Resolution-dependent estimates of multiple sclerosis lesion loads.

BACKGROUND: Changes in brain lesion loads assessed with magnetic resonance imaging obtained at 1.5 Telsa (T) are used as a measure of disease evolution in natural history studies and treatment trials of multiple sclerosis. METHODS: A comparison was made between the total lesion volume and individual lesions observed on 1.5 T images and on high-resolution 4 T images. Lesions were quantified using a computer-assisted segmentation tool. RESULTS: There was a 46% increase in the total number of lesions detected with 4 T versus 1.5 T imaging (p < 0.005). The 4 T also showed a 60% increase in total lesion volume when compared with the 1.5 T (p < 0.005). In several instances, the 1.5 T scans showed individual lesions that coalesced into larger areas of abnormality in the 4 T scans. The relationship between individual lesion volumes was linear (slope 1.231) showing that the lesion volume observed at 4 T increased with the size of the lesion detected at 1.5 T. The 4 T voxels were less than one quarter the size of those used at 1.5 T and there were no consistent differences between their signal-to-noise ratios. CONCLUSIONS: The increase in signal strength that accompanied the increase in field strength compensated for the loss in signal amplitude produced by the use of smaller voxels. This enabled the acquisition of images with improved resolution, resulting in increased lesion detection at 4 T and larger lesion volumes.

Agonistic behavior in groups of limbic epileptic male rats: pattern of brain damage and moderating effects from normal rats.

Different ratios of normal male rats and male rats in which limbic seizures had been induced by a single systemic injection of lithium and pilocarpine were housed in groups of six. The group ratios ranged along the continuum from all normal rats to all experimental rats. The average numbers of episodes of boxing, biting and mounting--thrusting per rat per hour per group were recorded by direct observation (red light) for 1 h during the midscotophase. Groups that contained less than two normal rats exhibited significantly elevated amounts of agonistic (boxing, biting) behavior but not mounting behavior. Multiple regression analyses showed that combinations of neuronal loss within only two to three areas accommodated at least 50% of the variance in the numbers of these behaviors.

Normal spatial memory following postseizure treatment with ketamine: selective damage attenuates memory deficits in brain-damaged rodents.

Within 30 min after the initiation of status epilepticus (SE) by lithium and pilocarpine, rats were injected with either acepromazine or ketamine. Compared to the rats that had received the acepromazine, the group that had received the ketamine displayed more accurate spatial memory. Their scores did not differ significantly from normal (non-seized) controls. Although the ketamine treatment did not significantly change the amount of neuronal loss within about 100 Paxinos and Watson structures, it was neuroprotective for several structures within the thalamus and portions of the temporal and parietal cortices. Ketamine-treated rats, however, displayed markedly more damage within the entorhinal cortices and amygdalohippocampal area.

Comparison of four automated hematology analyzers.

OBJECTIVE: To compare four automated hematology analyzers for efficiency and sensitivity. DESIGN: Four automated hematology analyzers were compared in a side by side study: Bayer ADVIA 120 (Bayer Diagnostic Division, Tarrytown, NY), Beckman Coulter GEN S (Beckman Coulter, Brea, CA), Abbott CELL DYN 3500 and CELL DYN 4000 (Abbott Diagnostics, Santa Clara, CA). 164 specimens were analyzed for cell counts, indices, and the automated WBC differential (DLC). Tallies were kept of all interventions, defined as any parameter necessitating examination of a stained blood smear by a clinical laboratory scientist. A 400-cell manual differential was performed on each specimen and used as the reference to prepare truth tables for each type of WBC. PATIENTS: Specimens comprised regular runs from this tertiary care teaching hospital. These included inpatients, outpatients, and oncology patients, including bone marrow transplant patients. MAIN OUTCOME MEASURES: Results from the truth tables were used for calculating sensitivity and efficiency for each analyzer. Each DLC parameter was analyzed for variance using the one-way ANOVA test. RESULTS: No intervention was required for 103 of 164 specimens for the CELL DYN 3500; the ADVIA gave 70 reportable DLCs without intervention, the GEN S provided 91 and the CELL DYN 4000 resulted in 117 of 164 DLCs without intervention. Agreement or efficiency was 65% for the CELL DYN 3500, 41% for the ADVLA, 58% for the GEN S, and 79% for the CELL DYN 4000. Sensitivity was 67% for the CELL DYN 3500, 86% for the ADVIA, 76% for the GEN S, and 71% for the CELL DYN 4000. Probability of significant variation was as follows for each parameter: % neutrophil 0.8747, % lymphocyte 0.8830, % monocyte 0.0296, % eosinophil 0.7903, and % basophil <.0001. CONCLUSION: The analyzers tested were acceptable for routine laboratory work. Selection would depend on individual need with respect to sensitivity and efficiency. The clinical significance of disagreement between the DLC and the manual differential remains to be determined.

