A pathological study of the tongues of rabid dogs in the Philippines.

During rabies virus infections, the minor salivary glands are one of the important organs for virus replication and excretion into the oral cavity. However, details of pathological findings and viral antigen distribution in the minor salivary glands remain poorly understood. In this study, we conducted pathological tests on the tongues of 71 rabid dogs in the Philippines; the minor salivary glands (von Ebner's glands, lingual glands), circumvallate papilla, autonomic ganglia, and skeletal muscles were evaluated. Inflammatory changes were observed in the von Ebner's glands of 20/71 dogs, in the circumvallate papilla of 10/71, and in the tongue muscle of 1/71. Conversely, no morphological changes were observed in the lingual glands and autonomic ganglia. Viral antigens were detected via immunohistochemistry-based methods in the cytoplasm of the acinar epithelium in the von Ebner's glands of all 71 dogs. Virus particles were confirmed in the intercellular canaliculi and acinar lumen via electron microscopy. In the autonomic ganglia, viral antigens were detected in 67/71 rabid dogs. Viral antigens were detected in the taste buds of all 71 dogs, and were distributed mainly in type II and III taste bud cells. In tongue muscle fibers, viral antigens were detected in 11/71 dogs. No virus antigens were detected in lingual glands. These findings suggest that rabies virus descends in the tongue along the glossopharyngeal nerve after proliferation in the brain, and von Ebner's glands and taste buds are one of the portals of virus excretion into the saliva in rabid dogs.

Balanced Caloric Restriction Minimizes Changes Caused by Aging on the Colonic Myenteric Plexus.

Aging can promote significant morphofunctional changes in the gastrointestinal tract (GIT). Regulation of GIT motility is mainly controlled by the myenteric neurons of the enteric nervous system. Actions that aim at decreasing the aging effects in the GIT include those related to diet, with caloric restriction (CR). The CR is achieved by controlling the amount of food or by manipulating the components of the diet. Therefore, the objective of this study was to evaluate different levels of CR on the plasticity of nicotinamide adenine dinucleotide phosphate- (NADPH-) reactive myenteric neurons in the colon of Wistar rats during the aging process using ultrastructural (transmission electron microscopy) and morphoquantitative analysis. Wistar male rats (Rattus norvegicus) were distributed into 4 groups (n = 10/group): C, 6-month-old animals; SR, 18-month-old animals fed a normal diet; CRI, 18-month-old animals fed a 12% CR diet; CRII, 18-month-old animals fed a 31% CR diet. At 6 months of age, animals were transferred to the laboratory animal facility, where they remained until 18 months of age. Animals of the CRI and CRII groups were submitted to CR for 6 months. In the ultrastructural analysis, a disorganization of the periganglionar matrix with the aging was observed, and this characteristic was not observed in the animals that received hypocaloric diet. It was observed that the restriction of 12.5% and 31% of calories in the diet minimized the increase in density and cell profile of the reactive NADPH neurons, increased with age. This type of diet may be adapted against gastrointestinal disturbances that commonly affect aging individuals.

Significance of Cholinergic and Peptidergic Nerves in Stress-Induced Ulcer and MALT Lymphoma Formation.

Backgound: The role of enteric nerves has previously been demonstrated in the formation of several gastric diseases. In the present review, the significance of the cholinergic nerves in stress-induced ulcer formation as well as the importance of substance P in the formation of gastric MALT lymphoma is discussed. METHODS: The stress-induced ulcer was induced by the plaster bandage methods in rats. The gastric MALT lymphoma was formed by the peroral infection of gastric mucosal homogenate of the infected mouse in C57BL/6 mice. For the stress-induced ulcer, the distribution of the cholinergic nerves and muscarinic acetylcholine receptors was investigated by acetylcholinesterase histochemistry and autoradiography of water soluble compounds using 3H-quinuclidinyl benzilate was performed. To the MALT lymphoma study, the distribution of the substance P and effect of substance P antagonist, spantide II, was investigated by immunohistochemical studies. RESULTS: The stress induced ulcer formation was shown to be related to the hyperactivity of the cholinergic nerves. The gastric MALT lymphoma was shown to be related to the increased localization of substance P. CONCLUSION: Stress-induced ulceration as a model of hyperactivity of the cholinergic nerves was proved to be a useful approach, while substance P and its role in MALT lymphoma formation may serve as a tool to clarify the neuroimmune modulation of chronic infectious diseases.

