Search

SFC is a work package in Eurad that investigates issues related to the properties of the spent nuclear fuel in the back-end of the nuclear fuel cycle. Decay heat, nuclide inventory, and fuel integrity (mechanical and otherwise), and not least the related uncertainties, are among the primary focal points of SFC. These have very significant importance for the safety and operational aspect of the back-end. One consequence is the operation economy of the back-end, where deeper understanding and quantification allow for significant optimization, meaning that significant parts of the costs can be reduced. In this paper, SFC is described, and examples of results are presented at about half-time of the work package, which will finish in 2024. The DisCo project started in 2017 and finished in November 2021 and was funded under the Horizon 2020 Euratom program. It investigated if the properties of modern fuel types, namely doped fuel, and MOX, cause any significant difference in the dissolution behavior of the fuel matrix compared with standard fuels. Spent nuclear fuel experiments were complemented with studies on model materials as well as the development of models describing the solid state, the dissolution process, and reactive transport in the near field. This research has improved the understanding of processes occurring at the interface between spent nuclear fuel and aqueous solution, such as redox reactions. Overall, the results show that from a long-term fuel matrix dissolution point of view, there is no significant difference between MOX fuel, Cr+Al-doped fuel, and standard fuels.

Diabetes-induced oxidative stress can lead to protein misfolding and degradation by the ubiquitin-proteasome system. This study examined protein ubiquitination in pancreatic sections from Zucker diabetic fatty rats. We observed large aggregates of ubiquitinated proteins (Ub-proteins) in insulin-expressing beta-cells and surrounding acinar cells. The formation of these aggregates was also observed in INS1 832/13 beta-cells after exposure to high glucose (30 mmol/l) for 8-72 h, allowing us to further characterize this phenotype. Oxidative stress induced by aminotriazole (ATZ) was sufficient to stimulate Ub-protein aggregate formation. Furthermore, the addition of the antioxidants N-acetyl cysteine (NAC) and taurine resulted in a significant decrease in formation of Ub-protein aggregates in high glucose. Puromycin, which induces defective ribosomal product (DRiP) formation was sufficient to induce Ub-protein aggregates in INS1 832/13 cells. However, cycloheximide (which blocks translation) did not impair Ub-protein aggregate formation at high glucose levels, suggesting that long-lived proteins are targeted to these structures. Clearance of Ub-protein aggregates was observed during recovery in normal medium (11 mmol/l glucose). Despite the fact that 20S proteasome was localized to Ub-protein aggregates, epoxomicin treatment did not affect clearance, indicating that the proteasome does not degrade proteins localized to these structures. The autophagy inhibitor 3MA blocked aggregate clearance during recovery and was sufficient to induce their formation in normal medium. Together, these findings demonstrate that diabetes-induced oxidative stress induces ubiquitination and storage of proteins into cytoplasmic aggregates that do not colocalize with insulin. Autophagy, not the proteasome, plays a key role in regulating their formation and degradation. To our knowledge, this is the first demonstration that autophagy acts as a defense to cellular damage incurred during diabetes. [ABSTRACT FROM AUTHOR]

Introducción: las enfermedades infecciosas son causa significativa de morbimortalidad y constituyen uno de los problemas de mayor importancia en salud pública a escala internacional. Los parásitos intestinales, como causantes de muchas de esas enfermedades, son también frecuentes y llama la atención que la información relacionada con la prevalencia e incidencia de esas infecciones en instituciones psiquiátricas es limitada. Objetivo: estimar la prevalencia de parásitos intestinales en pacientes con enfermedades mentales de larga evolución internados en el Hospital Provincial Psiquiátrico Docente Antonio Guiteras Holmes, de Matanzas, Cuba. Métodos: muestras de materia fecal de 56 internos fueron examinadas utilizando tres técnicas coproparasitológicas entre los meses de marzo y mayo de 2012. Resultados: 45 pacientes (80,4 %) presentaban al menos una especie de parásito en su aparato digestivo siendo Trichuris trichiura 44 (78,6%), el complejo Entamoeba histolytica/E. dispar (15/26,8 %) y Giardia lamblia (10/17,9 %) las mayormente identificadas. Ninguno de los pacientes presentaba síntomas o signos relacionados con el aparato digestivo, sin embargo, el 91,1 % presentó hábitos higiénico-dietéticos aberrantes. Conclusiones: la demostración de que más del 80 % de los pacientes con enfermedades mentales de larga evolución internados en el hospital psiquiátrico de Matanzas estaban infectados por alguna especie de parásitos intestinales, y alrededor del 10 % estaban poliparasitados evidencia la necesidad de acometer acciones encaminadas a crear un entorno saludable en el que se reduzca la posibilidad de transmisión de esas parasitosis.

