Functional Study of Chloride Currents of the Rat Carotid Body

SUMMARY Hipoxia, hipercapnia and pH are known to activate the chemoreceptor cell of the carotid body by inducing membrane depolarisation. It is widely accepted that stimulus-induced depolarisation is due to the inhibition of K + currents. Besides these kation currents that undoubtfully play significant role in the chemotransduction process, chemoreceptor cells also possess anion currents, which functions are hardly known. We aimed to study the functional role of the anion currents of rat chemoreceptor cell: the swelling-activated Cl – current and also the ClC-2-like Cl – current which latter one was previously characterised in the chemoreceptor cell by our group. Experiments were performed on primary cultures of chemoreceptor cells isolated from rats. Ionic currents were monitored by patch-clamp technique using the whole-cell mode, while of cytoplasmic Ca 2+ concentration ([Ca 2+ ] c) and pH (pH c) were detected by fluorescent technique with Indo-1 and carboxy-SNARF, respectively. In the first set of experiment we verified by patch-clamp measurements that hyposmotic challenge activates the swelling-activated Cl – current in our cell culture, as well. In the subsequent fluorescent measurements we examined the effect of hyposmotically induced cell-swelling on resting [Ca 2+ ] c. Decreasing osmolality by 50 mosmol/kg caused elevation of [Ca 2+ ] c in CO 2 /HCO 3 – buffered media, and more moderate decreases in osmolality (25 and 15 mosmol/kg) were capable to induce Ca 2+ response. By using Ca 2+-free medium and nifedipine, a blocker of the L-type Ca 2+ channel, we proved that hiposmosis-induced Ca 2+ response is due to the opening of high-voltage activated Ca 2+ channels that are activated by the swelling-induced depolarisation. This depolarisation is mediated by the swelling-activated Cl – current indeed, since niflumic acid, an inhibitor of the current, abolished the hyposmosis-induced Ca 2+ response. The cellular mechanism that is likely to underlie cytoplasmic Cl – accumulation which accounts for the Cl –-mediated depolarisation was tested by the withdrawal of CO 2 /HCO 3 – buffer. In our further investigations on the function the other, ClC-2-like Cl − current, we found that inhibition of current, induced slow, reversible cytoplasmic alkalinisation. Our results demonstrate that activation of the swelling-activated Cl – current results in the activation of the cell and this current underlies the osmotic sensitivity of the chemoreceptor cell. The phenomenon of Cl − mediated depolarisation was reported by us and this finding re-evaluates the significance of Cl – currents in …