Rabbit Superior Cervical Ganglion Research Articles

Rhythmic Discharge Induced by Temperature Variation and Drugs in Isolated Sympathetic Ganglia

Isolated sympathetic ganglia from various mammalian species fire spontaneous, rhythmic discharge when exposed to low temperatures. Extracellular recording from rat, guinea pig, and rabbit superior cervical ganglia as well as dog lumbar ganglion revealed large single potentials or bursts of potentials, occurring at regular intervals, when the bath temperature was kept between 15–30°C. When the temperature was reduced below 15°C or raised above 30°C, the rhythmic discharge decreased in frequency and finally stopped. Rhythmic discharge also appeared when ganglia were treated with emetine or the K+ channel blockers, cesium and 4-aminopyridine. The frequency and amplitude of potentials and the pattern of rhythm varied from ganglion to ganglion. The single potential or rhythmic burst firing seemed to originate from a single unit or multiple discharging units, as indicated by the amplitude and frequency of the potentials in a burst. The discharge was abolished by ganglionic blocking agents or by the absence of Ca2+, suggesting a presynaptic origin. The spontaneous rhythmic discharge may be important as a support mechanism for the cardiovascular system in victims of exposure to extreme cold temperature.

Involvement of N-cholinergic peripheral synapses in energy exchange within a sympathetic ganglion

The contents of ATP, ADP, and AMP, the energy level, and the ATP/ADP and ATP/AMP ratios were measured in rabbit superior cervical sympathetic ganglia under conditions of experimentally induced partial and maximum blockage of the N-cholinergic synapses. Synapse blockage proved to result in a significant deficiency in the macroergic phosphates. The content of ATP and ADP, as well as the total amount of macroergic phosphates, decreased linearly with the blockage increase. These changes were reversible: the content of macroergic phosphates, including AMP, also increased linearly with blockage decrease. The energy level was reduced only at the maximum blockage. The patterns of the ATP/ADP and ATP/AMP ratios were different during partial and maximum blockages and changed to reach the required level of the ganglion functional activity under conditions of total deficiency of macroergic phosphates. These results suggest that N-cholinergic synaptic processes regulate the adenylate pool of macroergic phosphates and the energy homeostasis in the sympathetic ganglion.

1479 LDH isoenzymes distribution in neurons and satellite glial cells under H-cholinergic synapses blockade in adult rabbit superior cervical ganglia: possibility evidence of existence ANLS mechanism in sympathetic ganglia

1479 LDH isoenzymes distribution in neurons and satellite glial cells under H-cholinergic synapses blockade in adult rabbit superior cervical ganglia: possibility evidence of existence ANLS mechanism in sympathetic ganglia

Carotid bifurcation pressure modulation of spontaneous activity in external and internal carotid nerves can occur in the superior cervical ganglion

Previous reports have suggested that a peripheral pressure-modulated reflex operates at the level of the superior cervical ganglion to alter evoked activity in the postganglionic nerves of the ganglion in both the cat and rabbit. In the present study we have examined if spontaneous activity of the external and internal carotid nerves of the rabbit superior cervical ganglion can be modulated during changes of the carotid bifurcation pressure (CBP), independent of central nervous system (CNS) integration. For external carotid nerve recordings increases in CBP resulted in a reduction in spontaneous activity while decreases in CBP were associated with an increase in spontaneous activity. For internal carotid nerve recordings similar effects were observed in the majority of recordings although a subset of recordings showed opposite effects or were not responsive to changes in pressure. To determine if vagus nerve afferents contribute to the observed pressure-modulated spontaneous activity effects, the influence of CBP on external carotid nerve recordings was examined before and after section of the vagus nerve rostral to the nodose ganglion. We found that even following section of the vagus nerve the external carotid nerve response to an increase in pressure remained intact. These results demonstrate that, after section of centrally-projecting afferent pathways from the carotid bifurcation to the CNS, changes in CBP can still modify spontaneous sympathetic activity of the rabbit superior cervical ganglion. The data reinforce previous findings related to evoked responses in the postganglionic nerves and also suggest that a pressure-modulated reflex, integrated at the level of the superior cervical ganglion, can influence ongoing sympathetic nervous outflow from the superior cervical ganglion in the rabbit.

