Nitric oxide (NO) is a signaling molecule with diverse and often opposing biological effects. Its biological function is determined by its concentration, with lower levels promoting cell survival and proliferation, while higher levels favor cell cycle arrest, apoptosis, and senescence. NO's effects are influenced by its chemical reactions, such as binding to the regulatory heme in soluble guanylate cyclase (sGC), rather than traditional protein receptor interactions. NO can directly react with biological targets or indirectly generate reactive nitrogen species (RNS), which can cause oxidative and nitrosative stress. The balance between these reactions depends on NO concentration, with lower levels favoring nitrosation and higher levels favoring oxidation. NO is produced by different isoforms of nitric oxide synthase (NOS): constitutive (nNOS, eNOS) and inducible (iNOS). These isoforms differ in their regulation, with iNOS producing high concentrations of NO under inflammatory conditions. NO concentrations in vivo range from sub-nM to μM, with different biological effects depending on the concentration and duration of exposure. For example, low concentrations of NO (10–30 nM) promote cell proliferation, while higher concentrations (400 nM or more) can induce apoptosis or senescence. NO also interacts with reactive oxygen species (ROS), which can reduce NO concentrations and alter its signaling. NO's concentration is influenced by factors such as diffusion, consumption, and substrate availability. These factors, known as kinetic determinants, shape cellular responses to NO. NO's ability to diffuse rapidly allows it to reach different cellular compartments and interact with various targets. However, its concentration is also affected by the local environment, such as the presence of other molecules or the proximity to blood vessels. NO plays a critical role in various physiological processes, including vascular tone control, immune responses, and neuronal function. It can also contribute to pathological conditions, such as inflammation and tissue injury. The complex interplay between NO and other reactive species, such as ROS and RNS, highlights the importance of understanding NO's chemical biology in both health and disease. The concentration-dependent effects of NO underscore its versatility as a signaling molecule, with different biological outcomes depending on its concentration and the cellular context.Nitric oxide (NO) is a signaling molecule with diverse and often opposing biological effects. Its biological function is determined by its concentration, with lower levels promoting cell survival and proliferation, while higher levels favor cell cycle arrest, apoptosis, and senescence. NO's effects are influenced by its chemical reactions, such as binding to the regulatory heme in soluble guanylate cyclase (sGC), rather than traditional protein receptor interactions. NO can directly react with biological targets or indirectly generate reactive nitrogen species (RNS), which can cause oxidative and nitrosative stress. The balance between these reactions depends on NO concentration, with lower levels favoring nitrosation and higher levels favoring oxidation. NO is produced by different isoforms of nitric oxide synthase (NOS): constitutive (nNOS, eNOS) and inducible (iNOS). These isoforms differ in their regulation, with iNOS producing high concentrations of NO under inflammatory conditions. NO concentrations in vivo range from sub-nM to μM, with different biological effects depending on the concentration and duration of exposure. For example, low concentrations of NO (10–30 nM) promote cell proliferation, while higher concentrations (400 nM or more) can induce apoptosis or senescence. NO also interacts with reactive oxygen species (ROS), which can reduce NO concentrations and alter its signaling. NO's concentration is influenced by factors such as diffusion, consumption, and substrate availability. These factors, known as kinetic determinants, shape cellular responses to NO. NO's ability to diffuse rapidly allows it to reach different cellular compartments and interact with various targets. However, its concentration is also affected by the local environment, such as the presence of other molecules or the proximity to blood vessels. NO plays a critical role in various physiological processes, including vascular tone control, immune responses, and neuronal function. It can also contribute to pathological conditions, such as inflammation and tissue injury. The complex interplay between NO and other reactive species, such as ROS and RNS, highlights the importance of understanding NO's chemical biology in both health and disease. The concentration-dependent effects of NO underscore its versatility as a signaling molecule, with different biological outcomes depending on its concentration and the cellular context.