Origins of the afferent fibers to the cat superior cervical ganglion.

The origins of the afferent fibers to the cat's superior cervical ganglion (SCG) were demonstrated by using the retrograde horseradish peroxidase tracing method. We found that the preganglionic neurons were located in the spinal segments C8-T5, particularly in T1-T3. These neurons were situated mainly in the intermediolateral column. The extra-SCG neurons along with the cervical sympathetic trunk originated ipsilaterally from the middle cervical and stellate ganglia, and contralaterally from the caudal part of the SCG. Labeled neurons also originated from the mandibular division of the trigeminal ganglion. Our results demonstrated that many fiber sources projected to the SCG, which plays a complicated synaptic role in controlling the visceral organs of the head and neck region.     

AF-DX 116 discriminates between muscarinic M2 receptors of the heart and vasculature

The newly developed muscarinic antagonist, AF-DX 116, has been reported to have a higher affinity in vivo for muscarinic receptors in the heart than in the vasculature. Therefore two in vitro preparations, the rabbit ear artery and spontaneously beating right atrium, were compared. AF-DX 116 had a 29 times greater affinity for muscarinic receptors in the cardiac preparation than in the ear artery, with a pA2 in the heart of 7.42 compared to a value of 5.95 in the ear artery. Thus AF-DX 116 shows promise as an approach to differentiating sub-classes of muscarinic M2 receptors.     

Effect of naloxone and prolonged preganglionic stimulation on noncholinergic transmission in the superior cervical ganglion of the cat

In cats anesthetized with sodium pentobarbital, a supramaximal 40-Hz, 30-s train to the cervical sympathetic trunk, during block of ganglionic cholinergic transmission with hexamethonium and scopolamine, produced a delayed, slow, small amplitude contraction of the nictitating membrane that persisted for several minutes after the end of the stimulus train. The post-stimulus component of the response was due to afterdischarge of the ganglion cells, since section of the post-ganglionic axons at the end of the train resulted in elimination of this component. The amplitude of the slow nictitating membrane response was enhanced in a dose-dependent manner by i.v. injection of naloxone. The enhancement was detectable at a dose as low as 1 microgram/kg and was maximal at 10 micrograms/kg. During continuous preganglionic stimulation at 40 Hz, the amplitude of the slow nictitating membrane response reached a peak in 2-4 min and then faded with time until it became undetectable. Time for 90% decay was 82 +/- 5 min (n = 18). The nictitating membrane response to postganglionic nerve stimulation was not modified by prolonged preganglionic stimulation. In three cats, the cervical sympathetic trunk was split into two bundles and one bundle was stimulated continuously at 40 Hz until the slow response disappeared. At this time stimulation of the unconditioned bundle evoked a slow response of normal appearance. This suggests that the process underlying the fade involves only the conditioned axons. Recovery from the fade was slow, the response approaching control by 24 h post-stimulus.(ABSTRACT TRUNCATED AT 250 WORDS)     

Conducting pathways of the superior cervical sympathetic ganglion of the cat

Individual nerves of the superior cervical sympathetic ganglion were stimulated in acute experiments on cats, and action potentials (AP) were recorded from other nerves of the ganglion in order to clarify whether or not there is transmission of excitation through the ganglion from one nerve to another and to establish whether this transmission is continuous or synaptic. The method of intracellular recording from neurons of the ganglion was also used. It is established that stimulation of the cervical sympathetic nerve evokes AP in all of the peripheral nerves of the ganglion, a circumstance that is the result of synaptic transmission of excitation. There is no transmission of excitation in the reverse direction or between any of the 12 peripheral nerves of the ganglion (including the four branches of the internal carotid nerve). Orthodromic excitation is recorded intracellularly from neurons of the ganglion during stimulation of the cervical sympathetic nerve, and antidromic excitation is recorded during stimulation of a peripheral nerve (the internal carotid nerve). It follows that the pathways through the ganglion which conduct excitation from the cervical sympathetic nerve into all of the remaining nerves of the ganglion are synaptic. Analysis of EPSP latent periods indicated that preganglionic fibers that differ sharply with respect to threshold and conduction rate (groups S₂ and S₄) converge on one and the same neurons of the ganglion.A. A. Bogomolets Institute of Physiology, Academy of Sciences of the Ukrainian SSR, Kiev. Translated from Neirofiziologiya, Vol. 2, No. 2, pp. 216–224, March–April, 1970.     

