Peptide 19 in the rat superior cervical ganglion

Peptide 19 is a 7.6 kDa polypeptide which can bind to calmodulin and inhibit calcium-calmodulin signaling. In this study, peptide 19-immunoreactivity was examined in the rat superior cervical ganglion. In the ganglion, 54.8% of postganglionic sympathetic neuron profiles were immunoreactive for peptide 19. These neuron profiles were small- to medium-sized and measured 87–845 μm² (mean±SD=343±111 μm²). Double immunofluorescence method revealed that 99.9% of peptide 19–containing neurons had neuropeptide Y in the superior cervical ganglion. Retrograde neuronal tracing and immunohistochemical studies also demonstrated that peptide 19 was common in postganglionic sympathetic neurons which innervated the facial skin and masseter but not the submandibular gland; 55.6% and 75.2% of cutaneous and muscular neuron profiles, respectively, contained peptide 19. Only 9.8% of glandular neurons were immunoreactive for peptide 19. These findings indicate that the content of peptide 19 in superior cervical ganglion neurons depends on their cell sizes and peripheral projections. On the other hand, colchicine injection into the superior cervical ganglion decreased the number of peptide 19–positive neurons (30.7%) compared to saline injection (53.3%). In contrast, the treatment induced nicotine adenine dinucleotide phosphate diaphorase activity in 12.7% of postganglionic sympathetic neurons. Double stain demonstrated that 56.3% of nicotine adenine dinucleotide phosphate diaphorase–positive neurons co-expressed peptide 19. These findings indicate that colchicine treatment causes decrease of peptide 19 expression and increase of nitric oxide synthase activity.     

Sodium and calcium currents of acutely isolated adult rat superior cervical ganglion neurons

Neurons enzymatically isolated from the adult rat superior cervical ganglion (SCG) were investigated using the whole-cell variant of the patch-clamp technique. Currentclamp studies revealed the following mean passive and active membrane properties: resting membrane potential, –54.9 mV; input resistance, 349 M; action potential (AP) threshold, –29.8 mV; AP overshoot, 53.3 mV; AP maximum rate of rise, 166.4 V/s; and AP duration, 3.2 ms. Chemosensitivity to acetylcholine remained intact following enzymatic dispersion. Voltage-clamp studies of a transient tetrodotoxin-sensitive Na⁺ current revealed activation and inactivation processes which could be fit to modified Boltzmann equations. Na⁺ current activation parameters for the half activation potential (Vh) and slope factor (K) were –23.3 mV and 5.3 mV, respectively. Inactivation parameters forVh andK were –59.3 mV and 7.6 mV, respectively. Voltage-clamp studies also revealed a high voltageactivated sustained inward current which was eliminated upon removal of external Ca²⁺, greatly reduced by 500 M Cd²⁺, and supported by Ba²⁺ or Sr²⁺. Tail current analysis of this Ca²⁺ current revealed a sigmoidal activation. A low voltage-activated transient Ca²⁺ current was not observed. We conclude that isolated SCG neurons retain the properties of neurons in intact ganglia and provide several advantages over conventional preparations for the study of voltagegated membrane currents.     

Somatostatin blocks a calcium current in acutely isolated adult rat superior cervical ganglion neurons

Somatostatin-like immunoreactivity has been reported to occur in the postganglionic neurons of sympathetic ganglia. We therefore investigated the effect of somatostatin (SOM) on the Ca2+ current in sympathetic neurons. Voltage-clamp recordings, using the whole-cell patch-clamp technique, were made from acutely isolated adult rat superior cervical ganglion (SCG) neurons in solutions (external and internal) designed to isolate Ca2+ currents. Application of 0.001-1.0 microM [D-Trp8]SOM resulted in a rapid, reversible and concentration-dependent decrease in the amplitude of the Ca2+ current evoked from a holding potential of -80 mV. The concentration-response relationship for SOM could be fitted to a single-site binding model with an apparent dissociation constant of 11 nM; the maximal attainable block of Ca2+ current by SOM was 50%. SOM also produced a pronounced slowing of the Ca2+ current rising phase, especially at more depolarized potentials. At higher concentrations (0.03-1.0 microM), prolonged application of SOM resulted in a progressive decrease in blocking ability. The results are consistent with a neurotransmitter and/or neuromodulator role for SOM in the sympathetic nervous system.     

