Circulatory deficits are common and pathophysiologically relevant in type 2 diabetes mellitus (T2DM). Perturbed red blood cell (RBC) homeostasis and diminished nitric oxide (NO) availability contribute to endothelial dysfunction, a hallmark of cardiometabolic disorders; however, underlying pathophysiological mechanisms remain elusive. Here, we investigated RBC signaling pathways in a murine model of metabolic disease, focused on NO. T2DM-RBCs had elevated levels of cytosolic NO, intracellular calcium ions (Ca2+), and reactive oxygen species. Acute stimulation with exogenous insulin had no effect on NO content. Whereas insulin exposure caused Ca2+ entry into healthy RBCs, T2DM-RBCs were insensitive. Using RBCs isolated from human blood, we confirmed that insulin had no effect on RBC-NO, despite prompting Ca2+ uptake. Ca2+ uptake with insulin exposure was sensitive to inhibition of mechanosensitive ion channels, as well as Ca2+ chelation. Furthermore, co-incubation of RBCs with the piezo-type mechanosensitive ion channel component 1 (Piezo1) channel agonist Yoda1 and insulin did not produce compounded Ca2+ uptake, raising the possibility of crosstalk between insulin and Piezo1. The hyperinsulinemia associated with T2DM may exacerbate normal Piezo1-dependent Ca2+ uptake into RBCs, contributing to RBC dysfunction and circulatory complications in T2DM. The significance of RBC signaling in the pathophysiology of cardiometabolic disorders is still emerging. Individuals carrying mutations in the PIEZO1 gene exhibit hematological aberrations and hereditary anemia, supporting the importance of Piezo1 in RBC homeostasis. Furthermore, a shift in RBC-NO metabolism favoring nitrosative stress may contribute to circulatory complications observed in metabolic diseases such as T2DM. Collectively, the emerging relevance of RBC signaling pathways may provide novel avenues for targeted drug development.