Parvalbumin-containing (PV + ) basket cells are specialized cortical interneurons that regulate the activity of local neuronal circuits with high temporal precision and reliability. To understand how the PV + interneuron connectivity underlying these functional properties is established during development, we used array tomography to map pairs of synaptically connected PV + interneurons and postsynaptic neurons from the neocortex of mice of both sexes. We focused on the axon-myelin unit of the PV + interneuron and quantified the number of synapses onto the postsynaptic neuron, length of connecting axonal paths, and their myelination at different time points between 2 weeks and 7 months of age. We find that myelination of the proximal axon occurs very rapidly during the third and, to a lesser extent, fourth postnatal weeks. The number of synaptic contacts made by the PV + interneuron on its postsynaptic partner meanwhile is significantly reduced to about one-third by the end of the first postnatal month. The number of autapses, the synapses that PV + interneurons form on themselves, however, remains constant throughout the examined period. Axon reorganizations continue beyond postnatal month 2, with the postsynaptic targets of PV + interneurons gradually shifting to more proximal locations, and the length of axonal paths and their myelin becoming conspicuously uniform per connection. These continued microcircuit refinements likely provide the structural substrate for the robust inhibitory effects and fine temporal precision of adult PV + basket cells. SIGNIFICANCE STATEMENT The axon of adult parvalbumin-containing (PV + ) interneurons is highly specialized for fast and reliable neurotransmission. It is myelinated and forms synapses mostly onto the cell bodies and proximal dendrites of postsynaptic neurons for maximal impact. In this study, we follow the development of the PV + interneuron axon, its myelination and synapse formation, revealing a rapid sequence of axonal reorganization, myelination of the PV + interneuron proximal axon, and pruning of almost two-thirds of the synapses in an individual connection. This is followed by a prolonged period of axon refinement and additional myelination leading to a remarkable precision of connections in the adult mouse cortex, consistent with the temporal precision and fidelity of PV + interneuron action., (Copyright © 2021 the authors.)

Parvalbumin-containing (PV+) basket cells in mammalian neocortex are fast-spiking interneurons that regulate the activity of local neuronal circuits in multiple ways. Even though PV+ basket cells are locally projecting interneurons, their axons are myelinated. Can this myelination contribute in any significant way to the speed of action potential propagation along such short axons? We used dual whole cell recordings of synaptically connected PV+ interneurons and their postsynaptic target in acutely prepared neocortical slices from adult mice to measure the amplitude and latency of single presynaptic action potential-evoked inhibitory postsynaptic currents. These same neurons were then imaged with immunofluorescent array tomography, the synapses between them identified and a precise map of the connections was generated, with the exact axonal length and extent of myelin coverage. Our results support that myelination of PV+ basket cells significantly increases conduction velocity, and does so to a degree that can be physiologically relevant., (© The Author(s) 2021. Published by Oxford University Press. All rights reserved. For permissions, please e-mail: journals.permission@oup.com.)

We investigated synaptic mechanisms in the hippocampus that could explain how loss of circadian timing leads to impairments in spatial and recognition memory. Experiments were performed in hippocampal slices from Siberian hamsters (Phodopus sungorus) because, unlike mice and rats, their circadian rhythms are easily eliminated without modifications to their genome and without surgical manipulations, thereby leaving neuronal circuits intact. Recordings of excitatory postsynaptic field potentials and population spikes in area CA1 and dentate gyrus granule cells revealed no effect of circadian arrhythmia on basic functions of synaptic circuitry, including long-term potentiation. However, dentate granule cells from circadian-arrhythmic animals maintained a more depolarized resting membrane potential than cells from circadian-intact animals; a significantly greater proportion of these cells depolarized in response to the cholinergic agonist carbachol (10 μM), and did so by increasing their membrane potential three-fold greater than cells from the control (entrained) group. Dentate granule cells from arrhythmic animals also exhibited higher levels of tonic inhibition, as measured by the frequency of spontaneous inhibitory postsynaptic potentials. Carbachol also decreased stimulus-evoked synaptic excitation in dentate granule cells from both intact and arrhythmic animals as expected, but reduced stimulus-evoked synaptic inhibition only in cells from control hamsters. These findings show that loss of circadian timing is accompanied by greater tonic inhibition, and increased synaptic inhibition in response to muscarinic receptor activation in dentate granule cells. Increased inhibition would likely attenuate excitation in dentate-CA3 microcircuits, which in turn might explain the spatial memory deficits previously observed in circadian-arrhythmic hamsters., (© 2021 The Authors. Hippocampus published by Wiley Periodicals LLC.)