Heterogeneous distribution of GABA-immunoreactive nerve fibers and axon terminals in the superior cervical ganglion of adult rat

The distribution of axons and axon varicosities containing GABA was studied in the superior cervical ganglion of rat by light and electron microscopic immunohistochemistry. Two different polyclonal antibodies were used, which had been made against GABA conjugated by glutardialdehyde to bovine serum albumin. GABA-like immunoreactivity occurred in many axons within the cervical sympathetic trunk and in axons and axon varicosities around the principal nerve cells in the superior cervical ganglion. GABA-positive axons were intermingled with non-stained axons, except for a small group of fibers in the trunk where the staining was absent. The rostral part of the ganglion and some scattered patches were more densely innervated by GABA-positive axons than the middle and caudal parts. Within dense areas, some of the large ganglion cells were abundantly surrounded by GABA-positive nerve fibers, while the vicinity of others was devoid of any immunoreactive axon terminals. None of the principal ganglion cells contained GABA-like immunoreactivity, although a class of small cells scattered within the ganglion was stained. Transection of the cervical sympathetic trunk for 11 days caused the disappearance of GABA-like positivity from most of the fibers, and only very little GABA-like staining was revealed in some small cells, which resembled satellite cells. Ultrastructurally, the GABA-positive nerve fibers were unmyelinated. However, their terminal branches and varicosities accumulated around the perikarya and dendrites of certain principal ganglion cells were partly wrapped in glial processes. The present results provide evidence that the superior cervical ganglion of adult rat receives a significant number of GABA-positive axons from the cervical sympathetic trunk and that these axons provide an innervation which is heterogeneously distributed within the superior cervical ganglion and on ganglionic cells. The source and function of the GABA-positive axons remain to be elucidated.     

Physiological classification of sympathetic neurons in the rat superior cervical ganglion

A new scheme is presented for identifying three sympathetic phenotypes in the rat superior cervical ganglion using electrophysiology and neuropeptide Y expression. Postganglionic compound action potentials recorded from the external and internal carotid nerves each contained two peaks, 1 and 2, with distinct preganglionic stimulus thresholds. Peak 2 in the external carotid response contained subpeaks 2a and 2b having a similar stimulus threshold. Neurons corresponding to peaks 1, 2a, and 2b were identified intracellularly by antidromic stimulation, graded preganglionic stimulation, injection with neurobiotin and immunostaining. Seventeen of 53 neurons studied this way had a low threshold for preganglionic stimulation of firing that corresponded to activation of extracellular peak 1. All low-threshold neurons were neuropeptide Y (NPY)-negative. The other 36 neurons had a high presynaptic stimulus threshold that corresponded to activation of extracellular peak 2, and 12 of these cells contained NPY. Together with other known features of ganglionic organization, the results indicate that low-threshold NPY-negative neurons are secretomotor cells projecting to salivary glands, that high-threshold NPY-negative neurons are pilomotor cells responsible for extracellular peak 2a, and that high-threshold, NPY-positive neurons are vasoconstrictor cells responsible for peak 2b. Secreto-, pilo-, and vasomotor neurons identified in this way had distinct axonal conduction velocities (0.52, 0.20, and 0.10 m/s) and diameters (33, 29, and 25 microm) but were indistinguishable in terms of preganglionic conduction velocities (0.30-0.34 m/s) and number of primary dendrites (8.4-8.6). The cell classification scheme presented here will allow future comparison of ganglionic integration in different sympathetic modalities.     

Competitive reinnervation of the rat superior cervical ganglion by foreign nerves

The efficacy of foreign nerves to form synapses was studied morphologically and physiologically in the superior cervical ganglion of the rat. The denervated ganglion was anastomosed to the central stump of either the vagus, the hypoglossal or both nerves together. The degree of reinnervation was assessed two to ten months later. We measured the strength of contraction of the nictitating membrane after each type of operation and compared this to the number, type and distribution of synapses in the same ganglion. Both the vagus and hypoglossal nerves preferentially reinnervated a population of neurones that are situated in the cranial pole of the superior cervical ganglion and supply the nictitating membrane. When both nerves were connected to the ganglion only the vagus nerve could be shown to reinnervate it, and no reinnervation by the hypoglossal nerve was detected. However, in this experiment neither foreign nerve did as well in competition as each did alone and the overall result was reduced functional efficiency.We conclude that not all sympathetic neurones are equivalent and that, just like sympathetic afferents, the foreign nerves are capable of selectively reinnervating preferred target cells.     

SHR鼠颈上交感神经节内NPY的差异表达研究

目的　通过对自发性高血压大鼠（SHR）颈上神经节形态学及其内神经肽酪氨酸（NPY）表达变化的研究,探讨NPY在高血压发生发展中的作用。 方法　随机选取成年SHR和WKY各20只,观测颈上交感神经节的位置、形状、大小及重量,采用Real-time PCR技术和免疫组织化学法,检测两组大鼠颈上神经节内NPY mRNA和蛋白的表达。 结果　与同周龄的WKY组大鼠相比较,SHR组大鼠血压明显升高（P<0.05）;Real-time PCR和免疫组化结果显示：SHR颈上神经节内NPY mRNA和蛋白水平均较WKY增加（P<0.05）。 结论　NPY在基因转录和蛋白表达等方面均较WKY上调,并参与高血压的形成。     