Distribution of 45Ca in the superior cervical ganglion of the cat

Superior cervical ganglia isolated from immature cats accumulated 0.9 ng atoms of ⁴⁵Ca per mg wet weight during 10-min incubations at 37°C; when expressed as an equivalent volume of medium the accumulation was four times the uptake of ³H-inulin. Orthodromic stimulation of the ganglia doubled ⁴⁵Ca accumulation, whereas excitation with 50 mM KCl, 5 mM glutamate, or antidromic stimulation increased accumulation by one-half. Hexamethonium reduced the increment in ⁴⁵Ca accumulation due to orthodromic stimulation only, but another ganglionic blocking agent, tetraethylammonium, did not reduce accumulation in any case. Both agents blocked ganglionic transmission monitored electrophysiologically. To resolve this discrepancy, and to approach the localization of ⁴⁵Ca within the ganglia, the efflux of previously accumulated ⁴⁵Ca was examined. The data could be fitted by an equation incorporating the sum of three exponentials, representing a rapidly exchanging compartment plus two more slowly exchanging ones. The latter two appeared to reflect the pre- and postganglionic elements in the ganglia: ⁴⁵Ca content of the “preganglionic” compartment was increased by orthodromic but not by antidromic stimulation, and was not decreased by either blocking agent; conversely, ⁴⁵Ca content of the “postganglionic” compartment was increased by both orthodromic and antidromic stimulation, and was decreased by both blocking agents after orthodromic stimulation. The lack of effect of tetraethylammonium on the whole ganglion resulted from an increase in “preganglionic” accumulation that offset the “postganglionic” decrease. After preganglionic denervation, the ⁴⁵Ca content of the “preganglionic” compartment was reduced by two-thirds, while the ⁴⁵Ca content of the “postganglionic” compartment was unchanged. Chemical stimulation increased ⁴⁵Ca accumulation in both compartments. Diphenylhydantoin, 0.1 mM, decreased the increment in ⁴⁵Ca accumulation due to electrical stimulation and to 50 m^M KCl; this inhibition occurred in the “preganglionic” compartment (and perhaps also in the “postganglionic”), and was accompanied by an increased efflux of ⁴⁵Ca.     

Electron microscope localization of acetylcholinesterase and butyrylcholinesterase in the superior cervical ganglion of the cat. I. Normal ganglion

The distributions of acetylcholinesterase (AChE) and butyrylcholinesterase (BuChE) in the superior cervical ganglion (SCG) of the cat were determined by electron microscopy (EM) with the bis- (thioacetoxy)aurate (I), or Au(TA)2, method. Before the infusion of fixative, one of the enzymes was selectively, irreversibly inactivated in vivo, as confirmed by light microscope (LM) examination of sections of the stellate ganglion stained by the more specific copper thiocholine method. Physostigmine-treated controls, for inhibition of AChE or BuChE, were stained concomitantly with tissue for enzyme localization by the Au(TA)2 method for EM examination in each experiment. It was concluded that most of the AChE of the cat SCG is present in the plasma membranes of the preganglionic axons and their terminals, and in the dendritic and perikaryonal plasma membranes of the postsynaptic ganglion cells. BuChE is confined largely to the postsynaptic neuronal plasma membranes. Reasons for the discrepancies between the localizations found by the present direct EM observations and those deduced earlier from LM comparisons of normal and denervated SCG are discussed. It is proposed that a trophic factor released by the preganglionic terminals is probably required for the synthesis of postsynaptic neuronal AChE, and that BuChE may serve as a precursor of AChE at that site.     

Origin of the long positive potential in the cat superior cervical ganglion

The sucrose gap technique was used to study the long positive potential (P-potential) in a curarized cat superior cervical ganglion. The frequency of stimulating the preganglionic trunk optimal for P-potential production was 30–40 impulses/sec at a stimulus series duration of 1 sec. Proserine in low concentrations (1–5 µg/ml) increased amplitude and especially duration of the P-potential. Atropine (0.5–2 µg/ml) blocked it completely. Adrenaline and noradrenaline (10–50 µg/ml inhibited both the negative potential (corresponding to the fast EPSP of neurons) and the P-potential in equal measure. The nature of dependence of P-potential amplitude on value of the membrane potential was also studied. It was found that the P-potential is inhibited in solutions with low potassium ion content, and that amplitude of the P-potential rises with an increase of intracellular sodium concentration. The rate of its increase rises with an increase of temperature. Under the influence of strophathin, the P-potential is inhibited. The data obtained support the hypothesis that the P-potential is determined by synaptic activation of the electrogenic sodium pump.A. A. Bogomol'ets Institute of Physiology, Academy of Sciences of the Ukrainian SSR, Kiev. Translated from Neirofiziologiya, Vol. 3, No. 1, pp. 76–83, January–February, 1971.     

