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.     

A comparison of the changes in the non-neuronal cell populations of the superior cervical ganglia following decentralization and axotomy

Transecting the axons of neurons in the adult superior cervical ganglion (SCG; axotomy) results in the survival of most postganglionic neurons, the influx of circulating monocytes, proliferation of satellite cells, and changes in neuronal gene expression. In contrast, transecting the afferent input to the SCG (decentralization) results in nerve terminal degeneration and elicits a different pattern of gene expression. We examined the effects of decentralization on macrophages in the SCG and compared the results to those previously obtained after axotomy. Monoclonal antibodies were used to identify infiltrating (ED1+) and resident (ED2+) macrophages, as well as macrophages expressing MHC class II molecules (OX6+). Normal ganglia contained ED2+ cells and OX6+ cells, but few infiltrating macrophages. After decentralization, the number of infiltrating ED1+ cells increased in the SCG to a density about twofold greater than that previously seen after axotomy. Both the densities of ED2+ and OX6+ cells were essentially unchanged after decentralization, though a large increase in OX6+ cells occurred after axotomy. Proliferation among the ganglion's total non-neuronal cell population was examined and found to increase about twofold after decentralization and about fourfold after axotomy. Double-labeling experiments indicated that some of these proliferating cells were macrophages. After both surgical procedures, the percentage of proliferating ED2+ macrophages increased, while neither procedure altered the proliferation of ED1+ macrophages. Axotomy, though not decentralization, increased the proliferation of OX6+ cells. Future studies must address what role(s) infiltrating and/or resident macrophages play in regions of decentralized and axotomized neurons and, if both are involved, whether they play distinct roles.     

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     

Transmission pathways in the rat superior cervical ganglion

On isolated preparations of the superior cervical ganglion (SCG, n = 8) taken from 21-day-old rats, we studied the intraganglion pathways and mechanisms underlying generation of synaptic responses of SCG neurons to antidromic stimulation. One of the three nerves connected with the SCG was stimulated, and compound action potentials were recorded simultaneously from the other two nerves; then, the order of stimulated and recorded nerves was changed. Orthodromic stimulation of the cervical sympathetic nerve (CSN) evoked responses in the internal carotid nerve (ICN), which could be completely blocked by hexamethonium, and responses in the external carotid nerve (ECN), which contained a component that was not blocked by this of the ECN caused responses in the CSN, which were not blocked by hexamethonium. Effects of superfusion of the SCG with a Ca²⁺-free solution allowed us to conclude that the hexamethonium-insensitive component of the responses in the CSN and ECN and ECN-CSN conduction can be explained by the presence of direct fibers going from the CSN to the ECN with no synaptic relay. Possible mechanisms underlying antidromic stimulation-induced synaptic responses in SCG neurons are discussed. Neirofiziologiya/Neurophysiology, Vol. 39, Nos. 4/5, pp. 396–399, July–October, 2007.     

Evidence for transsynaptic regulation of neuronal cell surface heparan sulfate proteoglycan in developing rat superior cervical ganglion

The effect of neonatal deafferentation on the expression of a neuronal cell surface heparan sulfate proteoglycan (HeS-PG) was investigated in the developing rat superior cervical ganglion. Two monoclonal antibodies, one directed against the core protein of HeS-PG, and one to a determinant associated with a heparan sulfate side-chain, were used to monitor postnatal increases of HeS-PG by radioimmunoassay. Following neonatal deafferentation by section of the cervical sympathetic trunk, total protein per ganglion was slightly reduced at survival times of 7, 14, and 30 days. Expression of the core protein determinant on HeS-PG was not altered in deafferented ganglia. In contrast, levels of side-chain determinant were significantly reduced at 14 and 30 days. These results suggest that processing of HeS-PG side-chains by principal ganglionic neurons is partially regulated by transsynaptic influences during development. Transsynaptic regulation of neuronal development may be a more general process than was believed previously, with effects not limited to molecules associated with synaptic development.     

