The spinal distribution of sympathetic preganglionic and visceral primary afferent neurons that send axons into the hypogastric nerves of the cat

The spinal distribution of sympathetic preganglionic neurons (PGN) and visceral primary afferent neurons sending axons into the hypogastric nerve of the cat has been studied with HRP tracing techniques. After application of HRP to the cat hypogastric nerve, labeled PGN were identified in segments L2-L5. Most of these neurons were oriented transversely and were divided approximately equally between two nuclei: the principal nucleus and the intercalated nucleus. Cells were distributed in clusters at 160-361-microns intervals along the length of the cord. Sensory neurons were labeled in dorsal root ganglia from T12 to L5. Central axons of these visceral afferents were observed in the medial half of Lissauer's tract from T13 to L7. Afferent axon collaterals extended through lamina I on both sides of the dorsal horn but were most prominent on the lateral side, where they continued into lateral lamina V and VII, often overlapping the dorsal dendrites of PGN in this region. Labeled afferent projections exhibited a periodic distribution in lamina I with clusters of axons occurring at 235-343-microns intervals in the rostrocaudal axis. The central projection of hypogastric nerve primary afferents was qualitatively similar to the distribution of visceral afferent projections at other levels of the spinal cord.     

An analysis of the sympathetic preganglionic neurons projecting from the upper thoracic spinal roots of the cat

The distribution of sympathetic preganglionic neurons that project via the right stellate ganglion has been studied quantitatively in adult cats. Retrograde transport of horseradish peroxidase (HRP) injected into the ganglion or applied to transected axons of the cervical sympathetic trunk (CST) resulted in labelling of neurons in the ipsilateral spinal cord over T1–T9 and T1–T7, respectively. Their distribution and morphology in the subnuclei of the intermediate zone were determined. Neurons within the principal part of the intermediolateral column (ILp) comprised the majority of labelled cells at all levels, irrespective of the site of HRP application, while more medially located neurons projected differentially. A combination of the application procedures labelled what appeared to be a sum of the individual projections, and few ILp cells remained unlabelled in the most rostral segments. From reconstructions of different segmental levels, most of the cells in the ILp were found to lie in a column ～200 μm in diameter composed of a series of cell aggregations of 20–150 neurons at intervals of ～300 μm. Less frequently, mediolaterally aligned cells extended toward the central canal, near which small cell clusters were also detected. After selective HRP application to individual rami of T1–T3, labelled cells were restricted to one segment's length, so that neurons located more caudally must project extraspinally to the stellate ganglion. Consideration of the maximum cell numbers labelled by each procedure suggests that preganglionic collaterals (probably unmyelinated) diverging from the pathway to the preferred target of their parent neuron were not labelled by this technique.     

Segmental organization of sympathetic preganglionic neurons in the mammalian spinal cord

We have used retrograde transport of horseradish peroxidase to determine the distribution of the preganglionic cell bodies whose axons join particular rami of the thoracic spinal cord in a series of guinea pigs, and in a small number of hamsters and cats. In contrast to other recent studies, our results show that the neurons sending axons to a ramus are confined to a single segment at the corresponding spinal level. This segmental organization supports the idea that the rostro-caudal position of preganglionic cell bodies is one determinant of selective synapse formation between preganglionic axons and sympathetic ganglion cells.     

Morphology of sympathetic preganglionic neurons in the thoracic spinal cord of the cat: an intracellular horseradish peroxidase study

Horseradish peroxidase was intracellularly injected into sympathetic preganglionic neurons (SPN) of the third thoracic segment in cats. Seven neurons were reconstructed from serial horizontal or parasagittal sections of the spinal cord. The cell bodies of all neurons were located in the n. intermediolateralis pars principalis (ILp). They were spindle-shaped with the long axis in craniocaudal direction or large and multipolar or small and oval in shape. Preferentially on the cranial and caudal pole of the cell body, five to eight primary dendrites arose from the cell body. Dendritic branches were traced to their terminations at distances up to 1,330 microns from the cell body. The dendritic fields of all SPNs were strictly oriented in the longitudinal direction with a total length of 1,500-2,540 microns. The cranial and caudal dendritic fields were about equal in length but, with one exception, the degree of branching was always greater in the cranial than in the caudal dendritic field. The dendritic fields of all SPNs were primarily restricted to the ILp. In the mediolateral direction it extended from 130 to 360 microns and in the dorsoventral direction from 50 to 180 microns. Only rarely, a higher-order dendrite left the boundaries of the ILp and projected dorsolaterally or laterally into the white matter or ventromedially or medially into the adjacent n. intercalatus. All dendrites showed various forms of spines. At a distance of 132-437 microns from the cell body the axon arose as a direct extension of a process which closely resembled a primary or second-order dendrite. The axons projected ventrally and mostly caudally along the lateral border of the gray matter until they turned laterally at the end of the ventral horn. No axon collaterals were observed.     

