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Shao X.M.,University of California at Los Angeles | Xu B.,University of California at Los Angeles | Xu B.,Tongji University | Liang J.,University of California at Los Angeles | And 3 more authors.
Nicotine and Tobacco Research | Year: 2013

Introduction: Nicotine is a heavily used addictive drug acquired through smoking tobacco. Nicotine in cigarette smoke is deposited and absorbed in the lungs, which results in a rapidly peaked slowly declining arterial concentration. This pattern plays an important role in initiation of nicotine addiction. Methods: A method and device were developed for delivering nicotine to rodents with lung alveolar region-targeted aerosol technology. The dose of delivery can be controlled by the nicotine aerosol concentration and duration of exposure. Results: Our data showed that, in the breathing zone of the nose-only exposure chamber, the aerosol droplet size distribution was within the respirable diameter range. Rats were exposed to nicotine aerosol for 2 min. The arterial blood nicotine concentration reached 43.2 ± 15.7 ng/ml (mean ± SD) within 1-4 min and declined over the next 20 min, closely resembling the magnitude and early pharmacokinetics of a human smoking a cigarette. The acute inhalation toxicity of nicotine: LC50 = 2.3 mg/L was determined; it was affected by pH, suggesting that acidification decreases nicotine absorption and/or bioavailability. Conclusions: A noninvasive method and toolkit were developed for delivering nicotine to rodents that enable rapid delivery of a controllable amount of nicotine into the systemic circulation and brain-inducing dose-dependent pharmacological effects, even a lethal dose. Aerosol inhalation can produce nicotine kinetics in both arterial and venous blood resembling human smoking. This method can be applied to studies of the effects of chronic intermittent nicotine exposure, nicotine addiction, toxicology, tobacco-related diseases, teratogenicity, and for discovery of pharmacological therapeutics. © The Author 2012. Published by Oxford University Press on behalf of the Society for Research on Nicotine and Tobacco. All rights reserved. Source

Toll L.,SRI International | Zaveri N.T.,Astraea Therapeutics, Llc | Polgar W.E.,SRI International | Jiang F.,SRI International | And 7 more authors.
Neuropsychopharmacology | Year: 2012

Genomic and pharmacologic data have suggested the involvement of the α3Β4 subtype of nicotinic acetylcholine receptors (nAChRs) in drug seeking to nicotine and other drugs of abuse. In order to better examine this receptor subtype, we have identified and characterized the first high affinity and selective α3Β4 nAChR antagonist, AT-1001, both in vitro and in vivo. This is the first reported compound with a Ki below 10 nM at α3Β4 nAChR and 90-fold selectivity over the other major subtypes, the α4Β2 and α7 nAChR. AT-1001 competes with epibatidine, allowing for 3 Hepibatidine binding to be used for structure-activity studies, however, both receptor binding and ligand-induced Ca 2 flux are not strictly competitive because increasing ligand concentration produces an apparent decrease in receptor number and maximal Ca 2 fluorescence. AT-1001 also potently and reversibly blocks epibatidine-induced inward currents in HEK cells transfected with α3Β4 nAChR. Importantly, AT-1001 potently and dose-dependently blocks nicotine self-administration in rats, without affecting food responding. When tested in a nucleus accumbens (NAcs) synaptosomal preparation, AT-1001 inhibits nicotine-induced 3 Hdopamine release poorly and at significantly higher concentrations compared with mecamylamine and conotoxin MII. These results suggest that its inhibition of nicotine self-administration in rats is not directly due to a decrease in dopamine release from the NAc, and most likely involves an indirect pathway requiring α3Β4 nAChR. In conclusion, our studies provide further evidence for the involvement of α3Β4 nAChR in nicotine self-administration. These findings suggest the utility of this receptor as a target for smoking cessation medications, and highlight the potential of AT-1001 and congeners as clinically useful compounds. © 2012 American College of Neuropsychopharmacology. All rights reserved. Source

Kulkarni S.,Stanford University | Zou B.,Stanford University | Zou B.,AfaSci Research Laboratory | Hanson J.,AfaSci Research Laboratory | And 8 more authors.
American Journal of Physiology - Gastrointestinal and Liver Physiology | Year: 2011

