiber bundles, while GAP-43-positive perikarya were restricted to the middle and caudal parts of the ganglia and showed an intensity that was stronger than at 1 day after axotomy. These GAP-43-positive neurons were also galanin positive. In the cranial part of the ganglia, S100-IR in satellite cells was weak at 18 h after axotomy. Peripheral to this area, S100-IR was stronger and co-localized with HSP-72-IR, preferentially located in satellite cells. HSP-72-IR was, however, occasionally observed also in principal neurons at 1 and 3 days after axotomy. In eosin-stained sections, neurons and satellite cells in the cranial part of 1 day axotomized ganglia were reduced in number, and a further loss was noted at 3 days. At 12 h some satellite cells in the cranial part of the ganglia were labelled by the in situ DNA 3'-end labelling method, indicating apoptosis, and at 18 h many cells were labelled. Some neuronal perikarya were also labelled in this region. Labelling was not observed at 1 day or later after axotomy, nor in control ganglia. The results may imply that not only neurons but also satellite cells react to neuronal axonal injury with apoptosis. Neurons in the middle and caudal part of the ganglia survived and showed increased content of GAP-43 and galanin, possibly a sign of regeneration/neuronal plasticity.     

Sympathetic neurons synthesize and secrete pro‐nerve growth factor protein

Postmitotic sympathetic neuronal survival is dependent upon nerve growth factor (NGF) provided by peripheral targets, and this dependency serves as a central tenet of the neurotrophic hypothesis. In some other systems, NGF has been shown to play an autocrine role, although the pervasiveness and significance of this phenomenon within the nervous system remain unclear. We show here that rat sympathetic neurons synthesize and secrete NGF. NGF mRNA is expressed in nearly half of superior cervical ganglion sympathetic neurons at embryonic day 17, rising to over 90% in the early postnatal period, and declining in the adult. Neuronal immunoreactivity is reduced when retrograde transport is interrupted by axotomy, but persists in a subpopulation of neurons despite diminished mRNA expression, suggesting that intrinsic protein synthesis occurs. Cultured neonatal neurons express NGF mRNA, which is maintained even when they are undergoing apoptosis. To determine which NGF isoforms are secreted, we performed metabolic labeling and immunoprecipitation of NGF‐immunoreactive proteins synthesized by cultured NGF‐dependent and ‐independent neurons. Conditioned medium contained high molecular weight NGF precursor proteins, which varied depending upon the state of NGF dependence. Mature NGF was undetectable by these methods. High molecular weight NGF isoforms were also detected in ganglion homogenates, and persisted at diminished levels following axotomy. We conclude that sympathetic neurons express NGF mRNA, and synthesize and secrete pro‐NGF protein. These findings suggest that a potential NGF‐sympathetic neuron autocrine loop may exist in this prototypic target‐dependent system, but that the secreted forms of this neurotrophin apparently do not support neuronal survival. © 2003 Wiley Periodicals, Inc. J Neurobiol 38–53, 2003     

Proliferation of neointimal smooth muscle cells after arterial injury. Dependence on interactions between fibroblast growth factor receptor-2 and fibroblast growth factor-9

The growth factor signaling mechanisms responsible for neointimal smooth muscle cell (SMC) proliferation and accumulation, a characteristic feature of many vascular pathologies that can lead to restenosis after angioplasty, remain to be identified. Here, we examined the contribution of fibroblast growth factor receptors (FGFRs) 2 and 3 as well as novel fibroblast growth factors (FGFs) to such proliferation. Balloon catheter injury to the rat carotid artery stimulated the expression of two distinctly spliced FGFR-2 isoforms, differing only by the presence or absence of the acidic box, and two distinctly spliced FGFR-3 isoforms containing the acidic box and differing only by the presence of either the IIIb or IIIc exon. Post-injury arterial administration of recombinant adenoviruses expressing dominant negative mutant forms of these FGFRs were used to assess the roles of the endogenous FGFR isoforms in neointimal SMC proliferation. Dominant negative FGFR-2 containing the acidic box inhibited such proliferation by 40%, whereas the dominant negative FGFR-3 forms had little effect. Expression of FGF-9, known to be capable of binding to all four neointimal FGFR-2/-3 isoforms, was abundant within the neointima. FGF-9 markedly stimulated both the proliferation of neointimal SMCs and the activation of extracellular signal-related kinases 1/2, effects which were abrogated by the administration of antisense FGF-9 oligonucleotides to injured arteries and the expression of the dominant negative FGFR-2 adenovirus in cultured neointimal SMCs. These studies demonstrate that, although multiple FGFRs are induced in neointimal SMCs following arterial injury, specific interactions between distinctly spliced FGFR-2 isoforms and FGF-9 contribute to the proliferation of these SMCs.     

Sympathetic neurons synthesize and secrete pro-nerve growth factor protein

Postmitotic sympathetic neuronal survival is dependent upon nerve growth factor (NGF) provided by peripheral targets, and this dependency serves as a central tenet of the neurotrophic hypothesis. In some other systems, NGF has been shown to play an autocrine role, although the pervasiveness and significance of this phenomenon within the nervous system remain unclear. We show here that rat sympathetic neurons synthesize and secrete NGF. NGF mRNA is expressed in nearly half of superior cervical ganglion sympathetic neurons at embryonic day 17, rising to over 90% in the early postnatal period, and declining in the adult. Neuronal immunoreactivity is reduced when retrograde transport is interrupted by axotomy, but persists in a subpopulation of neurons despite diminished mRNA expression, suggesting that intrinsic protein synthesis occurs. Cultured neonatal neurons express NGF mRNA, which is maintained even when they are undergoing apoptosis. To determine which NGF isoforms are secreted, we performed metabolic labeling and immunoprecipitation of NGF-immunoreactive proteins synthesized by cultured NGF-dependent and -independent neurons. Conditioned medium contained high molecular weight NGF precursor proteins, which varied depending upon the state of NGF dependence. Mature NGF was undetectable by these methods. High molecular weight NGF isoforms were also detected in ganglion homogenates, and persisted at diminished levels following axotomy. We conclude that sympathetic neurons express NGF mRNA, and synthesize and secrete pro-NGF protein. These findings suggest that a potential NGF-sympathetic neuron autocrine loop may exist in this prototypic target-dependent system, but that the secreted forms of this neurotrophin apparently do not support neuronal survival.     

