Morphology and electrophysiology of dentate granule cells in the rhesus monkey: comparison with the rat

The morphologic and electrophysiologic properties of dentate granule cells in the young adult rhesus monkey (Macaca mulatta) were examined for the first time with whole-cell patch clamp recordings and intracellular biocytin filling in in vitro hippocampal slice preparations. Data from monkeys were compared with data generated in an identical manner from adult Sprague-Dawley rats. Intracellularly filled monkey and rat granule cells were identical in numerous morphologic parameters, including area of somata, total dendritic length, dendritic spread, segment number and length, and branching pattern. The single statistically significant difference in morphology was the vertical extent of the dendritic tree (distance from soma to fissure), which was 20% greater in the monkey. The passive membrane properties (resting membrane potential, input resistance, and membrane time constant) measured under current clamp conditions were virtually identical. The thresholds and amplitudes of action potentials were the same, but significant differences were seen in the kinetics of single action potentials. Monkey granule cell action potentials were significantly longer in duration (with slower rise and fall times) than action potentials in rat cells. These differences were likely due to a much smaller fast after hyperpolarization in the monkey as compared with the rat cells. Thus, with the exception of action potential properties, the principal finding of this study is that there is significant conservation of both form and function in dentate granule cells in these two species, despite the enormous phylogenetic separation. This suggests that granule cell properties may be extremely stable across diverse mammalian species.     

A mathematical model for the dynamic behavior of melts subjected to electromagnetic forces: Part I. model development and comparison of predictions with published experimental results

A mathematical model was developed to predict electromagnetically driven flow and (particularly) free surface behavior in melts subject to electromagnetic forces. Such melts appear in electromagnetic casters, induction furnaces, and other metal processing units. The calculations started with Maxwell’s equations and Ohm’s law, which were solved by a novel “modified hybrid technique.” The instantaneous continuity and Navier-Stokes equations (rather than their time-averaged versions) were then solved with electromagrretic forces as input. The calculations allowed for the dynamic behavior of the free surface of the melt, and electromagnetic fields were recomputed as the free surface changed. In this first part of a two-part article, the model predictions are compared with the experimental measurements of induced current, magnetic field, melt velocity, and free surface deformation reported by others.     

Ethionine toxicity in vitro: the correlation of data from rat hepatocyte suspensions and monolayers with in vivo observations

The hepato-steatogenic compound ethionine has been used to investigate the correlations between in vivo and in vitro toxicity data. The aim was to find a suitable model of toxicity in hepatocyte suspensions or monolayers in vitro, which could predict the known toxicity of ethionine in vivo and which could be implemented in screening compounds of unknown toxicity. Thus a variety of markers of cytotoxicity, metabolic competence and liver-specific functions were investigated in rat hepatocyte suspensions and monolayers and compared with in vivo data in the rat. The following markers were measured in the appropriate system: (1) Neutral red uptake; 3-(4,5 dimethyl)thiazol-2-yl,-2,5-diphenyl tetrazolium bromide (MTT) reduction; lactate dehydrogenase (LDH), aspartate aminotransferase (AST) and alanine aminotransferase (ALT) leakage (cytotoxicity). (2) ATP levels, protein synthesis and glutathione (GSH) levels (metabolic competence). (3) Urea and triglyceride synthesis and beta-oxidation (liver specific functions). Ethionine (0-30 mM) did not affect the markers of direct cytotoxicity, except neutral red uptake, which was reduced by 18 and 30 mM ethionine after 20 h in culture. ATP and GSH depletion occurred in hepatocyte suspensions at the highest concentrations of ethionine (20 and 30 mM) after 1 h. In monolayers, GSH levels were reduced after 4 h, but not 20 h. Urea synthesis was increased in hepatocyte suspensions from 1 to 3 h by 10-30 mM ethionine and reduced after 20 h in cultured hepatocytes (18-30 mM). Protein synthesis was reduced and beta-oxidation was increased in ethionine-treated hepatocyte suspensions. Unfortunately, there was no measurable effect on triglyceride accumulation within cells (the major biochemical change in vivo) in either system. Ethionine treated hepatocytes in suspension showed the same rate of triglyceride synthesis and transportation out of cells as control cells. Thus, hepatocyte suspensions were able to mimic the early biochemical effects of ethionine in vivo (ATP and GSH depletion, inhibition of protein synthesis) and some effects on urea synthesis, but monolayer cultures appeared to be less sensitive to the toxicity of ethionine. However, neither in vitro system was able to model the effects of ethionine on the accumulation of triglycerides in vivo.     

