Hypoventilation recruits preganglionic sympathetic fibers with inspiration-related activity in the superior cervical trunk of the rat.

Activity in preganglionic sympathetic neurons projecting in the cervical sympathetic trunk (CST) of rats was analysed with respect to changes in the pattern of the respiratory modulation during a long lasting hypoventilation. Under normal acid-base status (pH: 7.36+/-0.04, pCO2: 42.1+/-6.1 mm Hg, pO2: 135.8+/-43 mm Hg) a maximum of activity during expiration (expiration-related activity) was observed in all nerve recordings (n = 27). No other pattern of respiratory modulation was observed under this condition. Under a hypoventilation a dissociation between the duration of phrenic nerve activity and that of the inspiratory inhibition in neurons with expiration-related activity was observed as the inhibition was significantly prolonged by 49+/-24.9% and outlasted inspiration in 5/7 multifibers. When acid-base status was systematically changed (pH: 7.15+/-0.05, pCO2: 80.4+/-11.8 mm Hg, pO2: 62.8+/-17.5 mm Hg [n = 7]) by a hypoventilation lasting for several hours activity with a maximum peak during central inspiration (inspiration-related activity) emerged and disappeared when control conditions were reestablished. Neurons with expiration-related activity showed a cardiac rhythmicity (CR) of 62.5+/-14.6% (n = 27) and were inhibited to baroreceptor stimulation whereas neurons with inspiration-related activity showed no discernible CR (23.1+/-5.1%; n = 7) and were not inhibited to baroreceptor stimulation. Furthermore, expiration-related neurons were inhibited by 32.5+/-18.3% (n = 27) during noxious cutaneous stimulation while neurons with inspiration-related activity were activated by 21.5+/-12.1% (n = 7). These findings suggest that the respiratory modulation of preganglionic sympathetic activity in the CST consists of expiration-related activity in normal acid-base status. During hypoventilation neurons with inspiration-related activity are recruited. These neurons show reflex patterns distinct from expiration-related neurons and probably constitute a subgroup of sympathetic neurons which is activated under increased respiratory drive.     

Acetylcholine-induced back-firing in the preganglionic trunk of the rat superior cervical ganglion

ACh (5 × 10⁻⁴ M), when applied to isolated ganglion preparations elicited an apparently antidromic discharge in the cervical sympathetic trunk. The intensity of this back-firing was found to be about 10 times lower than that of the postganglionic discharge evoked by ACh in the internal carotid nerve. Both responses, however, displayed a similar time course and consisted of an early and a late component. In the back-firing the early component died out in a few seconds, while the late one lasted for 20–30 s. The two components were cancelled by d-tubocurarine (5 × 10⁻⁶ M) and atropine (10⁻⁶ M), respectively, suggesting that both nicotinic and muscarinic cholinoceptive sites are involved. In chronically decentralized preparations ACh evoked a clear back-firing response not substantially different from that elicited in normal ganglia. Therefore it is likely that the back-firing phenomenon is not due to antidromic activation of preganglionic fibers. The back-firing observed in the rat superior cervical ganglion was interpreted as being due to activation of sympathetic neurons known to give rise to recurrent axons in the cervical sympathetic trunk.     

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

Responses of neurons of the rabbit superior cervical ganglion to stimulation of the fascicles of preganglionic fibres and tonic activity of these neurons were recorded with intracellular electrodes in vitro and in situ, respectively. Marked convergence of preganglionic fibres onto ganglion cells was found: 8.5 +/- 1.2 fibres, on the average, converged on each cell. It was found that each ganglionic neuron received two preganglionic inputs: single and multiple. The single input was formed by a single preganglionic fibre which evoked EPSP strong enough to trigger a postsynaptic spike. The multiple input was formed by two or more converging preganglionic fibres which could trigger a postsynaptic spike only through summation of the EPSPs they evoked. The interspike intervals for these two preganglionic inputs in tonic activity of neurons were described by distributions of different types: Gaussian and Poisonian ones. Those two distributions were typical of single and multiple inputs, respectively.     

Terminal distribution of mossy fibers traversing the superior cerebellar peduncle in the mouse.

