Long-term behavioral characterization of a rat model of intracerebral hemorrhage.

We tested the behavioral effects of intracerebral hemorrhage (ICH) in adult male rats. ICH was induced by collagenase injection into the basal ganglia and the rats were subjected to a longitudinal behavioral test battery. Both learning and memory deficits were detected shortly after injury. Two months after injury, there were still significant short- and long-term memory deficits. Rotarod testing also revealed long-term sensorimotor coordination deficits. No differences in activity levels were detected at any time. Thus, spontaneous ICH produced detectable cognitive and motor deficits that evolved over the course of 2 months. Along with histological analysis of infarct volume, this characterization provides a suitable baseline for the analysis of therapeutic interventions.

Long-term effects of melatonin after intracerebral hemorrhage in rats.

Free radical scavengers have been shown to improve short-term outcome after intracerebral hemorrhage (ICH). The purpose of this study was to evaluate whether melatonin (a potent free radical scavenger and an indirect antioxidant) can improve short- and/or long-term neurological function after ICH, which was induced by collagenase injection into the striatum of adult rats. Melatonin (15 mg/kg) was administered by intraperitoneal injection at 1, 24, 48, and 72 h. Neurological and behavioral testing was performed at several time points from 1 day to 8 weeks post-ICH. Neurological and behavioral deficits were observed in ICH rats at all time points, but the melatonin treatment regimen did not improve performance or level of brain injury.

Altered stress-induced anxiety in adenylyl cyclase type VIII-deficient mice.

Stress results in alterations in behavior and physiology that can be either adaptive or maladaptive. To define the molecular pathways involved in the response to stress further, we generated mice deficient (KO) in the calcium-stimulated adenylyl cyclase type VIII (AC8) by homologous recombination in embryonic stem cells. AC8 KO mice demonstrate a compromise in calcium-stimulated AC activity in the hippocampus, hypothalamus, thalamus, and brainstem. Hippocampal slices derived from AC8 KO mice fail to demonstrate CA1-region long-term depression after low-frequency stimulation, and AC8 KO mice also fail to activate CRE-binding protein in the CA1 region after restraint stress. To define the behavioral consequences of AC8 deficiency, we evaluated AC8 KO mice in the elevated plus-maze and open field. Although naive AC8 KO mice exhibit indices of anxiety comparable with that of wild-type mice, AC8 KO mice do not show normal increases in behavioral markers of anxiety when subjected to repeated stress such as repetitive testing in the plus-maze or restraint preceding plus-maze testing. These results demonstrate a novel role for AC8 in the modulation of anxiety.

A single day of ethanol exposure during development has persistent effects on bi-directional plasticity, N-methyl-D-aspartate receptor function and ethanol sensitivity.

To determine factors that contribute to the learning deficits observed in individuals with fetal alcohol syndrome, we examined the effects of early postnatal ethanol exposure on forms of synaptic plasticity thought to underlie memory. Treatment of rat pups with ethanol on postnatal day 7 impaired the induction of N-methyl-D-aspartate receptor-dependent long-term potentiation and abolished homosynaptic long-term depression in the CA1 region of hippocampal slices prepared at postnatal day 30. An N-methyl-D-aspartate receptor-independent form of long-term potentiation induced by very high frequency stimulation could be induced in slices from ethanol-treated rats. Defects in long-term depression correlated with a diminished contribution of ifenprodil-sensitive N-methyl-D-aspartate receptors to synaptic transmission and defects in a spontaneous alternation behavioral task. Rats exposed to ethanol on postnatal day 7 also exhibited diminished sensitivity of synaptic N-methyl-D-aspartate receptors to block by ethanol at postnatal day 30 and decreased behavioral sedation to systemic ethanol injections. These results indicate that changes in synaptic plasticity and N-methyl-D-aspartate receptor function are likely to provide a neural substrate for the cognitive and behavioral changes that follow developmental ethanol exposure.