[Acute Sensory Neuropathies and Acute Autonomic Neuropathies].

From the perspective of neuropathies with an acute onset mimicking that of Guillain-Barré syndrome (GBS), cases with profound sensory and/or autonomic impairment without any significant weakness have been reported. Although the possibility of infectious or toxic etiologies should be carefully excluded, immune mechanisms similar to those in GBS are suggested to be involved in these so-called acute sensory neuropathies and acute autonomic neuropathies. The types of neuropathy include those with predominant sensory manifestations, predominant autonomic manifestations such as autoimmune autonomic ganglionopathy, and both sensory and autonomic manifestations such as acute autonomic and sensory neuropathy. Neuronopathy in the sensory and/or autonomic ganglia (i.e., ganglionopathy) has been commonly suggested in patients with these types of neuropathies. The presence of Anti-GD1b antibodies has been reported in some of the patients with acute sensory neuropathy with deep sensory impairment, whereas anti-ganglionic acetylcholine receptor antibodies are reported to be present in half of the patients with autoimmune autonomic ganglionopathy. The discovery of anti-ganglionic acetylcholine receptor antibodies significantly expanded the spectrum of autoimmune autonomic ganglionopathy. This is because some of the patients with chronic progression mimicking neurodegenerative diseases such as pure autonomic failure were positive for these antibodies. In contrast, pathologically significant autoantibodies have not been identified in acute autonomic and sensory neuropathy. Further studies are needed to clarify the pathogenesis and the spectrum of these types of neuropathies.

[CHANGES IN THE PANCREATIC ISLETS AND NERVOUS ELEMENTS IN RATS DURING AGING (AN IMMUNOHISTOCHEMICAL STUDY)].

The neural apparatus and the endocrine part of the pancreas was studied in Wistar rats aged 3-4 and 19 months (n = 24) using the immunohistochemical reactions for synaptophysin (Syn), tyrosine hydroxylase (TH) and protein gene product 9.5 (PGP 9.5). Since Syn and PGP 9.5 are highly selective in detection of pancreatic islet (PI) endocrinocytes, it was possible to examine their topography and density in all parts of the pancreas. It was found that in rats aged 19 months, the total number of PI was decreased as compared to that in young animals. The study of PI size distribution has shown that the number of large islets decreased with age. Young animals showed rich innervation of the pancreas which was represented by three nerve plexuses: the first was a broadly-looped one, formed by small nerve trunks and bundles of unmyelinated and myelinated nerve fibers, the second consisted of thin bundles of postganglionic axons and microganglia, and the third (main terminal plexus) was formed by axons with varicosities and synapses of "en passant" type. In aged rats, marked degenerative changes in the neurons of intramural ganglia, nerve trunks and bundles were noted together with the reduction or complete absence of Syn- and TH-positive efferent parasympathetic and sympathetic terminals around blood vessels, excretory ducts, denervation of the exocrine and endocrine parts of the pancreas. Innervation disturbances in some lobules were accompanied by small inflammatory perivascular infiltrates.

Long-term potentiation in mammalian autonomic ganglia: an inclusive proposal of a calcium-dependent, trans-synaptic process.

Ganglionic synapses have the capability to express long-term potentiation (gLTP) after application of a brief high-frequency stimulus. It has been suggested a possible role of gLTP in some cardiovascular diseases. Although a number of characteristics of gLTP have been described, the precise locations and mechanisms underlying gLTP are not completely known. Current findings support two major conflicting presynaptic and postsynaptic hypotheses. The presynaptic hypothesis posits a presynaptic increase in acetylcholine (ACh) release, whereas the postsynaptic hypothesis proposes a long-lasting enhancement of the nicotinic response on the postsynaptic membrane. An alternative trans-synaptic hypothesis proposes the presynaptic release of a cotransmitter from large dense core vesicles, which postsynaptically enhances synaptic efficacy and accounts for gLTP. Here, we review the studies of LTP, with emphasis on gLTP in mammals, and we examine the findings that support the presynaptic, the postsynaptic and the trans-synaptic hypotheses. We then review our data on the contribution of calcium to gLTP as an approach to elucidate the mechanisms of gLTP. Data on the contribution of calcium to gLTP and on prolonged high-frequency stimulus-dependent fading of LTP have led us to support the trans-synaptic process as responsible for gLTP. Finally, we present a formal working model for the mechanisms of gLTP.

