Synaptic organisation of the pelvic ganglion in the guinea-pig

Summary A semi-quantitative electron-microscopic study of neuronal cell bodies, nerve profiles and synapses in the anterior pelvic ganglia of the guinea-pig has been carried out following in vivo labelling of adrenergic nerve endings with 5-hydroxydopamine. Ganglion cells of three main types have been distinguished: 1) the majority (about 70%) not containing granular vesicles, probably cholinergic elements; 2) those containing large granular vesicles of uniform size (80–110 nm), with granules of medium density and prominent halos; and 3) those containing vesicles of variable size (60–150 nm), with very dense eccentrically placed granular cores. Some non-neuronal granule-containing cells were present, mainly near small blood vessels. Some 95% of the total axon profiles within the ganglia were cholinergic, the remaining 5% were adrenergic. However, 99% of synapses (i.e. axons within 50 nm of nerve cell membrane with pre-and post-synaptic specialisations) were cholinergic, and 1 % were adrenergic. Only three examples of nerve cell bodies exhibiting both cholinergic and adrenergic synapses were observed. Unlike the para-and prevertebral ganglia, the pelvic ganglia contained large numbers of axo-somatic synapses. As many as 20% of the nucleated neuronal cell profiles displayed two distinct nuclei.     

Distribution of subgroups of noradrenaline neurons in the coeliac ganglion of the guinea-pig

Summary The distributions within the coeliac ganglion of different chemically coded subgroups of noradrenaline neurons, and the relationships between these neurons and nerve fibres projecting to the ganglion from the intestine, have been assessed quantitatively by use of an immunohistochemical double-staining method. Noradrenaline (NA) neurons made up 99% of all cell bodies. Of these, 21% were also reactive for somatostatin (NA/SOM neurons), 53% were also reactive for NPY (NA/NPY neurons), and 26% were not reactive for either peptide. NA neurons without reactivity for any of the peptides whose localization was tested have been designated NA/-. A small percentage, about 1%, of neurons were reactive for both NPY and SOM. The three major types of NA neurons were arranged in clumps or ribbons throughout the ganglia, with a tendency for NA/SOM neurons to be medial and NA/NPY neurons to be lateral in the ganglia. A small group of neurons (<1%) encoded with dynorphin, NPY and vasoactive intestinal peptide (VIP) was encountered. VIP-immunoreactive nerve terminals, projecting to the ganglion from cell bodies in the intestine, ended around NA/SOM and NA/neurons but not around NA/NPY neurons. Thus, the VIP axons from the intestine end selectively around neurons that modify intestinal function (NA/SOM and NA/-neurons) but not around neurons, the terminals of which supply blood vessels (NA/NPY neurons).     

The granule-containing cell somata in the superior cervical ganglion of the rat,as studied by a serial sampling method for electron microscopy

Summary The surface of 4 granule-containing cells, in a cluster within the rat superior cervical ganglion, was studied by a serial sampling technique for electron microscopy. The result shows that all the 4 cells receive one, or three afferent synaptic boutons from the preganglionic fibers impinging upon their somata, and a somatic efferent synapse exists at two locations on each soma of the 2 of these cells. The postsynaptic element of the efferent synapse is observed to be represented by non-vesiculated and vesiculated segments of dendrites, soma and a possible axon collateral of the adrenergic principal neuron of the ganglion. There is a remarkably constant development of the attachment plaque between the granule-containing cells themselves, representing 1.7–2.3% of surface area for each cell. The surface area exposed to the extracellular space (covered only by a basal lamina) varies from 0.1 to 2.3% of the total perikaryal surface of the 4 cells. A tendency is noted that those cells without efferent synapses possess a more extensive area exposed to extracellular space than those forming somatic efferent synapse to the postganglionic elements.It is a pleasure to acknowledge the advice and encouragement of Professor A. Yamauchi throughout this work. I thank Mr. K. Kumagai and Miss K. Tsushida for their technical assistance.     