Ganglionic neuronal mechanisms involved in circulatory control system.

Intracellular tonic activity and responses to orthodromic stimuli were recorded from the neurones of rabbit superior cervical ganglion and compared with those recorded from the nerves containing pre- and postganglionic nerve fibres, with the use of a coherent recording technique. It was found that: (1) firing of each ganglion neurone is triggered by two presynaptic inputs, single and multiple, formed by a single preganglionic nerve fibre whose discharges evoke the postsynaptic spikes correlated with cardiac rhythm, and by a few converging preganglionic nerve fibres which evoke the postsynaptic spikes, either correlated with cardiac rhythm or irregular, only if two or more of them discharge together, correspondingly; (2) about 240 neurones of the ganglion, on the average, fire synchronously during their tonic activity, being driven by only three preganglionic nerve fibres; (3) only about 9% of a "neural unit", the number of the ganglion neurones receiving innervation from the same preganglionic nerve fibre, are discharged during their tonic activity through a single input, while the rest of neurones are either discharged through a multiple input (17%), or generate only the excitatory postsynaptic potentials subthreshold for spike generation (73%). The results obtained suggest that the ganglionic neuronal mechanisms responsible for vasomotor control involve much more complex ganglionic integrative processes than it has been commonly thought.

Background activity of the rabbit superior cervical ganglion neurons: Soundness of random hypothesis of its formation

The soundness of hypotheses interpreting the mechanism of formation of background activity (BA) generated by the neurons of the rabbit superior cervical ganglion was analyzed, taking into account the statistical characteristics of this activity. These neurons are supplied with a multiple synapic input. According to the random hypothesis, the BA action potentials in the ganglion neurons result from occassional coincidence of the spikes arriving through independent presynaptic inputs. Our calculations showed that the frequency of BA action potentials observed in the experiments is too high to be explained by the random hypothesis and needs synchronization of firing in different presynaptic neurons, rather than their independent firing.

Synapses on axons of sympathetic preganglionic neurons in rat and rabbit thoracic spinal cord.

Axosomatic and axodendritic synapses occur on sympathetic preganglionic neurons, but it is not yet known whether their axons receive synaptic input, which could be particularly effective at regulating sympathetic outflow. Here, we examined retrogradely labelled sympathetic preganglionic axons to see if they received synapses. Cholera toxin B subunit (CTB) or CTB conjugated to horseradish peroxidase (CTB-HRP) was used to label neurons projecting to the rat or rabbit superior cervical ganglion, the rat adrenal medulla, or the rabbit stellate ganglion. At the light microscopic level, small groups of CTB-immunoreactive axons travelled through the ventral horn near its lateral boundary, with occasional axons taking a more medial course. The axons passed through the ventrolateral funiculus to exit at the ventral roots. In parasagittal section, a few axons branched within the ventral horn, sending processes rostrally and caudally for short distances before they turned ventrally to exit the spinal cord. At the ultrastructural level, CTB-immunoreactive rat and rabbit sympathetic preganglionic axons were almost exclusively unmyelinated. In contrast, labelling with CTB-HRP revealed both myelinated and unmyelinated axons in the ventral horn, the ventrolateral white matter, and the ventral roots. CTB-HRP also allowed the detection of the initial segment of a sympathetic preganglionic axon. Synapses, with vesicles clustered presynaptically and membrane specializations postsynaptically, were found on some unmyelinated CTB-immunoreactive axons. Occasional axons received several synapses. Synapses were most common on CTB-containing axons just ventral to the intermediolateral cell column. One synapse was found on an axon within 2 microns of its origin from a proximal dendrite. Rare synapses were found several hundred micrometers ventral to the intermediolateral cell column. One branching axon had synapses just below the branch point on both the main axon and the axonal branch. These findings indicate an extensive synaptic input to the axons of at least some sympathetic preganglionic neurons. These axoaxonic synapses could have a profound effect on sympathetic activity.

Kinetic and pharmacological examination of stimulation-induced increases in synaptic efficacy in the chick ciliary ganglion.