Ganglionar nervous cells and telocytes in the pancreas of Octodon degus: extra and intrapancreatic ganglionar cells and telocytes in the degus.

This study shows for the first time the presence of intra and extrapancreatic ganglionar neurons and telocytes in Octodon degus such as those described in human and guinea pig pancreas. Pancreatic ganglionar neurons were identified by their histological characteristics as well as their positive immunostaining with mouse anti-human neuron specific enolase (NSE) antibody. Somatostatin secreting delta cells (D cells) in the islets of Langerhans were identified by positive immunostaining with rabbit antihuman polyclonal somatostatin antibody. Electron microscopy evidenced the presence of some unmyelinated axons in the interlobular spaces or septa, usually located adjacent to blood vessels and the exocrine epithelial ducts. The presence of telocytes with at least 2 telopodes was observed in the interlobular space, frequently in close spatial relationship with blood vessels and nerve endings. Telocytes were often observed in the vicinity or even in close proximity with both secretory acini and exocrine epithelial ducts and regulatory nerves and blood vessel apparatuses. A possible framework has been put forward within which such structures might contribute to elicit physiological responses in the pancreas. Further studies of synaptic interactions within and between pancreatic neuron cells are needed to help clarify the morphological results reported here. A broad overview of the field of neurogastroenterology with focus on the pancreas of O. degus related to the enteric nervous system (ENS) is provided in order to help design future studies on the connections of specific neurons forming pancreatic pathways, their neurotransmission processes and how disruption of these pathways may contribute to pancreatic disease.

Diagnosis of mitochondrial neurogastrointestinal encephalopathy disease in gastrointestinal biopsies.

A 14-year-old boy with mitochondrial neurogastrointestinal encephalopathy (MNGIE) disease had a lifelong history of failure to thrive and gastrointestinal symptoms including vomiting, pain, and diarrhea, leading to progressive cachexia. At the age of 9 years, after an extensive workup, the diagnosis of Crohn disease was strongly suspected, and he underwent colonoscopy with multiple biopsies. At 11 years of age, vision change and poor balance lead to a diagnosis of leukodystrophy by magnetic resonance imaging. Investigations for metachromatic leukodystrophy, adrenal leukodystrophy, and globoid cell leukodystrophy were all negative. A diagnosis of MNGIE disease was suspected when he continued deteriorating with gastrointestinal symptoms, multiple neurologic deficits, and encephalopathy. Markedly diminished thymidine phosphorylase activity and increased thymidine plasma levels confirmed the diagnosis of MNGIE. At autopsy, megamitochondria were observed by light microscopy in submucosal and myenteric ganglion cells and in smooth muscle cells of muscularis mucosae and muscularis propria, along the entire gastrointestinal tract from the esophagus to the rectum. Megamitochondria in ganglion cells were also observed in a retrospective review of the endoscopic intestinal biopsies taken at age 9 and 13 years and in the appendectomy specimen obtained 1 month before his demise. This study corroborates the presence of megamitochondria in gastrointestinal ganglion cells in MNGIE disease, better illustrates their detailed morphology, and describes for the first time similar structures in the cytoplasm of gastrointestinal smooth muscle cells. Pathologists should be able to recognize these structures by light microscopy and be aware of their association with primary mitochondriopathies.

Neurochemical characterization of pterygopalatine ganglion branches in humans.

