Muscarine exerts negative inotropic effects in human atria when cAMP is elevated by various agonists

The heart's force of contraction (FOC) is tightly regulated by various neurohormonal systems, including muscarinic receptors. While acetylcholine's direct and indirect negative inotropic effects are known, the specific actions of muscarine, a muscarinic receptor agonist, on human atrial contractility remain less clear, especially when other pathways are activated. Current understanding often focuses on direct receptor activation, but indirect effects, particularly those involving secondary messengers like cyclic adenosine monophosphate (cAMP), are critical for a complete picture of cardiac regulation. This study aimed to elucidate muscarine's indirect negative inotropic effects in the presence of various cAMP-increasing agents, addressing a gap in understanding complex cardiac signaling interactions.

Researchers obtained human atrial preparations (HAP) from patients undergoing open heart surgery for severe coronary heart disease. They first quantified muscarinic receptor mRNA expression using digital polymerase chain reaction (qPCR). HAP were then mounted in organ baths, electrically stimulated at 1 Hz, and FOC was measured under isometric conditions. The study assessed the effect of 1 µM muscarine alone, and then after pre-stimulation with various cAMP-increasing agents: 1 µM isoprenaline (β-adrenoceptor agonist), 1 µM serotonin (5-HT4-serotonin receptor agonist), 1 µM histamine (H2-histamine receptor agonist), 10 µM fenoldopam (D1-dopamine receptor agonist), 100 nM semaglutide (GLP-1R agonist), 100 nM gastric inhibitory peptide (GIPR agonist), 1 µM forskolin (adenylyl cyclase activator), 1 µM cilostamide (cAMP degradation inhibitor), or 100 µM dibutyryl-cAMP (cAMP-dependent protein kinase activator).

Analysis of muscarinic receptor mRNA in HAP revealed a predominant expression of the M2-muscarinic receptor, contributing 96.34% of total M1, M2, and M3 mRNA, with M1 at 0.09% and M3 at 3.57%. When applied alone, 1 µM muscarine transiently reduced FOC. Crucially, when 1 µM muscarine was additionally applied after pre-stimulation with various cAMP-increasing agents, a consistent reduction in FOC was observed. This negative inotropic effect occurred following pre-treatment with 1 µM isoprenaline, 1 µM serotonin, 1 µM histamine, 10 µM fenoldopam, 100 nM semaglutide, and 100 nM gastric inhibitory peptide.

The negative inotropic effect of 1 µM muscarine was also evident after directly increasing cAMP levels or activating cAMP-dependent protein kinase with 1 µM forskolin, 1 µM cilostamide, or 100 µM dibutyryl-cAMP.