Locus coeruleus neurons and sympathetic nerves: Activation by visceral afferents

Previously brain norepinephrine (NE) neurons in the locus coeruleus (LC) have been shown to respond profoundly to external, environmental stimuli and are thought to be involved in behavioral functions such as vigilance, alarm and anxiety reactions to novel and, especially, threatening stimuli. Here we have used electrophysiological techniques to show that distension of the urinary bladder, the distal colon, rectum or the stomach causes pronounced activation responses of brain NE-LC neurons of the rat essentially without concomitant responses in splanchnic, sympathetic nerve activity (NE-SNA), thus indicating the non-noxious character of these internal stimuli. Our findings directly implicate the LC in micturition and, probably, defecation and we suggest that a high NE-LC activity may facilitate these phasic, vegetative events. In addition, the results implicate the LC as a pivotal system by which autonomic or visceral functions can affect behavior and, conversely, by which environmental stress can affect autonomic functions, for example in the opiate withdrawal syndrome.     

Reelin inhibits migration of sympathetic preganglionic neurons in the spinal cord of the chick

The present study examined the effects of Reelin in the migration of sympathetic preganglionic neurons (SPN) in the spinal cord of the chick. SPN in the chick first migrate from the neuroepithelium to the ventrolateral spinal cord. They then undergo a secondary migration to cluster adjacent to the central canal, forming the column of Terni (CT). During secondary migration, abundant Reelin is found in large areas of the ventral spinal cord; the only areas devoid of Reelin are areas occupied by SPN or somatic motor neurons and the pathway along which SPN migrate. Ectopic expression of Reelin in the pathway of SPN through electroporation of full-length Reelin DNA stopped SPN migration toward their destination. The spatiotemporal pattern of Reelin expression, along with the inhibition of SPN migration by exogenous Reelin, suggests that Reelin functions as a barrier to SPN migration during normal development of the spinal cord.     

Changes in synaptic inputs to sympathetic preganglionic neurons after spinal cord injury

Spinal cord injury (SCI) leads to plastic changes in organization that impact significantly on central nervous control of arterial pressure. SCI causes hypotension and autonomic dysreflexia, an episodic hypertension induced by spinal reflexes. Sympathetic preganglionic neurons (SPNs) respond to SCI by retracting and then regrowing their dendrites within 2 weeks of injury. We examined changes in synaptic input to SPNs during this time by comparing the density and amino acid content of synaptic input to choline acetyltransferase (ChAT)-immunoreactive SPNs in the eighth thoracic spinal cord segment (T8) in unoperated rats and in rats at 3 days or at 14 days after spinal cord transection at T4. Postembedding immunogold labeling demonstrated immunoreactivity for glutamate or gamma-aminobutyric acid (GABA) within presynaptic profiles. We counted the number of presynaptic inputs to measured lengths of SPN somatic and dendritic membrane and identified the amino acid in each input. We also assessed gross changes in the morphology of SPNs using retrograde labeling with cholera toxin B and light microscopy to determine the structural changes that were present at the time of evaluation of synaptic density and amino acid content. At 3 days after SCI, we found that retrogradely labeled SPNs had shrunken somata and greatly shortened dendrites. Synaptic density (inputs per 10-microm membrane) decreased on ChAT-immunoreactive somata by 34% but increased on dendrites by 66%. Almost half of the inputs to SPNs lacked amino acids. By 14 days, the density of synaptic inputs to dendrites and somata decreased by 50% and 70%, respectively, concurrent with dendrite regrowth. The proportion of glutamatergic inputs to SPNs in spinal cord-transected rats ( approximately 40%) was less than that in unoperated rats, whereas the GABAergic proportion (60-68%) increased. In summary, SPNs participate in vasomotor control after SCI despite profound denervation. An altered balance of excitatory and inhibitory inputs may explain injury-induced hypotension.     