Organization of common peroneal nerve motoneurones in the rat and monkey: a comparative study

The motoneurones with axons in the common peroneal nerve (CPN) of the rat and monkey were examined using retrograde labelling with horseradish peroxidase (HRP). In both species, the CPN motoneurone pool was localized in the dorsolateral part of the ventral horn of the spinal cord. In the rat, the labelled motoneurones were located between the L3 and L6 spinal segments whereas in the monkey, they extended from the caudal end of L4 to the L6 spinal segments. In both species the majority of the labelled neurones were located within the L5 segment. The mean number of the CPN motoneurones in the rat and monkey was 458 and 1148, respectively. A bimodal size distribution of motoneurones was found in both species.     

Use of precision-cut rat liver slices for studies of xenobiotic metabolism and toxicity: comparison of the Krumdieck and Brendel tissue slicers

1. In this study we have compared freshly cut and cultured precision-cut rat liver slices produced by the Krumdieck and Brendel-Vitron tissue slicers. 2. No significant differences were observed in levels of protein, potassium, total glutathione (i.e. GSH and GSSG), reduced glutathione (GSH) and cytochrome P450 and activities of 7-ethoxyresorufin O-deethylase and 7-benzoxyresorufin O-debenzylase in freshly cut rat liver slices produced by the two tissue slicers. However, levels of oxidized glutathione (GSSG) were significantly greater in liver slices produced with the Brendel-Vitron tissue slicer. 3. Precision-cut rat liver slices produced with both tissue slicers were cultured for 0 (i.e. a 1-h preincubation), 24 and 72 h in a dynamic organ culture system in an atmosphere of either 95% 02/5% CO2 or 95% air/5% CO2. 4. Apart from small differences in glutathione levels in 0 and 24 h cultured liver slices, no significant differences were observed in the parameters measured between liver slices prepared with both tissue slicers and cultured in both gas phases. 5. With liver slices produced by both tissue slicers 50 microM sodium arsenite produced a greater induction of heat shock protein 70 levels in slices cultured for 24 h in a high oxygen than in an air atmosphere. 6. These results suggest that both tissue slicers can readily produce precision-cut liver slices for studies of xenobiotic metabolism and toxicity. However, the data suggest that for any given application of precision-cut tissue slices it is desirable to establish optimal culture conditions     

Effects of xylose on monkey lenses in organ culture: a model for study of sugar cataracts in a primate

The beta subunit of DNA polymerase III is essential for negative regulation of the initiator protein, DnaA. DnaA inactivation occurs through accelerated hydrolysis of ATP bound to DnaA; the resulting ADP-DnaA fails to initiate replication. The ability of beta subunit to promote DnaA inactivation depends on its assembly as a sliding clamp on DNA and must be accompanied by a partially purified factor, IdaB protein. DnaA inactivation in the presence of IdaB and DNA polymerase III is further stimulated by DNA synthesis, indicating close linkage between initiator inactivation and replication. In vivo, DnaA predominantly takes on the ADP form in a beta subunit-dependent manner. Thus, the initiator is negatively regulated by action of the replicase, a mechanism that may be key to effective control of the replication cycle.     

Selective ablation of rat knee joint innervation with injected immunotoxin: a potential new model for the study of neuropathic arthritis

Neuropathic arthropathy is a severe chronic degenerative condition associated with decreased or absent sensory innervation of the involved joint. Existing animal models of neuropathic arthritis are limited by the technical difficulties of obtaining either highly selective or complete joint denervation in a minimally invasive fashion. We undertook experiments to determine the feasibility of using the newly described method of selective neuronal lesioning with injected immunotoxin as a means of creating a more tractable model of neuropathic arthritis. Retrograde tracing with fluorochrome revealed that the knee joint of the female Wistar rat is supplied by 581 +/- 31 (mean +/- SD) joint afferents. Immunohistochemistry confirmed that virtually all sensory neurons in the rat express the cell surface receptor Thy 1. Injection of rat knee joints with an immunotoxin targeted toward Thy 1 resulted in the selective ablation of an average of 88% of the joint afferents identified with fluorochrome that are normally found in the ipsilateral L3 and L4 ganglia.     

Subacute (4-wk) oral toxicity of a combination of four nephrotoxins in rats: comparison with the toxicity of the individual compounds

In a 4-wk study, 10-wk-old Wistar rats were fed the nephrotoxins hexachloro-1,3-butadiene (HCBD), mercuric chloride, d-limonene and lysinoalanine either alone or in combination. These nephrotoxins damage epithelial cells of the proximal tubules, but by different mechanisms. Each chemical was given alone at a Minimum-Nephrotoxic-Effect Level (MNEL), and at a No-Nephrotoxic-Effect Level (NNEL). The combination was given at the MNEL, the NNEL and one-quarter of the NNEL of the individual chemicals. The individual nephrotoxins caused slight growth depression in males at the MNEL, but not at the NNEL, whereas the combination depressed growth slightly at the NNEL and severely at the MNEL. In females at the MNEL, only HCBD retarded growth; in contrast to the effect in males this was not aggravated by combined treatment. Nephrotoxicity was more severe in males fed the combination than in males given the nephrotoxins alone. The former showed decreased renal concentrating ability and moderate histopathological changes in the kidneys at the MNEL, and a dose-dependent increase in kidney weight and number of epithelial cells in the urine at the NNEL and the MNEL. The males treated with a single agent showed slightly increased kidney weights, and/or slight histopathological changes in the kidneys at the MNEL, and (with d-limonene only) epithelial cells in the urine at the NNEL and MNEL. In females, renal changes induced by the combination were not more severe than those observed with individual compounds. No adverse changes attributable to treatment were observed in rats fed the combination at one-quarter of the NNEL. In the present study, combined exposure to four nephrotoxins at their individual NNEL did not constitute an obviously increased hazard, indicating absence of synergistic interaction, whereas at the MNEL clearly enhanced (renal) toxicity occurred in males, although not in females.     