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 M2-type antagonist AF-DX 116: it antagonized the s-ipsp with IC50 value of 1.5 X 10(-7) M, 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 M2-type muscarinic responses from those of M1-type in the nervous system.     

Electron microscope localization of acetylcholinesterase and butyrylcholinesterase in the superior cervical ganglion of the cat. II. Preganglionically denervated ganglion

Cat superior cervical ganglia (SCG), denervated preganglionically 6-8 d previously, were stained for acetylcholinesterase (AChE) and butyrylcholinesterase (BuChE) by the bis-(thioacetoxy)aurate (I), or Au(TA)2, method and compared by electron microscopy with normal SCG described previously (Davis, R., and G. B. Koelle. 1978. J. Cell Biol. 78:785-809). In confirmation of earlier light microscopic findings by the highly specific copper thiocholine method, there was nearly a total disappearance of AChE from the ganglion; no myelinated or unmyelinated axons with AChE-stained axolemmas were found, and only occasional traces of AChE staining were noted at dendritic and perikaryonal plasma membranes. Considerable staining for BuChE persisted at the latter sites, however. As in the normal SCG, physostigmine-resistant staining, caused by noncholinesterase enzymes plus the possible presence of very low concentrations of AChE or BuChE, was noted at external mitochondrial membranes, elements of the endoplasmic reticulum of neurites and Schwann cells, and also in lysosomes. These findings confirm the previous identification of AChE-stained myelinated fibers in the normal SCG as preganglionic and of the unstained myelinated fibers as postganglionic. It is proposed that the maintenance of AChE at postsynaptic sites in normal ganglia is caused by the release of a trophic factor(s) from presynaptic terminals. The source of the postsynaptic BuChE, which is apparently completely absent from the endoplasmic reticulum of the ganglion cells, remains unexplained.     

Effect of d-sparteine on nicotinic acetylcholine receptors in rat superior cervical ganglion neurons

Effects of d-sparteine (d-SP), a ganglionic blocking agent, on membrane currents evoked by iontophoretic applications of acetylcholine to rat superior cervical ganglion neurons, were studied using a whole-cell patch-clamp recording technique. Blocking effects of d-SP were enhanced by membrane hyperpolarization to potentials more negative than –50 mV. Analysis of the d-SP effect on the dose—response relationship suggests that d-SP at concentrations of 0.5–5.0 µM exerts both voltage-independent and voltage-dependent competitive actions on nicotinic acetylcholine receptors. No use-dependence of the d-SP-induced blockade was found using paired ACh applications at interpulse intervals longer than 0.5 sec. Inhibitory constantK _i estimated by the Dixon method was equal to 0.62±0.15 and 0.28±0.08 µM at membrane potential levels –30 and –90 mV, respectively. These characteristics of the d-SP blocking effects are compatible with a voltage-dependent competitive blocking mechanism. The possibility remains that an open channel-blocking mechanism with a comparatively fast kinetics contributes to the d-SP-induced blockade, but its contribution is small.Neirofiziologiya/Neurophysiology, Vol. 25, No. 4, pp. 266–272, July–August, 1993.     

Granule-containing cells in the human superior cervical ganglion.

The fine structure of granule-containing cells in the human superior cervical ganglion is described. These cells are larger than the typical SIF cells in mammals and exhibit green-yellow fluorescence. They are characterized by numerous granular vesicles (80-140 nm in diameter) in the cytoplasm, but have many features in common with ordinary ganglion cells. They emit several long processes which form bundles together with ordinary nerve fibers. No synapses are found where the cells are presynaptic, although a few synapses are observed there where nerves are prosynaptic on the perikarya and processes of the cells. No close topographical relations are seen between the cells and blood vessels. It is suggested that the granule-containing cells are a special type of postganglionic aminergic neurons.     