Glutamate-immunoreactive synapses on retrogradely-labelled sympathetic preganglionic neurons in rat thoracic spinal cord

Retrograde tracing with cholera toxin B subunit (CTB) combined with post-embedding immunogold labelling was used to demonstrate the presence of glutamate-immunoreactive synapses on sympathetic preganglionic neurons that project to the adrenal medulla or to the superior cervical ganglion in rat thoracic spinal cord. At the electron microscope level, glutamate-immunoreactive synapses were found on retrogradely labelled nerve cell bodies and on dendrites of all sizes. Two-thirds of the vesicle-containing axon profiles that were directly apposed to, or synapsed on, CTB-immunoreactive sympathoadrenal neurons were glutamate positive. The proportion of glutamate-immunoreactive contacts and synapses on sympathoadrenal neurons decreased to zero when the anti-glutamate antiserum was absorbed with increasing concentrations of glutamate from 0.1 mM to 10 mM. Double immunogold labelling for glutamate and gamma-aminobutyric acid (GABA) showed that glutamate-immunoreactive profiles did not contain GABA and that GABA-immunoreactive profiles did not contain glutamate. These results suggest that glutamate is the major excitatory neurotransmitter to sympathoadrenal neurons and possibly to other sympathetic preganglionic neurons in the intermediolateral cell column of the spinal cord.     

Ectopic expression of reelin alters migration of sympathetic preganglionic neurons in the spinal cord

Reelin, an extracellular matrix molecule, regulates neuronal positioning in the brain, brainstem, and spinal cord. Although Reelin was identified more than a decade ago, its function on neuronal migration is still poorly understood. Using a transgenic mouse that expressed reelin under the nestin promoter, we examined here the function of Reelin in control of sympathetic preganglionic neurons (SPN) migration in the spinal cord. SPN undergo primary and secondary migration to arrive at their final locations. In wildtype mice, postmitotic SPN undergo primary migration from the neuroepithelium to the ventrolateral spinal cord, and then undergo a secondary dorsal migration to their final location to form the intermediolateral column (IML). In reeler, which lacks Reelin, SPN also undergo primary migration to the ventrolateral spinal cord as in wildtype. However, during secondary migration, SPN migrate medially to cluster adjacent to the central canal. Our present study on transgenic rl/rl mutants (rl/rl ne‐reelin) shows that the initial migration of SPN (embryonic day [E]9.5–E12.5) was similar to reeler. SPN migrated from the neuroepithelium to the ventrolateral spinal cord and then back toward the central canal, despite strong reelin expression in the ventricular zone. However, SPN did not aggregate near the central canal when ectopic reelin was expressed. Only when the expression level of ectopic reelin in the ventricular zone became very weak (E18.5) were SPN found to cluster near the central canal. Postnatally, SPN in rl/rl ne‐reelin transgenic mice were located in both the IML and near the central canal. These results show that SPN position can change with location and level of reelin expression. Possible functions of Reelin on SPN migration are discussed. J. Comp. Neurol. 515:260–268, 2009. © 2009 Wiley‐Liss, Inc.     

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 μm 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. © 1995 Wiley-Liss, Inc.     

Neonate rat sympathetic preganglionic neurons intracellularly labelled with lucifer yellow in thin spinal cord slices

Sympathetic preganglionic neurons and interneurons were intracellularly labelled with lucifer yellow in thin transverse spinal cord slices of neonatal rats. Preganglionic neurons had spindle or oval shape somata and were located in the intermediolateral nucleus. The axons of these neurons coursed ventrally along the border of gray matter and exited the ventral horn; two to four long dendrites projected medially to the central canal and several relatively short dendrites oriented toward the lateral white matter. Interneurons were generally multipolar and located outside the immediate area of intermediolateral nucleus; their axons could sometimes be traced to the ventral funiculus. Interestingly, dye-coupled preganglionic neurons were observed for the first time. Our findings suggest that the dendritic domain of neonatal rat sympathetic preganglionic neurons is out-reaching and may represent potential sites of interaction with incoming segmental and/or descending inputs. In addition, the observation of dye-coupled preganglionic neurons raises the possibility that these neurons may have the capability of recruiting and/or synchronizing sympathetic outflow.     