Effect of preganglionic sympathectomy on the cholinesterase activity in the superior cervical ganglia of rats and hamsters

Summary By employing biochemical assay and histochemical enzyme techniques the effect of preganglionic sympathectomy on the cholinesterase (ChE) activity in the superior cervical ganglia of rats and hamsters was investigated. Biochemical assays indicate that the ChE activity in the superior cervical ganglia of adult rats and hamsters is 57.19 and 28.63 respectively (expressed in u moles acetylcholine hydrolyzed per min per g of tissue); two weeks after preganglionic denervation, about 50% and 60% of ChE activity are lost respectively. Histochemical enzyme examination reveals that in the rat superior cervical ganglion, the majority of the neurons are adrenergic with weak to moderate acetylcholinesterase (AChE) reaction and the minority of the neurons are cholinergic with strong AChE activity, while only one type of adrenergic neurons exhibits a weak AChE activity in the hamster superior cervical ganglion. The AChE activity is localized in the perinuclear area, in the cisternae of the rough surfaced endoplasmic reticulum, in the Golgi complex and on the plasma membrane of the hamster's neurons; it is mainly localized in the cisternae of the rough surfaced endoplasmic reticulum of the rat's neurons. AChE reaction product is also detected on the axolemmal membranes of the preganglionic nerve fibers in the sympathetic ganglia of rats and hamsters.After preganglionic sympathectomy, the AChE activity in the adrenergic neurons and in the preganglionic unmyelinated nerve fibers is markedly reduced, whereas the cholinergic neurons and preganglionic myelinated nerve fibers remain unchanged. On the basis of these results two conclusions have been reached: (1) The fact that strong AChE activity localized in the cholinergic neurons and preganglionic myelinated fibers is not influenced by denervation, suggests that these structures are able to produce AChE. (2) The reduction of AChE activity in the rat and hamster superior cervical ganglia two weeks after preganglionic denervation, observed by histochemical examination, can be correlated with a concomitant measurable reduction determined by biochemical assays.Supported in part by a grant from the National Science Council, Republic of China. The author wishes to express his gratitude to the Department of Pharmacology, College of Medicine, National Taiwan University, for the use of its equipment for biochemical assays     

Effects of hemicholinium-3 and sodium ions on choline uptake system in excised superior cervical sympathetic ganglia of rats

Active choline uptake by rat superior cervical sympathetic ganglia (SCG), which contain abundant cholinergic nerve terminals, was studied with respect to sensitivity to inhibition by hemicholinium-3 (HC-3) and dependence on extracellular Na⁺ under standard conditions of assay. Choline was taken up by a single saturable process with apparentK _m=3.07×10^–5 M and Vmax=286 pmoles/min/mg protein. Neither denervation followed by degeneration of cholinergic nerve terminals nor axotomy with successive neuronal degeneration significantly decreased in choline uptake by the ganglia in vitro. HC-3 dose-dependently inhibited ganglionic choline uptake more effectively at lower than at higher choline concentrations. HC-3 sensitive inhibition of ganglionic choline uptake was not seen in young rats one week after birth but appeared with maturity, attaining approximately 50% maximal inhibition in adult SCG. Extent of inhibition by HC-3 and Na⁺ dependence of ganglionic choline uptake was not altered by denervation or axotomy.Abbreviations used (HC-3) hemicholinium-3 - (HAChU) high affinity choline uptake - (LAChU) low affinity choline uptake - (SCG) superior cervical ganglia - (Ch) choline - (ACh) acetylcholine     

Synaptic Vesicle Proteins and Neuronal Plasticity in Adrenergic Neurons

The neurons in the superior cervical ganglion are active in plasticity and re-modelling in order to adapt to requirements. However, so far, only a few studies dealing with synaptic vesicle related proteins during adaptive processes have been published. In the present paper, changes in content and expression of the synaptic vesicle related proteins in the neurons after decentralization (cutting the cervical sympathetic trunk) or axotomy (cutting the internal and external carotid nerves) were studied. Immunofluorescence studies were carried out using antibodies and antisera against integral membrane proteins, vesicle associated proteins, NPY, and the enzymes TH and PNMT. For colocalization studies, the sections were simultaneously double labelled. Confocal laser scanning microscopy was used for colocalization studies as well as for semi-quantification analysis, using the computer software. Westen blot analysis, in situ 3'-end DNA labelling, and in situ hybridization were also employed. After decentralization of the ganglia several of the synaptic vesicle proteins (synaptotagmin I, synaptophysin, SNAP-25, CLC and GAP-43) were increased in the iris nerve terminal network, but with different time patterns, while TH-immunoreactivity had clearly decreased. In the ganglia, these proteins had decreased at 1 day after decentralization, probably due to degeneration of the pre-ganglionic nerve fibres and terminals. At later intervals, these proteins, except SNAP-25, had increased in the nerve fibre bundles and re-appeared in nerve fibres outlining the principal neurons.