Up-regulation of M1 muscarinic receptors expressed in CHOm1 cells by panaxynol via cAMP pathway

Loss of cholinergic neurons along with muscarinic acetylcholine receptors (mAChRs) in cerebral cortex and hippocampus is closely associated with Alzheimer's disease (AD). Recent drug development for AD treatment focuses heavily on identifying M1 receptor agonists. However, mAChRs undergo down-regulation in response to agonist-induced sustained activation. Therefore, therapeutic effectiveness wanes during continuous use. Thus, another potentially effective approach, which overcomes this drawback is to develop compounds, which instead up-regulate M1 receptor expression. In the present study, we took this alternative approach and contrasted in Chinese hamster ovary cells transfected with human m1 subtype gene (CHOm1 cells) changes of M1 receptor expression levels caused by muscarinic agonists and upregulators of its expression. The muscarinic agonists carbachol and pilocarpine reduced M1 receptor number in CHOm1 cells by 29 and 46%, respectively, at 100 μM, whereas panaxynol, a polyacetylene compound isolated from the lipophilic fraction of Panax notoginseng, concentration-dependently up-regulated the M1 receptor number after pre-incubation with CHOm1 cells for 48 h, reaching a plateau at 1 μM, and was accompanied by enhanced M1 mRNA levels. Moreover, the protein kinase A (PKA) inhibitor RP-adenosine-3′,5′-cyclic mono-phosphoro-thioate triethylamine salt (RP-cAMPs) 5 μM completely prevented panaxynol-induced up-regulation of M1 receptors. Panaxynol (1 μM) caused a significant and consistent stimulation of cAMP accumulation (27% increase above basal at 40 min). These results suggest that in CHOm1 cells panaxynol up-regulates M1 receptor number through cAMP pathway-mediated stimulation of gene transcription.