We have characterized the kinetic and pharmacological properties of stimulation-induced increases of synaptic efficacy in the embryonic chick ciliary ganglion. We found what appear to be four components of increased ganglionic efficacy with average time constants of decay of about 60 msec, 400 msec, 30 sec, and 200 sec. These time constants are similar to the those describing the decay of the four components of stimulation-induced increases in neurotransmitter release characterized at other synapses. These components have been termed first and second components of facilitation, augmentation, and potentiation. We found that the addition of small amounts of Ba2+ to the low Ca2+ bathing solution led to an increase in the magnitude of the augmentation-like component, whereas Sr2+ enhanced the magnitude and time course of the component resembling the second component of facilitation. These effects of Ba2+ and Sr2+ are similar to the effects of these same divalent cations on augmentation and the second component of facilitation, respectively, at the frog neuromuscular junction and rabbit superior cervical ganglion. Based on these similar kinetic and pharmacological properties, we conclude that the four components of stimulation-induced increases in release that have been described in other synaptic preparations also appear to be present in the chick ciliary ganglion.

Neuronal mechanisms responsible for ongoing activity of rabbit superior cervical ganglion neurons

The action potentials evoked by stimulation of single preganglionic nerve fibres or of single neurons of the ganglion were recorded from postganglionic nerves of rabbit superior cervical ganglion. The averaging technique was used to improve the nerve signal-to-noise ratio. It was found by dividing the area by the area that single preganglionic nerve fibre discharges, as an average, 15 neurons of the ganglion. Action potentials were also recorded from preganglionic nerves, either appearing synchronously with the intracellular spikes recorded during ongoing activity of the ganglion neuron, or being evoked by stimulation of a single preganglionic nerve fibre. It was found by dividing the area by the area that each ongoing spike in the ganglion neuron is preceded by firing, as an average, of three preganglionic nerve fibres. Thus, 45 ganglion neurons are expected to be discharged by a preganglionic volley during their ongoing activity. This number is much lower than that found in our previous experiments (about 100 neurons). The difference suggests that in the majority of the ganglion neurons the ongoing discharges need summation of excitatory effects produced by a few converging preganglionic nerve fibres (multiple input) rather than being evoked by a single preganglionic nerve fibre (single input). Methacine, a selective muscarinic antagonist (0.3–0.5 mg/kg, i.v.), decreased by about 10% the rate of the multiple input-induced ongoing spikes while not affecting the single input-induced ongoing spikes in the ganglion neurons. This effect is probably due to the increase in the duration of after-hyperpolarization observed after the application of methacine. The results obtained suggest that muscarinic receptors of the ganglion neurons are permanently activated during the ongoing activity maintaining the rate of firing at a somewhat increased level.

Introduction to slow synaptic potentials and their neuromodulation by dopamine.

The existence of two muscarinically mediated slow postsynaptic potentials (PSPs) and a noncholinergic (peptidergic) late-slow PSP was established in the 1960s. These have synaptic delays and PSP durations 100-10,000 times those for the nicotinic (fast) excitatory post-synaptic potential (EPSP). Evidence is reviewed for an against the proposal that, in rabbit superior cervical ganglia, the slow (s-) inhibitory postsynaptic potential requires a second transmitter, dopamine, released by muscarinic action on interneurones (the small, intensely fluorescent cells). The s-EPSP in frog ganglia appears only in already depolarized cells by a muscarinic closure of the M (voltage-sensitive K+) channels. But the large s-EPSP in mammalian neurones, not depolarized, is generated largely via other mechanisms, especially one involving cyclic GMP. Dopamine also produces a long-term enhancement (LTE) of the muscarinic slow PSPs in rabbit superior cervical ganglia, whether dopamine is applied exogenously or released intraganglionically by preganglionic nerve impulses at 10 s-1. LTE is producible heterosynaptically, and it persists well over 3 h; a noncholinergic (peptide?) transmitter may contribute to the initial 30 min of LTE. LTE is mediated by a D1 receptor coupled to cyclic AMP; it is blocked by cyclic GMP or low Ca2+ or calmidazolium (a calmodulin inhibitor). The modulatory process of LTE has certain similarities to, but also fundamental differences from, the long-term potentiation known in the hippocampus.