Recent studies have explored the potential of central nervous system-derived neural stem cells (CNSNSC) to repopulate the enteric nervous system. However, the exact phenotypic fate of gut-transplanted CNS-NSC has not been characterized. The aim of this study was to investigate the effect of the gut microenvironment on phenotypic fate of CNS-NSC in vitro. With the use of Transwell culture, differentiation of mouse embryonic CNS-NSC was studied when cocultured without direct contact with mouse intestinal longitudinal muscle-myenteric plexus preparations (LM-MP) compared with control noncocultured cells, in a differentiating medium. Differentiated cells were analyzed by immunocytochemistry and quantitative RT-PCR to assess the expression of specific markers and by whole cell patch-clamp studies for functional characterization of their phenotype. We found that LM-MP cocultured cells had a significant increase in the numbers of cells that were immune reactive against the panneuronal marker β-tubulin, neurotransmitters neuronal nitric oxide synthase (nNOS), choline acetyltransferase (ChAT), and neuropeptide vasoactive intestinal peptide (VIP) and showed an increase in expression of these genes, compared with control cells. Whole cell patch-clamp analysis showed that coculture with LM-MP decreases cell excitability and reduces voltage-gated Na + currents but significantly enhances A-current and late afterhyperpolarization (AHP) and increases the expression of the four AHP-generating Ca2 +-dependent K + channel genes (KCNN), compared with control cells. In a separate experiment, differentiation of LM-MP cocultured CNS-NSC produced a significant increase in the numbers of cells that were immune reactive against the neurotransmitters nNOS, ChAT, and the neuropeptide VIP compared with CNS-NSC differentiated similarly in the presence of neonatal brain tissue. Our results show that the gut microenvironment induces CNS-NSC to produce neurons that share some of the characteristics of classical enteric neurons, further supporting the therapeutic use of these cells for gastrointestinal disorders. © 2011 the American Physiological Society. Source

Villeda S.A.,University of California at San Francisco | Villeda S.A.,Eli and Edythe Broad Center for Regeneration Medicine and Stem Cell Research | Villeda S.A.,Stanford University | Plambeck K.E.,University of California at San Francisco | And 24 more authors.
Nature Medicine | Year: 2014

As human lifespan increases, a greater fraction of the population is suffering from age-related cognitive impairments, making it important to elucidate a means to combat the effects of aging. Here we report that exposure of an aged animal to young blood can counteract and reverse pre-existing effects of brain aging at the molecular, structural, functional and cognitive level. Genome-wide microarray analysis of heterochronic parabionts - in which circulatory systems of young and aged animals are connected - identified synaptic plasticity-related transcriptional changes in the hippocampus of aged mice. Dendritic spine density of mature neurons increased and synaptic plasticity improved in the hippocampus of aged heterochronic parabionts. At the cognitive level, systemic administration of young blood plasma into aged mice improved age-related cognitive impairments in both contextual fear conditioning and spatial learning and memory. Structural and cognitive enhancements elicited by exposure to young blood are mediated, in part, by activation of the cyclic AMP response element binding protein (Creb) in the aged hippocampus. Our data indicate that exposure of aged mice to young blood late in life is capable of rejuvenating synaptic plasticity and improving cognitive function. © 2014 Nature America, Inc. All rights reserved. Source

Kaufmann K.,Vanderbilt University | Romaine I.,Vanderbilt University | Days E.,Vanderbilt University | Pascual C.,AfaSci Research Laboratory | And 13 more authors.
ACS Chemical Neuroscience | Year: 2013

The G-protein activated, inward-rectifying potassium (K+) channels, "GIRKs", are a family of ion channels (Kir3.1- Kir3.4) that has been the focus of intense research interest for nearly two decades. GIRKs are comprised of various homo- and heterotetrameric combinations of four different subunits. These subunits are expressed in different combinations in a variety of regions throughout the central nervous system and in the periphery. The body of GIRK research implicates GIRK in processes as diverse as controlling heart rhythm, to effects on reward/addiction, to modulation of response to analgesics. Despite years of GIRK research, very few tools exist to selectively modulate GIRK channels' activity and until now no tools existed that potently and selectively activated GIRKs. Here we report the development and characterization of the first truly potent, effective, and selective GIRK activator, ML297 (VU0456810). We further demonstrate that ML297 is active in two in vivo models of epilepsy, a disease where up to 40% of patients remain with symptoms refractory to present treatments. The development of ML297 represents a truly significant advancement in our ability to selectively probe GIRK's role in physiology as well as providing the first tool for beginning to understand GIRK's potential as a target for a diversity of therapeutic indications. © 2013 American Chemical Society. Source

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