The carbocyanine membrane probe DiI was applied to cut fibers of the superior cerebellar peduncle in the fixed brains of adult mice. Labeled mossy fibers in the cerebellar cortex formed rosettes only in the most superficial aspect of the granular layer, just beneath the Purkinje cell layer. These results provide evidence for a functional division of the cerebellar granular layer into horizontal laminae, in addition to the commonly recognized lobular and sagittal divisions of the cerebellar cortex.     

Development of the rat superior cervical ganglion: ingrowth of preganglionic axons

The growth of sympathetic preganglionic axons has been studied in the fetal rat. Preganglionic axons first emerge from the spinal cord and enter the paravertebral chain of ganglia between days 12 and 13 of gestation (E12 to E13). By E14, hundreds of preganglionic neurons have sent axons into the sympathetic chain. Even at this age, the arrangement of the preganglionic axons conforms to the adult organization in a number of ways. As in maturity, these axons leave the spinal cord only through the ventral root of the segment in which their cell bodies reside. Furthermore, axons grow into the superior cervical ganglion only from the same set of spinal segments as supply this ganglion in maturity, and in correct segmental proportions, although there is a slight rostral-to-caudal lag along the spinal cord in the full establishment of the projection. The development of appropriate projections in this system owes little to the growth of axons along stereotyped pathways. Rather, preganglionic organization apparently reflects a more general instruction that governs the relative number of axons from each spinal segment which course rostrally or caudally within the sympathetic chain. The appropriate distribution of preganglionic axons early in development suggests that a fiber's segmental level of origin biases the axon to grow in a particular direction.     

A part of cholinergic fibers in mouse superior cervical ganglia contain GABA or glutamate

The localizations and functions of glutamate and GABA, the major amino acid neurotransmitters in the central nervous system, are still unclear in the peripheral nervous system. We immunohistochemically double-stained mouse superior cervical ganglia with antibodies for the vesicular acetylcholine transporter (VAchT), GAD65, the vesicular glutamate transporters 1-3 (VGluTs1-3), the marker of the sympathetic preganglionic neuron (SPN), GABAergic, and glutamatergic terminals, respectively. All GAD65-positive terminals showed VAchT immunoreactivity, indicating that GABAergic fibers originate from SPNs. VGluT2-immunoreactive terminals showing colocalization with VAchT were observed, but VGluT1 and 3 immunoreactive terminals were not. Colocalization of GAD65 and VGluT2 was rarely found. All VGluT2-immunopositive terminals were also immunopositive for neuronal nitric oxide synthase (nNOS), a marker for the subpopulation of the SPNs, while about half of the GABA-immunopositive fibers were immunopositive for nNOS. The origin of these fibers was discussed.     

Localization of spinal cord preganglionic neurons innervating the superior cervical ganglion in the golden hamster

The retrograde neuronal tracer Fluoro-Gold (FG) was used to determine the location and distribution of preganglionic sympathetic neurons in the spinal cord of the golden hamster. FG was injected unilaterally into the superior cervical ganglia. Labeled neurons were found only ipsilateral to the injection site in segments C8 to T5 of which the segments T1 to T3 contained about 98% of the labeled cells. Neurons were found in four regions of the spinal cord: the intermediolateral nucleus (43%), the lateral funiculus (55%), the central autonomic area (1%) and the intercalated region (less than 1%). In the intermediolateral nucleus, cells often were arranged in clusters of which several were seen in each spinal segment.     

Cyclic AMP synthesis in guinea pig superior cervical ganglia: response to pharmacological and preganglionic physiological stimulation

