Muscimol increases acetylcholine release by directly stimulating adult striatal cholinergic interneurons.

Because GabaA ligands increase acetylcholine (ACh) release from adult striatal slices, we hypothesized that activation of GabaA receptors on striatal cholinergic interneurons directly stimulates ACh secretion. Fractional [3H]ACh release was recorded during perifusion of acutely dissociated, [3H]choline-labeled, adult male rat striata. The GabaA agonist, muscimol, immediately stimulated release maximally approximately 300% with EC50 = approximately 1 microM. This action was enhanced by the allosteric GabaA receptor modulators, diazepam and secobarbital, and inhibited by the GabaA antagonist, bicuculline, by ligands for D2 or muscarinic cholinergic receptors or by low calcium buffer, tetrodotoxin or vesamicol. Membrane depolarization inversely regulated muscimol-stimulated secretion. Release of endogenous and newly synthesized ACh was stimulated in parallel by muscimol without changing choline release. Muscimol pretreatment inhibited release evoked by K+ depolarization or by receptor-mediated stimulation with glutamate. Thus, GabaA receptors on adult striatal cholinergic interneurons directly stimulate voltage- and calcium-dependent exocytosis of ACh stored in vesamicol-sensitive synaptic vesicles. The action depends on the state of membrane polarization and apparently depolarizes the membrane in turn. This functional assay demonstrates that excitatory GabaA actions are not limited to neonatal tissues. GabaA-stimulated ACh release may be prevented in situ by normal tonic dopaminergic and muscarinic input to cholinergic neurons.

Inter- and intraindividual variability of the response to intravenous glucose tolerance testing in cats.

OBJECTIVE: To evaluate inter- and intraindividual variation in results of the intravenous glucose tolerance test in cats. ANIMALS: 19 healthy specific-pathogen-free-derived cats were allotted to group A (n = 13), which was accustomed, and group B (n = 6), which was unaccustomed to having blood drawn. PROCEDURE: Blood samples were collected for glucose and insulin assays before and 5, 10, 15, 30, 45 and 60 minutes after i.v. administration of 500 mg of dextrose/kg of body weight. Glucose half-life (t1/2) and disappearance co-efficient (K), and the acute-phase insulin response (Ins0-10) were calculated. Inter- and intraindividual variability was assessed by calculating the coefficient of variation for test variables. RESULTS: Comparing the 2 tests, there were no significant differences in glucose and insulin concentrations prior to dextrose administration or in t1/2, K, or Ins0-10. However, compared with group-A cats, cats in group B had significantly (P < 0.05) longer t1/7 and lower K and Ins0-10 values, which was attributed to increased stress in these cats. Overall, the interindividual variability was 62.8% for K, 54.6% for t1/2, and 76.0% for Ins0-10. Mean intraindividual variability was 32.0 (range, 0.1 to 72.0)% for K and t1/2, and 45.8 (range, 4.0 to 179.5)% for Ins0-10. There was only a moderate correlation in results between the 2 tests (rs = 0.59 for t1/2 and K, rs = 0.58 for Ins0-10). CONCLUSION: The variability in results of intravenous glucose tolerance tests in cats suggests caution is necessary in interpreting results of a single test in individuals.

Cellular resistance to oxidative stress is accompanied by resistance to cisplatin: the significance of increased catalase activity and total glutathione in hydrogen peroxide-resistant fibroblasts.

Studies designed to better understand the involvement of cellular resistance to oxidative stress in mechanisms of cellular resistance to cisplatin were undertaken using H2O2-resistant variants of the HA1 Chinese hamster fibroblast cell line. H2O2-resistant cell lines were resistant to clonogenic inactivation mediated by cisplatin with dose modifying factors at 10% survival of 1.5-3.0, relative to HA1 cells. The most cisplatin resistant of these cell lines (OC5) also demonstrated fewer DNA-DNA crosslinks induced by cisplatin, relative to HA1. Since H2O2-resistant cells contained increased catalase activity as well as total glutathione (GSH) content, the involvement of these cellular antioxidants in the resistance to cisplatin toxicity was evaluated. Treatment of HA1 and H2O2-resistant cell lines (OC5, OC14) with 9 mM aminotriazole reduced catalase activity by 60-65% but had no effect on the cytotoxicity of cisplatin. In contrast, treatment with 5 mM buthionine sulfoximine reduced total GSH by 90% and sensitized the cells to cisplatin cytotoxicity. Furthermore, extracellular reaction of GSH with cisplatin prior to treating HA1 cells reduced the toxicity of the compound, indicating that this reaction is capable of participating in the detoxification of cisplatin. These results indicate that cellular adaptation to oxidative stress renders cells resistant to DNA damage as well as to cytotoxicity associated with cisplatin treatment. Furthermore, increases in total GSH content (but not catalase activity) appear to partially account for cisplatin resistance demonstrated by H2O2-resistant cells.