The major ganglion in the pelvic plexus of the male rat

Summary To further evaluate the role of autonomic ganglia in the regulation of pelvic visceral activity, the neural elements in the major pelvic ganglion of the male rat have been studied with histochemical and electron microscopic techniques. The principal findings are that the ganglion is composed of cholinergic and adrenergic ganglion cells as well as small intensely fluorescent (SIF) cells. Polarity in the ganglion is indicated by clustering of small ganglion cells which stain intensely for acetylcholinesterase (AChE) along the pelvic nerve while larger cells, with weak to moderate AChE activity, collect near small branches of the hypogastric nerve. Some cholinergic ganglion cells are enclosed by a plexus of adrenergic terminals. SIF cells appear to be in contact with both cholinergic and adrenergic cells, although many of the fluorescent beads around adrenergic neurons may be short dendrites of ganglion cells, rather than processes of SIF cells. Two types of SIF cells may be distinguished on the basis of size and morphology of their granulated vesicles. Afferent synapses of the cholinergic type were common on SIF cells of the large granule and small granule type. Portions of SIF cells with large granules occur within the capsule of ganglion cells. Contacts seen here were interpreted as efferent synapses from SIF cells to the dendrites of ganglion cells.     

Adrenergic innervation of the tunica media in the saphenous artery of the fetal and newborn guinea-pig

Summary Previous studies have demonstrated that adrenergic nerves are located in the medial-adventitial border of the muscular arteries. Observations made in this study have revealed that adrenergic nerves penetrate into the outer medial layer of the saphenous artery in fetal and newborn guinea-pigs, while in the adult these nerves are located in the medial-adventitial border. It is proposed that the adrenergic nerves located in the tunica media may have a trophic effect on the medial smooth muscle. It is further suggested that the final refinement of the dual control system of arterial walls, by nerves and circulating catecholamines, involves exclusion of adrenergic nerves from the tunica media.     

Distribution of enteric nerve cells that project to the coeliac ganglion of the guinea-pig

Summary The digestive tract of the guinea-pig, from the esophagus to the rectum, was examined in detail to determine the distribution and relative abundances of neurons in these organs that project to the coeliac ganglion and the routes by which their axons reach the ganglion. A retrogradely transported neuronal marker, Fast Blue, was injected into the coeliac ganglion. The esophagus, stomach, gallbladder, pancreas, duodenum, small intestine, caecum, proximal colon, distal colon and rectum were analysed for labelled neurons. Retrogradely labelled neurons were found only in the myenteric plexus of these organs, and in the pancreas. No labelled neurons were found in the gallbladder or the fundus of the stomach, or in the submucous plexus of any region. A small number of labelled neurons was found in the gastric antrum. An increasing density of labelled neurons was found along the duodenum. Similarly, an increasing density of labelled neurons was found from proximal to distal along the jejuno-ileum. However, the greates densities of labelled neurons were in the large intestine. many labelled neurons were found in the caecum, including a high density underneath its taeniae. An increasing density of labelled neurons was found along the length of the proximal colon, and labelled neurons were found in the distal colon and rectum. In total, more labelled cell bodies occurred in the large intestine than in the small intestine. The routes taken by the axons of viscerofugal neurons were ascertained by lesioning the nerve bundles which accompany vessels supplying regions of the digestive tract. Viscerofugal neurons of the caecum project to the coeliac ganglion via the ileocaeco-colic nerves; neurons in the proximal colon project to the ganglion via the right colic nerves, and neurons in the distal colon project to the ganglion via the mid colic and intermesenteric nerves. Neurons in the rectum project to the coeliac ganglion via the intermesenteric nerves. These nerves (except for the intermesenterics) all join nerve bundles from the small intestine that follow the superior mesenteric artery. All viscerofugal neurons of the caecum were calbindin-immunoreactive (calb-IR) and 94% were immunoreactive for vasoactive intestinal peptide (VIP-IR). In the proximal colon, 49% of labelled neurons were calb-IR and 85% were VIP-IR. In the distal colon, 80% of labelled neurons were calb-IR and 71% were VIP-IR.     

Different types of small granule-containing cells and neurons in the guinea-pig adrenal medulla