Distribution of sympathetic preganglionic neurons and monoaminergic nerve terminals in the spinal cord of the rat

A study was made of the distribution of sympathetic preganglionic neurons identified by retrograde labeling with horseradish peroxidase from various peripheral nerve trunks and of the distributions of monoaminergic terminals in the spinal cord of the rat. Nerve terminals were stained immunohistochemically by using antisera raised against tyrosine hydroxylase, phenylethanolamine-N-methyl-transferase, neuropeptide Y, and 5-hydroxytryptamine and by using formaldehyde-induced fluorescence. The three-dimensional distribution of sympathetic preganglionic neurons was described by using computer reconstruction and compared with the arrangement of each population of immunohistochemically stained terminals in the intermediate zone. Although monoaminergic terminals are associated with most sympathetic neurons, particularly in the intermediolateral column, the relationship of many terminals to sympathetic neuron somata in other parts of the intermediate zone is tenuous. Some of the descending innervation may terminate on interneurons. The data are consistent with the coexistence of phenylethanolamine-N-methyl-transferase and neuropeptide Y in terminals arising from cell bodies in the C1 region in the ventrolateral medulla and with the presence of at least two populations of catecholaminergic terminals as well as the adrenergic one. Serotoninergic terminals are denser and have a different arrangement from those of catecholaminergic terminals in the intermediate zone.     

Central projections of thoracic splanchnic and somatic nerves and the location of sympathetic preganglionic neurons in Xenopus laevis

The central and peripheral organization of thoracic visceral and somatic nervous elements was studied by applying dextran amines to the proximal cut ends of the thoracic splanchnic and somatic nerves in Xenopus laevis. Many labeled dorsal root ganglion cells of visceral afferents, and all somatic afferents, were located in a single ganglion of one spinal segment, and the two types of cells were distributed topographically within the ganglion. The labeled sympathetic preganglionic neurons were located predominantly in the same area of the thoracic spinal gray as in other frogs and in mammals. The labeled visceral afferents projected to Lissauer's tract and the dorsal funiculus. The visceral fibers of the tract ascended to the level of the subcerebellar area, supplying collateral branches to the lateral one-third of the dorsal horn and to the area of brainstem nuclei, including lateral cervical and descending trigeminal nucleus, and descended to the filum terminale. The visceral fibers of the dorsal funiculus were distributed to the dorsal column nucleus and the solitary tract. A similar longitudinal projection was also seen in the somatic afferents. The dual central pathway of thoracic primary afferents in the anuran spinal cord is a property held in common with mammals, but the widespread rostrocaudal projection through Lissauer's tract may be a characteristic of the anuran central nervous system. In frogs, the direct transmission of primary afferent information to an extremely wide area of the central nervous system may be important for prompt assessment of environmental factors and control of body functions.     

Spinal origin of sympathetic preganglionic neurons in the rat

The segmental distribution of sympathetic preganglionic neurons (SPNs) and dorsal root ganglion cells (DRGs) was studied after Fluoro-gold injections into the major sympathetic ganglia and adrenal gland in rats. A quantitative assessment of the segmental and nuclear locations was made. Four general patterns of innervation were apparent: (1) a large number of SPNs (1000–2000/ganglion) innervate the sympathetic ganglia which control head or thoracic organs and a relatively small number of SPNs (100–400/ganglion) innervate the sympathetic ganglia controlling the gut, kidney, and pelvic organs; this difference in density of innervation probably relates to the level of fine control that can occur in these end organs by the SPNs; (2) the reverse pattern is seen in the DRG labeling where a large number of DRGs were labeled after Fluoro-gold injections into the preaortic ganglia (celiac, superior, and inferior mesenteric) and a small number were labeled after injections into the cervical sympathetic ganglia; (3) the intermediolateral cell column is the main source of SPNs except for the inferior mesenteric ganglion which is innervated predominantly by SPNs originating in the central autonomic nucleus (75%); the lateral funiculus is a source of SPNs mainly for the cervical sympathetic ganglia; and (4) each sympathetic ganglion and the adrenal gland receives a multisegmental SPN and DRG input with one segment being the predominant source of the innervation. The adrenal gland shows an intermediate position in terms of the density of SPN input (800 cells) and dorsal root input (300 cells); it has a widespread segmental input (T₄-T₁₂) with the T₈ segment being the major source.