The effects of stimulation of guinea pig superior cervical ganglia (SCG) in vitro with 50 μM concentrations of dopamine,L-norepinephrine orL-isoproterenol in Eagle's Medium containing 5 mM theophylline were determined. Dopamine (50 μM) produced no stimulation of cyclic AMP levels, 50 μM norepinephrine produced a doubling of cyclic AMP levels, while 50 μM isoproterenol produced a 6-fold increase in cyclic AMP levels over control values. The increases were blocked by propranolol, indicating that they were due to stimulation of a β-adrenergic receptor-adenylate cyclase complex. In the rabbit SCG, by contrast, 50 μM dopamine produced an increase in cyclic AMP levels of 60% over control values. The possible involvement of the β-receptor complexin the modulation of ganglionic transmission was tested by subjecting ganglia to supramaximal stimulation for 8 min. No elevation of cyclic AMP levels occured in guinea pig SCG, but a doubling of cyclic AMP levels in rabbit SCG was noted. Current concepts of the function of cyclic AMP in neural transmission in the SCG involve a dopaminergic SIF cell, dopamine receptor-adenylate cyclase complex, and the generation of a slow inhibitory postsynaptic potential (s-IPSP). Previous research has indicated that the guinea pig SCG lacks a s-IPSP, and its SIF cells contain norepinephrine rather than dopamine. Since preganglionic stimulation of the guinea pig SCG has now been shown not to cause increased cyclic AMP levels, and since the cyclic AMP content of the guinea pig SCG is not increased following incubation with dopamine, we conclude that a dopamine receptor-adenylate cyclase complex is not involved in the modulation of neural transmission in the guinea pig SCG.     

Sympathoadrenal preganglionic neurons: their distribution and relationship to chemically-coded fibers in the kitten intermediolateral cell column

The location of those sympathetic preganglionic neurons in the spinal cord that project to the adrenal medulla--the sympathoadrenal preganglionic (SAP) neurons--was studied by the method of retrograde axonal transport of the fluorescent dye Fast Blue. The distribution of chemically-coded fibers and their relationship to the SAP neurons was also investigated using the indirect immunofluorescence technique. In kittens, 5 microliters of a 1% solution of Fast Blue was injected into the medulla of the left adrenal gland. After a survival period of 5 days, the spinal cords from C8 to L5 were sectioned and processed for the localization of enkephalin-, neurophysin-, oxytocin-, serotonin-, substance P- and somatostatin-like immunoreactivity. Retrogradely labeled neurons were found in the ipsilateral intermediolateral cell column (IML) (89.8% of all retrogradely labeled neurons) from T1 to L4, and in the contralateral IML (10.2%) from T1 to L4. The enkephalin, serotonin and substance P immunoreactive fibers appeared to surround both the retrogradely labeled and unlabeled IML neurons. The somatostatin immunoreactive fibers were observed only in proximity to the retrogradely labeled neurons. Only a sparse population of neurophysin and oxytocin immunoreactive fibers were observed in IML, and were not seen to be in apposition to the retrogradely labeled neurons.     

Long-term effects of preganglionic nerve stimulation on tyrosine hydroxylase activity in the rat superior cervical ganglion

The effects of increased synaptic stimulation of sympathetic neurons on the tyrosine hydroxylase activity of these cells were studied. Seventy-two hours after unilateral stimulation of the preganglionic cervical sympathetic trunk at 10 Hz for 30 min tyrosine hydroxylase activity was 32% higher in stimulated than in control superior cervical ganglia. Stimulation at 10 Hz for only 10 min increased enzyme activity by 25% when measured 72 h later, while stimulation for 60 min increased activity by 73%. No further change in enzyme activity was found after 90 min of stimulation although electrophysiological recordings from the ganglion demonstrated that synaptic transmission was maintained throughout the period of stimulation.Ganglionic neurons also follow high frequency trains of stimuli when they are interrupted by silent periods. Stimulation with 40 Hz trains (250 msec on/500 msec off) for 30 min and 90 min produced a 50% and a 92% increase in tyrosine hydroxylase activity, respectively. Stimulation of ganglia with the same number of pulses administered stered either continuously or in trains produced the same size increase in enzyme activity.The relationship between preganglionic nerve activity and tyrosine hydroxylase activity may represent an adaption of sympathetic neurons to situations requiring increased transmitter release.     

Development of the sex difference in neuron numbers of the superior cervical ganglion: effects of transection of the cervical sympathetic trunk

The number of superior cervical ganglion (SCG) neurons does not differ for males and females on the day of birth, but by 15 days, after most of the normal neuron death has occurred, males have significantly more neurons than females. This difference persists in the adult. The present study was undertaken to determine whether the presence of afferent input to the SCG is required for the development of this sex difference. Bilateral transection of the cervical sympathetic trunk, which deafferents the SCG neurons, or a sham operation was performed on male and female Sprague Dawley rats on the day of birth. Numbers of neurons were counted in SCGs of animals sacrificed on either postnatal day 4 or 15, before or after normal development of the SCG sex difference. At 4 days, the number of SCG neurons in sham-operated males and females were not different, but by 15 days, females had lost a significant number of neurons, whereas the males had not. Transection of the cervical sympathetic trunk led to a significant loss of over 6,000 SCG neurons by postnatal day 4 in both males and females. Whereas some of this loss is due to axotomy of caudally projecting SCG neurons, at least half of the neuron loss is due to removal of the afferent input. At 15 days, sham-operated males had significantly more SCG neurons than did sham-operated females, but the gender difference was not significant in animals with neonatally deafferented ganglia. Thus, the normal development of the gender difference in SCG neuron numbers requires an intact afferent input.     