Summary An electron microscopic, histoand biochemical study was carried out on the adrenal medulla of newborn and adult guinea-pigs giving special emphasis to small granule-containing (SGC) cells. Adrenaline (A) was the predominating catecholamine (CA) both in newborn (70–90 % of total CA) and adult (85–90%) guinea-pig adrenals. In analogy to the biochemical findings electron microscopy revealed a high predominance of A cells, which contained large granular vesicles with an average diameter of 180 nm. Most noradrenaline (NA) storing cells showed granular vesicles of a considerably smaller average diameter (80 nm) and had a higher nuclear-cytoplasmic ratio. These cells were termed SGC-NA cells. NA cells with large granular vesicles (average diameter 170 nm) were extremely rare. Another type of SGC cells contained granular vesicles with cores of low to medium electron-density (SGC-NA-negative cells). Biochemical determinations made it unlikely that these cells contained predominantly dopamine (DA). SGC cells were scarcely innervated by cholinergic nerves. They formed processes, which were found both in the adrenal cortex and medulla contacting blood vessels including sinusoid capillaries, steroid producing cells of the reticularis and fasciculata zone and processes, which were interpreted to belong to medullary nerve cells.Two types of neurons were present in the guinea-pig adrenal medulla, one resembling the principal neurons in sympathetic ganglia, the other, which, according to its morphology, occupied an intermediate position between principal neurons and SGC cells.In adrenomedullary grafts under the kidney capsule, which were studied three weeks after transplantation, ordinary A cells resembled SGC-NA negative cells with respect to their ultramorphology. Processes of transplanted principal neurons showed uptake of 5-hydroxydopamine and, hence, were considered to be adrenergic. Despite the lack of extrinsic nerves to the transplants, few principal neurons received cholinergic synapses, the origin of which is uncertain to date.Supported by a grant from Deutsche Forschungsgemeinschaft (Un 34/4)Dedicated to Professor H. Leonhardt in honor of his 60th birthday.     

Intramural neurons in the urinary bladder of the guinea-pig

Summary The urinary bladder of adult female guinea-pigs was stained histochemically to detect the presence of intramural ganglion neurons. Counts on wholemount preparations of entire bladders revealed the presence of 2000–2500 neurons per bladder, either as individual nerve cells or, more often, as ganglia containing up to 40 neurons. Both ganglia and single neurons lie along nerve trunks and are interconnected to form a plexus. Ganglia occur in every part of the bladder; they are more numerous on the dorsal than on the ventral wall, and they are especially abundant in an area within a radius of 800 m from the point of entry into the bladder wall of ureters and urinary arteries. The ganglia are located inside the muscle coat and close to muscle bundles; they usually lie nearer the mucosa than the serosa. Ultrastructurally, each ganglion is surrounded by a capsule; in addition to neurons and glial cells, the ganglia contain capillaries, collagen fibrils and fibroblasts; ganglion neurons are individually wrapped by glial cells and are separated from one another by connective tissue.     

NADPH-diaphorase activity in the nodose ganglion of normal and vagotomized guinea-pigs

The activity and distribution of reduced nico-tinamide adenine dinucleotide phosphate diaphorase (NADPH-diaphorase) in the nodose ganglion of normal and vagotomized guinea-pigs were examined by light and electron microscopy. Light microscopy confirmed a remarkable increase in the number of NADPH-diaphorase-reactive neurons in the nodose ganglion following unilateral cervical vagotomy. The increase was present at 5 days but became more prominent at 10 days and was sustained until at least 30 days after vagotomy when compared with the non-lesioned side. The NADPH-diaphorase reaction product was associated with the membrane of the rough endoplasmic reticulum, Golgi apparatus, mitochondria and nucleus of the nodose neurons. In animals killed 5 days post-operation, there was no noticeable degeneration in the nodose neurons. However, at 10 days, the mitochondria in some neurons appeared swollen and vacuolated with disrupted cristae. These changes were accentuated in some nodose neurons 20 and 30 days after vagotomy but there was no evidence of cell death. All the degenerating neurons exhibited NADPH-diaphorase activity. The increase in NADPH-diaphorase activity in the neuronal somata after vagotomy suggests that the enzyme is involved in either the retrograde degeneration or the recovery of the lesioned neurons. Received: 15 June 1995 / Accepted: 15 February 1996     

Sensory collaterals, intramural ganglia and motor nerves in the guinea-pig bladder: evidence for intramural neural circuits