Evidence for a blood-ganglion barrier in the superior cervical ganglion of the rat

The permeability of the blood vessels in the superior cervical ganglion of the rat was tested by intravenous injection of horseradish peroxidase (HRP). By light microscopy, peroxidase activity was found in three locations: in the capsule of the ganglion, in the lumina of the blood vessels, and within macrophages. Electron microscopy revealed that virtually all ganglionic blood vessels contained HRP 5 minutes following its administration. The intensity of peroxidase activity declined over the period of 15 minutes. The enzyme was localized on the luminal surface of the endothelial cells, attaching to the glycocalyx. Endothelial microvilli, projecting into the vessel lumen, were also covered with peroxidase. Micropinocytotic vesicles on the luminal surface of the endothelium contained reaction product. Some of these vesicles were free within the cytoplasm of the endothelium but none was observed on the abluminal surface. Peroxidase activity was not detected in the extracellular space even after 15 minutes. The majority of blood vessels in the superior cervical ganglion possess a continuous endothelium with tight junctions; features associated with the blood-brain barrier of the central nervous system and peripheral nerves. It is proposed that these vessels perform a barrier function between the capillary circulation and the superior cervical ganglion.     

Induction and maintenance of free postsynaptic membrane thickenings in the adult superior cervical ganglion

Summary The superior cervical ganglion (SCG) of adult rats was exposed to GABA, either by long lasting microapplication (implantation of glass bulbs for 1–24 days) or in short term experiments (external application up to 6 h). Autoradiography showed that [³H] GABA accumulated selectively in satellite cells. The GABA produced the following effects: (1) Specialized membrane thickenings-similar in fine structural appearance to those seen as postsynaptic membrane thickenings at Gray type I synapses — were formed at the extrasynaptic dendritic surface of principal ganglion cells. (2) Morphometry revealed that the surface to volume ratio of dendrites increased significantly corresponding to an enlargement of their extrasynaptic surface as a result of the formation of spine-like projections. (3) Electrophysiology confirmed that, at least after short term application, the action potentials induced by preganglionic stimulation were heavily suppressed. These results suggest that, in the course of depressed ganglionic activity, so-called free postsynaptic membrane thickenings are generated and maintained in the SCG of adult rats even in the absence of significant axonal degeneration. The discussion focuses on two points: (1) possible similarities between the conditions of neurons after denervation and under the influence of GABA; (2) a possible role of GABA and other substances with inhibitory action in synaptogenesis. On leave from: Laboratory of Molecular Neurobiology, Institute of Biophysics, Biological Research Centre, Szeged, Hungary. On leave from: Department of Comparative Physiology, József Attila University of Sciences, Szeged, Hungary.     

Division and migration of satellite glia in the embryonic rat superior cervical ganglion

Summary While distinct precursors committed to a neuronal or glial cell fate are generated from neural crest cells early in peripheral gangliogenesis, little is known about the subsequent generation and maturation of young satellite glia from restricted glial precursor cells. To examine the division and migration of glial precursor cells and their satellite cell progeny, morphological, immunocytochemical and culture techniques were applied to the developing rat superior cervical ganglion. At embryonic day (E)18.5, numerous clusters of nonneuronal cells appeared transiently in the ganglion. Individual cells with a similar morphology were present in E16.5 ganglia, and are likely to represent the precursor cells which generate these clusters. The clustered cells were distinguishable from neighbouring neurons as well as from endothelial cells and fibroblasts. Morphologically similar cells were present in nerve bundles at E18.5 and surrounding principal neurons and nerve bundles in the adult ganglion. Double-label studies of the E18.5 ganglion with tyrosine hydroxylase to identify noradrenergic neurons and propidium iodide counterstaining to visualize all cell nuclei revealed that the cells in clusters stained with propidium iodide but lacked tyrosine hydroxylase immunoreactivity. To determine if cell clusters arose from division, bromodeoxyuridine, a thymidine analogue, was administered to pregnant mothers between E16.5–E18.5, and ganglionic cells examined at E18.5 bothin vivo andin vitro. Numerous non-neuronal cells divided during this periodin situ and composed portions of clusters. When dissociated, superior cervical ganglion satellite glia reacted with an NGF-receptor antibody (MAb 217c) and possessed a flattened shape, in contrast to bipolar Schwann cells. Over half of the 217c-immunoreactive glia at E18.5 had incorporated bromodeoxyuridine during E16.5–18.5in vivo. At birth, non-neuronal cells were no longer grouped in clusters, but were associated with neuronal cell bodies and processes. These findings suggest that, between E16.5–E18.5, glial precursors divide rapidly to form clusters, and that, after the peak of neurogenesis, daughter cells migrate within the ganglion to associate with nerve cell bodies and processes where proliferation continues at a slower rate. Distinct cellular and molecular interactions are likely to trigger the initial rapid division of glial precursors, initiate their migration and association with neuron cell bodies, and control their subsequent slower division.     