Background: The ability to correlate plastic changes in synaptic physiology with changes in synaptic anatomy has been very limited in the central nervous system because of shortcomings in existing methods for recording the activity of specific CNS synapses and then identifying and studying the same individual synapses on an anatomical level., New Method: We introduce here a novel approach that combines two existing methods: paired neuron electrophysiological recording and array tomography, allowing for the detailed molecular and anatomical study of synapses with known physiological properties., Results: The complete mapping of a neuronal pair allows determining the exact number of synapses in the pair and their location. We have found that the majority of close appositions between the presynaptic axon and the postsynaptic dendrite in the pair contain synaptic specializations. The average release probability of the synapses between the two neurons in the pair is low, below 0.2, consistent with previous studies of these connections. Other questions, such as receptor distribution within synapses, can be addressed more efficiently by identifying only a subset of synapses using targeted partial reconstructions. In addition, time sensitive events can be captured with fast chemical fixation., Comparison With Existing Methods: Compared to existing methods, the present approach is the only one that can provide detailed molecular and anatomical information of electrophysiologically-characterized individual synapses., Conclusions: This method will allow for addressing specific questions about the properties of identified CNS synapses, even when they are buried within a cloud of millions of other brain circuit elements., (Copyright © 2016. Published by Elsevier B.V.)

Pair recordings involve simultaneous whole cell patch clamp recordings from two synaptically connected neurons, enabling not only direct electrophysiological characterization of the synaptic connections between individual neurons, but also pharmacological manipulation of either the presynaptic or the postsynaptic neuron. When carried out in organotypic hippocampal slice cultures, the probability that two neurons are synaptically connected is significantly increased. This preparation readily enables identification of cell types, and the neurons maintain their morphology and properties of synaptic function similar to that in native brain tissue. A major advantage of paired whole cell recordings is the highly precise information it can provide on the properties of synaptic transmission and plasticity that are not possible with other more crude techniques utilizing extracellular axonal stimulation. Paired whole cell recordings are often perceived as too challenging to perform. While there are challenging aspects to this technique, paired recordings can be performed by anyone trained in whole cell patch clamping provided specific hardware and methodological criteria are followed. The probability of attaining synaptically connected paired recordings significantly increases with healthy organotypic slices and stable micromanipulation allowing independent attainment of pre- and postsynaptic whole cell recordings. While CA3-CA3 pyramidal cell pairs are most widely used in the organotypic slice hippocampal preparation, this technique has also been successful in CA3-CA1 pairs and can be adapted to any neurons that are synaptically connected in the same slice preparation. In this manuscript we provide the detailed methodology and requirements for establishing this technique in any laboratory equipped for electrophysiology.

Recent studies have revealed the existence of stem cells in various human tissues including dental structures. We aimed to establish primary cell cultures from human dental pulp and periodontal ligament, to identify multipotential adult stem cells in these cultures, and to study the differentiation capacity of these cells to osteogenic and to neuronal fates. Dental pulp and the periodontal ligament were isolated from extracted human wisdom teeth. The extracellular matrix was enzymatically degraded to obtain isolated cells for culturing. Both dental pulp and periodontal ligament derived cultures showed high proliferative capacity and contained a cell population expressing the STRO-1 mesenchymal stem cell marker. Osteogenic induction by pharmacological stimulation resulted in mineralized differentiation as shown by Alizarin red staining in both cultures. When already described standard neurodifferentiation protocols were used, cultures exhibited only transient neurodifferentiation followed by either redifferentiation into a fibroblast-like phenotype or massive cell death. Our new three-step neurodifferentiation protocol consisting of (1) epigenetic reprogramming, then (2) simultaneous PKC/PKA activation, followed by (3) incubation in a neurotrophic medium resulted in robust neurodifferentiation in both pulp and periodontal ligament cultures shown by cell morphology, immunocytochemistry and real time PCR for vimentin and neuron-specific enolase. In conclusion, we report the isolation, culture and characterization of stem cell containing cultures from both human dental pulp and periodontal ligament. Furthermore, our data clearly show that both cultures differentiate into mineralized cells or to a neuronal fate in response to appropriate pharmacological stimuli. Therefore, these cells have high potential to serve as resources for tissue engineering not only for dental or bone reconstruction, but also for neuroregenerative treatments.