The afferent output from the bladder is important for triggering micturition. This study identifies different types of afferent nerve and explores the connections of their collateral fibres on intramural ganglia and potential ganglionic targets. The experiments were performed on tissues from male guinea-pigs (n=16). Fibres positive for choline acetyl transferase (ChAT⁺) were found to originate close to the urothelium, to transit the sub-urothelial interstitial cell layer and to pass into the lamina propria. A different population of fibres, immunopositive for calcitonin gene-related peptide (CGRP), capsaicin receptors or neurofilament protein (NF), were seen to intertwine with the ChAT⁺ fibres in the lamina propria. The ChAT⁺ fibres did not express NF. Ganglia with ChAT⁺ and NF⁺ neurones were found in the lamina propria and muscle. ChAT⁺ fibres, with pronounced terminal varicosities, were present on the nerve cell bodies. Two types were noted: NF⁺ terminals and those with little or no NF (NF⁻) suggesting that their origins were the ChAT⁺ afferent collaterals and the adjacent ganglia. Fibres containing CGRP or substance P were seen on the ganglionic cells. α1B adrenergic receptors were also found on the neurones indicative of adrenergic synapses. Thus, the ganglia had multiple inputs. Different types of ChAT⁺ nerves were seen in the muscle: NF⁺ and NF⁻. The ChAT⁺/NF⁺ nerves may represent a ganglionic output to the muscle. This complex neuronal network may therefore represent the elements generating and modulating bladder sensations. The role of such a scheme in bladder pathology and the therapeutic sites of action of anticholinergic and sympathomimetic drugs are discussed.We gratefully acknowledge the support of Pfizer. This work was supported by a grant from the Detrol Research Programme.     

Selective association of nerve fibres immunoreactive for substance P or bombesin with putative cholinergic neurons of the male rat major pelvic ganglion

The male rat major pelvic ganglion contains both sympathetic and parasympathetic neurons that supply the lower urinary and digestive tracts, and the reproductive organs. The aim of this study was to describe the distribution and identify potential targets of sensory and intestinofugal axons in this ganglion. Two putative markers of these projections were chosen, substance P for primary sensory axons and bombesin for myenteric intestinofugal projections. Varicose substance P-immunoreactive axons were associated only with non-noradrenergic (putative cholinergic) somata, and most commonly with those that contained vasoactive intestinal peptide. Immunoreactivity for substance P was also present in a small group of non-noradrenergic somata, many of which were immunoreactive for enkephalins, neuropeptide Y or vasoactive intestinal peptide. Bombesin immunoreactivity was found only in preterminal and terminal (varicose) axons, the latter of which were exclusively associated with non-noradrenergic somata that contain neuropeptide Y-immunoreactivity. Some varicose axons containing either substance P-or bombesin-immunoreactivity were intermingled with clumps of small, intensely fluorescent cells. These studies indicate that substance P-and bombesin-immunoreactive axons are likely to connect with numerically small, but discrete, populations of pelvic neurons.     

The ultrastructure of paraganglia associated with the inferior mesenteric ganglia in the guinea-pig

Summary The paraganglia of the inferior mesenteric ganglia in the guinea-pig are composed of small chromaffin cells containing an abundance of granule-containing vesicles. The chromaffin cells are almost completely surrounded by satellite cells. In areas in which satellite cell processes do not intervene, the membranes of adjacent chromaffin cells are closely apposed and often form specialized attachment zones. The paraganglia contain a dense capillary network, the endothelial cells of which are often extremely attenuated and show areas of fenestration. The processes of chromaffin cells approach close to the capillary walls and are often bare of satellite cells covering on the side facing the capillary. Evidence has been obtained for the exocytotic release of the contents of chromaffin cell vesicles into pericapillary spaces. Synapses of cholinergic and noradrenergic axons are seen on the chromaffin cells. The cholinergic axons degenerate when the praganglia are decentralized, but the noradrenergic axons, which appear to arise from the local inferior mesenteric ganglia, remain intact. The results suggest that the paraganglia have an endocrine function.     

Nuclear translocation and overexpression of GAPDH by the hyper-pressure in retinal ganglion cell

To investigate the effect of hyper-pressure on retinal ganglion cells (RGC-5), RGC-5 cells were exposed to an ambient hydrostatic pressure of 100 mmHg. Upon treatment, the proliferation of RGC-5 cells was inhibited and neuronal apoptosis was detected by specific apoptosis marker TUNEL (terminal deoxynucleotidyl transferase dUTP nick-end labeling). To probe into the mechanism mediating the apoptosis of RGC-5 cells in 100 mmHg, protein profile alterations following hyper-pressure treatment were examined using two-dimensional gel electrophoresis (2-DE) followed by MALDI-TOF. Out of the 400 protein spots of RGC-5 cells detected on 2-DE gels, 37 differentially expressed protein spots were further identified using in gel tryptic digestion and mass spectrometry. Among these proteins, glyceraldehyde-3-phosphate-dehydrogenase (GAPDH) was significantly expressed 10 times more in 100 mmHg than in normal pressure. The accumulation of GAPDH in the nucleus and its translocation from the cytosol to the nucleus in 100 mmHg were observed using a microscope. These results suggest that the hyper-pressure-induced apoptosis in RGC-5 cells may be involved with not only the increase of GAPDH expression, but also the accumulation and the translocalization of GAPDH to the nucleus.     

