The presence of chloride-formate anion exchange in vascular smooth muscle cells (VSMCs) and cardiac myocytes was investigated. Imposing an outward chloride gradient in sarcolemmal microsomes isolated from canine aorta stimulated [14C] formate uptake compared with the absence of a chloride gradient (24.3±2.33 versus 9.8±1.41 pmol/mg protein for 30 seconds, P<.03) and induced transient uphill [14C] formate uptake. The chloride-formate exchange was significantly inhibited in the presence of 1 mmol/L 4,4′-diisothiocyanostilbene-2,2′-disulfonic acid (DIDS) or furosemide (57% and 61%, respectively). Incubation of rat cultured VSMCs in a medium containing [14C] formate resulted in uptake of formate that was significantly DIDS and furosemide sensitive (79.34 ±2.47, 43.03±2.37, and 44.65±1.68 pmol/mg protein for 4 minutes in control, DIDS, and furosemide groups, respectively). Preincubation of the VSMCs in chloride-free medium significantly reduced the DIDS-sensitive (36.31 versus 16.85 pmol/mg protein for 4 minutes, P< .001) and furosemide-sensitive (34.72 versus 8.78 pmol/mg protein for 4 minutes, P<.001) [14C] formate uptake. These results are compatible with the presence of chloride-formate exchange in VSMCs. Influx of [14C] formate into sarcolemmal vesicles isolated from canine heart was significantly higher in the presence of an outward chloride gradient than in its absence (18.1 ±2.3 versus 9.6±1.7 pmol/mg protein for 30 seconds, P<.03). The chloride-formate exchange was significantly inhibited in the presence of 1 mmol/L DIDS or furosemide (41% and 52%, respectively). We conclude that the distribution of chloride-formate exchange may be more universal than previously suggested. The physiological significance of this anion exchanger remains to be determined.