Angiotensin II binding sites in the superior cervical ganglia of spontaneously hypertensive and Wistar-Kyoto rats after preganglionic denervation

Angiotensin II binding was higher in superior cervical ganglia of adult spontaneously hypertensive rats (SHR) when compared to ganglia of Wistar-Kyoto (WKY) rats (571 +/- 29 and 375 +/- 9 fmol/mg protein, SHR and WKY, respectively, P less than 0.05). Unilateral preganglionic denervation reduced binding site density in ganglia of WKY (-39%, P less than 0.05 vs sham operated ganglia in WKY), and the decrease of binding sites was larger in SHR (-59%, P less than 0.01, operated vs sham operated ganglia in SHR). Part of the binding sites in the superior cervical ganglia may be present in or be associated to preganglionic nerves, and the number of these sites is higher in SHR.     

Depletion by preganglionic stimulation and post-stimulus recovery of large dense core vesicles in synaptic boutons of the cat superior cervical ganglion.

Large dense core vesicles (LDCV), a possible site for neuropeptide storage, were counted within synaptic boutons of the superior cervical ganglion of anaesthetized cats in the unstimulated condition and 2 h, 1 day or 10 days after 40 Hz 2 h stimulation of the cervical sympathetic trunk. The number of LDCV decreased markedly after stimulation. It was lowest at 2 h and then recovered slowly. Ten days post-train the number was close to, but still significantly lower than, control. This time course of recovery is much longer than for agranular vesicles number, which is known to be back to normal within 2 h of the end of a comparable stimulus train. The post-stimulus loss of LDCVs is consistent with the hypothesis that LDCVs are released by the nerve terminal. The slow recovery is consistent with the hypothesis of replenishment by axoplasmic transport.     

Ultrastructural relationships between noradrenergic nerve fibers and non-neuronal elements in the rat cerebral cortex

Pharmacological and biochemical data suggest that noradrenaline (NA)-containing fibers not only regulate the activity of cortical neurons but also influence the functional state of non-neuronal elements. In the present study, immunocytochemistry with an antiserum against NA, followed by silver-gold intensification of the immunoreaction end-product, was employed to examine the ultrastructural relationships between the NA fiber system and the intraparenchymal blood vessels, oligodendrocytes, and astrocytes in the rat visual cortex. Electron microscopy revealed a large number of fine varicose NA fibers to be in intimate contact with cortical capillaries. Examination of single thin sections showed that NA boutons were usually separated from the capillary wall by a fine astroglial sleeve. However, serial section analysis revealed that the continuity of the astrocytic end feet was interrupted at sites, resulting in direct apposition of the perivascular NA fibers to the capillary basal lamina. Noradrenergic fibers were found to contact both types of macroglial cells. Single or clustered oligodendrocytes in intimate contact with NA fibers were observed throughout the cortical depth. Individual contacts could be followed in more than six successive thin sections, and oligodendrocyte plasma membrane frequently exhibited a light thickening at the sites of the NA fiber apposition. NA fiber-astroglial relationships were largely encountered in supragranular layers. In these layers, astrocytic cell bodies were characteristically outlined by fine varicose NA fibers. However, no plasma membrane differentiations were observed at the sites of intimate NA fiber apposition. The present ultrastructural findings provide the anatomical substrate for the control exerted by the NA fiber system over cortical microvasculature and macroglia. © 1996 Wiley-Liss, Inc.     