Brain injuries are too common. Most people are unaware of the incidence of and horrendous consequences of traumatic brain injury (TBI) and mild traumatic brain injury (MTBI). Research and the advent of sophisticated imaging have led to progression in the understanding of brain pathophysiology following TBI. Seminal evidence from animal and human experiments demonstrate links between TBI and the subsequent onset of premature, psychiatric syndromes and neurodegenerative diseases, including Alzheimer's disease (AD) and Parkinson's disease (PD). Objectives of this summary are, therefore, to instill appreciation regarding the importance of brain injury prevention, diagnosis, and treatment, and to increase awareness regarding the long-term delayed consequences following TBI.

The effects of exercise training on hypothalamo-pituitary-adrenal (HPA) function are unclear. We investigated whether pituitary-adrenal adaptation during exercise training is mediated by changes in neuropeptide and corticosteroid receptor gene expression in the brain and pituitary. Sprague-Dawley rats were subject to either daily swimming (DS) or sham exercise (SE) for 45 min/day, 5 days/week, for 2 (2W), 4 (4W), or 6 wk (6W) (n = 7-10/group). Corticosterone (Cort) and catecholamine responses during swimming were robust at 6W compared with 2W and 4W, indicating that HPA response to exercise during training is not attenuated when absolute intensity is progressively increased. In DS, basal (morning) plasma ACTH and Cort levels increased from 2W to 4W but plateaued at 6W, whereas in SE, they increased from 4W to 6W, with 6W values higher than in DS. In DS, there was a transient decrease in glucocorticoid receptor (GR) mRNA in the paraventricular nucleus (PVN) and pituitary and a transient increase in corticotrophin-releasing hormone (CRH) mRNA. In contrast, hippocampal mineralocorticoid receptor mRNA and PVN GR mRNA decreased from 4W to 6W in SE, with 6W values lower than in DS. These findings suggest that exercise training prevents an elevation in basal pituitary-adrenal activity potentially via transient alterations in the gene transcription of PVN and pituitary GR as well as CRH to suppress central drive to the HPA axis. In contrast, the increase in basal pituitary-adrenal activity with repeated sham exercise appears to be associated with decreases in hippocampal MR and PVN GR mRNA expression.

Background: In Alzheimer's disease (AD) the olfactory system, including the olfactory bulb, a limbic paleocortex is severely damaged. The occurrence of early olfactory deficits and the presence of senile plaques and neurofibrillary tangles in olfactory bulb were reported previously by a few authors. The goal of the present study was to analyze the occurrence of AD-type degenerative changes in the peripheral part of the olfactory system and to answer the question whether the frequency and severity of changes in the olfactory bulb and tract are associated with those of the cerebral cortex in AD., Material and Methods: In 110 autopsy cases several cortical areas and the olfactory bulb and tract were analyzed using histo- and immunohistochemical techniques. Based on a semiquantitative analysis of cortical senile plaques, neurofibrillary tangles and curly fibers, the 110 cases were divided into four groups: 19 cases with severe (definite AD), 14 cases with moderate, 58 cases with discrete and 19 control cases without AD-type cortical changes., Results: The number of cases with olfactory involvement was very high, more than 84% in the three groups with cortical AD-type lesions. Degenerative olfactory changes were present in all 19 definite AD cases, and in two of the 19 controls. The statistical analysis showed a significant association between the peripheral olfactory and cortical degenerative changes with respect to their frequency and severity (P < 0.001). Neurofibrillary tangles and neuropil threads appear in the olfactory system as early as in entorhinal cortex., Conclusion: The results indicate a close relationship between the olfactory and cortical degenerative changes and indicate that the involvement of the olfactory bulb and tract is one of the earliest events in the degenerative process of the central nervous system in AD.