Rapid and protracted phases of retinal ganglion cell loss follow axotomy in the optic nerve of adult rats

To investigate the short-and long-term effects of axotomy on the survival of central nervous system (CNS) neurons in adult rats, retinal ganglion cells (RGCs) were labelled retrogradely with the persistent market diI and their axons interrupted in the optic nerve (ON) by intracranial crush 8 or 10 mm from the eye or in intraorbital cut 0.5 or 3 mm from the eye. Labelled RGCs were counted in flat-mounted retinas at intervals from 2 weeks to 20 months after axotomy. Two major patterns of RGC loss were observed: (1) an inital abrupt loss that was confined to the first 2 weeks after injury and was more severe when the ON was cut close to the eye; (2) a slower, persistent decline in RGC densities with one-half survival times that ranged from approximately 1 month after intraorbital ON cut to 6 months after intracranial ON crush. A small population of RGCs (approximately 5%) survived for as long as 20 months after intraorbital axotomy. The initial loss of axotomized RGCs presumably results from time-limited perturbations related to the position of the ON injury. A. persistent lack of terminal connectivity between RGCs and their targets in the brain may contribute to the subsequent, more protracted RGC loss, but the differences between intraorbital cut and intracranial crush suggest that additional mechanisms are involved. It is unclear whether the various injury-related processes set in motion in both the ON and the retina exert random effects on all RGCs or act preferentially on subpopulations of these neurons. © 1993 John Wiley & Sons, Inc.     

Expression of vitamin D receptors in the superior cervical ganglia of rats

We investigated the role of vitamin D in the sympathetic nervous system including the distribution of vitamin D receptors (VDR), 1α-hydroxylase and 24-hydroxylase (CYP24) in neuronal subpopulations and satellite glia in the superior cervical ganglia (SCGs) of rats using immunohistochemistry. VDR immunoreactivity was observed in the cytoplasm and nucleus of nearly all neurons in the SCG. Intensity of VDR fluorescence was significantly greater in the cytoplasm of neuropeptide Y (NPY) negative somata compared to NPY positive neurons. Immunoreactivity for 1α-hydroxylase also was observed in the cytoplasm of all neurons of the SCG, but the intensity of fluorescence was less in the nuclei. To the contrary, the immunoreactivity for CYP24 was stronger in the nuclei, although it was present at lower intensity also in the cytoplasm of neurons. VDR and 1α-hydroxylase immunofluorescence was observed in many non-neuron cells, except satellite glial cells, which exhibited weak CYP24 immunofluorescence. Expression of VDRs and key metabolizing enzymes indicated the importance of vitamin D in the autonomic nervous system and the ability of sympathetic neurons to activate and deactivate vitamin D for its autocrine and paracrine roles.     

Changes in the intracellular calcium concentration upon activation of rat superior cervical ganglion neurons

Changes in the intracellular calcium concentration induced by activation of neurons of the isolated intact rat superior cervical ganglion were recorded. It is concluded that stimulation within the physiological range of frequencies can effectively increase the intracellular calcium concentration in these neurons. Neirofiziologiya/Neurophysiology, Vol. 39, Nos. 4/5, pp. 400–402, July–October, 2007.     

Morphometric study of the superior cervical and stellate ganglia of spontaneously hypertensive rats during the prehypertensive stage

To compare the functional state of the superior cervical (SCG) and stellate sympathetic ganglia (SG) of spontaneously hypertensive rats (SHR) with those of age-matched normotensive Wistar Kyoto rats (WKY), ganglion cell volume and area occupied by ganglion cells relative to each whole ganglionic area were morphometrically examined using the Texture Analyse System (TAS) in rats at 0, 10 and 30 days of age. The weight of each ganglion relative to animal weight was also measured. The ganglion cell volume and the relative area of ganglionic cells in both ganglia of SHR were significantly larger (P<0.05) than those of age-matched WKY at ages 0 and 10 days after birth. The relative ganglionic weights of SHR were significantly larger (P<0.01) compared with those of WKY at all ages examined, except for SG at 0 days after birth. These results show that the relative volume of sympathetic ganglion cells is greater in both SCG and SG of SHR than that of WKY, suggesting that hyperfunction of sympathetic ganglia occurs at the prehypertensive stage as a primary factor in the development of hypertension in SHR.     

Induction of tyrosine hydroxlase synthesis in rat superior cervical ganglia in vitro by nerve growth factor and dexamethasone.