Suppression of experimental allergic encephalomyelitis is specific to the frequency and intensity of nocturnally applied, intermittent magnetic fields in rats.

Female Lewis rats (n=72) were inoculated with an emulsion of spinal cord and complete Freund's adjuvant. They were then exposed for approximately 6 min every hour between midnight and 08:00 h for 2 weeks to either 7 or 40 Hz amplitude-modulated magnetic fields whose temporal pattern was designed to simulate a (geomagnetic) storm sudden commencement. The peak strengths of the fields averaged between either 30-50 nT (low intensity) or 500 nT (high intensity). Rats exposed to the 7 Hz, low intensity magnetic fields displayed significantly less severe overt signs of experimental allergic encephalomyelitis than rats exposed to either of the two intensities of the 40 Hz fields, the high intensity 7 Hz field, or the reference (<10 nT) condition. The latter groups did not differ significantly from each other. Predicted severity based upon the numbers of foci of infiltrations of lymphocytes within the brains of the rats also demonstrated the ameliorating effects of the low intensity, 7 Hz exposures. These results suggest very specific characteristics of complex, weak magnetic fields within the sleeping environment could affect the symptoms of autoimmunity.

Differential effects of low frequency, low intensity (<6 mG) nocturnal magnetic fields upon infiltration of mononuclear cells and numbers of mast cells in Lewis rat brains.

Immediately after inoculation to induce experimental allergic encephalomyelitis, 64 female Lewis rats were exposed to either a reference condition (<10 nT) or to one of two frequencies (7 Hz, 40 Hz) of magnetic fields whose two intensities (either 50 nT or 500 nT) were amplitude-modulated for 6 min once per hour between midnight and 8 h for 15 nights. Rats that had been exposed to the 7 Hz, low intensity fields displayed fewer numbers of foci of infiltrations of mononuclear cells compared to all other groups that did not differ significantly from each other. Rats exposed to the 5 mG (500 nT), 40 Hz magnetic fields displayed more foci in the right thalamus while those exposed to the 5 mG, 7 Hz fields displayed more foci in the left thalamus. Numbers of mast cells within the thalamus were also affected by the treatments. These results suggest that weak magnetic fields can affect the infiltration of immunologically responsive cells and the presence of mast cells in brain parenchyma. Implications for the potential etiology of 'electromagnetic sensitivity' symptoms are discussed.

Suppression of experimental allergic encephalomyelitis in rats exposed nocturnally to magnetic fields.

After inoculation with spinal cord and complete Freund's adjuvant, female Lewis rats were exposed to weak, 7 Hz complex magnetic fields or to the control condition. The computer-generated magnetic field, whose amplitude varied from 15 nT to 60 nT every 6 to 12 sec, was presented for 6 min every hour between midnight and 0800 of the scotophase. In two replicates of the experiment, the rats exposed to the field displayed statistically significant suppression relative to sham-field controls (effect size = 55%) of the overt symptoms of experimental allergic encephalomyelitis which included hindleg paralyses.

Ventricular enlargement and the lithium/pilocarpine seizure model: possible explanation for agonistic behaviour.

Rats treated with the NMDA-blocker, ketamine hydrochloride 30 minutes after the induction of seizures by lithium pilocarpine exhibited statistically smaller lateral ventricles in the left hemisphere compared to rats that had received acepromazine after the induction of these seizures. In addition, the ketamine-treated rats had more neurons and glial cells surrounding the ventricles. These results suggest a neuroprotective effect of ketamine, such that there is less atrophy surrounding the ventricles and therefore, a smaller degree of dilatation. The possibility that insidious neuronal atrophy and death associated with the ventricular enlargement encourages the marked aggression in the epileptic rats not treated by ketamine is discussed.