BACKGROUND: Pterygopalatine ganglion (PPG) branches, seem to be involved in the pathophysiology of facial pain. The functions of these branches, including a recently discovered orbital branch, are not completely known but could be of clinical significance. This study was designed to characterize PPG branches through immunohistochemical stain and study their anatomy, specifically the orbital branches. METHODS: In a cadaver study of four specimens, the pterygopalatine fossa (PPF) was dissected out of its bony surroundings as a tissue block. Subsequently, cryostat sectioning of these blocks was performed. In one specimen the PPF was microscopically dissected. Recently discovered neural structures were identified, dissected out of the tissue block, and cryosectioned. All cryostat sectionings were immunohistochemically stained for protein gene product (PGP) 9.5, nitric oxide synthase (NOS), and tyrosine hydroxylase (TH). RESULTS: A recently discovered neural connection between the PPG and the ophthalmic nerve could be confirmed and classified as an orbital PPG branch. The connection stained throughout for PGP 9.5 and partially stained for NOS. In other orbital branches, both NOS and TH(+) nerve fibers were found. The PPG contained NOS(+) cells. TH labeling was also found in nerve fibers running through the PPG and the vidian nerve. CONCLUSION: The recently discovered orbital PPG branch is of a mixed parasympathetic and sensory nature. In the other orbital branches, sympathetic fibers were shown as well. This knowledge may add to understanding the symptomatology and therapies of headache syndromes such as nerve block.

[Distribution and structural organization of autonomic neural apparatus of the rat pancreas (an immunohistochemical study)].

Using the immunohistochemical methods for detection of neurofilaments (NF), peripherin (PRF), synaptophysin (SF), tyrosine hydroxylase (TH) and Nissl-staining with toluidine blue, thick sections (10-20 microm) prepared through all parts of pancreas of adult Wistar rats (n=7) were studied. The topography and the density of distribution of pancreatic islets in the various parts of pancreas were defined. The greatest density of the islets was found in the body of pancreas. The dense innervation of the organ was detected which included several nervous plexuses: the big-looped one consisting of nervous bundles and small trunks of NF-positive myelinated and unmyelinated nervous fibers, the second one was formed by PRF-positive thin posganglionic bundles of axons and microganglia, and the third one, detected with SF-staining, was main terminal plexus consisting of varicose axons with en passant synapses. Interactions of synaptophysin-positive terminals (distant en passant synapses) with blood vessels, endocrine (islet) and exocrine cells, excretory ducts of the pancreatic lobules are described in details. Peculiarities of the structure of parasympathetic ganglia, their neurons, and pericellular synaptic apparatus are described; problems of the innervation and the nature of pancreatic insular endocrine cells are discussed. Attention is drawn to the fact that in no case neurons were found in the islets in the rat pancreas.

Morphoquantitative study of the submucous plexus (of Meissner) of the jejunum-ileum of young and old guinea pigs.

OBJECTIVE: To study the aging of submucous plexus of the small intestine (jejunum-ileum) of the guinea pigs from the quantitative, structural and ultrastructural perspective. METHOD: Chemical preparations of membrane of the jejunum-ileum of old and young animals with the use of light and electronic microscope. RESULTS: The ganglia of young animals presented between 1 and 56 neurons and the old animals presented from 1 to 30 neurons. The mean density of the ganglia by cm(2) in the young jejunum-ileum was of 551±36.89 and in the old one 413±11.86. The density of the neurons was 5011±291.11 neurons/cm(2) average in young animals and 2918±120.70 neurons/cm(2) in the old ones. The size of the neurons varied in both age groups. The collagen fibers in the ganglia of old animals they were condensed. Degenerated mitochondrias in the interior of the cell were frequent in the old animals. CONCLUSION: In submucous plexus of the jejunum-ileum there is a loss of 38% of the neurons with aging.

Morphoquantitative study of the submucous plexus (of Meissner) of the jejunum-ileum of young and old guinea pigs / Estudo morfoquantitativo do plexo submucoso (de Meissner) do jejuno-íleo de cobaias jovens e velhas

OBJECTIVE: To study the aging of submucous plexus of the small intestine (jejunum-ileum) of the guinea pigs from the quantitative, structural and ultrastructural perspective. METHOD: Chemical preparations of membrane of the jejunum-ileum of old and young animals with the use of light and electronic microscope. RESULTS: The ganglia of young animals presented between 1 and 56 neurons and the old animals presented from 1 to 30 neurons. The mean density of the ganglia by cm² in the young jejunum-ileum was of 551±36.89 and in the old one 413±11.86. The density of the neurons was 5011±291.11 neurons/cm² average in young animals and 2918±120.70 neurons/cm² in the old ones. The size of the neurons varied in both age groups. The collagen fibers in the ganglia of old animals they were condensed. Degenerated mitochondrias in the interior of the cell were frequent in the old animals. CONCLUSION: In submucous plexus of the jejunum-ileum there is a loss of 38 percent of the neurons with aging.