3H]AF-DX 116 labels subsets of muscarinic cholinergic receptors in rat brain and heart

The in vitro binding properties of the novel muscarinic antagonist [3H]AF-DX 116 were studied using a rapid filtration technique. Association and dissociation rates of [3H]AF-DX 116 binding were rapid at 25 degrees C (2.74 and 2.70 X 10(7) min-1 M-1 for K+1; 0.87 and 0.93 min-1 for k-1) but 20-40 times slower at 0-4 degrees C (0.13 and 0.096 X 10(7) min-1 M-1 for k+1; 0.031 and 0.022 min-1 for k-1 in cerebral cortical and cardiac membranes, respectively). Kinetic dissociation constants (Kds) were estimated to be 31.8 nM and 30.9 nM at 25 degrees C; 23.1 nM and 0-4 degrees C for the cerebral cortex and heart, respectively. In saturation studies, [3H]AF-DX 116 labeled 29 percent of the total [3H](-)QNB binding sites in the cerebral cortical membranes and 87 percent in the cardiac membranes, with Kd values of 28.9 nM and 17.9 nM, respectively. Muscarinic antagonists inhibited [3H]AF-DX 116 binding in a rank order of potency of atropine greater than dexetimide greater than AF-DX 116 greater than PZ greater than levetimide in both tissues. Except for PZ/[3H]AF-DX 116 and AF-DX 116/[3H]AF-DX 116 in the cerebral cortex, all the antagonist competition curves had Hill coefficients close to one. Carbachol and oxotremorine produced shallow inhibition curves against [3H]AF-DX 116 binding in both tissues. Regional distribution studies with [3H](-)QNB, [3H]PZ and [3H]AF-DX 116 showed that most of the muscarinic receptors in the cerebral cortex, hippocampus, nucleus accumbens and corpus striatum are of the M1 subtype while those in the brainstem, cerebellum and other lower brain regions are of the M2 subtype. These results indicate that [3H]AF-DX 116 is a useful probe for the study of heterogeneity of muscarinic cholinergic receptors.     

Ionic mechanisms involved in the release of 3H-norepinephrine from the cat superior cervical ganglion

It has previously been reported that in the isolated cat superior cervical ganglion (SCG) labeled with tritiated norepinephrine (3H-NE), the stimulation of the preganglionic trunk at 10 Hz as well as the exposure to 100 microM exogenous acetylcholine (ACh), produced a Ca++-dependent release of 3H-NE. The present results show that a Ca++-dependent release of 3H-NE was produced also by exposure to either 50 microM veratridine or 60 mM KCl. Tetrodotoxin (0.5 microM) abolished the release of 3H-NE induced by preganglionic stimulation, ACh and veratridine but did not modify the release evoked by KCl. The metabolic distribution of the radioactivity released by the different depolarizing stimuli showed that the 3H-NE was collected mainly unmetabolized. In the cat SCG neither the release of 3H-NE evoked by KCl nor the endogenous content of NE was modified by pretreatment with 6-OH-dopamine (6-OH-DA). On the other hand, this chemical sympathectomy depleted the endogenous content of NE in the cat nictitating membrane, whose nerve terminals arise from the SCG. The data presented suggest that the depolarization-coupled release of NE from the cat SCG involves structures that are different to nerve terminals and that contain Na+ channels as well as Ca++ channels.     

Voltage-dependent effect of tetraethylammonium on the nicotinic acetylcholine receptors of rat superior cervical ganglion neurons

The effect of tetraethylammonium (TEA) on the currents evoked in neurons of the rat superior cervical ganglion by iontophoretic application of acetylcholine (ACh) was studied using a whole-cell patch-clamp recording technique. Tetraethylammonium was used at a concentration of about 20 µM, providing no blocking effect on the ACh-induced membrane currents at a range of positive membrane potentials and reducing these currents recorded at a range of negative membrane potentials by about half. The blocking effect of TEA increased with hyperpolarization within the –50 to –90 mV membrane potential range, and did not depend on the membrane potential level within a range of 0 to –50 mV. The analysis of dose dependence showed that both the voltage-dependent and the voltage-independent blocking effects are due to TEA competitive action on the ganglionic nicotinic acetylcholine receptors (nAChR). The results suggest that the TEA-induced competitive blockade is voltage-dependent.Neirofiziologiya/Neurophysiology, Vol. 27, No. 1, pp. 63–66, January–February, 1995.