Fulminant CNS perivascular lymphocytic proliferation: association with sargramostim, a hematopoietic growth factor.

Sargramostim (GM-CSF) therapy was instituted in a 49-year-old woman with hepatitis C on chronic interferon alpha-2b therapy. Within two weeks, she developed progressive confusion, lethargy, and gait disturbance. At autopsy 4 months later, diffuse perivascular nonmonoclonal lymphoid infiltrates were demonstrated throughout the central nervous system (CNS). As the use of hematopoietic growth factors in clinical practice increases, potential adverse effects, such as the fulminant CNS lymphocytic proliferation in this patient, are more likely to be encountered.

The Schizosaccharomyces pombe hst4(+) gene is a SIR2 homologue with silencing and centromeric functions.

Although silencing is a significant form of transcriptional regulation, the functional and mechanistic limits of its conservation have not yet been established. We have identified the Schizosaccharomyces pombe hst4(+) gene as a member of the SIR2/HST silencing gene family that is defined in organisms ranging from bacteria to humans. hst4Delta mutants grow more slowly than wild-type cells and have abnormal morphology and fragmented DNA. Mutant strains show decreased silencing of reporter genes at both telomeres and centromeres. hst4(+) appears to be important for centromere function as well because mutants have elevated chromosome-loss rates and are sensitive to a microtubule-destabilizing drug. Consistent with a role in chromatin structure, Hst4p localizes to the nucleus and appears concentrated in the nucleolus. hst4Delta mutant phenotypes, including growth and silencing phenotypes, are similar to those of the Saccharomyces cerevisiae HSTs, and at a molecular level, hst4(+) is most similar to HST4. Furthermore, hst4(+) is a functional homologue of S. cerevisiae HST3 and HST4 in that overexpression of hst4(+) rescues the temperature-sensitivity and telomeric silencing defects of an hst3Delta hst4Delta double mutant. These results together demonstrate that a SIR-like silencing mechanism is conserved in the distantly related yeasts and is likely to be found in other organisms from prokaryotes to mammals.

Infiltration of lymphocytes in the limbic brain following stimulation of subclinical cellular immunity and low dosages of lithium and a cholinergic agent.

This experiment was designed to investigate the hypothesis that single small dosages of lithium (1.5 mEq/kg), the muscarinic agent pilocarpine (15 mg/kg) and spinal cord emulsion encourage perivascular infiltration of lymphocytes into the brain even when overt symptoms of experimental allergic encephalomyelitis are not apparent. The brains of rats that had received this small dosage of lithium and pilocarpine exhibited discernable infiltrations of lymphocytes within limbic tracts but no discernable neuronal loss. Although the brains of the rats that displayed overt seizures following larger dosages of lithium (3 mEq/kg) and pilocarpine (30 mg/kg) exhibited the usual pattern of neuronal loss within multiple thalamic and limbic structures and conspicuous foci of lymphocytic infiltration (particularly within the hippocampal formation) the correlation between the numbers of foci and the proportions of neuronal damage in these structures was not significant statistically. These results indicate that infiltrations of lymphocytes into brain parenchyma are not simple artifacts of the amount of neuronal damage and may be sensitive toxicological markers for subclinical interactions between drugs and immune responses.

The conserved core of a human SIR2 homologue functions in yeast silencing.

Silencing is a universal form of transcriptional regulation in which regions of the genome are reversibly inactivated by changes in chromatin structure. Sir2 (Silent Information Regulator) protein is unique among the silencing factors in Saccharomyces cerevisiae because it silences the rDNA as well as the silent mating-type loci and telomeres. Discovery of a gene family of Homologues of Sir Two (HSTs) in organisms from bacteria to humans suggests that SIR2's silencing mechanism might be conserved. The Sir2 and Hst proteins share a core domain, which includes two diagnostic sequence motifs of unknown function as well as four cysteines of a putative zinc finger. We demonstrate by mutational analyses that the conserved core and each of its motifs are essential for Sir2p silencing. Chimeras between Sir2p and a human Sir2 homologue (hSir2Ap) indicate that this human protein's core can substitute for that of Sir2p, implicating the core as a silencing domain. Immunofluorescence studies reveal partially disrupted localization, accounting for the yeast-human chimeras' ability to function at only a subset of Sir2p's target loci. Together, these results support a model for the involvement of distinct Sir2p-containing complexes in HM/telomeric and rDNA silencing and that HST family members, including the widely expressed hSir2A, may perform evolutionarily conserved functions.