Presence of neurons with GABA-like immunoreactivity in the superior cervical ganglion of the rat

Using antibodies raised against gamma-aminobutyric acid (GABA)-glutaraldehyde complexes, we have found neurons with GABA-like immunoreactivity in the superior cervical ganglion of adult rats. The processes of these neurons formed pericellular networks around the principal ganglion cells. Electron microscopy revealed that the immunoreactive dendrites were innervated by non-reactive axon terminals which formed asymmetrical synapses and probably originated from the preganglionic nerve. Axons with GABA-like immunoreactivity, especially axonal varicosities filled with synaptic vesicles, were found in direct apposition to principal ganglion cells. The GABA-positive axons and axon varicosities persisted in experimentally decentralized (deafferented) ganglia, suggesting that the perikarya of the immunoreactive neurons were intrinsic to the superior cervical ganglion. Taken together with data on inhibitory effects of GABA in sympathetic ganglia, these findings suggest that the superior cervical ganglion of rats contains a subpopulation of inhibitory interneurons which is GABAergic. This would indicate that GABAergic neurons do not only occur in the central but also in the peripheral nervous system.     

Expression of preproenkephalin mRNA in rat superior cervical ganglion during postnatal development

The number of principal neurons in the rat superior cervical ganglion (SCG) exhibiting enkephalin-peptide immunoreactivity is reported to be limited. To better determine the degree of enkephalinergic phenotype in sympathetic neurons, sections of SCGs from rats aged newborn to adult were processed for in situ hybridization histochemistry, using a [³⁵S]cRNA probe directed against preproenkephalin (PPENK). > 50% of principal ganglion neurons express mRNA for PPENK in adults. Striking variability in labeling intensity is observed. PPENK mRNA is detected in developing ganglia beginning at postnatal days 4–7. Both the number of cells and intensity of labeling increases with postnatal development. These results indicate that expression of PPENK mRNA is more widespread than expression of enkephalin peptides and develops postnatally.     

Deafferentation-induced increases in a synaptic vesicle protein in the adult rat superior cervical ganglion are associated with new protein synthesis

Previous studies have shown that deafferentation of the adult rat superior cervical ganglion results in a transient increase in levels of a 65 kDa synaptic vesicle membrane protein (SV). The present study indicates that the observed increase in SV after deafferentation is the result of new protein synthesis. Treatment with cycloheximide, a protein synthesis inhibitor, for 8 h at selected times after surgery produces decreases in SV which are greater than that observed after treatment of unoperated animals. The results suggest that an increased rate of synthesis of this protein is induced by deafferentation. Transsynaptic factors may play important roles in regulation of protein synthesis in sympathetic ganglia.     

5-Hydroxytryptamine-immunoreactive neurons and nerve fibers in the superior cervical ganglion of the rat

Indirect immunofluorescence method was used to localize 5-hydroxytryptamine-immunoreactive structures in the superior cervical ganglion of adult rats. In the ganglia of normal rats, 5-hydroxytryptamine immunoreactivity was localized in the small intensely fluorescent cells, but not in principal nerve cells. In the superior cervical ganglion of rats, pretreated with nialamide, a monoamine oxidase inhibitor, 150 mg/kg i.p., and the 5-hydroxytryptamine precursor, L-tryptophan, 45 mg/kg i.p., a large number of 5-hydroxytryptamine-immunoreactive principal nerve cells and small intensely fluorescent cells were detected. The immunoreactive principal nerve cells had long processes, and 5-hydroxytryptamine-immunoreactive nerve fibers were observed traversing the ganglion. In ganglia of rats pretreated with colchicine, occasional 5-hydroxytryptamine-immunoreactive principal nerve cells and several small intensely fluorescent cells were detected. Ligation of the main postganglionic nerve trunks of the superior cervical ganglion of normal rats resulted in the appearance of several 5-hydroxytryptamine-immunoreactive principal nerve cells and nerve fibers in the ganglion. To study whether the 5-hydroxytryptamine immunoreactivity in the superior cervical ganglion represented uptake or synthesis of 5-hydroxytryptamine, rats were injected with a specific 5-hydroxytryptamine uptake inhibitor, fluoxetine, 10 mg/kg i.p. twice a day for 5 days, and then they were treated with nialamide and L-tryptophan, as described above. In the superior cervical ganglion of fluoxetine-treated rats, a few 5-hydroxytryptamine-immunoreactive principal nerve and small intensely fluorescent cells, as well as some nerve fibers, were detected.(ABSTRACT TRUNCATED AT 250 WORDS)