Cyclic dinucleotides are important messengers for bacteria and protozoa and are well-characterized immunity alarmins for infected mammalian cells through intracellular binding to STING receptors. We sought to investigate their unknown extracellular effects by adding cyclic dinucleotides to the culture medium of freshly isolated human blood cells in vitro. Here we report that adenosine-containing cyclic dinucleotides induce the selective apoptosis of monocytes through a novel apoptotic pathway. We demonstrate that these compounds are inverse agonist ligands of A2a, a G ␣s -coupled adenosine receptor selectively expressed by monocytes. Inhibition of monocyte A2a by these ligands induces apoptosis through a mechanism independent of that of the STING receptors. The blockade of basal (adenosine-free) signaling from A2a inhibits protein kinase A (PKA) activity, thereby recruiting cytosolic p53, which opens the mitochondrial permeability transition pore and impairs mitochondrial respiration, resulting in apoptosis. A2a antagonists and inverse agonist ligands induce apoptosis of human monocytes, while A2a agonists are antiapoptotic. In vivo, we used a mock developing human hematopoietic system through NSG mice transplanted with human CD34؉ cells. Treatment with cyclic di-AMP selectively depleted A2a-expressing monocytes and their precursors via apoptosis. Thus, monocyte recognition of cyclic dinucleotides unravels a novel proapoptotic pathway: the A2a G ␣s protein-coupled receptor (GPCR)-driven tonic inhibitory signaling of mitochondrion-induced cell death. P urine and pyrimidine-based signaling is a fundamental and conserved mode of intercellular communication. In the body, mononucleotides are the major mediators of tissue protection and regeneration, for example, adenosine, which is released from damaged cells. In contrast, microorganisms use cyclic dinucleotides, such as cyclic di-AMP (c-di-AMP) and cyclic di-GMP (cdi-GMP), which are secreted by bacteria and protozoa, respectively (1, 2). As a defense mechanism, mammalian cells have evolved a means to sense cyclic dinucleotides. When infected with retroviruses (3) or carrying cytosolic DNA (4), cells assemble an endogenous 2=,3= cyclic GMP-AMP (cGAMP) dinucleotide which is recognized by the intracellular protein STING to induce type I interferon (IFN) responses (5). In this way, intracellular cyclic dinucleotides represent important alarmins in immunity.Extracellular cyclic dinucleotides are released from infected dying cells or damaged tissues and also represent important signals, but whether and how mammalian cells can detect these and respond is presently unknown. We hypothesized that human peripheral blood cells might be capable of detecting extracellular cyclic dinucleotides. We found that the extracellular 3=,5= cyclic dinucleotides c-di-AMP and cGAMP are specifically recognized by human monocytes expressing the A2a adenosine receptor and can selectively induce their apoptosis. Analysis of the role of A2a in this apoptotic response showed that it controls the s...