The unfolded protein response (UPR) counteracts stress caused by unprocessed ER client proteins. A genome-wide survey showed impaired induction of many UPR target genes in xbp-1 mutant Caenorhabditis elegans that are unable to signal in the highly conserved IRE1-dependent UPR pathway. However a family of genes, abu (activated in blocked UPR), was induced to higher levels in ER-stressed xbp-1 mutant animals than in ER-stressed wild-type animals. RNA-mediated interference (RNAi) inactivation of a representative abu family member, abu-1 (AC3.3), activated the ER stress marker hsp-4::gfp in otherwise normal animals and killed 50% of ER-stressed ire-1 and xbp-1 mutant animals. Abu-1(RNAi) also enhanced the effect of inactivation of sel-1, an ER-associated protein degradation gene. The nine abu genes encode highly related type I transmembrane proteins whose lumenal domains have sequence similarity to a mammalian cell surface scavenger receptor of endothelial cells that binds chemically modified extracellular proteins and directs their lysosomal degradation. Our findings that ABU-1 is an intracellular protein located within the endomembrane system that is induced by ER stress in xbp-1 mutant animals suggest that ABU proteins may interact with abnormal ER client proteins and this function may be particularly important in animals with an impaired UPR.

In Caenorhabditis elegans, let-60 Ras controls many cellular processes, such as differentiation of vulval epithelial cells, function of chemosensory neurons, and meiotic progression in the germ line. Although much is known about the let-60 Ras signaling pathway, relatively little is understood about the target genes induced by let-60 Ras signaling that carry out terminal effector functions leading to morphological change. We have used DNA microarrays to identify 708 genes that change expression in response to activated let-60 Ras., ((c) 2002 Elsevier Science (USA).)

The nematode worm Caenorhabditis elegans and its relatives are unique among animals in having operons. Operons are regulated multigene transcription units, in which polycistronic pre-messenger RNA (pre-mRNA coding for multiple peptides) is processed to monocistronic mRNAs. This occurs by 3' end formation and trans-splicing using the specialized SL2 small nuclear ribonucleoprotein particle for downstream mRNAs. Previously, the correlation between downstream location in an operon and SL2 trans-splicing has been strong, but anecdotal. Although only 28 operons have been reported, the complete sequence of the C. elegans genome reveals numerous gene clusters. To determine how many of these clusters represent operons, we probed full-genome microarrays for SL2-containing mRNAs. We found significant enrichment for about 1,200 genes, including most of a group of several hundred genes represented by complementary DNAs that contain SL2 sequence. Analysis of their genomic arrangements indicates that >90% are downstream genes, falling in 790 distinct operons. Our evidence indicates that the genome contains at least 1,000 operons, 2 8 genes long, that contain about 15% of all C. elegans genes. Numerous examples of co-transcription of genes encoding functionally related proteins are evident. Inspection of the operon list should reveal previously unknown functional relationships.

We have assembled data from Caenorhabditis elegans DNA microarray experiments involving many growth conditions, developmental stages, and varieties of mutants. Co-regulated genes were grouped together and visualized in a three-dimensional expression map that displays correlations of gene expression profiles as distances in two dimensions and gene density in the third dimension. The gene expression map can be used as a gene discovery tool to identify genes that are co-regulated with known sets of genes (such as heat shock, growth control genes, germ line genes, and so forth) or to uncover previously unknown genetic functions (such as genomic instability in males and sperm caused by specific transposons).

We have constructed DNA microarrays containing 17,871 genes, representing about 94% of the 18,967 genes currently annotated in the Caenorhabditis elegans genome. These DNA microarrays can be used as a tool to define a nearly complete molecular profile of gene expression levels associated with different developmental stages, growth conditions, or worm strains. Here, we used these full-genome DNA microarrays to show the relative levels of gene expression for nearly every gene during development, from eggs through adulthood. These expression data can help reveal when a gene may act during development. We also compared gene expression in males to that of hermaphrodites and found a total of 2,171 sex-regulated genes (P < 0.05). The sex-regulated genes provide a global view of the differences between the sexes at a molecular level and identify many genes likely to be involved in sex-specific differentiation and behavior.

Regulation of calbindin and calretinin expression by brain-derived neurotrophic factor (BDNF) was examined in primary cultures of cortical neurons using immunocytochemistry and northern blot analysis. Here we report that regulation of calretinin expression by BDNF is in marked contrast to that of calbindin. Indeed, chronic exposure of cultured cortical neurons for 5 days to increasing concentrations of BDNF (0.1-10 ng/ml) resulted in a concentration-dependent decrease in the number of calretinin-positive neurons and a concentration-dependent increase in the number of calbindin-immunoreactive neurons. Consistent with the immunocytochemical analysis, BDNF reduced calretinin mRNA levels and up-regulated calbindin mRNA expression, providing evidence that modifications in gene expression accounted for the changes in the number of calretinin- and calbindin-containing neurons. Among other members of the neurotrophin family, neurotrophin-4 (NT-4), which also acts by activating tyrosine kinase TrkB receptors, exerted effects comparable to those of BDNF, whereas nerve growth factor (NGF) was ineffective. As for BDNF and NT-4, incubation of cortical neurons with neurotrophin-3 (NT-3) also led to a decrease in calretinin expression. However, in contrast to BDNF and NT-4, NT-3 did not affect calbindin expression. Double-labeling experiments evidenced that calretinin- and calbindin-containing neurons belong to distinct neuronal subpopulations, suggesting that BDNF and NT-4 exert opposite effects according to the neurochemical phenotype of the target cell.

