Biomedical subjects

R C Cannon

Publications and source records attributed to R C Cannon.

8 recordsLinked to original sources

PubMed2000

Distribution of spontaneous currents along the somato-dendritic axis of rat hippocampal CA1 pyramidal neurons.

Excitatory and inhibitory pathways have specific patterns of innervation along the somato-dendritic axis of neurons. We have investigated whether this morphological diversity was associated with variations in the frequencies of spontaneous and miniature GABAergic and glutamatergic synaptic currents along the somato-dendritic axis of rat hippocampal CA1 pyramidal neurons. Using in vitro whole cell recordings from somata, apical dendrites and basal dendrites (for which we provide the first recordings) of CA1 pyramidal neurons, we report that over 90% of the spontaneous currents were GABAergic, <10% being glutamatergic. The frequency of spontaneous GABAergic currents was comparable in the soma and in the dendrites. In both somata and dendrites, the Na(+) channel blocker tetrodotoxin abolished more than 80% of the spontaneous glutamatergic currents. In contrast, tetrodotoxin abolished most dendritic (>90%) but not somatic (<40%) spontaneous GABAergic currents. Computer simulations suggest that in our experimental conditions, events below 40pA are electrotonically filtered to such a degree that they are lost in the recording noise. We conclude that, in vitro, inhibition is massively predominant over excitation and quantitatively evenly distributed throughout the cell. However, inhibition appears to be mainly activity-dependent in the dendrites whereas it can occur in the absence of interneuron firing in the soma. These results can be used as a benchmark to compare values obtained in pathological tissue, such as epilepsies, where changes in the balance between excitation and inhibition would dramatically alter cell behaviour.

Action Potentials↗

PubMed1999

Dendrites of classes of hippocampal neurons differ in structural complexity and branching patterns.

Dendrites of reconstructed hippocampal neurons were analyzed for morphometric, topologic, and fractal parameters (n = 32 quantities) to investigate neuronal groupings and growth characteristics with a common set of assumptions. The structures studied included CA1 and CA3 pyramidal cells, interneurons, and granule cells from young animals (71 cells in total). Most of the cells showed no characteristic fractal dimension; rather, the scaling relation could be well represented by a two-parameter fit, of which one parameter showed a significant difference between cell classes. Other significant quantities that differentiated cell classes were related to the complexity of the dendritic tree (number of branch points and maximal terminal branch order) and the cell's electrical properties such as the mean attenuation between the soma and terminals. Principal components analysis produced combined measures of only slightly greater discriminative power than the best individual measures, indicating that the elementary quantities capture most of the structural variation between hippocampal cell groups. Another finding was that for all cells the mean segment length increased with dendritic branch order, which is consistent with decreasing branching probability as a function of the path distance from the soma. Analysis of another set of CA1 pyramidal neurons from aged animals (n = 15; 22-24 months) showed only a few significant differences than those from young animals (n = 11; a subset of n = 71) of which the most important was a straightening of the paths between terminals and the soma. The quantities analyzed in these reconstructed hippocampal neurons may reflect both intrinsic neuronal characteristics and extrinsic influences. Hippocampal cell groupings (i.e., pyramidal cells as opposed to dentate granule cells and interneurons) were significantly differentiated by most parameters. These differences and parameter values may be critical for understanding and generating synthetic neuronal populations for modelling studies.

Aging↗

PubMed1999

Radioactivity-based and spectrophotometric assays for isoorotate decarboxylase: identification of the thymidine salvage pathway in lower eukaryotes.

A few organisms, notably some fungi, have the ability to metabolize thymidine to uracil, thus conserving the pyrimidine ring for subsequent metabolic use. Neurospora crassa possesses this pathway, termed the thymidine salvage pathway, and can utilize thymidine as a total pyrimidine source. The enzyme isoorotate decarboxylase (IDCase) completes this pathway via the enzymatic removal of the carboxylate from isoorotate to yield uracil. We describe in this communication two assays for IDCase and their application to determine activity levels, kinetic constants, and inhibitory properties. One uses [carboxy-14C]isoorotate from which the enzymatically generated 14CO2 is collected and quantitated. The second assay utilizes the spectral difference between 2-thioisoorotate and its decarboxylated product, 2-thiouracil. The spectral difference is greatest at 334 nm, out of the range of absorbance of total protein and thus usable for a spectrophotometric assay. The assays are sufficiently sensitive and accurate to be used in the measurement of Km values for both substrates. IDCase activity is found to be significantly higher in N. crassa strains lacking uc-1, a putative regulatory gene, suggesting a degree of metabolic control over this pathway. 5-Nitrouracil is found to inhibit IDCase with an estimated Ki value that is too low for accurate determination.

