Sazetidine-A has been recently proposed to be a "silent desensitizer" of ␣4␤2 nicotinic acetylcholine receptors (nAChRs), implying that it desensitizes ␣4␤2 nAChRs without first activating them. This unusual pharmacological property of sazetidine-A makes it, potentially, an excellent research tool to distinguish between the role of activation and desensitization of ␣4␤2 nAChRs in mediating the central nervous system effects of nicotine itself, as well as those of new nicotinic drugs. We were surprised to find that sazetidine-A potently and efficaciously stimulated nAChR-mediated dopamine release from rat striatal slices, which is mediated by ␣4␤2* and ␣6␤2* subtypes of nAChR. The agonist effects on native striatal nAChRs prompted us to re-examine the effects of sazetidine-A on recombinant ␣4␤2 nAChRs in more detail. We expressed the two alternative stoichiometries of ␣4␤2 nAChR in Xenopus laevis oocytes and investigated the agonist properties of sazetidine-A on both ␣4(2)␤2(3)and ␣4(3)␤2(2) nAChRs. We found that sazetidine-A potently activated both stoichiometries of ␣4␤2 nAChR: it was a full agonist on ␣4(2)␤2(3) nAChRs, whereas it had an efficacy of only 6% on ␣4(3)␤2(2) nAChRs. In contrast to what has been published before, we therefore conclude that sazetidine-A is an agonist of native and recombinant ␣4␤2 nAChRs but shows differential efficacy on ␣4␤2 nAChRs subtypes.

Pharmacological manipulation of specific neural circuits to optimize therapeutic index is an unrealized goal in neurology and psychiatry. AMPA receptors are important for excitatory synaptic transmission, and their antagonists are antiepileptic. Although efficacious, AMPA-receptor antagonists, including perampanel (Fycompa), the only approved antagonist for epilepsy, induce dizziness and motor impairment. We hypothesized that blockade of forebrain AMPA receptors without blocking cerebellar AMPA receptors would be antiepileptic and devoid of motor impairment. Taking advantage of an AMPA receptor auxiliary protein, TARP γ-8, which is selectively expressed in the forebrain and modulates the pharmacological properties of AMPA receptors, we discovered that LY3130481 selectively antagonized recombinant and native AMPA receptors containing γ-8, but not γ-2 (cerebellum) or other TARP members. Two amino acid residues unique to γ-8 determined this selectivity. We also observed antagonism of AMPA receptors expressed in hippocampal, but not cerebellar, tissue from an patient with epilepsy. Corresponding to this selective activity, LY3130481 prevented multiple seizure types in rats and mice and without motor side effects. These findings demonstrate the first rationally discovered molecule targeting specific neural circuitries for therapeutic advantage.

Regulatory CD4+CD25+ T cells (Tregs) suppress autoimmune and inflammatory diseases through mechanisms that are only partly understood. Previous studies suggest that Tregs can suppress bacterially triggered intestinal inflammation and respond to LPS through TLRs with enhanced suppressive activity. In this study, we have used murine cecal ligation and puncture as a model of polymicrobial sepsis to explore the effects of adoptive transfer of Tregs on septic outcome. Adoptive transfer of in vitro-stimulated Tregs in both prevention and therapeutic modes significantly improved survival of cecal ligation and puncture mice. Furthermore, the effect was dependent on both the number of Tregs adoptively transferred and the presence of host T cells. Animals that received stimulated Tregs had significantly increased peritoneal mast cells and peritoneal TNF-α production. More importantly, adoptive transfer of in vitro-stimulated Tregs significantly improved bacterial clearance, which resulted in improved survival. Our results suggest a novel role for Tregs in sepsis.

Transmembrane AMPA receptor regulatory proteins (TARPs) are a family of scaffolding proteins that regulate AMPA receptor trafficking and function. TARP γ-8 is one member of this family and is highly expressed within the hippocampus relative to the cerebellum. A selective TARP γ-8-dependent AMPA receptor antagonist (TDAA) is an innovative approach to modulate AMPA receptors in specific brain regions to potentially increase the therapeutic index relative to known non-TARP-dependent AMPA antagonists. We describe here, for the first time, the discovery of a noncompetitive AMPA receptor antagonist that is dependent on the presence of TARP γ-8. Three major iteration cycles were employed to improve upon potency, CYP1A2-dependent challenges, and in vivo clearance. An optimized molecule, compound (-)-25 (LY3130481), was fully protective against pentylenetetrazole-induced convulsions in rats without the motor impairment associated with non-TARP-dependent AMPA receptor antagonists. Compound (-)-25 could be utilized to provide proof of concept for antiepileptic efficacy with reduced motor side effects in patients.

The data from this study indicate that an early decrease in protein C concentration predicts poor outcome in a rat sepsis model. The data further indicate that increases in the CXC chemokines macrophage inflammatory protein-2 and KC precede poor outcome.