OBJETIVO: Estudar o envelhecimento do plexo submucoso do intestino delgado (jejuno-íleo) das cobaias do ponto de vista quantitativo, estrutural e ultra-estrutural. MÉTODO: Preparados de membrana do jejuno-íleo de animais jovens e velhos com a utilização de microscopia de luz e eletrônica. RESULTADOS: Os gânglios de animais jovens apresentaram entre 1 e 56 neurônios e os animais velhos apresentaram de 1 a 30 neurônios. A densidade média dos gânglios por cm² no jejuno-íleo jovem foi de 551±36,89 e no velho foi de 413±11,86. A densidade dos neurônios foi de 5011±291,11 neurônios/cm² em média nos animais jovens e 2918±120,70 neurônios/cm² nos velhos. O tamanho dos neurônios variou em ambos os grupos etários. As fibras colágenas nos gânglios de animais velhos estavam mais condensadas. Mitocôndrias degeneradas no interior da célula foram freqüentes nos animais velhos. CONCLUSÃO: No plexo submucoso do jejuno-íleo há uma perda de 38 por cento dos neurônios com o envelhecimento.

Syncytial cytoplasmic anastomoses between neurites in caudal mesenteric ganglion cells in adult cats.

The fact that most published data on syncytial cytoplasmic anastomoses are based on the autonomic nervous system in the early postnatal period of development, when many nerve fibers are poorly ensheathed by glia or have no glial sheaths at all, has led to the assumption that these anastomoses do not exist in adults because of the significant development of the glia and glial insulation of individual neurites from each other. We tested this assumption using electron microscopic studies of the caudal mesenteric ganglion in adult cats. A high level of glial ensheathing of neurites was observed. However, syncytial pores were seen between contacting neurites lacking glial sheaths in almost every specimen. This is the first report describing axodendritic synapses with perforations in the presynaptic zone outside the synaptic specializations in the autonomic nervous system. It is suggested that although syncytial cytoplasmic connections are seen in adult animals, they do not contradict the neuron theory.

[Syncytial cytoplasmic anastomoses between the neurites of caudal mesenteric ganglion cells in adult cats].

Since the majority of the published data on syncytial cytoplasmic anastomoses relate to the autonomic nervous system in the early postnatal period of development, when many nerve fibers are still poorly covered by glia or have no glial sheaths at all, it was suggested that such anastomoses were not present in adults due to the significant development of glia separating individual neurites from each other. To check this assumption, we have performed an electron microscopic study of the adult cat dorsal caudal mesenteric ganglion. The cell neurites were found to be frequently covered by glial sheaths. However, almost in every sample, the syncytial pores were detected between the contacting neurites lacking glial covering layers. Sometimes serial syncytially connected neurites were seen. Axo-dendritic synapses with presynaptic perforations outside the synaptic specializations were described in the autonomic nervous system for the first time. These observations therefore provide evidence of syncytial cytoplasm connections in normal adult animals, however this does not reject the neuronal doctrine.

The problem of neuronal syncytical connection in disease.

The membranes of neuron profiles adjacent to each other in the caudal mesenteric ganglion were examined electron microscopically during the first 3 days after crossing of the preganglionar or postganglionar branches. The contacting membranes were thinned and perforated, and neuron-to-neuron syncytial connections were forming. Multiple connections were formed between the nerve processes and terminals in the synapse. The pre- and even postsynapses were clear in pronounced perforations.

Silent synapses sit and wait for a better day.

Synaptic activity is thought to be critical for synaptic stabilization. In this issue of Neuron, Krishnaswamy and Cooper show that nicotinic synapses on autonomic neurons remain intact without synaptic activity. Postsynaptic responses are required, however, for presynaptic terminals to acquire the high-affinity choline transporter necessary for high-frequency transmission.

Peripheral nervous system pathology in fragile X tremor/ataxia syndrome (FXTAS).