Ammonium Sulfate↗

PubMed1999

Reduced Mg2+ blockade of synaptically activated N-methyl-D-aspartate receptor-channels in CA1 pyramidal neurons in kainic acid-lesioned rat hippocampus.

Unilateral kainic acid lesion in the hippocampus caused a long-term change in the balance between excitatory and inhibitory drive onto CA1 pyramidal cells, making these cells hyperexcitable several weeks post-lesion. In this study, we have shown an enhanced N-methyl-D-aspartate receptor-mediated component in the excitatory synaptic transmission together with a reduced GABA(A) receptor-mediated inhibition in CA1 pyramidal cells one-week post kainic acid lesion. In these cells, pharmacologically isolated N-methyl-D-aspartate receptor-mediated whole-cell excitatory postsynaptic currents were significantly larger at negative holding potentials, and the voltage-dependence of N-methyl-D-aspartate receptor channels was shifted in the hyperpolarizing direction. The plot of relative conductance (g/gMax) shifted significantly (P<0.01) to more negative holding potentials by 19 mV (-28+/-4 mV in control slices and -47+/-4 mV in kainic acid slices) at the half maximal conductance point (g/gMax =0.5). This shift gives a larger N-methyl-D-aspartate receptor-mediated component in the excitatory synaptic transmission at resting membrane potentials (around -60 mV). The shifted voltage dependence is highly sensitive to extracellular Mg2+ ions. Moderate increases in [Mg2+]o from 1 mM to 2.6 mM more than compensated for the negative shift and effectively suppressed the population epileptiform bursting activity. Fitting the voltage dependence to an ionic block model revealed a higher dissociation constant of N-methyl-D-aspartate receptor channels for Mg2+ in kainic acid-lesioned slices (52 mM at 0 mV; 330 microM at -60 mV) than in control slices (7.7 mM at 0 mV; 93 microM at -60 mV). While a simple single site model adequately fitted the control data for [Mg2+]o at 1 mM and 2.6 mM, no consistent model of this form was found for the kainic acid-lesioned slices. These results revealed changed properties of N-methyl-D-aspartate receptor channels in the kainic acid-lesioned model of epilepsy. The reduced Mg2+ blockade of N-methyl-D-aspartate receptor channels contributed significantly to the epileptiform bursting activity.

2-Amino-5-phosphonovalerate↗

PubMed1998

An on-line archive of reconstructed hippocampal neurons.

We have developed an on-line archive of neuronal geometry to encourage the use of realistic dendritic structures in morphometry and for neuronal modeling, located at web address www.neuro.soton.ac.uk. Initially we have included full three-dimensional representations of 87 neurons from the hippocampus, obtained following intracellular staining with biocytin and reconstruction using Neurolucida. The archive system includes a structure editor for correcting any departures from valid branching geometry and which allows simple errors in the digitisation to be corrected. The editor employs a platform-independent file format which enforces the constraints that there should be no isolated branches and no closed loops. It also incorporates software for interconversion between the archive format and those used by various neuronal reconstruction and modelling packages. The raw data from digitisation software can be included in the archive as well as edited reconstructions and any further information available. Cross-referenced tables and indexes are updated automatically and are sorted according to a number of fields including the cell type, contributor, submission date and published reference. Both the archive and the structure editor should facilitate the quantitative use of full three-dimensional reconstructions of neurons from the hippocampus and other brain regions.

Animals↗

PubMed1998

Pro-epileptic changes in synaptic function can be accompanied by pro-epileptic changes in neuronal excitability.

Repetitive sensory input, stroboscopic lights or repeated sounds can induce epileptic seizures in susceptible individuals. In order to understand the process we have to consider multiple factors. The output of a set of neurones is determined by the amount of excitatory synaptic input, the degree of positive feedback and their inherent electrical excitability, which can be modified by synaptic inhibition. Recent research has shown that it is possible to separate these phenomena, and that they do not always behave in unison.