The superior cervical ganglion: Origin of sympathetic fibers in the facial and hypoglossal nerves in the cat

Location of superior cervical ganglion (SCG) neurons, sending axons into the facial and hypoglossal nerves, was investigated in the cat by means of retrograde axonal transport of horseradish peroxidase (HRP). After wheat germ agglutinin conjugated HRP (WGA-HRP) was injected into these nerves, many retrogradely labeled neurons were found widely in the ipsilateral SCG, particularly around the caudal half of the SCG. These neurons were round or oval in shape and 70-80% of these were medium in size. In fluorescent experiments, fast blue (FB) was used in combination with diamidino yellow (DY). After injections of FB into the facial nerve and DY into the hypoglossal nerve, a few FB-DY double-labeled neurons occurred in the SCG ipsilaterally.     

Effect of preganglionic application of vinblastine-impregnated silastic cuffs on acetylcholinesterase,butyrocholinesterase, and acetylcholine contents of cat superior cervical ganglion

Vinblastine sulfate in silastic cuffs was applied to the preganglionic vagosympathetic trunk of the cat superior cervical ganglion in an attempt to assess the role of acetylcholine as a possible neurotrophic factor required for maintaining postsynaptic ganglionic acetylcholinesterase and butyrocholinesterase. Following implantation of 1% (w/w) vinblastine cuffs, specific activities of acetylcholinesterase and butyrocholinesterase in the ganglion decreased to approximately 20 and 50% of control activities, respectively, within 1 week. The acetylcholine content of the superior cervical ganglion of most cats showed a moderate to marked decrease within this period as well. Histochemical findings were indistinguishable from those seen following preganglionic denervation. Because of the similar fall in ganglionic acetylcholine and enzyme contents, results with this technique do not eliminate acetylcholine as a possible neurotrophic factor for the maintenance of postsynaptic ganglionic acetyl- and butyrocholinesterases.     

Effects of glucagon on axoplasmic transport in mouse superior cervical ganglion cells.

The effects of glucagon on the axoplasmic transport of cultured mouse superior cervical ganglion cells were analyzed with the video-enhanced DIC microscope system. Glucagon increased the rate of fast axoplasmic transport by 30% in both anterograde and retrograde directions. The average velocity was increased from 1.36 +/- 0.48 microns/s to 1.74 +/- 0.43 microns/s (anterograde, n = 60) and from 1.37 +/- 0.48 microns/s to 1.62 +/- 0.39 microns/s (retrograde, n = 60). The stimulatory effect of glucagon on the axoplasmic transport was reversed in a glucose-free medium, whereas blocking the citrate cycle by pretreating neuronal cells with malonate did not alter the effect of glucagon. Together with our previous findings, our data suggest that neurotransmitters and hormones play a major role in the regulation of fast axoplasmic transport.     

Immunohistochemical study of synaptophysin distribution in the superior cervical ganglion of newborn and adult rats.

Intense synaptophysin immunoreactivity was observed around neuronal cell bodies and in the neuropil of the superior cervical ganglion of adult rats. In newborn rats synaptophysin was comparatively less concentrated and restricted to small interstitial spaces. In contrast, in newborns, consistent traces of positivity were found in the Golgi-like area of larger neurons, in agreement with the higher neonatal rate of synaptophysin synthesis. A few clusters of small neurons, numerically more expanded in adult rats, exhibited a dense reaction product filling the whole cytoplasm. No positivity was found in intraganglionic fibres.     

Corticotropin-releasing factor-like immunoreactive nerve fibers in the rat superior cervical ganglion and their fine structures

The existence of nerve fibers containing corticotropin-releasing factor (CRF)-like immunoreactivity (CRFI) in the rat superior cervical ganglion (SCG) was demonstrated by using immunocytochemistry. They were found to be extrinsic in origin, because no CRFI neurons were seen in the SCG and decentralization resulted in the disappearance of CRFI fibers in the SCG on the operated side. These findings were also confirmed by immunoelectron microscopic analysis; CRFI fibers contained a number of small clear synaptic vesicles but were devoid of large granular and agranular vesicles. These morphological characteristics are identical to those of the preganglionic fibers. The present immunoelectron microscopic analysis revealed that most of the CRFI fibers in the SCG make synaptic contact predominantly with the dendrites of the principal cells, partly with their somas and rarely with a non-CRFI terminal. Thus, the present study provides direct morphological evidence that CRF directly influences the function of the principal cells of the SCG and that CRFI fibers are preganglionic.