Rat superior cervical ganglia infected with herpes virus suis (pseudorabies virus) display a spontaneous bursting activity of still unknown origin. Previous intracellular recordings from the ganglionic neurons combined with pharmacological studies showed that the postganglionic action potentials are induced by acetylcholine release spontaneously from the preganglionic nerve. In this study we investigated whether the acetylcholine release is caused by mechanisms which are dependent on action potentials spontaneously generated on the preganglionic nerve or by mechanisms which occur without any changes in the excitability of presynaptic fibers. Simultaneous intra- and extracellular recordings from the ganglion cells and from the preganglionic nerve, respectively, were performed 32-38 h after the inoculation of herpes virus suis (strain Aujeszky) into the anterior chamber of one eye of the rat. Tetrodotoxin, well known to prevent the generation of action potentials by blocking the fast sodium channels, completely and reversibly abolished, whereas the potassium channel blockers 4-aminopyridine and apamin, enhanced the spontaneous, bursting activity at pre- and postsynaptic levels. The nicotinic receptor antagonist hexamethonium abolished the postsynaptic discharges and reduced the preganglionic activity by 50%. Pre- and postsynaptic electrical activities were suppressed in low calcium Krebs' solution, demonstrating that extracellular calcium is required not only for acetylcholine release but also for triggering the presynaptic action potentials. It is concluded that in the infected ganglia the spontaneous acetylcholine release is due to the generation of action potentials in the preganglionic nerve. Voltage-gated sodium and calcium channels contribute to the presynaptic electrogenesis, while the latter appears to be damped by the activation of voltage- and calcium-dependent potassium channels. Possible factors as well as mechanisms inducing such an increase in excitability are discussed.

Neuropharmacological and histochemical evidence presented here indicates that serotonin (5-HT) is the transmitter mediating one of the postsynaptic potentials in the guinea pig celiac ganglion. Repetitive nerve stimulation elicited in celiac neurons, in addition to the nicotinic fast excitatory postsynaptic potential (EPSP), a slow EPSP that was resistant to cholinergic antagonists. Application of 5-HT caused a depolarization with membrane characteristics similar to those of the slow EPSP; furthermore, the latter was reversibly suppressed by 5-HT. The slow depolarization evoked by either nerve stimulation or 5-HT was augmented by fluoxetine, a 5-HT reuptake blocker, and depressed by cyproheptadine, a 5-HT receptor blocker; in addition, tryptophan, a precursor of 5-HT, enhanced differentially the slow EPSP. Lastly, histochemical study revealed dense networks of 5-HT immunoreactive nerve fibers encircling many ganglionic neurons.

The effects of capsaicin (0.5-100 microM) on neurones of the isolated inferior mesenteric ganglia (i.m.g.) of the guinea-pig were investigated by means of intracellular recording techniques. When applied to neurones of the i.m.g. that exhibited a slow non-cholinergic excitatory potential (Dun & Jiang, 1982), capsaicin caused in the large majority of these cells a long lasting depolarization accompanied by intense neuronal discharges. During and immediately following the depolarization, repetitive presynaptic stimulation consistently failed to elicit the non-cholinergic depolarization; a partial recovery was observed in relatively few neurones. The fast (nicotinic) excitatory post-synaptic potentials (e.p.s.p.s) were not suppressed by capsaicin in any of these cells. The membrane potential, input resistance and the amplitude of fast e.p.s.p.s in neurones of the i.m.g. that generated no detectable non-cholinergic depolarizations were not affected by capsaicin. Post-synaptic membrane sensitivity to exogenous application of substance P was not altered following capsaicin superfusion even though the latter effectively abolished the non-cholinergic depolarization in the same neurones. Superfusing the i.m.g. with a Ca-free Krebs solution markedly attenuated or abolished the depolarizing effect of capsaicin whereas tetrodotoxin (1 microM) was without effect. Capsaicin was without effect in a few neurones that generated a non-cholinergic depolarization; the latter was not desensitized by bath application of substance P. Capsaicin caused no appreciable effects in neurones of the bullfrog sympathetic ganglia; thus, the fast and slow post-synaptic potentials including the late slow e.p.s.p. the transmitter of which is the peptide luteinizing hormone-releasing hormone (Jan & Jan, 1982) were not affected. The results suggest that the depolarizing effect of capsaicin in neurones of the guinea-pig i.m.g. is due to a selective release of ganglionic substance P or a substance P-like peptide in a Ca-dependent manner, and that the non-cholinergic potentials elicited in capsaicin-insensitive neurones may be generated by a transmitter(s) other than substance P.