"Subclinical" dosages of lithium and pilocarpine that do not evoke overt seizures affect long-term spatial memory but not learning in rats.

After training is an automated radial maze, 11 male rats were injected with either "subclinical" dosages of lithium and pilocarpine or saline and then tested 5 days or 4 months later. When employed as their own controls or when compared with a saline-injected reference group, the rats that had received the lithium and pilocarpine displayed memory deficits but not learning learning deficits after the longest of the two delays (effect size was 41%). These results suggest that subtle disruption in memory but not learning to criterion could be associated with "subclinical electrical seizures" or the micromorphological changes associated with this activity.

Emergent properties following brain injury: the claustrum as a major component of a pathway that influences nociceptive thresholds to foot shock in rats.

Flinch (pain) thresholds for electric current delivered to the feet were correlated with the amount of necrosis within the diencephalon and telencephalon for rats in which seizures had been induced by lithium and pilocarpine about two months before the testing. The shared variance of the quantitative damage within the claustrum, the anterior part of the paraventricular nucleus of thalamus, (central) mediodorsal thalamus, and lateral amygdala (ventromedial part) explained 81% of the variance in the nociceptive (flinch) thresholds. A primary role of the claustrum within the neuropathways that mediate the response to the interoceptive and "painful" characteristics of stimuli is indicated. The concept of primary pathways versus "emergent" pathways subsequent to excitotoxic damage within the neuromatrix is discussed.

Long-term consequences of subtle stimuli during the first twenty-four hours of seizure-induced brain injury.

Chronically epileptic (induced by a single systemic injection of lithium and pilocarpine about 30 days before the experiment began) male rats were trained within a radial maze while they were administered either GABA-pentin (Neurontin), or prednisolone or given no treatment. There was no significant improvement in learning or memory between the groups. Numbers of trials per day were positively correlated with the time required to display the overt stereotyped forelimb clonus after the single pilocarpine injection. The numbers of correct trials completed during the first few days of acquisition were significantly greater for the rats that had receive weak (1 microT) complex, pulsed magnetic fields over the right hemisphere during the first 24 hr. after seizure induction than for those who received the same field over the left hemisphere or that had been exposed to reference conditions. Implications of the enhanced sensitivity of limbic neurons to subtle electromagnetic interaction during electrical lability are discussed.

Demands during maze. learning in limbic epileptic rats: selective damage in the thalamus?

A qualitatively evident enhancement of chromolytic neurons within the lateral posterior thalamus of rats in which limbic seizures had been induced by lithium and pilocarpine and who were later trained for spatial memory was assessed quantitatively. The significant increase in the numbers of chromolytic neurons and the decrease in the numbers of normal neurons for these rats compared to the reference brains suggested these morphological changes were recent. The hypothesis that excessive stimulation of the lateral posterior nucleus by daily training in a radial maze may have facilitated the necrosis was supported by the inverse relationship between a linear combination of the numbers of normal neurons and oligodendroglia and the rate of learning during the earlier but not the later sessions. An implication for iatrogenic effects from rehabilitation of humans following brain injury was suggested.

Interhemispheric modulation of dopamine receptor interactions in unilateral 6-OHDA rodent model.

