Postnatal ontogenic development of the adrenergic innervation pattern in the rat portal vein

Summary The postnatal development of the adrenergic innervation pattern in the rat portal vein has been studied with the histochemical fluorescence method of Hillarp and Falck.Stretch preparations and transverse freeze-dried sections of intact portal veins were studied from rats during the first 5 weeks of life and from adult rats. Orientation of undifferentiated smooth muscle cells into two layers was observed at 4 days of age. Dominance of the thick outer longitudinal muscle layer was apparent at two weeks of age. A terminal adrenergic nerve plexus with some varicosities was restricted outside the media at the end of the first week. Ingrowth of penetrating non-terminal adrenergic nerve fibers through the longitudinal muscle layer occurred during the second week of age when the main terminal nerve plexus was developing between the two muscle layers. After 3 weeks of age the adult pattern of a two-dimensional adrenergic plexus between the muscle was established. In the adult rat pharmacological treatment with nialamide and noradrenaline revealed the thin, penetrating non-terminal adrenergic nerve fibers in the longitudinal muscle layer which were poorly visible otherwise.The present observations strongly indicate that the main adrenergic plexus between the two muscle layers emanates directly from the outer axonal plexus. These findings are discussed regarding possible trophic interactions between ingrowing sympathetic adrenergic vasomotor nerves and maturing vascular smooth muscle.Supported by grants from the Swedish Medical Research Council (grants No. 14X-2207, O4P-4173, 3884), Magn. Bergwall's Foundation, G. & M. Lindgren's Foundation, the Medical Faculty of University of Göteborg. The technical assistance of Miss Serney Bööj, Mr. Pär-Anders Larsson and Miss Ann Kjellstedt is gratefully acknowledged     

The adrenergic innervation of the pulmonary vasculature,the lung and the thoracic aorta,and on the presence of aortic bodies in the domestic fowl (Gallus gallus domesticus L.)

Summary The Falck-Hillarp technique for the localisation of biogenic amines has been used to examine the adrenergic innervation of the thoracic vasculature and lung, and to demonstrate the occurrence of aortic bodies in the domestic fowl. The proximal pulmonary vein is very densely innervated but distally the innervation becomes sparse. The pulmonary artery is sparsely innervated over its whole length. The bronchial muscle of the lung has little adrenergic innervation and fluorescent cell bodies are absent from the lung. The thoracic aorta receives a moderate adrenergic innervation. In the region of the aortic arch and pulmonary arteries groups of fluorescent cells are common. Extramedullary chromaffin cells and small, intensely fluorescent cells occur within these groups. In the media of the aorta and pulmonary artery other types of fluorescent cells are found. These results are discussed in the light of previous observations.Part of this work was performed while the author was a postdoctoral research fellow of the National Heart Foundation of Australia. His thanks are due to Prof. G. Burnstock for use of laboratory facilities.     

Ovarian innervation develops before initiation of folliculogenesis in the rat

Summary Sympathetic neurotransmitters have been shown to be present in the ovary of the rat during early postnatal development and to affect steroidogenesis before the ovary becomes responsive to gonadotropins, and before the first primordial follicles are formed. This study was undertaken to determine if development of the ovarian innervation is an event that antedates the initiation of folliculogenesis in the rat, Rattus norvegicus. Serial sections of postnatal ovaries revealed a negligible frequency of follicles 24 h after birth (about 1 primordial follicle per ovary). Twelve hours later there were about 500 follicles per ovary, a number that more than doubled to about 1300 during the subsequent 12 h, indicating that an explosive period of follicular differentiation occurs between the end of postnatal days 1 and 2. Electron microscopy demonstrated that before birth the ovaries are already innervated by fibers containing clear and dense-core vesicles. Immunohistochemistry performed on either fetal (day 19) or newborn (less than 15h after birth) ovaries showed the presence of catecholaminergic nerves, identified by their content of immunoreactive tyrosine hydroxylase (TH), the rate-limiting enzyme in catecholamine biosynthesis. While some of these fibers innervate blood vessels, others are associated with primordial ovarian cells, thereby suggesting their participation in non-vascular functions. Since prefollicular ovaries are insensitive to gonadotropins, the results suggest that the developing ovary becomes subjected to direct neurogenic influences before it acquires responsiveness to gonadotropins.     