The nature of the putative transmitter(s) mediating the non-cholinergic excitatory post-synaptic potential (e.p.s.p.) described in the preceding paper was investigated by means of electrophysiological, pharmacological and immunohistochemical methods. Serotonin (1-10 microM) when applied by superfusion caused a slow depolarization that closely mimicked the synaptic response in about 60% of the coeliac neurones that exhibited a non-cholinergic e.p.s.p. The serotonin depolarization evoked in low-Ca2+, high-Mg2+ solution or in a Krebs solution containing cholinergic antagonists was quantitatively similar to that elicited in normal Krebs solution. When compared in the same neurones the membrane resistance change during the course of the serotonin depolarization and of the non-cholinergic e.p.s.p., as well as their respective responses to conditioning polarization, were similar. The non-cholinergic e.p.s.p. was reversibly abolished during serotonin-induced depolarization; the blockade persisted when the membrane potential was restored to the resting level by hyperpolarizing current. The serotonin depolarization as well as the non-cholinergic e.p.s.p. were reversibly suppressed by cyproheptadine (20-50 microM), a serotonin antagonist, and enhanced by fluoxetine (30-50 microM), a serotonin reuptake inhibitor. On the other hand, pre-treating the ganglia with L-tryptophan (50 microM), a precursor of serotonin, preferentially augmented the synaptically induced response. A portion of the neurones (15%) were depolarized by substance P (1 microM) which also reversibly desensitized the non-cholinergic e.p.s.p. elicited in these neurones. The remaining neurones (25%) were insensitive to either serotonin or substance P, and the non-cholinergic e.p.s.p.s elicited in these cells were likewise not appreciably affected by these two agents. Furthermore, cyproheptadine, fluoxetine and L-tryptophan had no significant effect on the non-cholinergic e.p.s.p.s elicited in serotonin-insensitive neurones. Using the immunohistofluorescent techniques, dense but unevenly distributed serotonin immunoreactive nerve fibres could be observed surrounding many coeliac neurones. Immunoreactivity was not observed in the ganglia incubated with antisera pre-absorbed with excess serotonin. Collectively our results suggest that serotonin is the mediator of non-cholinergic e.p.s.p.s. elicited in about 60% of coeliac neurones sampled in this study, and that in the remaining neurones the slow depolarization may be generated by substance P and/or some unknown transmitter(s).

The actions of arginine-vasopressin (AVP) and oxytocin (OXT) were investigated in the rat superior cervical ganglion (SCG). At micromolar concentrations AVP decreased the amplitude of fast excitatory postsynaptic potentials (f-EPSPs) evoked by preganglionic stimulation and in many cells depolarized the postsynaptic membrane. Both effects were reversibly abolished by a potent vasopressor antagonist. The peptide decreased the frequency of spontaneous miniature EPSPs and the quantal content of the f-EPSPs without affecting the sensitivity of the ganglion cells to acetylcholine. OXT exerted the same effects as AVP but was less powerful. It was concluded that neurohypophysial peptides exert a dual pre- and post-synaptic action mediated by specific receptors.

Binding of radioactive vasopressin--but not of oxytocin--was detected by autoradiography and by labeling of membranes obtained from the rat superior cervical ganglion. In both instances binding could be displaced by V1 (smooth muscle-type) but not by V2 (kidney-type) agonists, indicating that the ganglionic vasopressin receptors are similar to those present on hepatocytes and vascular smooth muscle. In accordance with the V1 character of the receptors, vasopressin activated the turnover of membrane inositol lipids, and this effect was abolished by a structural analogue known to act as a vasopressor antagonist. A possible physiological role of vasopressin was suggested by intracellular recordings obtained from ganglion cells in vitro. Vasopressin induced a reduction in the amplitude of the fast excitatory postsynaptic potential evoked by electrical stimulation of the preganglionic nerve. This reduction in ganglionic transmission was antagonized by the same synthetic structural analogue that blocked the effect of vasopressin on inositol lipids. This study provides evidence for the presence of functional vasopressin receptors in a rat sympathetic ganglion and thus suggests that vasopressin may play a role in peripheral autonomic function.