Establishment of a hydrogen peroxide resistant variant of renal tubular epithelial cells: role of calcium-independent phospholipase A2 in cell damage.

Renal epithelial cells resistant to oxidant stress mediated by hydrogen peroxide have been isolated and characterized. African green monkey kidney epithelial cell line, BSC-1 cells, chronically exposed to 50 microM hydrogen peroxide for 15 passages exhibited increased catalase (1.5-fold) and glutathione peroxidase (2.4-fold) activity, as well as increased total cellular glutathione (1.6-fold). This was associated with the acquisition of resistance to hydrogen peroxide cytotoxicity, as judged by nuclear staining with ethidium homodimer and clonogenic survival assay. H2O2-adapted and wild-type BSC-1 cells were used to examine the role of elevated cytosolic calcium concentration and the activation of phospholipase A2 in the development of lethal cell injury. Despite dramatic differences in resistance to oxidative stress, both cell types showed similar kinetics of cytosolic calcium increase in response to challenge with hydrogen peroxide. In contrast to this, oxidant-induced release of arachidonic acid correlated with the resistance of both types of BSC-1 cells to oxidative stress. A mechanism-based inhibitor of calcium-independent phospholipase A2 (Hazen et al., J. Biol. Chem. 266, 7227, 1991) reduced oxidant-induced lethal cell injury, suggesting that this class of phospholipases contributes to damage of BSC-1 cells exposed to hydrogen peroxide. H2O2-adapted BSC-1 cells may represent a valuable tool to study adaptation to oxidative stress and various mechanisms of cell injury.

Differences in innovation-related characteristics between singleskilled and multiskilled health practitioner educational programs.

A survey of educational programs preparing single- and multiskilled health practitioners was conducted to determine the predictive power of organizational formalization and centralization and individual program director cosmopolitanism (ie, orientation to their larger profession as opposed to local environment) upon readiness to innovate for the educational units. Questionnaires were sent to 45 directors of programs preparing multiskilled health practitioners and 49 directors of programs preparing singleskilled health practitioners. Results indicated no significant differences between program directors of single- and multiskilled programs in their perceptions of the readiness to innovate for their educational units. Regression analysis indicated that a decentralized organization was a predictor of an educational unit's readiness to innovate. This finding held for both single- and multiskilled programs. Although cosmopolitanism was not a significant predictor of readiness to innovate, program directors at four-year institutions had a significantly higher level of that characteristic than directors at two-year institutions. The results of this study, although limited by individual and not multiple responses from each institution, do not indicate any differences in the ability of organizational formalization and centralization to predict the innovative characteristics of single- and multiskilled allied health programs. Additionally, the cosmopolitanism of program directors also does not predict these same characteristics.

Mechanisms of cellular resistance to hydrogen peroxide, hyperoxia, and 4-hydroxy-2-nonenal toxicity: the significance of increased catalase activity in H2O2-resistant fibroblasts.

An H2O2-resistant variant (OC14) of the HA1 Chinese hamster fibroblast cell line which demonstrates a 20-fold increase in catalase activity was utilized in the study of mechanisms responsible for cellular resistance to hydrogen peroxide, oxygen, and 4-hydroxy-2-nonenal toxicity. HA1 and OC14 cells were treated with 9 mM aminotriazole which resulted in a 60 to 80% reduction in catalase activity. Pretreatment with aminotriazole resulted in significant sensitization to the toxicity of 1-h exposures to exogenously applied H2O2, which was proportional to the reduction in catalase activity. Treatment with aminotriazole produced significant sensitization to the toxicity of 95% O2 after 45 h of O2 exposure but no sensitization to the toxicity of a 1-h exposure to 50 microM 4-hydroxy-2-nonenal. Inhibition of catalase activity by aminotriazole had no effect on the metabolism of 4-hydroxy-2-nonenal by either cell line tested. These results support the conclusion that in H2O2-resistant cells, catalase activity is a major determinant of cellular resistance to H2O2 toxicity, whereas catalase activity has a limited role in cellular resistance to an acute exposure to 95% O2 and is unrelated to cellular resistance to 4-hydroxy-2-nonenal.