Deletion of the N-terminus of murine map2 by gene targeting disrupts hippocampal ca1 neuron architecture and alters contextual memory.

Microtubule-associated protein-2 (MAP2) is a brain specific A-kinase anchoring protein that targets the cyclic AMP-dependent protein kinase holoenzyme (PKA) to microtubules. Phosphorylation of MAP2 by different protein kinases is crucial for neuronal growth. The N-terminus of MAP2 contains the binding site for regulatory subunit II of cAMP-dependent protein kinase (PKA-RIIbeta). Using homologous recombination, we created a mutant line of mice (delta1-158) that express truncated MAP2 lacking the N-terminal peptide and the PKA binding site. Deletion of the PKA binding site from the MAP2 gene resulted in decreased efficiency of MAP2 phosphorylation. Biochemical and immunohistochemical studies demonstrate major changes in the morphology of hippocampal neurons in delta1-158 mice. Behavioral tests indicate that delta1-158 mice were impaired (exhibited less conditioned freezing) relative to Wild-Type (WT) controls during a test of contextual, but not during auditory cue, fear conditioning when tested at 8 weeks or 8 months of age. The delta1-158 mice displayed a heightened sensitivity to shock at 8 weeks, but not at 8 months of age. We conclude that PKA binding to MAP2 and MAP2 phosphorylation is essential for the selective development of contextual memory.

Carbon dioxide fixation by extracts of Streptococcus faecalis var. liquefaciens.

Extracts of cells of Streptococcus faecalis var. liquefaciens strain 31 incorporated (14)CO(2) into aspartate. Dialyzed extracts produced radioactive oxalacetate in the absence of exogenously added glutamate and pyridoxal-5'-phosphate and produced radioactive aspartate in the presence of these components. Reduced nicotinamide adenine dinucleotide or reduced nicotinamide adenine dinucleotide phosphate could not be substituted for adenosine triphosphate (ATP); phosphoenolpyruvate even in the presence of nucleoside diphosphates could not replace pyruvate plus ATP; propionate plus coenzyme A (CoA) could not replace pyruvate in supporting CO(2) fixation by cell extracts. Fixation by dialyzed cell extracts required pyruvate, ATP, MgSO(4), and was stimulated by biotin, KCl, 2-mercaptoethanol, CoA, and acetyl CoA. Inhibition of fixation occurred when avidin, NaCl, oxalacetate, or aspartate was added to dialyzed extracts. On the basis of the products formed and the effects of substrates and cofactors on the fixation reaction, it was concluded that pyruvate carboxylase is responsible for CO(2) fixation in this microorganism.

Carbon dioxide fixation in Helminthosporium cynodontis.

A mycelial suspension of Helminthosporium cynodontis (ATCC24938), grown on glucose-peptone-yeast extract broth and exposed to NaH14CO3 for 5 h, fixed significant quantities of 14C into the following fractions (%): small molecular weight components, 7-4; lipid and lipoproteins, 3-9; nucleic acids, 59; the residual protein and cell wall fragments, 29-2. The labelled protein components were (%): aspartate, 39; glutamate, 18; cystine, 15; threonine, 9. Radioactive nucleic acid components were (%): adenine, 18; guanine, 18; cytidylate, 34; uridylate, 30. When the mycelium was grown in Czapek-Dox glucose medium and incubated in this medium plus NaH14CO3, the nucleic acid fraction contained 29-9% and the residual protein 49-5% of the cellular radioactivity. The removal of CO2 from the atmosphere did not reduce growth. Pyruvate carboxylase (PC) and phosphoenolypyruvate carboxykinase (PEPCK) activities were demonstrated in extracts of H. cynodontis. Synthesis of PEPCK was stimulated under conditions promoting gluconeogenesis and was reduced under conditions promoting glycolysis, while PC synthesis was similar under both conditions.

Carbon dioxide fixation by cells of Streptococcus faecalis var. liquefaciens.