Receptor binding sites for cholecystokinin, galanin, somatostatin, substance P and vasoactive intestinal polypeptide in sympathetic ganglia

Sympathetic ganglia are innervated by neuropeptide-containing fibers originating from pre- and postganglionic sympathetic neurons, dorsal root ganglion neurons, and in some cases, myenteric neurons. In the present report receptor autoradiography was used to determine whether sympathetic ganglia express receptor binding sites for several of these neuropeptides including bombesin, calcitonin gene-related peptide-α, cholecystokinin, galanin, neurokinin A, somatostatin, substance P, and vasoactive intestinal polypeptide. The sympathetic ganglia examined included the rat and rabbit superior cervical ganglia and the rabbit superior mesenteric ganglion. High levels of receptor binding sites for cholecystokinin, galanin, somatostatin, substance P, and vasoactive intestinal polypeptide were observed in all sympathetic ganglia examined, although only discrete neuronal populations within each ganglion appeared to express receptor binding sites for any particular neuropeptide. These data suggest that discrete populations of postganglionic sympathetic neurons may be regulated by neuropeptides released from pre- and postganglionic sympathetic neurons, dorsal root ganglion neurons, and myenteric neurons.

Organization of the tonically active pathways through the superior cervical ganglion of the rabbit

Responses of the neurons of the rabbit superior cervical ganglion to stimulation of the fascicles of preganglionic nerve fibres dissected from the cervical sympathetic trunk, and tonic activity of these neurons were recorded with intracellular electrodes in vitro and in vivo, respectively. It was found that on average 8.5 + 1.2 preganglionic fibres converge on each ganglion cell. In each preparation a single input spike could be evoked in a ganglion cell by stimulation of only one of all tested fascicles. The interspike intervals for single and multiple preganglionic inputs in tonic activity of ganglion neurons displayed Gaussian and Poisson distributions, respectively. The frequencies of spikes showing a Gaussian distribution correspond in most cases to pulse and respiratory frequencies. These data suggest that (1) single input is formed by a single preganglionic fibre which evokes EPSP of an amplitude high enough to trigger a postsynaptic spike, and that (2) regular activity of the ganglion neuron triggered by single input is probably due to a regular afferent drive. A cross-correlation analysis between the intracellular tonic activity and the tonic activity simultaneously recorded from the postganglionic nerve was made. The results of the analysis were compared with those obtained when intracellular spikes were evoked by direct stimulation of one particular neuron. From this comparison it has been concluded that about 100 ganglion cells, on the average, discharge synchronously within the 20-ms range during their tonic activity.

Postsynaptic long-term enhancement (LTE) by dopamine may be mediated by Ca 2+ and calmodulin

Long-term enhancement (LTE), of postsynaptic slow depolarizing responses to a muscarinic agonist (MCh), follows a brief exposure of the rabbit superior cervical ganglion to another transmitter, dopamine (DA). Either reduction of external Ca 2+ (to 1.0 mM or 0.2 mM) or presence of a specific calmodulin antagonist (calmidazolium at 5 μM) blocked DA induction of this LTE. However, unlike LTP in hippocampus, induction of LTE is not mediated by depolarization-dependent influx of Ca 2+.

Acethylcholine elictits metabolically mediated M2-muscarinic hyperpolarization in isolated rabbit sympathetic neurons.