The addition of dexamethasone and nerve growth factor to organ cultures of superior cervical ganglia from young rats induces the synthesis of tyrosine hydroxylase. The combination of nerve growth factor and dexamethasone $\text{in vitro}$ produces a differential rate of tyrosine hydroxylase synthesis which approaches that obtained by the $\text{in vivo}$ administration of nerve growth factor.     

3′,5′‐Cyclic adenosine monophosphate regulates expression of synaptotagmin in neonatal sympathetic ganglia in vitro

The expression of the synaptic vesicle protein, synaptotagmin, in developing rat superior cervical ganglia is influenced by transsynaptic factors associated with membrane depolarization. The present study examines the role of cyclic AMP in the regulation of synaptotagmin in neonatal superior cervical ganglia maintained in explant culture. Ganglia were treated for 48 h in vitro with the Na⁺‐channel ionophore, veratridine, or with pharmacological agents that alter cyclic AMP levels. Levels of cyclic AMP and synaptotagmin were determined by radioimmunoassay. Veratridine treatment significantly increased cyclic AMP in cultured ganglia, with a long time course, and also increased synaptotagmin levels. Drugs that elevate cyclic AMP levels significantly increased synaptotagmin levels, with similar magnitude to that produced by veratridine treatment. These pharmacological agents did not alter neuron survival or total ganglionic protein content. No additive effects were observed after combined treatment with veratridine and pharmacological agents that increased cyclic AMP. Agents that blocked adenylyl cyclase blocked the veratridine‐induced increase in synaptotagmin levels. The results suggest that regulation of expression of synaptotagmin in neonatal sympathetic neurons is mediated partially by cyclic AMP. © 2001 John Wiley & Sons, Inc. J Neurobiol 46: 281–288, 2001     

大鼠坐骨神经切断后外源性bcl-2对脊髓运动神经元的保护作用

采用大鼠坐骨神经切断损伤模型,行神经外膜端端对线缝合,术中依不同组别,动物于神经缝合处远端0．5cm处分别注射人的正义和反义bcl-2重组腺病毒(Ad／s-bcl-2、Ad／as-bcl-2),报道基因重组腺病毒(Ad／lacZ)和生理盐水。术后48h,7d,15d和30d常规灌注固定大鼠,取L4-L6脊髓节段,应用X-gal染色、bel-2原位杂交和免疫组化染色、TUNEL染色以及乙酰胆碱酯酶(AChE)组织化学染色方法,观察到外源基因能在脊髓中表达,同时外源性Ad／s-bcl-2能显著减少L4到L6节段脊髓前角运动神经元凋亡的数目,减少脊髓前角运动神经元中因坐骨神经切断导致的AChE活性的降低幅度,并加快其恢复。而Ad／as-bcl-2可显著增加坐骨神经切断诱导的脊髓前角运动神经元凋亡数目以及AChE活性降低幅度,并延缓其恢复。这些观察结果表明,外源性bcl-2能保护周围神经切断后引起的脊髓运动神经元损伤。     

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     

Effects of oxotremorine and RMI 12330 A on [3H]acetylcholine release and adenylate cyclase activity in guinea pig superior cervical ganglion

There is considerable evidence that adenosine 3, 5-cyclic monophosphate (cAMP) is involved in the modulation of synaptic transmission in the guinea pig superior cervical ganglion (SCG). Presynaptic muscarinic receptors are known to attenuate, when activated, acetylcholine (ACh) release in the periphery as well as in the brain. Thus, the possible relationship between ganglionic adenylate cyclase activity and the output of ACh from electrically stimulated ganglia, preloaded with [³H]choline, was investigated. The muscarinic agonist oxotremorine significantly reduced in a dose-dependent manner the electrically evoked neurotransmitter release. The adenylate cyclase inhibitor N-(cis-2-phenylcyclopentyl)azacyclotridecan-2-imine hydrochloride (RMI 12330 A) also decreased ACh output. The inhibitory effects of these two drugs were additive. In crude ganglion membrane fractions oxotremorine significantly inhibited adenylate cyclase activity. The results indicate that drugs capable of inhibiting adenylate cyclase, significantly decrease ACh output from preganglionic nerve terminals in guinea pig SCG.