A predictive in vitro model of the impact of drugs with anticholinergic properties on human neuronal and astrocytic systems

The link between off-target anticholinergic effects of medications and acute cognitive impairment in older adults requires urgent investigation. We aimed to determine whether a relevant in vitro model may aid the identification of anticholinergic responses to drugs and the prediction of anticholinergic risk during polypharmacy. In this preliminary study we employed a co-culture of human-derived neurons and astrocytes (NT2.N/A) derived from the NT2 cell line. NT2.N/A cells possess much of the functionality of mature neurons and astrocytes, key cholinergic phenotypic markers and muscarinic acetylcholine receptors (mAChRs). The cholinergic response of NT2 astrocytes to the mAChR agonist oxotremorine was examined using the fluorescent dye fluo-4 to quantitate increases in intracellular calcium [Ca2+]i. Inhibition of this response by drugs classified as severe (dicycloverine, amitriptyline), moderate (cyclobenzaprine) and possible (cimetidine) on the Anticholinergic Cognitive Burden (ACB) scale, was examined after exposure to individual and pairs of compounds. Individually, dicycloverine had the most significant effect regarding inhibition of the astrocytic cholinergic response to oxotremorine, followed by amitriptyline then cyclobenzaprine and cimetidine, in agreement with the ACB scale. In combination, dicycloverine with cyclobenzaprine had the most significant effect, followed by dicycloverine with amitriptyline. The order of potency of the drugs in combination frequently disagreed with predicted ACB scores derived from summation of the individual drug scores, suggesting current scales may underestimate the effect of polypharmacy. Overall, this NT2.N/A model may be appropriate for further investigation of adverse anticholinergic effects of multiple medications, in order to inform clinical choices of suitable drug use in the elderly

Pinning the Blame in Cyberspace: Towards a Coherent Theory for Imposing Vicarious Copyright, Trademark and Tort Liability for Conduct Ocurring over the Internet

Vicarious liability, or the principle that under certain circumstances it is fair and just to hold unrelated third parties liable for conduct which they did not initiate or perhaps even condone, is both a logical outgrowth of, and impediment to, the ongoing rapid expansion of the Internet. Online infringers and tortfeasors may be more likely than others to be effectively judgment proof, because their conduct was undertaken anonymously, they cannot satisfy a damages award, or they are located beyond the jurisdiction of a convenient and economical U.S. venue for litigation. As a consequence, and as Internet use has increased and the Internet has made detection of small scale fraud easier, a natural pressure has been building to impose vicarious liability on more financially solvent defendants amenable to suit closer to home; namely Internet and other online service providers. Holding Internet providers liable for conduct which they cannot reasonably prevent, however, is irrational, and could discourage companies from providing Internet service. This Article analyzes the current standards for imposing direct, contributory, and vicarious copyright, trademark, and tort liability on online providers. The author concludes that, in light of the standard protocols under which data is transmitted over the Internet, it is unreasonable to impose indirect liability on online providers in cases where they did not know of or condone third party torts or acts of infringement. Specifically, the author challenges the notion, first advanced by the Clinton Administration in its National Information Infrastructure White Paper, that online providers may be held liable for vicarious copyright infringement under the so-called dance hall cases. The author also argues that uncertainty surrounding the scope of vicarious liability in Cyberspace stems from the wooden application of case law developed on terra firma to the world of Cyberspace. Among other things, the Article suggests that because of unique attributes of the online world, time, rather than content, must be given greater weight in analyzing misconduct and copyright fair use in Cyberspace