Topically applied acetylcholine elicited a hyperpolarizing (ACh-HP) response in about 2/3 of the principal neurons of rabbit superior cervical ganglia isolated in a culture medium for several days. Membrane changes responsible for the ACh-HP were multiple and varied with the level of membrane potential (Vm): At, or less negative than, resting Vm (about -60 mV), membrane conductance (Gm) was increased; at Vm more negative than -70 mV, Gm either increased, decreased, or remained unchanged. The effect of altering extracellular K+ and Ca2+ concentrations suggest that a Ca2(+)-dependent increase in K+ conductance contributes to ACh-HP; however, drugs reported to block such channels (D-tubocurarine at 30 microM or apamin at 200 nM) shortened duration of the ACh-HP but did not depress peak amplitude. The ACh-HP was depressed by the M2-muscarinic antagonist AF-DX 116 and blocked by an intracellular administration of guanosine-5'-o-(2-thiodiphosphate) (GDP-beta-S) or by preincubation with pertussis toxin. Intracellularly injected inositol-1,4,5-triphosphate (IP3) elicited a hyperpolarization associated with an increase in Gm at any Vms. Activators of protein kinase C applied extracellularly, 1,2-oleoylacetylglycerol (OAG) or phorbol-12,13-dibutyrate (Pb(Bu)2) elicited a hyperpolarization with a decrease in Gm at Vms more negative than -50 mV and a reversal potential close to Ec1. In the presence of Li+, ACh-HP was smaller after a repetitive stimulation with ACh. These results suggest that, in these neurons, ACh can directly activate M2-muscarinic receptors coupled to GTP-binding proteins, which leads to both an increase in Ca2(+)-sensitive K+ conductance mediated by the intracellular messenger IP3 and a decrease in a conductance, possibly Gc1, through another phosphatidylinositide pathway.

Convergence of preganglionic fibers on superior cervical ganglion neurons in the rabbit

Intracellular recording techniques were used to record electrical response from neurons of the rabbit (isolated) superior cervical ganglion to single stimuli applied to bundles of preganglionic fibers as well as tonic electrical neuronal activity in this ganglion during acute experiments in situ. A review of the findings obtained confirms that neurons of the ganglion receive preganglionic synaptic inputs of two types, the first of a single pattern, formed by a preganglionic fiber; excitatory action of the latter on ganglion units suffices to induce postsynaptic action potentials (AP I) and the second of a multiple pattern, formed by several preganglionic fibers with relatively faint excitatory action, capable of evoking postsynaptic action potentials (AP II) only when excited in unison. Interspike intervals for AP I and AP II in tonic neuronal activity conformed to a normal distribution and a random distribution pattern respectively.

Cyclic GMP‐Dependent Protein Kinase and Phosphorylation of the Endogenous Substrate Proteins in the Rabbit Superior Cervical Ganglion

When the homogenate of rabbit superior cervical ganglia (SCG) was incubated in the presence of [gamma-32P]ATP and Mg2+, two specific proteins were strongly labeled. Their apparent molecular weights were 90,000 and 54,000, respectively. The phosphorylation of the latter was significantly stimulated by 10-50 nM cyclic GMP but to a lesser extent by cyclic AMP, whereas that of the former was not stimulated significantly by either of the cyclic nucleotides. The purified protein kinase inhibitor from rabbit skeletal muscle did not inhibit the phosphorylation. These results indicated that the observed phosphorylation of 54K protein was dependent on cyclic GMP but not on cyclic AMP. When intact SCG was incubated in the presence of 32Pi, phosphorylation of 90K protein was stimulated by cyclic GMP, dibutyryl cyclic GMP, and 8-bromo-cyclic GMP (10 microM), whereas phosphorylation of 54K protein was not significantly stimulated by any of these substances. The present demonstration of endogenous cyclic GMP-dependent protein kinase activity and its endogenous substrate proteins raises a possibility that the physiological actions of cyclic GMP in SCG are mediated by the phosphorylation of these proteins.

Secondary late components of the muscarinic postsynaptic potentials, in rabbit superior cervical ganglion

