CO2 chemosensing is a vital function for the maintenance of life that helps to control acid–base balance. Most studies have reported that CO2 is measured via its proxy, pH. Here we report an inwardly rectifying channel, in outside-out excised patches from HeLa cells that was sensitive to modest changes in PCO2 under conditions of constant extracellular pH. As PCO2 increased, the open probability of the channel increased. The single-channel currents had a conductance of 6.7 pS and a reversal potential of –70 mV, which lay between the K+ and Cl– equilibrium potentials. This reversal potential was shifted by +61 mV following a tenfold increase in extracellular [K+] but was insensitive to variations of extracellular [Cl–]. The single-channel conductance increased with extracellular [K+]. We propose that this channel is a member of the Kir family. In addition to this K+ channel, we found that many of the excised patches also contained a conductance carried via a Cl–-selective channel. This CO2-sensitive Kir channel may hyperpolarize excitable cells and provides a potential mechanism for CO2-dependent inhibition during hypercapnia.