Fine structure and fluorescence morphology of adrenergic nerves after 6-hydroxydopamine in vivo and in vitro

Summary In parallel fine structural, fluorescence histochemical and biochemical experiments the effect of 6-OH-DA administered in vivo and in vitro on the adrenergic nerves in the mouse iris was studied. As seen in the electron microscope, in vivo administration of 6-OH-DA causes a selective, rapid degeneration of the adrenergic axon terminals similar to that found after axotomy, whereas the cholinergic nerves are unaffected at all time intervals studied. Already 1 hr after the injection of 6-OH-DA the axonal enlargements swell and the size of the dense core of the granular vesicles is strongly reduced. Since the NA stores are almost completely depleted at this time interval, the small core present may be due to a reaction between 6-OH-DA and the fixative. From 2–4 hr after the injection increasing numbers of axonal enlargements with a high electron density are observed in the Schwann cell cytoplasm, which later are digested and completely absent about 48–72 hr after the 6-OH-DA injection. During the following weeks adrenergic axons reappear. This time course of degeneration obtained is considerably faster than that seen after axotomy in other studies. After incubation in 6-OH-DA containing media similar changes were observed in the axonal enlargements, starting already after 30 min of incubation. At this time-point there is a considerable reduction of endogenous NA and a severe damage of the membrane pump uptake mechanism. Incubation with 6-OH-DA and subsequent rinsing for 2 hr caused marked changes, including partly swelling of axons and partly shrinking of the axons into electron dense bodies.The fluorescence histochemical and biochemical results are in good agreement with the ultrastructural studies demonstrating a rapid loss of NA from the adrenergic nerve terminals and main axons and a long lasting depletion of the NA, with a gradual recovery to 75% 6 weeks after the injection.The investigation has been supported by research grants from the Swedish Medical Research Council (14X-2295, 14X-2887 and 04X-3881) Karolinska Institutet, Magnus Bergvalls and Carl-Berthel Nathorst Stiftelser. For generous supplies of drugs we are indebted to the following companies: AB Hässle (6-OH-DA, through Dr H. Corrodi), Pfizer (Niamid^®), Ciba (Serpasil^®). The skilful technical assistance of Miss Bodil Flock, Mrs Waltraut Hiort and Mrs Eva Lindqvist is gratefully acknowledged.     

Adrenergic innervation of pancreatic islets and modulation of insulin secretion by the sympatho-adrenal system

Summary Morphological changes in the adrenergic innervation of pancreatic islets after chemical sympathectomy by use of 6-hydroxydopamine and the influence of the sympatho-adrenal system on insulin secretion were investigated in the mouse and rat.Fluorescence histochemistry revealed a clear-cut reduction in the number of adrenergic nerve fibers in the pancreatic islets 2 days after administration of 6-hydroxydopamine; the reduction was more pronounced in the rat than in the mouse. In the rat, a partial regeneration was seen after 6 weeks. In the pancreas of the mouse, after administration of 6-hydroxydopamine, a severe damage of unmyelinated nerve fibers was revealed electron microscopically. However, no ultrastructural or immunohistochemical alterations could be demonstrated in the endocrine cells of the islets.6-Hydroxydopamine induced a depression of basal plasma insulin concentrations in mice and an elevation in rats. Adrenalectomy depressed basal plasma insulin levels in mice.The -adrenoceptor antagonist phentolamine enhanced insulin secretion in normal mice. The secretory response of insulin to phentolamine was diminished by chemical sympathectomy and almost abolished by adrenalectomy or the combination of chemical sympathectomy and adrenalectomy. Thus, the effect of phentolamine is probably mediated by liberated catecholamines.It is concluded that basal insulin secretion is partially regulated by the sympatho-adrenal system and that species differences exist in this respect. In addition, the results suggest that endogenous catecholamines have the ability to promote insulin secretion.     