Brain regional substrates for the actions of the novel wake-promoting agent modafinil in the rat: comparison with amphetamine

Modafinil is a novel wake-promoting compound for which the mechanism and sites of action are unknown. We examined the neural substrates in the brain for the actions of modafinil using 2-deoxyglucose autoradiography and compared the findings to those obtained with amphetamine. Modafinil showed a relatively restricted pattern of changes in brain regional metabolic activity, while amphetamine altered glucose utilization in a wide variety of brain regions. Both modafinil and amphetamine increased glucose utilization in all subregions of the hippocampus (subiculum, CA1-CA3 and dentate gyrus) and in the centrolateral nucleus of the thalamus. Modafinil also increased glucose utilization in the central nucleus of the amygdala, but amphetamine had no effect in this region. Brain structures in which amphetamine increased metabolic rate but modafinil had no effect included regions of the basal ganglia, other nuclei of the thalamus, the frontal cortex, the nucleus accumbens, the ventral tegmental area and the pontine reticular fields. These findings suggest that, while both modafinil and amphetamine promote wakefulness, they act via distinctly different mechanisms. Modafinil appears to act on a specific subset of brain pathways which regulate sleep and wakefulness, whereas amphetamine affects a greater number of cerebral structures involved in the regulation of these behavioral states. Modafinil also lacks the pronounced effects on the extrapyramidal motor system which are characteristic of amphetamine and other psychomotor stimulants, implying that the effects of modafinil are not mediated by the dopamine system and that modafinil may selectively increase wakefulness with fewer side effects.     

Gait analysis in the rat as a model for the study of peripheral vascular disease

A number of problems are associated with investigation of the pathophysiology of peripheral vascular disease (PVD) in humans and unilateral ligation of the common iliac artery in the rat has been extensively used as an experimental model of PVD. However, techniques so far applied to monitor the effects of PVD on muscular function are invasive and difficult to relate to natural locomotion. The present study aimed to investigate gait during spontaneous, unforced locomotion in the animal model. No differences were observed between control and experimental rats in velocity or stride lengths. Although stride time was unchanged, its internal balance was disturbed with reduced stance, prolonged swing, for the ligated limb (LL) relative to the nonligated limb (NLL). Gait symmetry was also affected with NLL strike occurring earlier during LL stance. Analysis of footfall patterns showed a reduction of stance area and pressures exerted via LL relative to NLL. It is concluded that gait and footfall analysis could be useful for assessing limb function under natural exercise conditions.     

Trigeminal-reticular connections: possible pathways for nociception-induced cardiovascular reflex responses in the rat

Cardiovascular regulatory neurons of the ventral medulla and pons are thought to have an important role in the mediation of trigeminal nociception-induced reflex cardiovascular responses. However, the neural pathways that link the spinal trigeminal nucleus with ventral medullary and pontine autonomic cell groups are poorly understood. The present study utilized injections of the highly sensitive anterograde tracer substance biotinylated dextran combined with immunocytochemistry for tyrosine hydroxylase, the synthesizing enzyme for catecholamines, to investigate the distribution and morphology of projections from the spinal trigeminal subnucleus caudalis to ventral medullary and pontine catecholaminergic cell groups. Injection of biotylinated dextran into the dorsal subnucleus caudalis produced dense anterograde labeling in dorsal regions of the medullary and pontine reticular formation including the dorsal medullary reticular field, the parvicellular reticular field, and the parvicellular reticular field pars anterior. In the ventral medullary and pontine reticular formation, light anterograde labeling tended to be distributed in close proximity to the distal dendrites of catecholaminergic neurons located in the C1, A1, and A5 regions. Injections of anterograde tracer into the dorsal medullary reticular field produced dense anterograde labeling in the ventral medullary and pontine reticular formation. Numerous terminal-like varicosities were observed in close proximity to catecholaminergic neurons located in the C1, A1, and A5 regions. These data suggest that trigeminal pain-induced reflex cardiovascular responses involve indirect projections that terminate in the dorsal medullary and pontine reticular formation before reaching ventral medullary and pontine catecholaminergic cell groups known to be involved in cardiovascular regulation.