Re-establishment of neurochemical coding of preganglionic neurons innervating transplanted targets.

We investigated the effect on neurochemical phenotype of changing the targets innervated by sympathetic preganglionic neurons. In neonatal rats, the adrenal gland was transplanted into the neck, to replace the postganglionic neurons of the superior cervical ganglion. Transplanted adrenal glands survived, and contained noradrenergic and adrenergic chromaffin cells, and adrenal ganglion cells. Retrograde tracing from the transplants showed that they were innervated by preganglionic neurons that would normally have supplied postganglionic neurons of the superior cervical ganglion. The neurochemical phenotypes of preganglionic axons innervating transplanted chromaffin cells were compared with those innervating the normal adrenal medulla or superior cervical ganglion neurons. As in the normal adrenal gland, preganglionic nerve fibres apposing transplanted chromaffin cells were cholinergic. The peptide and calcium-binding protein content of preganglionic fibres was similar in normal and transplanted adrenal glands. In both cases, cholinergic fibres immunoreactive for enkephalin targeted adrenergic chromaffin cells, whilst cholinergic fibres with co-localised calretinin-immunoreactivity innervated noradrenergic chromaffin cells and adrenal ganglion cells. In contrast to the innervation of normal adrenal glands, these axons lacked immunoreactivity to nitric oxide synthase. In a set of control experiments, the superior cervical ganglion was subjected to preganglionic denervation in rat pups the same age as those that received adrenal transplants, and the ganglion was allowed to be re-innervated over the same time course as the adrenal transplants were studied. When the superior cervical ganglion was re-innervated by preganglionic nerve fibres, we observed that all aspects of chemical coding were restored, including cholinergic markers, nitric oxide synthase, enkephalin, calcitonin gene-related peptide and calcium binding proteins in predicted combinations, although the density of nerve fibres was always lower in re-innervated ganglia. These data show that the neurochemical phenotypes expressed by preganglionic neurons re-innervating adrenal chromaffin cells are selective and similar to those seen in the normal adrenal gland. Two explanations are advanced: either that contact of preganglionic axons with novel target cells has induced a switch in their neurochemical phenotypes, or that there has been target-selective reinnervation by pre-existing fibres of appropriate phenotype. Regardless of which of these alternatives is correct, the restoration of normal preganglionic codes to the superior cervical ganglion following denervation supports the idea that the target tissue influences the neurochemistry of innervating preganglionic neurons.

[Changes of the nerve fibers innervating the minor salivary glands in Sjögren syndrome]. / A kis nyálmirigyek beidegzésében résztvevó idegelemek változása Sjögren-szindrómában.

INTRODUCTION: A large number of nerve fibres containing different neuropeptides/transmitters are also found in the salivary glands. The number and the distribution of nerve fibres is altered in many diseases, including in Sjögren's syndrome. AIM: Therefore in the present study the distribution and precise localisation of the nerve fibres containing the frequently observed neuropeptides were studied in the minor salivary glands. METHODS: Vasoactive intestinal polypeptide, neuropeptide Y, substance P, calcitonin gene-related peptide, somatostatin, nitric oxide synthase and tyrosine beta-hydroxylase antibodies were used as primary antisera, and then by the aid of avidin-biotin-peroxidase complex method the immunoreactive fibers in human labial glands (control and with Sjögren's syndrome) and in minor glands of the root of the rat's tongue were detected. RESULTS: Large number of vasoactive intestinal polypeptide and nitric oxide synthase immunoreactive nerve fibres were seen around the acini. The neuropeptide Y and tyrosine beta-hydroxylase positive nerve fibres were mainly found around the blood vessels. Some of the IR fibers were also found around the excretory ducts. In the biopsy of patients with Sjögren's syndrome, the acini were destroyed and only few excretory ducts were seen. The number of the nerve fibres was significantly decreased and many degenerated fibres were also observed among the acini. The electron-microscopic examinations showed that the immunoreactive nerve fibres were in close association to the secretory cells, to the smooth muscle cells of blood vessels and to the immunocells. The synaptic gap between the nerve fibres and the target cells were 40-200 nm. CONCLUSIONS: On the bases of the distribution of the different transmitters containing nerve fibres and their relationship to effector cells, the authors suppose that these transmitters control the function of the gland and regulate the blood flow. The close association to immunocells and decreasing the nerve fibres in Sjögren's syndrome imply that they may have also a role in the neuroimmunologic processes.

Chemically distinct preganglionic inputs to iris-projecting postganglionic neurons in the rat: A light and electron microscopic study.