The well known muscarinic slow excitatory polysynaptic potential (s-EPSP) of rabbit superior cervical ganglion (SCG) peaking at about 1–2 s and lasting 5–10 s, is immediately followed by an abrupt change in slope to a longer, lower depolarizing phase. A brief dip in the level of depolarization (DP) often separates the two depolarizing phases. The secondary phase of s-EPSP rises to its own peak at about 25 s; total duration 60–120 s. With repetition of orthodromic volleys secondary s-EPSP builds up more gradually than initial s-EPSP, but more rapidly than slow-slow (ss-) EPSP. The later ‘secondary’ depolarizing phase along with the antecedent ‘dip in DP’ are, like the ‘initial’ s-EPSP, eliminated by a muscarinic antagonist, quinuclinidyl benzilate hydrochloride (QNB). This distinguishes secondary s-EPSP from the even slower rising non-cholinergic ss-EPSP. The ss-EPSP, although relatively small in the responses to the usual 3-pulse test stimuli, rises to an extraordinary amplitude (equal to the compound action potential) during a 10 s–120 s train of pulses. Gallamine blocked most of the slow IPSP component in test responses but not initial or secondary s-EPSP. A preganglionic conditioning train (10/s for 2 min) induced a long-term-enhancement (LTE) of secondary s-EPSP lasting > 3 h, with maximum postconditioning percentage increases greater than for initial s-EPSP. Also enhanced was the dip in DP, now forming a deeper notch between initial and secondary s-EPSPs; this attains a maximum at about 30 min postconditioning but thereafter progressively loses the enhancement by about 90 min. The dip in DP appears to represent some delayed, perhaps hyperpolarizing process that may overlap with the initial and secondary phases of s-EPSP and affect their forms and peak amplitudes. This would not exclude the possibility that the initial and secondary phases of s-EPSP reflect multiple muscarinically activated electrogenic mechanisms having differing time-courses.

Dual synaptic effects of activating M 1-muscarinic receptors, in superior cervical ganglia of rabbits

Postsynaptic potentials elicited by various muscarinic agonists and by preganglionic stimuli in the presence of such agonists were recorded from rabbit superior cervical ganglia using sucrose-gap and air-gap methods. While methacholine and bethanechol (both at 10 −4 M) induced biphasic potential changes, McN-A-343 and a novel synthetic compound AF-102B (10 −7 M-10 −5 M) produced only a depolarizing response which was depressed by the M 1-antagonist pirenzepine (10 −7 M), but not by the M 2 antagonist AF-DX 116 (same concentration), indicating that these compounds act purely as M 1-muscarinic agonists in this system. These agonists selectively depressed the orthodromic slow excitatory postsynaptic potential (EPSP) in a dose-dependent manner without substantially affecting the fast EPSP; this is in accord with the view that their depolarizing action is on the same postsynaptic muscarinic receptor that mediates the slow EPSP. The slow inhibitory post synaptic potential (IPSP), on the other hand, was found potentiated in the presence of these agonists. This potentiation was antagonized not only by pirenzepine but also by yohimbine; the potentiation was itself enlarged by nomifensine (a dopamine-uptake inhibitor). We postulate that M 1-muscarinic receptors are present not only on the postganglionic principal cells but also on the interneurons; the former were already known to be responsible for the generation of slow EPSP, but the latter may be on terminals of dopamine-containing small intensely fluorescent cells and regulate the orthodromic release of dopamine and are to be distinguished from the M 2-receptors.

The actions of histamine in cat and rabbit superior cervical ganglia.

Depolarization of the cat superior cervical ganglion (SCG) evoked by histamine was antagonized by mepyramine. Histamine-induced depolarization, indicated by changes in the negative and positive afterpotentials of ganglionic action potentials, was decreased by pretreatment with aminophylline. In the cat SCG, histamine evoked stimulus bound decremental oscillatory potentials (SBDOP). In the rabbit SCG, only depolarization and no SBDOP were observed after the administration of histamine. Histamine-induced SBDOP were increased by isoprenaline and decreased by Leu-enkephalin pretreatment. Our results indicate that histamine evokes depolarization and an increase in excitability of the cat SCG which seems to be mediated by H1-receptors.

A novel muscarinic receptor antagonist AF-DX 116 differentially blocks slow inhibitory and slow excitatory postsynaptic potentials in the rabbit sympathetic ganglia

Muscarinic, slow postsynaptic potentials (s-epsp and s-ipsp) in the rabbit superior cervical ganglia were shown to be differentially depressed by a novel cardioselective M 2-type antagonist AF-DX 116: it antagonized the s-ipsp with IC 50 value of 1.5 × 10 −7M, which is 16-fold more potent in depressing the s-ipsp than the s-epsp. A hyperpolarizing component in the biphasic potential changes induced by a muscarinic agonist, methacholine, was selectively eliminated by this antagonist. AF-DX 116 was thus shown to be an useful tool for discriminating the M 2-type muscarinic responses from those of M 1-type in the nervous system.