Evidence for exclusive adrenergic innervation of feather muscles (mm. pennati) in the chicken

Summary Feather follicles in the avian skin are interconnected by well-defined bundles of smooth muscle cells, which are responsible for the erection and depression of feathers and thus play an important role in thermoregulation. The depressing and erecting muscle bundles were found to receive a very dense supply of unmyelinated nerve fibres that displayed ultrastructural and histochemical characteristics of noradrenergic axons (formaldehyde- and glyoxylic acid-induced catecholamine fluorescence; uptake to 5-hydroxydopamine). No nerve fibres were encountered showing histochemical acetylcholinesterase activity. There was no indication of the presence of peptidergic or purinergic nerve endings.The neuromuscular space usually ranged from 40–60 nm in width and contained a basal lamina. Occasionally, this space was reduced to approximately 20 nm. At such close neuromuscular contacts a basal lamina was lacking, and focal densities beneath the pre- and postsynaptic plasma membrane were observed. Since no gap junctions between muscle cells were detected, the dense supply with noradrenergic nerve fibres indicates a high amount of directly innervated smooth muscle cells.An additional finding of the present study was the observation that high local concentrations of 5-hydroxydopamine led to degeneration of noradrenergic nerve endings.Supported by a grant from the Deutsche Forschungsgemeinschaft (Dr. 91)     

Ultrastructure and innervation of the swimbladder of Tetractenos glaber (Tetraodontidae)

Summary The general structure, ultrastructure and innervation of the swimbladder of the smooth toadfish, Tetractenos glaber, were examined with light-microscopic, fluorescence-histochemical, and transmission electron-microscopic techniques. The structure of the swimbladder is similar to that of other euphysoclists. Fluorescence histochemistry showed adrenergic fibres in both the secretory and resorptive areas of the swimbladder. Transmission electron microscopy revealed two morphologically distinct axon profiles type-I profiles containing many small, flattened vesicles; type-II profiles containing both large, granular vesicles and rounded, small clear vesicles in varying proportions.The gas-gland cells and surrounding muscularis mucosae are innervated by both type-I and type-II fibres. Type-I fibres also innervate pre-rete arteries. The rete- and gas-gland capillaries do not appear to be innervated. Arteries running to the resorptive area are innervated by type-I fibres. Both type-I and type-II profiles make contact with the muscularis mucosae in the resorptive area. Only type-I fibres innervate the radial dilator muscle in the oval sphincter region, whereas only type II fibres innervate the circular muscle of the oval sphincter.Type-I fibres took up -methyl-noradrenaline, and could not be found after pre-treatment with 6-hydroxydopamine. They are, therefore, assumed to be adrenergic. Type-II fibres were tentatively identified, by exclusion, as cholinergic.     

Fine structure and peroxidatic activity of rat blood monocytes

Summary Transmission and scanning electron microscopy of the rat epithalamus shows a regional variation in the distribution of Supraependymal nerves (SN) which correlates well with Supraependymal yellow fluorescence reported by Richards et al. (1974). The medial habenular nucleus, the intercommissural and suprahabenular recesses, the habenular commissure and the fibrae periventriculares thalami have the greatest density of SN/100 m of ependymal surface. The floor of the suprahabenular and intercommissural recesses is covered by non-ciliated ependyma.The significance of these findings is discussed with respect to (1) a direct functional relationship of SN with ependyma, and (2) a possible participation of the non-ciliated ependyma of the suprahabenular and intercommissural recesses in secretory activity whereby the CSF serves as a vehicle for neuroendocrine communication.The author thanks Drs. Sanford Palay, Milton Brightman, and Constantino Sotelo for their advice in the preparation of nervous tissue for electron microscopy; Dr. Gunter Bahr for the use of scanning electron microscopy facilities; and Messrs. Cyril Wingfield and Walter Engler and Mrs. Michie Vane for their technical support     

Intraepithelial blood vessels in the vomeronasal neuroepithelium of the rat

Summary Epithelial-vascular relationships are established during the development of the vomeronasal neuroepithelium of the rat. Special attention is given to the fine structure of the endothelial wall of intra-epithelial vessels, to ultrastructural aspects of the neuronal-vascular relationships, and to the appearance of inclusion bodies in the neuronal cells adjacent to these vessels. The neuronal perikarya surrounding the blood vessels are filled with highly developed smooth endoplasmic reticulum. Possible functional implications of the vascularization of the neuroepithelium of the vomeronasal organ in mediating olfacto-endocrine relationships are discussed. It is suggested that the intra-epithelial blood vessels are at least supportive and nutritive in nature, while their implication in an olfacto-endocrine connection remains obscure.On sabattical leave: Visiting Professor, Department of Anatomy, University of Kentucky, Lexington, Ky., USA     