Animals↗

PubMed1997

Model of spatio-temporal propagation of action potentials in the Schaffer collateral pathway of the CA1 area of the rat hippocampus.

There is a sharp contrast between the profuse in vivo axonal arborization of CA3 pyramidal cells in the CA1 area and the low probability of finding pairs of connected CA3-CA1 pyramidal cells in vitro. These anatomical differences contribute to a connectivity argument for discrepancies between electrophysiological data recorded in vitro and in vivo. In order to investigate this issue, we have developed a realistic computer model of the Schaffer collateral pathway of the hippocampus and analyzed the spatio-temporal distribution of action potentials along this pathway following three different types of electrical test stimulus. Direct activation of mossy fibers, CA3 pyramidal cells and focal stimulation of CA1 stratum radiatum were investigated. The parameters of the model were selected from available biological data. Spikes in Schaffer collaterals were followed from their onset in the CA3 pyramidal cell initial segment to the last order branches of their axonal tree in two types of configuration: the whole hippocampus and the slice configuration. The anatomical and electropysiological characteristics of the mossy fibre and Schaffer collateral pathways were found to impose strong constraints on the spatio-temporal distribution of action potentials in the CA1 area. Specific projection zones are determined by the spatial localization of the emitting CA3 pyramidal cells. Their position also defines precise time windows during which some CA1 projection zones receive a large number of correlated signals. Moreover, the variability of the delay at the mossy fibre/CA3 pyramidal cell synapse seems to provide the CA1 projection zones with a background level of excitation. Finally, we show how the patterns of activation obtained in the whole hippocampus are different from those obtained in the slice.

Action Potentials↗

PubMedDate not supplied

New fluorescence polarization immunoassays for analysis of barbiturates and benzodiazepines in serum and urine: performance characteristics.

The performance of the new fluorescence polarization immunoassay reagents Cassette COBAS INTEGRA Serum Benzodiazepines assay (SBENZ) and Cassette Serum Barbiturates assay (SBARB) was evaluated as compared to other immunoassays (Abbott TDx Serum Benzodiazepines, Abbott TDx Urine Benzodiazepines, Behring EMIT Serum Benzodiazepines, Abbott ADx Serum Barbiturates, Behring EMIT Serum Barbiturates, and the COBAS INTEGRA Barbiturates (BARB) urine assay) and gas chromatography-mass spectrometry (GC-MS). Recoveries of nordiazepam and secobarbital using the SBENZ and SBARB assays, respectively, were equivalent for serum, plasma, and urine. Cross-reactivities of structurally related benzodiazepines, barbiturates, and their metabolites were very similar in serum and urine for the SBENZ and SBARB assays. Precision was within 5.4% for SBENZ serum and within 11% from 10 to 100 ng/mL for urine. Precision was within 5% for SBARB serum and within 7% from 136 to 277 ng/mL for the urine application. The standard curves for SBENZ and SBARB were stable for at least 16 weeks with the reagents stored open on the COBAS INTEGRA analyzer. Clinical comparison of the SBENZ serum assay indicated an increased pickup rate, as confirmed by GC-MS, compared to TDx and EMIT. The diagnostic sensitivities of the SBENZ serum application, TDx, and EMIT versus GC-MS were 100%, 89%, and 36%, respectively. The diagnostic specificities were 71%, 79%, and 100%, respectively. The diagnostic sensitivities of the SBENZ urine application and TDx versus GC-MS were 100% and the diagnostic specificities were 88%. The increased positive pick-up of the SBENZ assay compared to the other immunoassays is most probably due to the difference in the limit of detection (LOD) and the increased cross-reactivity for the low-dose benzodiazepines. Clinical comparison of the SBARB serum assay indicated an increased positive pick-up rate, as confirmed by GC-MS. The diagnostic sensitivities of the SBARB serum application, ADx, and EMIT versus GC-MS were 96%, 65%, and 35%, respectively. The diagnostic specificities were all 100%. The diagnostic sensitivities for the SBARB urine application and BARB versus GC-MS were all 100%, and the diagnostic specificities were all 91%. The SBENZ and SBARB kits demonstrated increased sensitivity for the detection of benzodiazepines and barbiturates in both serum and urine compared to the other immunoassays.

Barbiturates↗