A neural network dynamic model reproducing the output signal of the ganglion cell

A functional model of a neural network reproducing the output signal of the ganglion cell is proposed. The model assumes that receptive fields with antagonistic center and periphery are formed.     

Excitatory GABAA receptor in autonomic pelvic ganglion neurons innervating bladder

Major pelvic ganglia (MPG) are relay centers for autonomic reflexes such as micturition and penile erection. MPG innervate the urogenital system, including bladder. γ-Aminobutyric acid (GABA) is the major inhibitory neurotransmitter in the mammalian central nervous system, and may also play an important role in some peripheral autonomic ganglia, including MPG. However, the electrophysiological properties and function of GABA_A receptor in MPG neurons innervating bladder remain unknown. This study examined the electrophysiological properties and functional roles of GABA_A receptors in bladder-innervating neurons identified by retrograde Dil tracing. Neurons innervating bladder showed previously established parasympathetic properties, including small membrane capacitance, lack of T-type Ca²⁺ channel expression, and tyrosine-hydroxylase immunoreactivity. GABA_A receptors were functionally expressed in bladder innervating neurons, but GABA_C receptors were not. GABA elicited strong depolarization followed by increase of intracellular Ca²⁺ in neurons innervating bladder, supporting the hypothesis GABA may play an important role in bladder function. These results provide useful information about the autonomic function of bladder in physiological and pathological conditions.     

Patterns of co-existence of peptides and differences of nerve fibre types associated with noradrenergic and non-noradrenergic (putative cholinergic) neurons in the major pelvic ganglion of the male rat

Summary The pelvic ganglia supply cholinergic and noradrenergic nerve pathways to many organs. Other possible transmitters are also present in these nerves, including peptides. Multiple labelling immunofluorescence techniques were used in this study of the male rat major pelvic ganglion (MPG) to examine: (1) the peptides present in noradrenergic (tyrosine hydroxylase (TH)-positive) and non-noradrenergic (putative cholinergic) neurons, and (2) the types of peptide-containing nerve fibres closely associated with these two groups of neurons. The distribution of the peptide galanin (GAL) within the MPG was also investigated. All of the TH-neurons contained neuropeptide Y (NPY), but none of the other tested peptides. However, many NPY neurons did not contain TH and may have been cholinergic. TH-negative neurons also displayed vasoactive intestinal peptide (VIP), enkephalin (ENK) or GAL. VIP and NPY formed the most common types of putative cholinergic pelvic neurons, but few cells contained both peptides. Many ENK neurons exhibited VIP, NPY or GAL. Varicose nerve terminals surrounding ganglion cells contained ENK, GAL, somatostatin (SOM) and cholecystokinin (CCK). These peptide-immunoreactive fibres were more often associated with the non-noradrenergic (putative cholinergic) than the noradrenergic neurons; two types (SOM and CCK) were preferentially associated with the non-noradrenergic NPY neurons. GAL was distributed throughout the MPG, in small neurons, scattered small, intensely fluorescent (SIF) cells, and both varicose and non-varicose nerve fibres. The nerve fibres were concentrated near the pelvic and penile nerves; most of the varicose fibres formed baskets surrounding individual GAL-negative somata.     

Culture of intramural cardiac ganglia of the newborn guinea-pig

Summary The present study describes the ultrastructure of non-neuronal cells and their interrelationships with intracardiac neurones present in cultures dissociated atria and interatrial septum from newborn guinea-pig. When compared with the in situ preparation, most of these features in culture were similar to those observed in situ, but some differences were also apparent. Both mature and immature Schwann cells were observed in culture, and as in situ, the latter were closely associated with intracardiac neurones, whilst the former were more widely separated. The ultrastructure of satellite cells was more variable in culture than in situ: three general types were distinguished on the basis of their 10-nm filament content. This variation could be due to conditions of culture. Interstitial cells were present in culture and closely resembled those described in situ, although there was less space between cultured interstitial cells and their associated cells. Many fibroblasts, some myoblasts and a few mast cells were also found in the culture preparations.