In this study, human platelets were used as a cellular model for exploring cytosolic free Ca (Ca i ) regulation in non-insulin-dependent diabetes mellitus (NIDDM). Ca i levels were monitored in resting and thrombin-stimulated platelets from obese females with NIDDM; obese, nondiabetic women, and nonobese, nondiabetic women. All subjects were black. Significant and marked elevation of basal Ca i levels was observed in platelets from the diabetic subjects when no aspirin was used during platelet isolation. However, no significant differences were observed in Ca i between aspirin-treated platelets from women with NIDDM and platelets from nondiabetic women. The rate of the Ca i return to basal level after thrombin stimulation was significantly lower in platelets from the diabetic subjects, suggesting an abnormality in platelet Ca extrusion or sequestration in NIDDM. Platelet Ca i levels positively correlated with low-density lipoprotein cholesterol/high-density lipoprotein cholesterol ratio (LDL) and fasting blood glucose. These findings suggest abnormalities in platelet Ca i homeostasis in NIDDM that are influenced by the serum lipid profile and perhaps glucose.