A critical assumption in the unilateral 6-hydroxydopamine (6-OHDA) model is that interactions between the intact and denervated hemispheres do not influence the response to insult. The present study examined this issue by assessing the effects of unilateral substantia nigra 6-OHDA lesions in rats that previously had received corpus callosum transections, a treatment designed to minimize interhemispheric influences. Quantitative autoradiography in the caudate-putamen ipsilateral to the lesion revealed that corpus callosum transection did not alter the increase in D2-like receptors ([125I]-epidepride-labeled sites) that is induced by unilateral 6-OHDA lesion. There were no effects of either 6-OHDA lesion or transection on D1 receptor density ([125I]-SCH23982 autoradiography). As a functional endpoint, dopamine-stimulated cAMP efflux was measured in superfused striatal slices. In this paradigm, the net effect of dopamine (DA) represents a combination of D1 receptor-mediated stimulation and D2 receptor-mediated inhibition. 6-OHDA lesion increased cAMP efflux induced by exposure to 100 microM DA alone; corpus callosum transection did not alter this effect. An interaction between 6-OHDA lesion and transection status was revealed, however, by comparison of results obtained with DA alone vs. DA plus the D2 antagonist sulpiride (to block the D2 inhibitory effects of 100 microM DA). This comparison revealed two important effects of 6-OHDA lesion in rats with an intact corpus callosum: 1) a moderate decrease in dopamine D1 receptor-mediated stimulation; and 2) a dramatic decrease in the ability of D2 receptors to inhibit this stimulation. Corpus callosum transection prevented these effects of 6-OHDA. These results provide a biochemical demonstration of D1:D2 receptor uncoupling in unilateral 6-OHDA lesioned rats, and suggest that interhemispheric influences (e.g., contralateral cortico-striatal glutamatergic projections) may contribute to lesion-induced alterations in D1:D2 receptor interactions.

A role for recombination junctions in the segregation of mitochondrial DNA in yeast.

In S. cerevisiae, mitochondrial DNA (mtDNA) molecules, in spite of their high copy number, segregate as if there were a small number of heritable units. The rapid segregation of mitochondrial genomes can be analyzed using mtDNA deletion variants. These small, amplified genomes segregate preferentially from mixed zygotes relative to wild-type mtDNA. This segregation advantage is abolished by mutations in a gene, MGT1, that encodes a recombination junction-resolving enzyme. We show here that resolvase deficiency causes a larger proportion of molecules to be linked together by recombination junctions, resulting in the aggregation of mtDNA into a small number of cytological structures. This change in mtDNA structure can account for the increased mitotic loss of mtDNA and the altered pattern of mtDNA segregation from zygotes. We propose that the level of unresolved recombination junctions influences the number of heritable units of mtDNA.

Dihydrexidine, a novel full efficacy D1 dopamine receptor agonist.

The present work provides a detailed pharmacological characterization of dihydrexidine (DHX) (trans-10,11-dihydroxy- 5,6,6a,7,8,12b-hexahydrobenzo[a]phenanthridine), the first high-potency, full efficacy, bioavailable D1 dopamine receptor agonist. DHX represents a new conformationally rigid structural class of dopamine receptor ligands. It competes stereoselectively and potently for D1 binding sites in rat striatal membranes labeled with [3H]SCH23390 with an IC50 of about 10 nM compared to about 30 nM for the prototypical D1 agonist SKF38393. Like other dopamine agonists, DHX has a shallow competition curve (nH = ca. 0.7) that can be fitted by a two-site model consisting of high-affinity (63%; KD = 3 nM) and low-affinity (37%; KD = 75 nM) sites. DHX was screened for activity against 40 other binding sites, and was inactive (IC50 greater than 10 microM) against all except D2 dopamine receptors (IC50 = 130 nM) and alpha 2 adrenoreceptors (IC50 = ca. 230 nM). Functionally, DHX is a full efficacy dopamine D1 agonist. In homogenates of rat striatum, DHX or dopamine doubles the rate of cyclic AMP synthesis, whereas SKF38393 only causes a maximal increase of about 50%. These effects of DHX are blocked by the selective D1 antagonist SCH23390, but are not affected by D2, 5-hydroxytryptamine2, muscarinic, or alpha or beta adrenergic antagonists. Because DHX is known to cause D2-like behavioral effects at high doses, the nature of its D2 activity was characterized using prolactin release as an end-point. DHX and the prototypical D2 agonist quinpirole both caused a significant inhibition of the prolactin release induced by 5-hydroxytryptophan. These effects of DHX are not due to "indirect" alterations at the presynaptic terminal, because DHX is essentially inactive at inhibiting the dopamine uptake system, and does not cause the release of dopamine. These data demonstrate the utility of DHX for probing the biochemistry and function of D1 dopamine receptors.