Individual autonomic postganglionic neurons are surrounded by pericellular baskets of preganglionic terminals that are easily identifiable with the light microscope. It has been assumed that the target cell of a pericellular basket of preganglionic terminals is the neuron at the centre of the basket. This assumption has enabled the connectivity of preganglionic neurons to be determined at the light microscopic level. However, if the preganglionic terminals in a pericellular basket make synapses with the dendrites of nearby, but functionally different, postganglionic neurons, then the conclusions of light microscopic studies are far less certain. We have used a serial section ultrastructural study to determine the target of the preganglionic pericellular basket in a situation where the apparent target cell is surrounded by neurons of dissimilar function. In the rat superior cervical ganglion, postganglionic neurons projecting to the iris were identified, using retrograde tracers, as single neurons (i.e., not in clusters). We have used immunohistochemistry to show that iris-projecting neurons are surrounded by preganglionic nerve terminals containing calcitonin gene-related peptide (CGRP). We have demonstrated that the pericellular basket of CGRP-immunoreactive preganglionic terminals provides inputs only to the soma at the centre of the basket and not to the dendrites of surrounding neurons. This suggests that, in autonomic ganglia, light microscopic identification of the preganglionic terminal baskets is likely to be a reliable method for identifying the targets of subclasses of preganglionic neurons.

Origin and neurochemical characteristics of nerve fibres supplying the mammalian vas deferens.

The present paper deals with the origin and neurochemical characteristics of autonomic postganglionic and sensory nerve fibres supplying the mammalian vas deferens. The vas deferens is innervated by postganglionic nerve fibres originating primarily from neurons in pelvic ganglia and, to a lesser extent, from neurons in the inferior mesenteric ganglion and sympathetic chain ganglia as well as by sensory nerve fibres arising from dorsal root ganglia. Three major populations of nerve terminals innervating the organ can be distinguished: (1) noradrenergic fibres; (2) cholinergic fibres containing vasoactive intestinal polypeptide, neuropeptide Y, nitric oxide synthase, and (in the pig) somatostatin, supplying particularly the lamina propria; and (3) non-noradrenergic, presumably sensory fibres, containing calcitonin gene-related peptide and/or substance P. The population of noradrenergic nerves is the most common. In the pig, it can be divided into three subpopulations: a somatostatin-containing, a Leu-enkephalin-containing and a subpopulation immunonegative to these peptides, in descending order of magnitude. In the rat, guinea-pig, and man, NPY seems to be the most common peptide occurring in the noradrenergic axons. In the pig, coexistence patterns of the substances existing within nerve fibres supplying the vas deferens blood vessels are clearly different from those found in nerve fibres innervating the organ wall. The majority of the noradrenergic fibres associated with blood vessels contain neuropeptide Y only, while non-noradrenergic perivascular nerves contain predominantly vasoactive intestinal polypeptide. The possibility of different sources of origin of the particular nerve fibre subpopulations supplying the mammalian vas deferens and its blood vessels is discussed.

Colocalization of acetylcholinesterase and vasoactive intestinal peptide (VIP) in nicotinamide adenine dinucleotide phosphate diaphorase (NADPH-d) positive neurons in the intralingual ganglia and perivascular nerve fibers around lingual arteries in the porcine, monkey and canine tongue.

Distribution of nitric oxide synthase in the intrinsic ganglia in the porcine, monkey and canine tongue was histologically investigated using the reduced nicotinamide adenine dinucleotide phosphate diaphorase (NADPH-d) method, acetylcholinesterase histochemistry and vasoactive intestinal peptide (VIP) immunohistochemistry. The majority of intralingual ganglionic cells showed intense NADPH-d reactivity with positive acetylcholinesterase reaction or positive VIP immunohistochemistry. The NADPH-d positive, acetylcholinesterase-rich and the NADPH-d positive, VIP immunoreactive nerve fibers are particularly conspicuous around intralingual blood vessels. These fibers around the arteries in the tongue may be partly derived from the intralingual ganglion cells, because some bundles associated with these nerve cells were easily traced on the wall of blood vessels. The present study suggests the view that the three markers coexist in the axons and nerve terminals of these intralingual neurons.

Proportions of renal and splenic postganglionic sympathetic populations containing galanin and dopamine beta hydroxylase.

Galanin is a 29-amino acid neuropeptide found in rat spinal cord, autonomic ganglia and gastrointestinal tract, as well as in other areas of the nervous system in rats and other species. As part of an overall objective to determine if peptides contribute to target-specific control of visceral function, this study was designed to determine the percentages of populations of renal and splenic postganglionic neurons that contain galanin, and to determine if these neurons were likely to be adrenergic. Retrogradely transported fluorescent dyes were placed on renal and splenic nerves in male Wistar rats anaesthetized with sodium pento-barbital. Four days post-operatively, rats were perfused transcardially with fixative, and T12-L1 thoracolumbar chain ganglia, splanchnic ganglia and the solar plexus were removed. Immunocytochemical methods were then used to determine the proportions of the retrogradely labelled renal and splenic neurons containing galanin-like immunoreactivity and dopamine beta hydroxylase-like immunoreactivity. In seven rats, 24 +/- 3% of 2838 renal neurons were found to contain galanin-like immunoreactivity; in six rats, 32 +/- 5% of 5102 splenic neurons were found to contain galanin-like immunoreactivity. These proportions of the two populations were not significantly different from one another. In three rats, 94 +/- 2% of 684 renal neurons were found to contain dopamine beta hydroxylase-like immunoreactivity, and 95 +/- 2% of 2597 splenic neurons in three rats also showed dopamine beta hydroxylase-like immunoreactivity. These experiments indicate that subpopulations of both renal and splenic postganglionic sympathetic neurons contain the neuropeptide galanin and that these neurons are likely to be adrenergic in function. These findings suggest a role for galanin in control of the kidney and the spleen by the sympathetic nervous system.