Fragile X tremor/ataxia syndrome (FXTAS) occurs in individuals with moderate CGG expansion of the fragile X mental retardation 1 (FMR1) gene and is associated with intranuclear inclusions in neurons and astrocytes. Although the neuropathologic findings in the brain and spinal cord were described, pathological features in the peripheral nervous system were not reported. Here, we report on novel neuropathological findings in the peripheral nervous system and especially in autonomic ganglia at autopsy in a man with FXTAS. In addition to the characteristic brain and spinal cord findings, typical intranuclear inclusions were identified in the ganglion cells of adrenal medulla, dorsal root ganglia, paraspinal sympathetic ganglia, myenteric ganglia of the stomach and subepicardial autonomic ganglion of the heart. Our findings indicate that FXTAS diffusely involves the central and peripheral nervous systems, which explains the protean neurological symptoms ranging from dementia to dysautonomia.

Structure of peripheral synapses: autonomic ganglia.

Final motor neurons in sympathetic and parasympathetic ganglia receive synaptic inputs from preganglionic neurons. Quantitative ultrastructural analyses have shown that the spatial distribution of these synapses is mostly sparse and random. Typically, only about 1%-2% of the neuronal surface is covered with synapses, with the rest of the neuronal surface being closely enclosed by Schwann cell processes. The number of synaptic inputs is correlated with the dendritic complexity of the target neuron, and the total number of synaptic contacts is related to the surface area of the post-synaptic neuron. Overall, most neurons receive fewer than 150 synaptic contacts, with individual preganglionic inputs providing between 10 and 50 synaptic contacts. This variation is probably one determinant of synaptic strength in autonomic ganglia. Many neurons in prevertebral sympathetic ganglia receive additional convergent synaptic inputs from intestinofugal neurons located in the enteric plexuses. The neurons support these additional inputs via larger dendritic arborisations together with a higher overall synaptic density. There is considerable neurochemical heterogeneity in presynaptic boutons. Some synapses apparently lack most of the proteins normally required for fast transmitter release and probably do not take part in conventional ganglionic transmission. Furthermore, most preganglionic boutons in the ganglionic neuropil do not form direct synaptic contacts with any neurons. Nevertheless, these boutons may well contribute to slow transmission processes that need not require conventional synaptic structures.

Exocytosis unbound.

The accepted theory of vesicular release of neurotransmitter posits that only a single vesicle per synapse can fuse with the membrane following action potential invasion, and this exocytotic event is limited to the ultrastructurally defined presynaptic active zone. Neither of these dictums is universally true. At certain synapses, more than a single vesicle can be released per action potential, and there is growing evidence that neuronal exocytosis can occur from sites that are unremarkable in electron micrographs. The first discrepancy extends the dynamic range of synapses, whereas the second enables faster and more robust chemical transmission at sites distant from morphologically defined synapses. Taken together, these attributes expand the capabilities of cellular communication in the nervous system.

Number and distribution of intraganglionic laminar endings in the mouse esophagus as demonstrated with two different immunohistochemical markers.

Intraganglionic laminar endings (IGLEs) represent the only vagal mechanosensory terminals in the tunica muscularis of the esophagus. Two specific markers for IGLEs were recently described in mouse: the purinergic P2 x 2 receptor and the vesicular glutamate transporter 2 (VGLUT2). This study aimed at comparing both markers with respect to their suitability for quantitative analysis. We counted IGLEs immunostained for VGLUT2 and P2 x 2, respectively, and mapped their distribution in esophageal wholemounts of C57Bl/6 mice. Numbers and distribution of IGLEs were compared with those of myenteric ganglia as demonstrated by cuprolinic blue histochemistry. Whereas the distribution of VGLUT2-immunopositive IGLEs closely matched that of myenteric ganglia, P2 x 2-immunopositive IGLEs were rarely found in upper and middle esophagus but increasingly in its lower parts. P2 x 2 stained only half the number of IGLEs found with VGLUT2 immunostaining. We also investigated the correlation between anterograde tracing and immunohistochemistry for identifying IGLEs. Confocal microscopy revealed colocalization of all three markers in approximately 50% of IGLEs. The remaining IGLEs showed only tracer and VGLUT2 labeling but no P2 x 2 immunoreactivity. Thus, VGLUT2 and P2 x 2 represent two specific markers for qualitative demonstration of esophageal IGLEs. However, VGLUT2 may be superior to P2 x 2 as a quantitative marker for IGLEs in the esophagus of C57Bl/6 mice.