Detection of multiple H(3) receptor affinity states utilizing [(3)H]A-349821, a novel, selective, non-imidazole histamine H(3) receptor inverse agonist radioligand

1. A-349821 is a selective histamine H(3) receptor antagonist/inverse agonist. Herein, binding of the novel non-imidazole H(3) receptor radioligand [(3)H]A-349821 to membranes expressing native or recombinant H(3) receptors from rat or human sources was characterized and compared with the binding of the agonist [(3)H]N-α-methylhistamine ([(3)H]NαMH). 2. [(3)H]A-349821 bound with high affinity and specificity to an apparent single class of saturable sites and recognized human H(3) receptors with 10-fold higher affinity compared to rat H(3) receptors. [(3)H]A-349821 detected larger populations of receptors compared to [(3)H]NαMH. 3. Displacement of [(3)H]A-349821 binding by H(3) receptor antagonists/inverse agonists was monophasic, suggesting recognition of a single binding site, while that of H(3) receptor agonists was biphasic, suggesting recognition of both high- and low-affinity H(3) receptor sites. 4. pK(i) values of high-affinity binding sites for H(3) receptor competitors utilizing [(3)H]A-349821 were highly correlated with pK(i) values obtained with [(3)H]NαMH, consistent with labelling of H(3) receptors by [(3)H]A-349821. 5. Unlike assays utilizing [(3)H]NαMH, addition of GDP had no effect on saturation parameters measured with [(3)H]A-349821, while displacement of [(3)H]A-349821 binding by the H(3) receptor agonist histamine was sensitive to GDP. 6. In conclusion, [(3)H]A-349821 labels interconvertible high- and low-affinity states of the H(3) receptor, and displays improved selectivity over imidazole-containing H(3) receptor antagonist radioligands. [(3)H]A-349821 competition studies showed significant differences in the proportions and potencies of high- and low-affinity sites across species, providing new information about the fundamental pharmacological nature of H(3) receptors

G protein-dependent pharmacology of histamine H3 receptor ligands: Evidence for heterogeneous active state receptor conformations

ABSTRACT Previously reported pharmacological studies using the imidazole-containing histamine H 3 receptor ligands GT-2331 (Cipralisant) and proxyfan resulted in a range of classifications (antagonist, agonist, and protean) for these compounds. We examined the role that the signaling system, with particular emphasis on the type of G protein, had on the pharmacology observed for H 3 ligands. Ligands were assessed using assays measuring neurotransmitter release, cAMP, and guanosine 5Ј- binding. Whereas clobenpropit and ciproxifan were consistently antagonists, GT-2331, proxyfan, and imetit exhibited differential activity. Although GT-2331 and proxyfan exhibited little agonist activity in neurotransmitter release assays, both demonstrated full agonism relative to (R)-␣-methylhistamine in cAMP assays. In [ 35 S]GTP␥S binding assays, GT-2331 and proxyfan demonstrated partial agonism. Imetit showed full agonism in most assays, but it was slightly less efficacious in a neurotransmitter release assay and in [ 35 S]GTP␥S binding at the human H 3 receptor. To further examine these ligands, we coexpressed G␣16 or chimeric G␣q/i 5 in human embryonic kidney cells expressing the human H 3 receptor and assayed intracellular calcium and cAMP levels. GT-2331, proxyfan, and imetit demonstrated full agonism in all assays of cAMP activity. However, in cells expressing G␣16, they exhibited minimal agonism in calcium mobilization assays, whereas imetit showed partial agonism. When G␣q/i 5 was used, the activity of both GT-2331 and proxyfan increased, whereas imetit became a full agonist. These results demonstrate that GT-2331 and proxyfan's differential pharmacology at the H 3 receptor depends on the type of G protein used and provide indirect evidence for differential ligand-bound active states that mediate signaling by the H 3 receptor

α7 nicotinic receptor agonist enhances cognition in aged 3xTg-AD mice with robust plaques and tangles

Alzheimer disease (AD) is a progressive neurodegenerative disorder with associated memory loss, spatial disorientation, and other psychiatric problems. Cholinergic system dysfunction is an early and salient feature of AD, and enhancing cholinergic signaling with acetylcholinesterase inhibitors is currently the primary strategy for improving cognition. The beneficial effects of acetylcholinesterase inhibitors, however, are typically short-lived and accompanied by adverse effects. Recent evidence suggests that activating alpha(7) nicotinic acetylcholine receptors (alpha(7) nAChR) may facilitate the specific modulation of brain cholinergic signaling, leading to cognitive enhancement and possibly to amelioration of AD pathologic findings. In the present study, we determined the effect of long-term treatment with the selective alpha(7) nAChR agonist A-582941 in aged 3xTg-AD mice with robust AD-like pathology, which is particularly significant not only because this is the only mouse model that co-develops amyloid plaques and neurofibrillary tangles but also because it enabled us to explore whether A-582941 is able to restore brain function after the severe damage associated with AD. Analysis of beta-amyloid deposits, tau phosphorylation, and inflammatory cells revealed that, overall, pathologic findings were unchanged. Rather, alpha(7) nAChR activation induced expression of c-Fos and brain-derived neurotrophic factor and phosphorylation of cyclic adenosine monophosphate response element binding and neurotrophic tyrosine receptor kinase type 2. More important, A-582941 completely restored cognition in aged 3xTg-AD mice to the level of that in age-matched nontransgenic mice. These novel findings indicate that activating alpha(7) nAChR is a promising treatment for cognitive impairment in AD

Improving Electrodeposition of Mg through an Open Circuit Potential Hold

We used <i>in situ</i> X-ray diffraction, XPS, SEM, and electrochemical methods to interrogate the mechanism of Mg electrodeposition from PhMgCl/AlCl₃ (APC) and EtMgCl electrolytes. An open circuit potential (OCP) pause following Mg deposition led to retained enhancement of Mg deposition and stripping kinetics along with lowered overpotentials for both. <i>In situ</i> X-ray diffraction demonstrated that the OCP pause led to a more polycrystalline deposit relative to that without the pause, while SEM presented micrographs that showed smaller deposits with an OCP hold. The improvement is attributed to an “enhancement layer” that formed on the electrode during the OCP hold. Analysis of XPS data suggests that the “enhancement layer” consists of Mg and Cl retained on the electrode surface, possibly following electrode depassivation