Central nervous system innervation of the penis as revealed by the transneuronal transport of pseudorabies virus.

Transneuronal tracing techniques were used in order to identify putative spinal interneurons and brainstem sites involved in the control of penile function. Pseudorabies virus was injected into the corpus cavernosus tissue of the penis in rats. After a four day survival period, rats were perfused with fixative and virus-labelled neurons were identified by immunohistochemistry. Postganglionic neurons were retrogradely labelled in the major pelvic ganglia. In the spinal cord, sympathetic and parasympathetic preganglionic neurons were labelled transneuronally. Presumptive interneurons were also labelled in the lower thoracic and lumbosacral spinal cord in locations consistent with what is currently known about such interneurons. In the brainstem, transneuronally labelled neurons were found in the medulla, pons and hypothalamus. Regions consistently labelled included the nucleus paragigantocellularis, parapyramidal reticular formation of the medulla, raphe pallidus, raphe magnus, A5 noradrenergic cell group, Barrington's nucleus and the paraventricular nucleus of the hypothalamus. This study confirmed previous studies from our lab and others concerning the preganglionic and postganglionic neurons innervating the penis. The number, morphology and location of these neurons were consistent with labelling seen following injection of conventional tracers into the penis. The brainstem nuclei labelled in this study were also consistent with what is currently known about the brainstem control of penile function. The labelling appeared to be highly specific, in that descending systems involved in other functions were not labelled. These results provide further evidence that the pseudorabies virus transneuronal tracing technique is a valuable method for identifying neural circuits mediating specific functions.

Vasomotor, pilomotor and secretomotor neurons distinguished by size and neuropeptide content in superior cervical ganglia of mice.

Populations of postganglionic sympathetic neurons projecting to cranial targets from the superior cervical ganglia of mice were identified by retrograde axonal tracing with Fast blue combined with double-labelling immunofluorescence to detect immunoreactivity to tyrosine hydroxylase and neuropeptide Y. Nearly all neurons in the ganglion contained tyrosine hydroxylase immunoreactivity, but only about 50% of them also contained immunoreactivity to neuropeptide Y. The maximum diameter of cells with immunoreactivity to neuropeptide Y was significantly smaller than that of cells without it. Terminal axons containing immunoreactivity to both neuropeptide Y and tyrosine hydroxylase occurred around blood vessels supplying most cranial tissues, including the skin. Axons with immunoreactivity to tyrosine hydroxylase but not to neuropeptide Y innervated the piloerector muscles and the acini of the salivary glands. After injection of Fast blue into the skin or the submandibular salivary gland, populations of vasomotor, pilomotor and secretomotor neurons could be distinguished by soma size and by neuropeptide Y immunoreactivity. Neurons projecting to the salivary glands were the largest (mean diameter: 32 microns) and lacked immunoreactivity to neuropeptide Y; neurons projecting to cutaneous blood vessels were the smallest (mean diameter: 19 microns) and contained immunoreactivity to neuropeptide Y; neurons projecting to piloerector muscles were intermediate in size (mean diameter: 23 microns) and lacked neuropeptide Y immunoreactivity. A cluster analysis procedure confirmed that soma size and peptide content together identify major functional populations of neurons in the superior cervical ganglia of mice.

Distribution of postganglionic parasympathetic fibers originating in the pterygopalatine ganglion in the maxillary and ophthalmic nerve branches of the trigeminal nerve; HRP and WGA-HRP study in the guinea pig.

The distribution of the postganglionic parasympathetic fibers originating in the pterygopalatine ganglion (PTPG) has been traced in the guinea pig by means of the HRP and WGA-HRP methods. The greatest number of labeled cells were observed when WGA-HRP was injected in the lacrimal gland. After applying HRP to all the ramifications of the maxillary and ophthalmic divisions of the trigeminal nerve, labeled neurons were found in the PTPG. Numerous PTPG fibers were detected in the ethmoidal and sphenopalatine nerves. The presence of PTPG fibers in the supraorbital, infratrochlear, zygomaticotemporal, zygomaticofacial-inferior palpebral, sphenopalatine and infraorbital-superior alveolar nerves has not hitherto been reported in mammals.