Fixation of NaH(14)CO(3) by a heavy cell suspension of Streptococcus faecalis var. liquefaciens was studied. Several nutrients, pyridoxal, riboflavine, adenine, uracil, and O(2) stimulated (14)CO(2) incorporation into cells only under conditions that were adequate for synthesis of cell macromolecules. Biotin increased CO(2) incorporation in the absence of extensive synthesis of macromolecules, whereas O(2) inhibited incorporation under these conditions. When (14)CO(2) fixation was occurring during synthesis of macromolecules, 71% of the (14)C was incorporated into cells and 29% occurred extracellularly. Ninety-three per cent of the cellular (14)C was in protein and 5.5% was in nucleic acid. Aspartic acid was the only amino acid in the protein fraction that was radioactive. Eighty-three per cent of the extracellular (14)C was resistant to precipitation by trichloroacetic acid. When (14)CO(2) fixation was occurring in cells that were not carrying on extensive synthesis of macromolecules, 38% of the (14)C was incorporated into cells and 59% occurred in the supernatant fluid. Sixty-nine per cent of the cellular (14)C was in protein, 21% was in low-molecular-weight compounds, and 9% was in nucleic acid. Addition of unlabeled aspartate to the medium inhibited incorporation of (14)CO(2). Based on studies of the rate of (14)CO(2) fixation, the cells fix CO(2) into a pool of intermediates which are either used for synthesis, primarily protein, or are excreted into the medium.

Effect of carbon dioxide on the aspartic acid requirement for proteinase biosynthesis by Streptococcus faecalis var. liquefaciens.

Non-proliferating cells of Streptococcus faecalis var. liquefaciens required aspartic acid for proteinase biosynthesis in the absence of CO(2) but not in the presence of CO(2).

Evaluation of the efficacy of minocycline therapy for staphylococcal soft-tissue infection.

Ten patients with soft-tissue infections due to Staphylococcus aureus were treated with minocycline, a semisynthetic tetracycline with potent in vitro antistaphylococcal effects. Serum concentrations averaged three to five times the concentration of minocycline required to inhibit growth of S. aureus in vitro. Clearing of the infecting organism was slow (less than 50% of lesions were sterile on day 10 of therapy), but clinical improvement was noted in 8 of 10 patients.

Behavioral phenotyping of GFAP-apoE3 and -apoE4 transgenic mice: apoE4 mice show profound working memory impairments in the absence of Alzheimer's-like neuropathology.

For the purpose of studying the potential neurobehavioral effects of different human apolipoprotein E (apoE) isoforms produced within the brain, transgenic (TG) mice were generated in which human apoE3 or apoE4 isoforms were under control of an astrocyte-specific, glial fibrillary acidic protein promoter and these TG mice were bred back to apoE knockout (KO) mice. Behavioral phenotypes of apoE3 and apoE4 TG mice were derived by conducting a longitudinal study in which apoE3 and apoE4 TG mice were compared with apoE KO and wild-type (WT) mice (all male) on several behavioral measures. Analysis of locomotor activity, "open-field" behaviors, acoustic startle/prepulse inhibition, and elevated plus maze data suggested that the apoE TG/KO groups were more "emotionally reactive" than WT mice, with apoE4 mice typically being the most reactive. The absence of performance differences among groups on the rotating holeboard and water navigation tasks suggested intact reference memory processing in apoE TG/KO mice. However, apoE4 mice were profoundly impaired on a working memory-based protocol in the radial arm maze (11-14 months). Nonassociative factors (sensorimotor capacities or emotionality differences) did not appear to confound interpretation of the learning/memory results. Western blot analysis revealed no alterations in the level of synaptic, neuronal, or glial markers in neocortex or hippocampus and histologic analysis revealed no evidence of Abeta deposition or neuritic plaques in the apoE KO/TG mice. Our findings suggest that apoE4 expression in the brain may have selective deleterious effects on memory function in the absence of typical Alzheimer's-like neuropathology.