Adrenergic innervation of the umbilical vessels

Summary The intra- and extrafetal portions of the umbilical vessels in the guinea-pig and the umbilical cord of man, mouse and rabbit have been investigated by means of the Falck-Hillarp method for the fluorescence microscopical demonstration of catecholamines.The umbilical cord was found to be devoid of nerves in all species investigated. Adrenergic nerves are present only in the immediate vicinity of the umbilicus.The intrafetal portions of the umbilical artery and umbilical vein receive adrenergic nerves, the distribution pattern of which is different for each vessel. In the guinea-pig the ductus venosus is an intrahepatic branch of the vena umbilicalis. No adrenergically innervated sphincter has been detected in the initial segment of the ductus venosus. Regional variations in the pattern of innervation of the intrafetal portion of the umbilical vein are paralleled by regional differences in the construction pattern of the vessel's wall. Regional differences in the noradrenaline concentration (measured by fluorometry) which correspond to the fluorescence microscopical findings have been detected in umbilical vessels: low noradrenaline content of the umbilical cord, high concentrations in the intrafetal sections of the umbilical vessels. The noradrenaline concentration of the guinea-pig umbilical artery is three times that of the umbilical vein.Supported by the Joachim Jungius-Gesellschaft der Wissenschaften, Hamburg.For continuous advice and constructive criticism I am indebted to Prof. Dr. Dr. E. Horstmann.     

The tanycyte of the rat median eminence

Summary The present ultrastructural study proves the existence of nerve terminals closely apposed to the plasmalemmata of tanycytes in the rat median eminence. Several of these axo-tanycytic endings display remarkable accumulations of agranular endoplasmic reticulum in the form of pleomorphic vesicles which are closely apposed on either side of the plasma membrane of each cell compartment. Some of these vesicular profiles give the impression of structural continuity across both membrane systems. This phenomenon is discussed in the context of being a potential substratum for communication between both cell compartments.Supported by NIH Grant NS13717 and NSF Grant BNS 78-11820     

The ultrastructure of the accessory sex organs of the male rat

Summary The fine structure of the nuclei of epithelial cells of the dorsal lobe of the rat prostate were studied 2, 3, 5, 7 and 21 days after castration. The nucleolus appears to undergo a progressive disorganisation with partial fragmentation and dispersion of its normal components.Changes in the nucleoplasm were primarily reflected by a condensation of chromatin, particularly along the nuclear membrane and adjacent to the nucleolus. Later, different types of intranuclear inclusions were observed.After 21 days, the nuclei were characterized by an irregular outline with large indentation. Within the nucleoplasm aggregates of coarse granular chromatin were found. No cell necrosis was observed, indicating that androgen deprivation results in a remodeling of the cell to a less active state with marked cellular alterations and cessation of secretion, but apparently with some of their basic functions still intact.Injections of testosterone completely reverse the castrated-induced alterations.The changes observed are assumed to be due to the withdrawal of the androgenic stimulus, with a direct influence on the secretory function of the cell. The findings support the view that the stimulating secretory effect of androgen is mediated via an intranuclear androgen receptor, probably located in the nucleolus-associated-chromatin. It is also proposed that the secretory function of the epithelial cells of the prostatic complex, initiated by androgens, may be regulated by an intranuclear secretory center.     

Studies on the juxtaglomerular apparatus VI. Sympathetic innervation,catecholamines and the renin-angiotensin-system in rats and tree-shrews (Tupaia belangeri)

Summary It has previously been reported that the primitive primate Tupaia belangeri develops a renal failure when exposed to psychosocial stress. In order to learn if this high susceptibility to stress of the Tupaia kidney can be correlated with morphological and functional parameters of the Juxtaglomerular apparatus (JGA) and the renin-angiotensin system, comparative experiments were performed on Tupaia and rat.Our results reveal an outstandingly high potency of the JGA and the renin-angiotensin system in Tupaia as evident from the following findings: The Tupaia JGA contains a great number of epithelioid cells abounding in renin granules (electron microscopy). The renin content of the Tupaia kidney is considerably higher than in the rat (radio-immunoassay). The sympathetic innervation of the kidney and especially of the JGA is abundant in Tupaia (fluorescence and electron microscopy). Catecholamine contents of the kidney and other organs are significantly higher in Tupaia than in rats (spectrophotofluorometry).Our results support the previously developed concept of a potent intrarenal neuroendocrine interaction at the JGA level favouring, under certain conditions of social stress, the development of acute renal failure in Tupaia belangeri.The studies were supported by the German Research Foundation within the SFB90 Cardiovasculäres System