Electrophysiologically identified neurons of rat superior cervical ganglion were intracellularly injected with horseradish peroxidase and processed for light and electron microscopic observation. At light microscope level, neurons could be classified according to their dendritic arborization pattern in the vicinity of the soma into radiate, tufted and intermediate types. Upon electrical stimulation of the internal and external carotid nerves it was observed that radiate and intermediate neurons sent their axons into one or the other of these nerve trunks, whereas a majority of tufted neurons gave no response to stimulation of either of these postganglionic nerves. Electron microscopic exploration of horseradish peroxidase-labelled neurons revealed a surprisingly high prevalence of interconnectivity between ganglionic neurons. These contacts were both dendrosomatic and dendrodendritic, and were a universal feature of the labelled neurons explored. Twenty-two of the 23 labelled cells were found to receive direct dendritic appositions on their somata, and 13 of these 23 cells were seen each to send their dendrites into contact with at least one unlabelled neuronal soma. Dendrodendritic contacts were observed for 87% of the labelled neurons, and most of the cells (80%) were seen to form triadic contacts which included two dendrites and a preganglionic nerve ending. All these figures represent minimum incidences. None of the dendrosomatic or dendrodendritic appositions observed was overtly synaptic although several morphological features indicated the possibility of somatic and or dendritic release and uptake at sites of apposition. It is suggested that the observed appositions provide anatomical substrates for modulatory interactions between the ganglionic neurons, possibly involving slow potentials or the switching of metabolic pathways.

The bioelectric activity of the rat superior cervical ganglion (SCG) infected with pseudorabies virus (PRV) was examined in vitro 30-38 h after inoculation. Simultaneous intra- and extracellular recordings on the internal (ICN) and external carotid nerves (ECN) revealed a synchronized spontaneous activity. This synchronization can be ascribed either to the functional organization of the ganglion or to the mechanism of initiation itself. In the infected ganglia two categories of cells were observed: cells displaying abnormal spontaneous discharges, and silent cells whose electrophysiological behavior was similar to control cells. Spontaneously active cells showed intermittent spiking and bursting activity. The discharge pattern was associated with the firing rate of the emitting cell: sporadically active cells emitted single spikes whereas highly active cells fired bursts of action potentials (APs). Long lasting intracellular recordings demonstrated that the cells undergo gradual changes evolving from sporadic on to high activity. Spontaneous APs usually rode on prepotentials similar to the excitatory postsynaptic potentials (EPSPs). A comparative study of spontaneous prepotentials and orthodromically evoked EPSPs in the same cell demonstrated that the spontaneous prepotentials are real synaptic potentials. No pace-maker potentials were observed. The passive and active electrical membrane properties of spontaneously active neurons were not different from those of silent cells or control cells impaled in uninfected ganglia. D-Tubocurarine abolished the spontaneous activity in the whole ganglion. Ortho- and antidromic electrical stimulations of suprathreshold intensity elicited an evoked response in neurons displaying spontaneous activity, followed by a delayed burst whose shape was similar to the spontaneous burst of the cell. Stimuli of subthreshold intensities induced this delayed burst independently from the evoked response. We conclude that the spontaneous bioelectrical activity is of presynaptic, but not necessarily of preganglionic origin. The possible existence of a cholinergic intraganglionic pathway revealed by the viral infection is discussed.

Serotonin (5-HT) in the guinea pig celiac-superior mesenteric plexus was quantitatively measured by HPLC and visualized by an immunohistochemical method. Preincubation of the ganglia in a Krebs solution containing L-tryptophan and pargyline markedly elevated the content of 5-HT and K+ solution caused a release of 5-HT into the incubation medium. 5-HT immunoreactivity was localized to dense but unevenly distributed nerve fibers throughout the plexus and to small diameter cells commonly referred to as small intensely fluorescent cells. These findings provide evidence of an extensive network of 5-HT-containing neural elements in the guinea pig prevertebral ganglia.